Krüppel-like zinc-finger transcription factor 5 (KLF5) is highly expressed in large and giant unruptured cerebral aneurysms.

Krüppel-like zinc-finger transcription factor 5 (KLF5), known as BTEB2 and IKLF, has several biological functions that involve cell proliferation, development, and apoptosis. In human cerebral aneurysms, macrophage infiltration is profoundly associated with growth and rupture, but the role of KLF5 remains unclear. We examined the significance of KLF5 expression in cerebral aneurysms. Unruptured (n=15) and ruptured (n=12) aneurysms obtained at surgery or autopsy were divided into 3 size groups: small (<10 mm); large (10 mm but <25 mm); and giant (25 mm). Control samples comprised 5 cerebral arteries obtained from surgery or autopsy subjects. The expression of KLF5-, -smooth muscle actin-, and KP-1 (macrophages) -positive cells were counted and compared between groups. Media of control arteries was negative for KLF5. In the luminal layers, KLF5 in unruptured small aneurysm was also negative; KLF5 expression was higher in unruptured large/giant aneurysms than other groups (P<0.05). KP-1 expression in unruptured large/giant aneurysms, ruptured small aneurysms, and ruptured large/giant aneurysms was higher than in unruptured small aneurysms (P<0.05). In the unruptured large/giant aneurysms, KP-1-positive cells were lower than KLF5-positive cells. On the other hand, irrespective of size, KLF5 positivity tended to be lower than KP-1 in the luminal and abluminal layers of all ruptured aneurysms. This represents the first documentation that KLF5 is highly expressed in large and giant unruptured aneurysms and that in ruptured aneurysmal wall KLF5 expression was scarce. These findings suggest that the KLF5 expression and macrophage infiltration play essential roles on aneurysmal growth or rupture.


Analysis of expression and structure of the rat GH-secretagogue/ghrelin receptor (Ghsr) gene: Roles of epigenetic modifications in transcriptional regulation.

In the current study, to elucidate the molecular basis of cell type-specific expression of the GH-secretagogue/ghrelin receptor type 1A (GHSR1A), we characterized the structure and putative promoter region of the rat Ghsr gene. We identified an alternative 5'-untranslated first exon that contains multiple transcription start sites, and confirmed a 200-bp sequence proximal to this exon to be sufficient for basal promoter activity. A promoter-associated CpG island conserved across different species was found to be hypomethylated in Ghsr1a-expressing cell lines, while being heavily methylated in non-expressing cells. In cells with low or absent Ghsr1a expression, treatment with demethylating agents activated Ghsr1a transcription. Chromatin immunoprecipitation assays demonstrated Ghsr1a-expressing cells to display active histone modifications, whereas repressive modifications were present exclusively in other cell types. These results suggest epigenetic modifications at GHSR to play important roles in determining GHSR1A expression and abundance, and therefore the consequent sensitivity of cells to ghrelin.


DNA methyltransferases 1, 3a, and 3b overexpression and clinical significance in gastroenteropancreatic neuroendocrine tumors.

The alteration of DNA methylation is one of the most common epigenetic changes in human cancers. Three genes, namely, DNA methyltransferase 1, 3a, and 3b, which code for DNA methyltransferases that affect promoter methylation status, are thought to play an important role in the development of cancers and may be good anticancer therapy targets. The methylation of tumor suppressor genes has been reported in gastroenteropancreatic neuroendocrine tumors; however, there have been no studies about DNA methyltransferase protein expression and its clinical significance in gastroenteropancreatic neuroendocrine tumors. In this study, the expression of DNA methyltransferase 1, 3a, and 3b was studied in 63 gastroenteropancreatic neuroendocrine tumors by immunohistochemistry. The expression of DNA methyltransferase 1, 3a, and 3b was frequently detected in gastroenteropancreatic neuroendocrine tumors (87%, 81%, and 75%, respectively). The DNA methyltransferase 3a expression level was significantly higher in poorly differentiated neuroendocrine carcinomas than in well-differentiated neuroendocrine tumors or well-differentiated neuroendocrine carcinomas (P < .01 and P < .05, respectively). The expression of DNA methyltransferase 1, 3a, and 3b showed significantly higher levels in stage IV tumors than in stage I or II tumors. In addition, the expression levels of DNA methyltransferase 1, 3a, and 3b were positively correlated with the MIB-1 labeling index in gastroenteropancreatic neuroendocrine tumors (R = 0.293, P = .019; R = 0.457, P = .001; and R = 0.249, P = .049; respectively). In addition, the expression levels and positive immunostaining frequencies of DNA methyltransferase 3a and 3b were significantly lower in midgut neuroendocrine tumors than in foregut or hindgut neuroendocrine tumors. Our findings suggest that the overexpression of DNA methyltransferase 1, 3a, and 3b is related to tumorigenesis and the progression of gastroenteropancreatic neuroendocrine tumors.


Pituitary adenomas that show a faint GH-immunoreactivity but lack fibrous body: Pit-1 adenoma with endocrinologically low activity.

Growth hormone (GH)-producing pituitary adenomas have been classified into densely and sparsely granulated adenomas. The latter are chromophobic with weak GH-positivity and characteristically possess fibrous body (FB), aggregation of cytokeratin filaments. We report eight cases of unusual chromophobic adenomas. GH-immunoreactivity was detected in most adenoma cells in five cases and scattered in three cases. However, it appeared much weaker than that seen in ordinary GH-producing adenomas because of spotty immunoreactivity. Although intracytoplasmic organelles were well-developed, secretory granules were small and sparse. FB was not identified in any cases. Thyroid-stimulating hormone was positive in four cases. Pit-1 protein was positive in all eight cases. A weak labeling with GH probe was detected in two of two cases examined by in situ hybridization. Acromegalic features were evident in four cases, while mild or absent in four cases. GH levels were below 5 microg/l in four cases and 5-10 microg/l in the remaining cases. Macroadenomas and invasive adenomas were seen in seven and six cases, respectively.Pituitary adenomas that show a faint GH-immunoreactivity but lack FB do not fit the established classification. These adenomas may be a distinct pituitary adenoma type of Pit-1 lineage with endocrinologically low activity.


High expression of Toll-like receptor 4/myeloid differentiation factor 88 signals correlates with poor prognosis in colorectal cancer.

BACKGROUND: The Toll-like receptor (TLR) 4 signalling pathway has been shown to have oncogenic effects in vitro and in vivo. To demonstrate the role of TLR4 signalling in colon tumourigenesis, we examined the expression of TLR4 and myeloid differentiation factor 88 (MyD88) in colorectal cancer (CRC). METHODS: The expression of TLR4 and MyD88 in 108 CRC samples, 15 adenomas, and 15 normal mucosae was evaluated by immunohistochemistry, and the correlations between their immunoscores and clinicopathological variables, including disease-free survival (DFS) and overall survival (OS), were analysed. RESULTS: Compared with normal mucosae and adenomas, 20% cancers displayed high expression of TLR4, and 23% cancers showed high expression of MyD88. The high expression of TLR4 and MyD88 was significantly correlated with liver metastasis (P=0.0001, P=0.0054). In univariate analysis, the high expression of TLR4 was significantly associated with shorter OS (hazard ratio (HR): 2.17; 95% confidence interval (95% CI): 1.15-4.07; P=0.015). The high expression of MyD88 expression was significantly associated with poor DFS and OS (HR: 2.33; 95% CI: 1.31-4.13; P=0.0038 and HR: 3.03; 95% CI: 1.67-5.48; P=0.0002). The high combined expression of TLR4 and MyD88 was also significantly associated with poor DFS and OS (HR: 2.25; 95% CI: 1.27-3.99; P=0.0053 and HR: 2.97; 95% CI: 1.64-5.38; P=0.0003). Multivariate analysis showed that high expressions of TLR4 (OS: adjusted HR: 1.88; 95% CI: 0.99-3.55; P=0.0298) and MyD88 (DFS: adjusted HR: 1.93; 95% CI: 1.01-3.67; P=0.0441; OS: adjusted HR: 2.25; 95% CI: 1.17-4.33; P=0.0112) were independent prognostic factors of OS. Furthermore, high co-expression of TLR4/MyD88 was strongly associated with both poor DFS and OS. CONCLUSION: Our findings suggest that high expression of TLR4 and MyD88 is associated with liver metastasis and is an independent predictor of poor prognosis in patients with CRC.


Expression of p18(INK4C) is down-regulated in human pituitary adenomas.

Cyclin-dependent kinase inhibitors represented by the INK4 family comprising p16(INK4A), p15(INK4B), p18(INK4C), and p19(INK4D) are regulators of the cell cycle shown to be aberrant in many types of cancer. Mice lacking p18(Ink4c) exhibit a series of phenotypes including the development of widespread organomegaly and pituitary adenomas. The objective of our study is to examine the role of p18(INK4C) in the pathogenesis of human pituitary tumors. The protein and mRNA levels of p18(INK4C) were examined by immunohistochemistry and real-time reverse transcription-polymerase chain reaction, respectively. The methylation status of the p18(INK4C) gene promoter and somatic mutations of the p18(INK4C) gene were also investigated. p18(INK4C) protein expression was lost or significantly reduced in 64% of pituitary adenomas compared with levels in normal pituitary glands. p18(INK4C) mRNA levels were low in all ACTH adenomas and non-functioning (NF)-FSH and in 42%, 70% and 66% of GH, PRL, and subtype 3 adenomas, respectively. p18(INK4C) mRNA levels were significantly associated with p18(INK4C) protein levels. Neither methylated promoters in pituitary adenomas, except in one NF-FSH adenoma, nor somatic mutations of the p18(INK4C) gene in any pituitary adenomas were detected. The down-regulation of p18(INK4C) expression may contribute to the tumorigenesis of pituitary adenomas.


Idiopathic phlebosclerosis: an atypical presentation of ischemic colitis treated by laparoscopic colectomy.


Frequent overexpression of HMGA1 and 2 in gastroenteropancreatic neuroendocrine tumours and its relationship to let-7 downregulation.

The molecular pathogenesis of gastroenteropancreatic (GEP) neuroendocrine tumours (NETs) remains to be elucidated. High-mobility group A (HMGA) proteins play important roles in the regulation of transcription, differentiation, and neoplastic transformation. In this study, the expression of HMGA1 and HMGA2 was studied in 55 GEP NETs. Overexpression of HMGA1 and 2 was frequently detected in GEP NETs compared with normal tissues. Nuclear immunostaining of HMGA1 and 2 was observed in GEP NETs (38 of 55, 69%; 40 of 55, 73%, respectively). High-mobility group A2 expression increased from well-differentiated NET (WNET) to well-differentiated neuroendocrine carcinoma (WNEC) and poorly differentiated NEC (PNEC) (P<0.005) and showed the highest level in stage IV tumours (P<0.01). In WNECs, the expression of HMGA1 and 2 was significantly higher in metastatic tumours than those without metastasis (P<0.05). Gastroenteropancreatic NETs in foregut showed the highest level of HMGA1 and 2 expressions. MIB-1 labelling index (MIB-1 LI) correlated with HMGA1 and 2 overexpression (R=0.28, P<0.05; R=0.434, P<0.001; respectively) and progressively increased from WNETs to WNECs and PNECs (P<0.001). Let-7 expression was addressed in 6 normal organs, 30 tumour samples, and 24 tumour margin non-tumour tissues. Compared with normal tissues, let-7 downregulation was frequent in NETs (19 of 30, 63%). Higher expression of HMGA1 and 2 was frequently observed in tumours with let-7 significant reduction (53, 42%, respectively). The reverse correlation could be detected between HMGA1 and let-7 (P<0.05). Our findings suggested that HMGA1 and 2 overexpression and let-7 downregulation might relate to pathogenesis of GEP NETs.


Overexpression of HMGA2 relates to reduction of the let-7 and its relationship to clinicopathological features in pituitary adenomas.

High-mobility group A2 is highly expressed during embryogenesis and in various benign and malignant tumors. Recent studies report that high-mobility group A2 is negatively regulated by the let-7 microRNAs (miRNAs) family in vitro. The development of pituitary adenomas in high-mobility group A2 transgenic mice showed that high-mobility group A2 may be involved in pituitary tumorigenesis. However, no studies have investigated the clinical significance of high-mobility group A2 and its relationship to the let-7 miRNA family in human pituitary adenomas. Using immunohistochemistry, we analyzed high-mobility group A2 expression with respect to various clinicopathologic factors in 98 pituitary adenomas. Overexpression of high-mobility group A2 was observed in 39% (38/98) of pituitary adenomas compared with normal adenohypophysial tissue and was frequently found in adenomas including prolactin (PRL), adrenocorticotrophic hormone, or follicle-stimulating hormone/luteinizing hormone and in null cell adenomas, but relatively rare in growth hormone (GH) and mixed GH/PRL adenomas. High-mobility group A2 expression was significantly associated with tumor invasion (P<0.05) and was significantly higher in grade IV than in grades I, II, and III adenomas (P<0.05). High levels of high-mobility group A2 expression were more frequently observed in macroadenomas than in microadenomas (P<0.05). High levels of high-mobility group A2 expression also significantly correlated with the proliferation marker Ki-67 (P<0.0001). Real-time quantitative RT-PCR analysis was carried out to evaluate the expression of let-7 in 55 pituitary adenomas. Subsequently, decreased expression of let-7 was confirmed in 23 of 55 (42%) adenomas and was correlated with high-grade tumors (P<0.05). An inverse correlation between let-7 and high-mobility group A2 expression was evident (R=-0.33, P<0.05). These findings support a causal link between let-7 and high-mobility group A2 whereby loss of let-7 expression induces high-mobility group A2 upregulation that represents an important mechanism in pituitary tumorigenesis and progression.


Altered fracture callus formation in chondromodulin-I deficient mice.

Chondromodulin-I (Chm-I) is a glycoprotein that stimulates the growth of chondrocytes and inhibits angiogenesis in vitro. Mice lacking the Chm1 gene show abnormal bone metabolism and pathological angiogenesis in cardiac valves in the mature stage although they develop normally without aberrations in endochondral bone formation during embryogenesis or in cartilage development during growth. These findings indicate that Chm-I is critical under conditions of stress such as bone repair through endochondral ossification of a fracture callus. We carried out the present study to examine the expression and role of Chm-I in bone repair using a stabilized tibial fracture model, and compared fracture healing in Chm1 knockout (Chm1(-/-)) mice with that in wild-type mice. Chm-I mRNA and protein localized in the external cartilaginous callus in the reparative phase of fracture healing. Radiological examination showed a delayed union in Chm1(-/-) mice although the fracture site was covered with both external and internal calluses. Chm1 null mutation reduced external cartilaginous callus formation as judged by marked decrease of type X collagen alpha 1 (Col10a1) expression and the total amount of cartilage matrix. Interestingly, the majority of chondrocytes in the periosteal callus failed to differentiate into mature chondrocytes in Chm1(-/-) mice, while the hypertrophic maturation of chondrocytes between the cortices was not affected. These results suggest that Chm-I is involved in hypertrophic maturation of periosteal chondrocytes. Although a direct effect of Chm-I on bones is still unclear, bony callus formation was increased while external cartilaginous callus decreased in Chm1(-/-) mice. We conclude that in the absence of Chm1, predominant primary bone healing occurs due to an indirect effect induced by reduction of cartilaginous callus rather than to a direct effect on osteogenic function, resulting in a delayed union.


Clinicopathological features of growth hormone-producing pituitary adenomas: difference among various types defined by cytokeratin distribution pattern including a transitional form.

Pituitary adenomas producing almost exclusively growth hormones (GH) have been ultrastructurally classified into two distinct types: densely granulated somatotroph (DG) adenomas and sparsely granulated (SG) adenomas. Fibrous body (FB), an intracytoplasmic globular aggregation of cytokeratin (CK) filaments, is a hallmark of SG adenomas. Under light microscope, FB could be identified by CK immunohistochemistry as a dot-pattern immunoreaction versus a perinuclear pattern for cells without FB. However, it has been noted that numerous adenomas contain mixed populations of the two patterns. To clarify clinicopathological characteristics of the adenomas with mixed populations ("intermediate type" adenomas) and to confirm clinicopathological differences between strictly defined DG-type and SG-type adenomas, we performed this study on 104 GH cell adenomas. Having segregated "intermediate-type" adenomas (26 cases), we found significant differences between typical DG-type (47 cases) and SG-type adenomas (31 cases); SG-type adenomas had younger ages (44 vs. 50), higher frequency of macroadenomas (86% vs. 58%), invasiveness (65% vs. 38%), advanced grades (3 or 4) in Knosp's classification (50% vs. 24%), and weaker immunoreaction for GH, beta-TSH, alpha-subunit, E-cadherin, and beta-catenin. Clinicopathological characteristics of "intermediate-type" adenomas were identical to those of DG-type adenomas. These findings confirm that SG-type adenoma is a distinct section of GH cell adenomas with special properties and biological behavior, and suggest that intermediate-phenotype adenomas are enrolled in DG-type adenomas. Special properties and biological behavior of SG-type adenomas may appear after the majority of tumor cells possess a fully developed fibrous body.


Reduction of GSTP1 expression by DNA methylation correlates with clinicopathological features in pituitary adenomas.

Pi-class glutathione-S-transferase (GSTP1) located on chromosome 11q13 encodes a phase II metabolic enzyme that detoxifies reactive electrophilic intermediates. GSTP1 plays an important role in the protecting cells from cytotoxic and carcinogenic agents and is expressed in normal tissues at variable levels in different cell types. Altered GSTP1 activity and expression have been reported in many tumors and this is largely due to GSTP1 DNA hypermethylation. The role of GSTP1 in pituitary tumorigenesis has not been investigated. In this study, we evaluated the GSTP1 expression level and GSTP1 DNA methylation status in a series of pituitary adenomas. Using immunohistochemistry, we identified expression of GSTP1 in all of the various normal hormone-producing adenohypophysial cell types. In pituitary adenomas, loss or reduced expression of GSTP1 was detected in 27 of 53 tumors (50.9%). Expression of GSTP1 was significantly lower in invasive adenomas than in noninvasive adenomas (P<0.05). Using methylation-specific polymerase chain reaction (MS-PCR), GSTP1 DNA promoter hypermethylation was detected in adenomas (38 of 53, 71.7%) but not in normal tissues. GSTP1 methylation was more frequent in grade II, III, and IV tumors (66.7, 85, and 83%, respectively) than in grade I tumors (33%, P<0.05). In addition, the frequency of GSTP1 methylation was higher in invasive tumors (85%) than in noninvasive tumors (59%; P<0.05). Methylation status correlated with significant downregulation of GSTP1 expression; the frequency of GSTP1 methylation was higher in tumors with reduced-GSTP1 expression (85%) than in tumors with normal or high GSTP1 expression (54%; P<0.05). These data indicate that GSTP1 inactivation through CpG hypermethylation is common in pituitary adenomas and may contribute to aggressive pituitary tumor behavior.


Prognostic significance of HIF-2alpha expression on tumor infiltrating macrophages in patients with uterine cervical cancer undergoing radiotherapy.

Hypoxia-inducible factor (HIF)-2alpha, a basic helix-loop-helix (bHLH)-PAS protein, is the principal regulator of the hypoxic transcriptional response. An immunohistochemical study reported strong HIF-2alpha expression in the cytoplasm of tumor infiltrative macrophages (TIMs). Thus we assessed the expression of HIF-2alpha in human cervical cancer tissue before radiation therapy and its relationship to the clinical outcome. Seventy three patients with histologically proven primary advanced squamous cell carcinoma of the uterine cervix underwent radiotherapy in Tokushima University Hospital after biopsy specimens were taken. Among 73 specimens stained for HIF-2alpha, 53 (72.6%) exhibited HIF-2alpha immunoreactivity in the TIMs. In only 5 of 73 cases, HIF-2alpha immunoreactivity was observed in the nuclei of tumor cells. The HIF-2alpha positive cell count ratio in TIMs was associated with disease-free survival (DFS) with the worst DFS (p=0.024) being in cases in the group with a high positive cell count ratio. A high HIF-2alpha positive cell count ratio in TIMs increased the risk of local recurrence (p=0.0142). These findings might suggest that the ratio of the HIF-2alpha positive cell in TIMs may be a new predictive indicator for prognosis before radiation therapy for uterine cervical cancer.

The Journal of Medical Investigation : JMI


Pathomechanism of loss of elasticity and hypertrophy of lumbar ligamentum flavum in elderly patients with lumbar spinal canal stenosis.

A histologic, biologic, and immunohistochemical assessment using human samples of lumbar ligamentum flavum. To clarify the pathomechanism of loss of elasticity and hypertrophy of the lumbar ligamentum flavum (LF) in the elderly population. The most common spinal disorder in elderly patients is lumbar spinal canal stenosis, causing low back and leg pain, and paresis. Canal narrowing, in part, results from hypertrophy of the LF. Although histologic and biologic literature on this topic is available, the pathomechanism of loss of elasticity and hypertrophy of the LF is still unknown. One fetus, 5 young, and 5 elderly LF were obtained for histologic study. Hematoxylin and eosin, Alcian blue, Masson Trichrome, and Elastica Van Gieson stains were performed for each LF. Nine LF were collected and were used for biologic study of real time RT-PCR to quantitatively measure mRNA expression of Type I collagen and elastin in each LF. In the LF of the fetus, elastic fibers accounted for about 75% of the entire area. In the dural aspect of the LF in the young and elderly group, the ratio was also around 75%; however, the ratio of the dorsal aspect decreased with age. Almost half of the area showing loss of elastic fibers was shown to be converted to cartilaginous tissue producing Type II collagen and proteoglycan by Alcian blue and Type II collagen immunohistochemistry. The area, which did not stain black with EV nor blue with AB stain, was positively stained blue with T stain, indicating scarring. The area of the normal dural layer was 18.0 +/- 2.3 and 33.8 +/- 4.3 (mm2), for young and elderly group, respectively. Accordingly, it was 3.2 +/- 0.8 and 18.0 +/- 10.2 (mm2), for the dorsal abnormal layer. Elastin mRNA showed a relatively strong correlation (r = 0.44) with age; however, the slope was very gentle. Type I collagen mRNA showed a very strong correlation (r = 0.80) with age. The slope was steeper, and the value reached at 1000% (10-fold) around 65 years old when compared with the LF from younger patient. Elastin mRNA showed a weak correlation (r = 0.36) with thickness, and the slope was gentle. Type I collagen mRNA showed relatively strong correlation (r = 0.52) with thickness. The slope was steeper, and the line reached at 1000% (10-fold) around 6.5 (mm) when compared with a thin LF. Decreased elasticity of LF in the elderly is due to the loss of elastic fibers and a concomitant increase of collagenous fibers in the dorsal aspect. LF hypertrophy could be due to the thickening of the normal elastic layer as well as of the abnormal collagenous layer.


Callus formation during healing of the repaired tendon-bone junction. A rat experimental model.

This study was undertaken to elucidate the mechanism of biological repair at the tendon-bone junction in a rat model. The stump of the toe flexor tendon was sutured to a drilled hole in the tibia (tendon suture group, n = 23) to investigate healing of the tendon-bone junction both radiologically and histologically. Radiological and histological findings were compared with those observed in a sham control group where the bone alone was drilled (n = 19). The biomechanical strength of the repaired junction was confirmed by pull-out testing six weeks after surgery in four rats in the tendon suture group. Callus formation was observed at the site of repair in the tendon suture group, whereas in the sham group callus formation was minimal. During the pull-out test, the repaired tendon-bone junction did not fail because the musculotendinous junction always disrupted first. In order to understand the factors that influenced callus formation at the site of repair, four further groups were evaluated. The nature of the sutured tendon itself was investigated by analysing healing of a tendon stump after necrosis had been induced with liquid nitrogen in 16 cases. A proximal suture group (n = 16) and a partial tenotomy group (n = 16) were prepared to investigate the effects of biomechanical loading on the site of repair. Finally, a group where the periosteum had been excised at the site of repair (n = 16) was examined to study the role of the periosteum. These four groups showed less callus formation radiologically and histologically than did the tendon suture group. In conclusion, the sutured tendon-bone junction healed and achieved mechanical strength at six weeks after suturing, showing good local callus formation. The viability of the tendon stump, mechanical loading and intact periosteum were all found to be important factors for better callus formation at a repaired tendon-bone junction.

The Journal of Bone and Joint Surgery. British Volume


Tumor-specific down-regulation and methylation of the CDH13 (H-cadherin) and CDH1 (E-cadherin) genes correlate with aggressiveness of human pituitary adenomas

The gene products of CDH13 and CDH1, H-cadherin and E-cadherin, respectively, play a key role in cell-cell adhesion. Inactivation of the cadherin-mediated cell adhesion system caused by aberrant methylation is a common finding in human cancers, indicating that the CDH13 and CDH1 function as tumor suppressor and invasion suppressor genes. In this study, we analyzed the expression of H-cadherin mRNA and E-cadherin protein in 5 normal pituitary tissues and 69 primary pituitary adenomas including all major types by quantitative real-time RT-PCR (qRT-PCR) and immunohistochemistry, respectively. Reduced expression of H-cadherin was detected in 54% (28/52) of pituitary tumors and was significantly associated with tumor aggressiveness (P<0.05). E-cadherin expression was lost in 30% (21 of 69) and significantly reduced in 32% (22 of 69) of tumors. E-cadherin expression was significantly lower in grade II, III, and IV than in grade I adenomas (P=0.015, P=0.029, and P=0.01, respectively). Using methylation-specific PCR (MSP), promoter hypermethylation of CDH13 and CDH1 was detected in 30 and 36% of 69 adenomas, respectively, but not in 5 normal pituitary tissues. Methylation of CDH13 was observed more frequently in invasive adenomas (42%) than in non-invasive adenomas (19%) (P<0.05) and methylation of CDH1 was more frequent in grade IV adenomas compared with grade I adenomas (P<0.05). Methylation of either CDH13 or CDH1 was identified in 35 cases (51%) and was more frequent in grade IV invasive adenomas than in grade I non-invasive adenomas (P<0.05 and P<0.05, respectively). Downregulation of expression was correlated with promoter hypermethylation in CDH13 and CDH1. In conclusion, the tumor-specific downregulation of expression and methylation of CDH13 and CDH1, alone or in combination, may be involved in the development and invasive growth of pituitary adenomas.


Usefulness of cytokeratin immunoreactivity pattern for distinction of Barrett's esophagus from intestinal metaplasia of the stomach

The histological distinction between Barrett's esophagus involving the distal esophagus and intestinal metaplasia of the stomach has important clinical implications and can be difficult even with the use of histochemical stains. Cytokeratin (CK) 7 and 20 are cytoplastic structural proteins that show restricted expression in normal and malignant epithelia of the gastrointestinal tract. CK7 and 20 immunostaining were performed on a 67-year-old male with cardiac cancer with reflux esophagitis due to sliding hernia. The CK7/20 immunoreactivity pattern of cancer and reflux esophagitis in this case showed superficial CK20 staining and strong CK7 staining of both superficial and deep glands. In intestinal metaplasia of the stomach, strong CK20 immunostaining in superficial and deep glands and absent CK7 immunoreactivity were noted. Neither CK7 nor CK20 immunoreactivity was noted in squamous cell epithelium. Therefore, we concluded that in this patient intestinal metaplasia of the esophagus was BE. The CK7/20 reactivity pattern is useful for identifying the intestinal metaplasia of the esophagus from the stomach using histological materials from biopsy and surgically resected specimens.


A study of the correlation between morphological findings and biological activities in clinically nonfunctioning pituitary adenomas.

The aims of this study are to review the histology of the clinically nonfunctioning pituitary adenomas (CNFPAs) we have observed and to determine whether or not the frequency of cavernous sinus invasion is different among each type of morphology. In addition, several proliferative markers, including Ki67, p53, E-cadherin, matrix metallo-proteinase 9 and pituitary tumor derived fibroblast growth factor receptor 4 (ptd-FGFR4), were also investigated in invasive and non-invasive tumors. Our consequent 213 CNFPAs were diagnosed as follows: 64% were silent gonadotroph adenomas, 18% were null cell adenomas, 12% were silent corticotroph adenomas, 4% were silent Subtype 3 adenomas, and 1% were other types of adenomas. Female patients or younger patients showed a significant preponderance in silent corticotroph adenomas and in silent Subtype 3 adenomas, respectively. Cavernous sinus invasion occurs most frequently in silent corticotroph adenomas (85%) followed by Subtype 3 adenomas (67%), null cell adenomas (38%), and silent gonadotroph adenomas (11%). There were no significant differences in the Ki67, p53, E-cadherin, matrix metallo-proteinase 9, and ptd-FGFR4 expression between tumors with and without cavernous sinus invasion. From a clinical standpoint, it is quite important to differentiate morphological type in CNFPAs to aid the clinician in assessing the clinical behavior and prognosis of the tumor. Therefore, we suggest that all CNFPAs be examined not only by conventional light microscopy but also by immunohistochemistry, preferably by electron microscopy, to achieve a correct morphological diagnosis.


A Growth Hormone Receptor Mutation Impairs Growth Hormone Autofeedback Signaling in Pituitary Tumors.

Pituitary tumors are a diverse group of neoplasms that are classified based on clinical manifestations, hormone excess, and histomorphologic features. Those that cause growth hormone (GH) excess and acromegaly are subdivided into morphologic variants that have not yet been shown to have pathogenetic significance or predictive value for therapy and outcome. Here, we identify a selective somatic histidine-to-leucine substitution in codon 49 of the extracellular domain of the GH receptor (GHR) in a morphologic subtype of human GH-producing pituitary tumors that is characterized by the presence of cytoskeletal aggresomes. This GHR mutation significantly impairs glycosylation-mediated receptor processing, maturation, ligand binding, and signaling. Pharmacologic GH antagonism recapitulates the morphologic phenotype of pituitary tumors from which this mutation was identified, inducing the formation of cytoskeletal keratin aggresomes. This novel GHR mutation provides evidence for impaired hormone autofeedback in the pathogenesis of these pituitary tumors. It explains the lack of responsiveness to somatostatin analogue therapy of this tumor type, in contrast to the exquisite sensitivity of tumors that lack aggresomes, and has therapeutic implications for the safety of GH antagonism as a therapeutic modality in acromegaly.


The aryl hydrocarbon receptor-interacting protein gene is rarely mutated in sporadic GH-secreting adenomas

Recently, germline mutations of aryl hydrocarbon receptor-interacting protein (AIP) gene located on 11q13 were identified in patients with pituitary adenoma predisposition. AIM/PATIENTS AND METHODS: We investigated the involvement of the AIP gene in one family with isolated familial somatotropinomas (IFS). To investigate the role of AIP in sporadic GH-secreting adenomas, we first analysed somatic mutations in 40 tumours. Second, DNA from corresponding leucocytes was analysed in tumours showing genetic changes of the AIP gene. Germline mutation of AIP was found in an IFS family. Bi-allelic inactivation of AIP by a combination of germline mutation and loss of heterozygosity were confirmed in two pituitary adenomas. Mutation analysis of the AIP gene in the 40 sporadic GH-secreting adenomas showed no mutations except for a missense mutation, suggesting that germline mutations in patients diagnosed with sporadic acromegaly or gigantism were rare. In a patient with gigantism, a missense mutation of V49M was identified at the germline level. Based on these results, we conclude that the loss of function of AIP contributes to IFS, but not for most Japanese sporadic GH-secreting adenomas.


A case of parathyroid hormone-related peptide producing gallbladder carcinoma presenting humoral hypercalcemia of malignancy

Humoral hypercalcemia of malignancy(HHM)を来したPTHrP産生胆嚢癌の1例

Humoral hypercalcemia of malignancy (HHM) in neoplastic syndrome has been most commonly reported in squamous cell carcinoma. Gallbladder carcinoma with HHM is uncommon. In this report, we describe a male case of gallbladder carcinoma with marked hypercalcemia and a high level of serum parathyroid hormone-related peptide (PTHrP). An immunohistochemical examination using PTHrP was also positive.

腫瘍随伴症候群は扁平上皮由来の癌で報告が多いが胆嚢癌に合併した報告は少ない．筆者らは経過中に高カルシウム血症および血中PTHrP高値を認め，病理組織標本においてPTHrP免疫染色が陽性であった胆嚢癌の1剖検例を経験したので報告する．

The Japanese Journal of Gastro-enterology


Diversity of ACTH-immunoreactive cells in the human adenohypophysis: an immunohistochemical study with special reference to cluster formation and follicular cell association.

Horvath et al. called relatively small ACTH-immunoreactive (ACTH-IR) cells observed in basophil invasion "proopiomelanocortin (POMC) cells," and these cells were supposed to be different from larger ACTH-IR cells in human adenohypophysis. To clarify the existence of "anterior POMC cells," we examined 21 autopsy cases taking note of follicular cells (FCs). We found that smaller ACTH-IR cells were clustered forming small areas without FC association, clustered small cell island (CSCI). CSCI was found in all the specimens we examined using immunohistochemical staining for ACTH and cytokeratin (AE1/AE3) in serial sections. ACTH-IR cells outside CSCI were of various sizes, mainly consisting of larger ACTH-IR cells that were diffusely scattered among other hormone-secreting cells and closely associated with FCs. By immunohistochemistry, ACTH-IR cells within CSCI and basophil invasion showed uniformly weak reactivity and common patterns for cytokeratins (CAM5.2 and 35 beta H11), carboxypeptidase D, and LeuM1, whereas those outside CSCI showed a great variety of immunoreactivity. The similarity in the immunoreactivity of ACTH-IR cells between CSCI and basophil invasion suggests that ACTH-IR cells in these two areas have common characteristics and ACTH-IR cells in CSCI are most likely "POMC cells." The clustered "anterior POMC cells" may be distinguishable by light microscopic immunohistochemistry.


嚢胞変性により特異な形態を呈した胃GISTの一例

嚢胞変性により特異な形態を呈した胃GISTの一例


Colonic pseudolipomatosis, microscopically classified into two groups

Colonic pseudolipomatosis is rare and the pathogenesis is controversial. The purpose of the present paper was to clarify endoscopic and histological characteristics of colonic pseudolipomatosis and to discuss the etiology. A total of 15 lesions from 14 patients was reviewed. They were able to be histologically classified into two groups on the basis of variety in size of the vacuoles: Group A, the ratio of largest vacuole to smallest vacuole in size is less than three, Group B, the ratio is more than four. Four of 15 lesions were group A, and were endoscopically polypoid or flat lesions covered with normal-looking mucosa. They were microscopically characterized by (i) predominant location in the upper portion of the lamina propria; (ii) no submucosal involvement; (iii) less variation in vacuolar size; and (iv) no association with lymph follicles. The vacuoles of group A contained proteinaceous materials in two of four lesions. Group B (11 lesions) had small elevated mucosa with normal-looking surface or non-elevated reddish mucosa. Microscopically, the lesions were mainly located in the lower portion of the lamina propria, occasionally also in the submucosa, had variable-sized vacuoles, and were related to lymph follicles. It is suggested that the vacuoles in group A contain fluid, and may indicate an abnormal stagnation of interstitial fluid. Microscopic appearance of group B is essentially similar to that of pneumatosis coli. It is thought that group B probably results from penetration of gas from the crypts into the mucosa during colonoscopy. It is unclear why group B had a preference for ileocecal valve and an association with lymph follicles.

Journal of Gastroenterology and Hepatology


Comments on the WHO histological classification of pituitary tumors.


Duodenal lymphangitis carcinomatosa: endoscopic characteristics and clinical significance.

Duodenal lymphangitis carcinomatosa has been sporadically described, but so far little attention has been paid to duodenal lymphangitis carcinomatosa. Four cases with duodenal lymphangitis carcinomatosa were endoscopically and histologically examined. The four cases exhibited multiple polypoid lesions along the Kerckring's folds and/or were covered by characteristically granular, non-ulcerated mucosa upon thickening. The granularity seems to been caused by dilated lymph vessels containing the carcinoma cells. The lesions were microscopically characterized by: (i) involvement of lymph vessels located in the upper portion of the lamina propria; (ii) no inflammatory changes; and (iii) no desmoplastic changes. Primary sites were thought to be the stomach in case 1, the pancreas in cases 2 and 4, and unknown in case 3. All patients died within 6 months after admission or endoscopic examination. As duodenal lymphangitis carcinomatosis shows characteristic endoscopic appearance, endoscopic diagnosis is not difficult. We should realize that the lesion represents extremely poor prognosis, and it should be distinguished from ordinary metastatic duodenal carcinoma.


Genetic analyses in patients with familial isolated hyperparathyroidism and hyperparathyroidism-jaw tumour syndrome.

A subset of familial isolated primary hyperparathyroidism (FIHP) is a variant of hyperparathyroidism-jaw tumour syndrome (HPT-JT). AIM/PATIENTS AND METHODS: We investigated the involvement of the HRPT2, MEN1 and CASR genes in 11 provisional FIHP families and two HPT-JT families. Germline mutations of HRPT2 were found in two of the 11 FIHP families and one of the two HPT-JT families. One FIHP family with parathyroid carcinoma and atypical adenomas and another FIHP family with cystic parathyroid adenoma had novel frameshift mutations of 518-521del and 62-66del, respectively. In a patient with HPT-JT, a de novo germline mutation of 39delC was detected. Novel somatic HRPT2 mutations of 70-73del and 95-102del were found in two of five parathyroid tumours in a family with a 518-521del mutation. Biallelic inactivation of HRPT2 by a combination of germline and somatic mutation was confirmed in the parathyroid tumours. The finding that two families diagnosed with FIHP carried HRPT2 mutations suggests that they have occult HPT-JT. In the remaining 10 families, one family had a missense MEN1 mutation. No mutations of CASR were detected. Our results confirm the need to test for HRPT2 in FIHP families, especially those with parathyroid carcinomas, atypical adenomas or adenomas with cystic change.


Malignant B-lymphoid cells with bone lesions express receptor activator of nuclear factor-kappaB ligand and vascular endothelial growth factor to enhance osteoclastogenesis.

Receptor activator of nuclear factor-kappaB ligand (RANKL) is a key mediator of osteoclastogenesis. Because certain types of tumor cells aberrantly express RANKL, and because bone destruction also develops in B-cell lymphomas of bone origin, we investigated RANKL expression and the mechanisms of osteoclastogenesis in B-lymphoid neoplasms. Immunohistochemistry of bone specimens resected from patients with primary B-cell lymphoma of bone with bone destruction revealed that lymphoma cells express RANKL as well as vascular endothelial cell growth factor (VEGF). The tumor cells isolated from the bone specimens enhanced osteoclastogenesis in vitro. In contrast, B-cell lymphoma infiltrating to the bone marrow without bone destruction did not express RANKL. Both RANKL and VEGF were expressed by a portion of B-lymphoid cell lines, including Daudi and IM-9. These RANKL-expressing tumor cells enhanced osteoclastogenesis from RAW264.7 cells and human monocyte-derived preosteoclasts in the absence of stromal cells/osteoblasts in a RANKL-dependent manner. Furthermore, conditioned media from Daudi cells enhanced transmigration of preosteoclasts that was inhibited by anti-VEGF antibody, suggesting that tumor cell-derived VEGF mediates recruitment of osteoclast precursors. Moreover, cocultures of B-lymphoid cell lines with osteoclasts enhanced the growth of B-lymphoid cells. Some malignant B cells aberrantly express functional RANKL as well as VEGF to enhance osteoclastogenesis. The coexpression of RANKL and VEGF may also contribute to the close cellular interactions with osteoclastic cells, thereby forming a vicious cycle between osteoclastic bone destruction and tumor expansion in bone.


Histological and immunohistochemical study of composite neuroendocrine-exocrine carcinomas of the stomach.

Composite neuroendocrine-exocrine carcinomas (NEECs) with two distinct components of adenocarcinoma and neuroendocrine (NE) carcinoma within the same tumor are rare but may have a clue for clarifying the pathogenesis of NE tumors arising from non-endocrine organs. This study was done to characterize histological and immunohistochemical features of NEECs of the stomach comparing with pure NE tumors of the gastrointestinal (GI) tract. Microscopically, adenocarcinoma components in 6 of 8 NEECs were well differentiated and located superficially, whereas NE components were poorly differentiated and located deeply. In the remaining two cases, smaller NE components were intermingled within adenocarcinoma components. Immunohistochemically, neural cell adhesion molecule (NCAM) was positive in 5 NE components, of which 3 cases were homogeneously positive, and 2 adenocarcinoma components of 8 NEECs, while 19 of 21 pure NE tumors of GI tract were homogeneously positive for NCAM. Ghrelin-immunoreactivity was found in 4 NE components and 2 adenocarcinoma components of NEECs, although 20 pure NE tumors were positive for ghrelin. Smad4 was positive in both components of 7 NEECs. These findings suggest that composite NEECs and pure NE tumors of GI tract may have different NE properties and that most NE components of composite NEECs of the stomach may originate from an adenocarcinoma precursor cell and occasional tumors from a pluripotent cell.

The Journal of Medical Investigation : JMI


筋ジストロフィー心臓病理

筋ジストロフィー心臓病理

デュシェンヌ型筋ジストロフィーの心臓病変について述べた．

デュシェンヌ型筋ジストロフィーの心臓病変について述べた．


immunohistochemical study on so-called follicular cells and folliculostellate cells in the human adenohypophysis.

Non-hormone-secreting cells in human adenohypophysis have been designated as either follicular cells (FC) or folliculostellate cells (FSC). They have similarly long cytoplasmic processes, and the difference between FC and FSC remains unclear. An immunohistochemical study for S-100 protein, cytokeratin (CK, detected by AE1/AE3) and glial fibrillary acidic protein (GFAP) was performed in autopsy pituitaries. Double immunohistochemistry for S-100 protein and CK revealed that there were numerous coexpressed cells. The most frequent type ('CK-type cell') was cells weakly positive for S-100 protein in the nucleus and for CK-immunoreactivity in the cytoplasm. The next numerous type ('S-100 protein cell') was cells strongly positive for S-100 protein and weakly positive or negative for CK. The CK-type cells were frequently observed in the vicinity of follicular structures and in neighborhood of adrenocorticotropic hormone -immunoreactive cells, and were most likely the cells termed FC. They were often observed around necrotic areas. The S-100 protein cells were individually found in the circumference of endocrine cell nest, and seemed to be the so-called stellate cells. GFAP-positive cells were rare. It is implied that S-100 protein-positive FSC could be divided into at least two main subtypes: FC (CK-type cells) and stellate cells (S-100 protein cells).


Overexpression of a novel superoxide-producing enzyme,NADPH oxidase 1,in adenoma and well differentiated adenocarcinoma of the human colon

A superoxide-producing enzyme, NADPH oxidase 1 (Nox1), dominantly expressed in the colon, is implicated in the pathogenesis of colon cancer. Immunohistochemistry showed that Nox1 was constitutively expressed in surface mucous cells. Adenomas and well differentiated adenocarcinomas up-regulated Nox1 expression. Ki-67-negative, well differentiated tumor cells contained abundant Nox1, whereas Ki-67-positive, proliferating cells did not express it. This differentiation-dependent expression in normal as well as tumor tissues suggests distinct roles of Nox1 besides mitogenic function. Nuclear factor (NF)-kappaB was predominantly activated in adenoma and adenocarcinoma cells expressing abundant Nox1, suggesting that Nox1 may stimulate NF-kappaB-dependent antiapoptotic pathways in colon tumors.


Inactivation of RASSF1A tumor suppressor gene by aberrant promoter hypermethylation in human pituitary adenomas.

Aberrant promoter methylation and resultant silencing of several tumor suppressor genes play an important role in the pathogenesis of many tumor types. The human Ras association domain family 1A gene (RASSF1A), recently cloned from the lung tumor locus at 3p21.3, was shown to be frequently inactivated by hypermethylation of its promoter region in a number of malignancies. We have investigated the expression and epigenetic changes of this novel universal tumor suppressor gene in pituitary adenomas and correlated the data with clinicopathologic findings. Fresh frozen normal pituitary tissues and 52 primary pituitary adenomas including all major types were examined. Methylation-specific polymerase chain reaction (MSP), combined bisulfite restriction analysis (COBRA), bisulfite sequencing and semiquantitative reverse transcription-polymerase chain reaction were used to analyze DNA promoter methylation status and the mRNA expression of RASSF1A, respectively. High levels of RASSF1A transcript and no methylation of the RASSF1A promoter were found in normal pituitary tissues. RASSF1A promoter methylation was detected in 20 of 52 (38%) adenomas including all major types of pituitary adenomas. However, a lower frequency of methylation of the RASSF1A promoter was found in gonadotroph cell adenomas (15%) compared with growth hormone cell, prolactin cell, or adrenocorticotropic hormone cell adenomas (54, 46 and 50%, respectively). Methylation frequency was higher in the most aggressive adenomas (87% in grade IV tumors, P=0.0163). In addition, methylation of the RASSF1A promoter potentially correlated with higher labeling index of the proliferation marker Ki-67 (P=0.1475). Loss or significant reduction of RASSF1A messenger RNA transcripts was identified in 18 of 20 (90%) adenomas with hypermethylation of RASSF1A (P<0.0001). Our data suggest that promoter hypermethylation of RASSF1A and resultant alterations of RASSF1A expression may play a critical role in pituitary tumorigenesis and may be involved in tumor progression.

Laboratory Investigation; a Journal of Technical Methods and Pathology


Microsatellite instability and protein expression of the DNA mismatch repair gene, hMLH1, of lung cancer in chromate-exposed workers.

Our previous studies of lung cancer in chromate-exposed workers (chromate lung cancer) have revealed that the frequency of replication error (RER) in chromate lung cancer is very high. We examined whether the RER phenotype of chromate lung cancer is due to an abnormality of DNA mismatch repair protein. We investigated the expression of a DNA mismatch repair gene, hMLH1, and hMSH2 proteins using immunohistochemistry and microsatellite instability (MSI) in 35 chromate lung cancers and 26 nonchromate lung cancers. Lung cancer without MSI or with MSI at one locus was defined as "RER(-)," lung cancer with MSI at two loci was defined as "RER(+)," and lung cancer with MSI at three or more loci was defined as "RER(++)." The repression rate of hMLH1 and hMSH2 proteins in chromate lung cancer was significantly more than that of nonchromate lung cancer (hMLH1: 56% vs. 20%, P = 0.006, hMSH2: 74% vs. 23%, P < 0.0001). In chromate lung cancer, the repression rate for hMLH1 was 43% in RER(-), 40% in RER(+), and 90% in the RER(++) group. The repression rate of hMLH1 protein in the RER(++) group was significantly higher than that in the RER(-) and RER(+) groups (P = 0.039). The inactivation of hMLH1 expression strongly correlated with the microsatellite high instability phenotype in chromate lung cancer. The genetic instability of chromate lung cancer is due to the repression of hMLH1 protein.


Hypoplasia of endocrine and exocrine pancreas in homozygous transgenic TGF-β1

We generated the homozygous transgenic mice with expression of the active form of TGF-beta1 by the glucagon promoter (homozygous NOD-TGF-beta1). The homozygous NOD-TGF-beta1 showed severe diabetes in 84.6%, impaired glucose tolerance, and low serum insulin levels. The final size of endocrine and whole pancreas decreased, respectively, to 6 and 34%, compared to wild-type mice. The homozygous N(2) backcross to C57BL/6 (B6-TGF-beta1) showed no diabetes, but impaired glucose tolerance and low serum insulin levels. In homozygous NOD-TGF-beta1, the expression of p15(INK4b) was induced by 3.4-fold in pancreatic islets than that in wild-type mice. Based on these, we conclude first that excessive paracrine TGF-beta1 signaling in islets results in endocrine and exocrine pancreatic hypoplasia, second that TGF-beta1decrease the final size of endocrine and exocrine pancreas presumably through regulating cell cycle via p15(INK4b) at least in endocrine pancreas, and third that hypoplastic action of TGF-beta1 of pancreatic islets is independent of the genetic background.


Co-localization of honeycomb golgi and ACTH granules in a giant ACTH-producing pituitary adenoma

We document the co-localization of honeycomb golgi and ACTH-immunopositive granules in giant ACTH-producing pituitary adenoma cells. A 42-yr-old woman presented with Cushing's disease and a giant adenoma that invaded the sphenoid and cavernous sinus. She underwent transsphenoidal surgery followed by radiation therapy. Some of the adenoma cells were ACTH-positive and upon electron-microscopic (EM) study most were found to contain sparse granules and no type-I filaments. In many cells the golgi complex had undergone partial or total vacuolar transformation that resulted in the appearance of honeycomb golgi. Immunohistochemical study of mirror sections of portions containing cells with honeycomb golgi revealed that the cells with honeycomb golgi showed ACTH-immunopositivity. Honeycomb golgi, which was formerly considered a morphological marker of gonadotroph adenomas in females, has previously been identified in large ACTH-producing pituitary adenomas but there has been no direct evidence that individual cells with honeycomb golgi are cells that produce ACTH. Our immunohistochemical documentation of ACTH-immunoreactivity in individual adenoma cells containing honeycomb golgi clearly confirms that honeycomb golgi is not confined only to gonadotroph adenomas in females. Rather, the existence of honeycomb golgi in cells of other adenoma types may be due to their low hormone production and/or to disturbances in the regulation of the exocytotic pathway.


Clinical and Molecular Features of a TSH-Secreting Pituitary Microadenoma

We describe a case of a thyroid stimulating hormone (TSH)-secreting pituitary microadenoma, and report the systematic gene expression profile of the surgically- removed tumor. A 50-year-old woman was referred to our hospital because she had high TSH, free-T4, and free-T3 levels, and a pituitary tumor that was visualized with magnetic resonance imaging. Her basal TSH level was high even after a high T3 loading dose, and increased following administration of thyroid releasing hormone (TRH) even after administration of a high dose of exogenous T3. Her clinical symptoms and peripheral markers for T3 were responsive to exogenous T3. There was no thyroid hormone receptor (TR) beta gene mutation. The patient was diagnosed with a TSH-secreting pituitary adenoma, and trans-sphenoid surgery was performed. The histologic features and immunophenotype were consistent with a TSH-secreting pituitary adenoma. Reverse transcription-polymerase chain reaction analysis of pituitary hormones, pituitary-specific transcription factors, receptors, and transcriptional cofactors of clinical significance was performed on the removed tumor. The tumor expressed TSH, growth hormone, prolactin, alpha-subunit, pituitary transcription factor-1 (pit-1) but not proopiomelanocortin (POMC), prophet of pit-1 (prop-1) and pituitary cell-restricted T box factor (Tpit). TRbeta and TRH-receptor gene expression was normal. Three steroid receptor coactivators (SRC)-1, SRC-2, and SRC-3 were expressed. Nuclear receptor corepressor (N-CoR)2 was absent in the tumor, whereas nuclear receptor corepressor (N-CoR1) was expressed. Somatostatin receptor type 1 expression was significantly decreased, whereas type 4 receptor was expressed, which are unusual characteristics for pituitary tumors. The gene expression pattern in the tumor might have a role in the clinical features of this case.


A Treatment Using ST1571 for Jejunal Gastrointestinal Stromal Tumor (GIST) Accompanied with Liver Metastas and Peritoneal Dissemination

A 64-year-old man with a chief complaint of melena visited our emergency outpatient clinic. After several examinations, he was diagnosed as a gastrointestinal stromal tumor (GIST) with liver metastasis. Surgical resection of the jejunal lesion and postoperative adjuvant therapy with STI571 for one year was performed. Due to recent immunohistological studies and introduction of STI571, the diagnosis, treatment, and prognosis of GIST are about to change profoundly. Further accumulation of cases is necessary to investigate the diagnosis, treatment, and prognosis of GIST.


Expression of adhesion molecules and cytokeratin 20 in merkel cell carcinomas.Endocr Pathol

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine carcinoma of the skin. MCCs often show characteristic paranuclear dot-like immunopositivity for cytokeratin 20 (CK20), a globular aggregation of CK20 intermediate filaments. These aggregates typically form rhabdoid features and fibrous bodies and may be associated with a down-regulation in adhesion molecules (AMs). To date, the relationship between the expression of AMs and CK20 and clinicopathological findings in MCC has not been well examined. In this immunohistochemical study, we assessed the expression of AMs, CK20, and chromogranin A (CgA) on MCCs in 8 men and 23 women with this disease, and also characterized their clinicopathological features. This study is the largest of its kind that has been undertaken to date in Japanese patients. Compared to normal tissue, E-cadherin and alpha- and beta-catenins showed reduced membranous expression in 95.7%, 46.7%, and 45.2% of MCCs, respectively. Nuclear E-cadherin localization was seen in four tumors, all of which predominantly showed a CK20 dot pattern. However, there was no significant relationship between the membranous expression of AMs and a CK20 dot pattern. E-cadherin expression was significantly lower in tumors of > or =2 cm, and tumors negative for E-cadherin more frequently developed outside of the head and neck than within those regions. CgA was more intensely expressed in tumors with uniform nuclei and a dense lymphocytic infiltrate than in those that showed pleomorphisms and that had few, if any, infiltrating lymphocytes. These findings suggest that MCCs have a reduced expression of AMs and that down-regulation of E-cadherin expression may correlate with increased tumor aggressiveness. The fact that no significant relationship was demonstrable between the membranous expression of AMs and the CK20 expression pattern suggests that the mechanism of aggregation of intermediate filaments may be different in different types of tumors.


Vitiligo as a paraneoplastic syndrome preceding pituitary adenoma and subsequent acute lymphoblastic leukemia.

We present an unusual case of rapid onset of vitiligo heralding blindness due to pituitary apoplexy secondary to a non-secreting pituitary adenoma in a 49-year-old man. Deterioration in vision and skin condition coincided with tumor progression over 3 years. He subsequently died of acute lymphoblastic leukemia after pituitary radiation. The possible pathological associations between the 2 malignancies and vitiligo are discussed.


Preoperative identification of clearly separated double pituitary adenomas

Double pituitary adenomas are extremely rare. They can be divided into contiguous and clearly separated types. Most contiguous tumours are surgically removed as one tumour and the co-existence of different adenoma types can be confirmed by histological methods. In contrast, detailed preoperative neuroimaging studies can suggest the co-existence of separated multiple adenomas. In patients with multiple adenomas, surgical failure may result when one adenoma is missed during surgery. Among 600 surgical cases we encountered four patients with clearly separated double pituitary adenomas; all were highly suspect on preoperative MRI studies. All four patients manifested acromegalic symptoms; one patient also exhibited hyperprolactinemia and two had familial pituitary adenomas unrelated to multiple endocrine neoplasia type I (MEN-1). All underwent transsphenoidal surgery and histology confirmed the diagnosis of GH-producing plus gonadotroph adenoma in two cases and of two GH-producing adenomas each in the other two patients. Although the pathogenesis of our double adenomas remains unknown, genetic abnormalities may be involved because two patients had familial pituitary adenomas unrelated to MEN-1. When preoperative MRI is suggestive of double adenomas, careful surgical exploration is necessary to avoid missing the other adenoma because the risk of surgical failure is high, especially in patients with functioning adenomas.


Expression of adhesion molecules and Ki-67 in female adnexal tumor of probable Wolffian origin (FATWO): report of two cases and review of the literature.

Female adnexal tumor of probable Wolffian origin (FATWO) is a rare entity which is believed to originate from mesonephric (Wolffian) remnants on the basis of its location where the remnants are abundant. Its behavior is usually indolent, although some cases can recur or metastasize. The authors present the clinicopathological features of two cases of FATWO arising in the broad ligament, and focus on the expression of adhesion molecules and proliferative marker. Mesonephric duct remnants are also examined in an attempt to elucidate the histogenesis of FATWOs. The two FATWOs were well-circumscribed solid masses arising in the leaves of the broad ligament and histological examination revealed a mixture of cysts and tubules imparting a sieve-like pattern and mucin-negative eosinophilic secretion within these tubules. Immunohistochemically, the tumors showed the expression of cytokeratin 7 and 20, high-molecular-weight cytokeratin, and calretinin, which closely resembled that of the mesonephric duct remnants. Regarding CK 20, CD 10, EMA, S-100 protein, and vimentin their expression was in part not identical with previous studies. E-cadherin, alpha and beta-catenin were strongly expressed along the cell membrane of the tumor cells. The Ki-67 labeling index of FATWO was 0% and 3.2% in each case. The preservation of the E-cadherin-catenin complex and low Ki-67 labeling index could explain the indolent behavior and low malignant potential of this tumor.


Morules in cribriform-morular variant of papillary thyroid carcinoma: Immunohistochemical characteristics and distinction from squamous metaplasia.

Morules are a diagnostic clue to the cribriform-morular variant (C-MV) of papillary thyroid carcinoma, and are superficially similar to squamous metaplasia. In order to clarify the histogenesis of morules and differentiate them from squamous metaplasia, we immunohistochemically compared the morules in five cases of C-MV with squamous metaplasia in six cases of diffuse sclerosing variant (DSV) of papillary thyroid carcinoma. The squamous metaplastic cells were immunopositive for low- and high-molecular-weight cytokeratin, whereas the morular cells were negative or focally positive. Vimentin-positive cells were observed focally in the morules and squamous metaplasia, except for one case of CMV that showed intense positivity. The morular cells showed weak cytoplasmic positivity for beta-catenin, and the cell membrane was not highlighted. Some nuclei of the morular cells were also positive for this antibody. Beta-catenin was intensively positive along the cell membrane of the metaplastic cells, and did not react against the nuclei or cytoplasm. Bcl-2 was positive in the morular cells, but negative in the metaplastic cells. S-100 protein-positive dendritic cells were observed in the metaplastic nests, but not in the morules. We argue that morules appear in connection with nuclear and cytoplasmic aberrant localization of beta-catenin, and are not an early form of squamous metaplasia.


Cytoplasmic expression of fibroblast growth factor receptor-4 in human pituitary adenomas: relation to tumor type, size, proliferation, and invasiveness.

The pathogenesis of pituitary adenomas remains unknown. A pituitary tumor-derived (ptd) isoform of fibroblast growth factor receptor-4 (ptd-FGFR4) has been implicated in the neoplastic process. To further understand the expression of FGFR4 in sporadic human pituitary adenomas, we studied 137 pituitary adenomas of various types (102 adenomas from Japanese patients and 35 adenomas from Canadian patients) and 10 nontumorous pituitaries using a polyclonal antiserum that recognizes the C terminus of FGFR4 and analyzed possible relationships among expression of FGFR4, patient nationality, tumor type, size, invasion, and the labeling index of the proliferation marker Ki-67 using the MIB-1 antibody. Cytoplasmic expression of FGFR4 protein was observed in 57.8% of Japanese cases and 62.8% of Canadian cases. FGFR4 reactivity was absent in all 10 normal adenohypophysial tissues examined. FGFR4 expression in pituitary adenomas was restricted mainly to the cytoplasm, a pattern similar to that seen in rat pituitary cells transfected with human ptd-FGFR4 but different from that of cells transfected with wild-type FGFR4, which displayed membrane localization of staining. Protein from primary human adenomas migrated as a 65-kDa species consistent with the predicted size of ptd-FGFR4. FGFR4 protein expression was frequently found in adenomas containing GH, ACTH, or FSH/LH and was also found in null cell adenomas, but reactivity was relatively rare in prolactin-containing adenomas in both Japanese and Canadian groups. The expression of FGFR4 protein was stronger in macroadenomas than in microadenomas (P = 0.02) and high levels of FGFR4 expression (moderate or greater density staining) were more frequently observed in macroadenomas than in microadenomas (P < 0.05). High levels of FGFR4 expression also correlated significantly with the proliferation marker Ki-67 (P = 0.002) and tended (but not significantly) to be found in invasive tumors. These data are consistent with a role for ptd-FGFR4 in pituitary tumorigenesis in a majority of human pituitary adenomas. Moreover, detection of FGFR4 cytoplasmic staining may provide an ancillary diagnostic tool in the diagnosis of pituitary adenoma, particularly in equivocal cases.

The Journal of Clinical Endocrinology and Metabolism


Vertebral forward slippage in immature lumbar spine occurs following epiphyseal separation and its occurence is unrelated to disc degeneration. Is the pediatric spondylolisthesis a physis stress fracture of vertebral body?

Radiographic and histologic evaluation of a rat model of lumbar spine slippage. To clarify the pathomechanism of slippage in the immature spine. There are controversial hypotheses regarding the pathogenesis of slippage of the pediatric spine with pars defects. Some studies supported that disc degeneration was its cause, while others indicated the growth plate injury was the cause. An immature lumbar spine slippage model in 4-week-old rats was used. Following posterior destabilizing surgery, the lumbar spine was radiographically and histologically examined at 1, 3, 5, and 7 days after surgery. Radiographically, slippage occurred about 7% in the % slip on day 7, and no slippage was observed before day 5. Histologically, epiphyseal separation also appeared on day 7; before day 5, the growth plate showed no abnormalities. Within 7 days after the operation, the anulus fibrosus did not show any sign indicating degeneration. The nucleus pulposus was also normal up to day 7. The findings of this study support the hypothesis that vertebral forward slippage of the immature spine occurs following epiphyseal separation and its occurrence is unrelated to disc degeneration.


Unique cell membrane expression if topoisomerase-ll alpha as a useful diagnostic marker of liposarcoma

Topoisomerase-II alpha (Topo-II alpha) is known as a cell cycle-related intranuclear marker. To the best of our knowledge, the expression of Topo-II alpha on extranuclear sites has not been reported. The aim of the present study was to determine the usefulness of Topo-II alpha immunostaining for detecting the lipoblasts that are essential to diagnosing liposarcoma. Surgical specimens, including benign lipomatous tumors (four cases), well-differentiated liposarcomas (three cases), myxoid liposarcomas (six cases), pleomorphic liposarcomas (two cases), dedifferentiated liposarcomas (two cases), myxoid malignant fibrous histiocytomas (six cases), and one case of mesenteric panniculitis, were studied. Samples were immunostained using antibodies for Topo-II alpha, S-100 protein and Ki-67. In addition, we used the western blot method to investigate immunohistochemical-affinity in adipocytes. Mature adipocytes and lipoblasts in all of the benign and malignant lipomatous tumors intensively expressed cell contours positivity for Topo-II alpha. Cytoplasm of the lipoblasts occasionally reacted to the antibody and highlighted intracytoplasmic small unilocular, multivacuolated, or bubble-like patterns. Western blot analysis confirmed a 70 kDa product reactive to Topo-II alpha in the cell membrane fragment of mature adipocytes. S-100 protein expressed adipocytes and lipoblasts, but the detection of lipoblasts was not as easy as in Topo-II alpha immunostaining. Immunoreactivity of Ki-67 was limited to the nuclei, and the nuclear labeling index of Ki-67 correlated with that of Topo-II alpha. The immunoreactivity of Topo-II alpha for lipoblasts was more sensitive and obvious than those of S-100 protein. Immunostaining using the antibody for Topo-II alpha seems to be useful in recognizing lipoblasts that have been overlooked in hematoxylin-eosin-stained preparations, and is a useful marker for diagnosing liposarcoma.


Melanotic oncocytic metaplasia of the nasopharynx: a report of seven cases and review of the literature.


Hypopituitarism and diabetes insipidus with localized hypertrophic pachymeningitis (Tolosa-Hunt syndrome) associated with Hashimoto thyroiditis.

We report a 69-year-old woman with intracranial pachymeningitis showing hypopituitarism, diabetes insipidus, and Tolosa-Hunt syndrome associated with Hashimoto thyroiditis confirmed by autopsy. A large tumorous lesion of the hypothalamo-pituitary gland was revealed on magnetic resonance imaging, after the patient complained of gait and visual field disturbance. These symptoms subsided after thyroid hormone supplementation. Hypopituitarism and diabetes insipidus were diagnosed after cessation of the treatment by the patient herself. Multiple cranial nerve palsies and orbito-frontalgia appeared. Methylprednisolone pulse therapy improved the symptoms, but they recurred when the dose of glucocorticoid was decreased. The patient died of brain thrombosis. Autopsy revealed typical findings of Hashimoto thyroiditis and intracranial pachymeningitis involving the cranial base and pituitary gland. The high titer of rheumatoid factor and Hashimoto thyroiditis in this patient suggest an immunological role in the pathogenesis of pachymeningitis.

The American Journal of the Medical Sciences


Colorectal xanthomas with polypoid lesion:report of 25 cases.

Little attention has been paid to colorectal xanthoma. To clarify the clinical and pathological features of colorectal xanthoma, we report 28 colorectal xanthomas biopsied from 25 patients. All were composed of typical xanthoma cells and showed polypoid configuration. Median age of the patients was 64 years and there were 15 men and 10 women. Diabetes mellitus, constipation, and hyperlipidemia were found in two, one, and seven patients, respectively. Seventeen (60.7%) of the 28 polyps were located in the sigmoid colon and the remaining 11 in the rectum. Twenty-three polyps (82.1%) were sessile. Twelve (60.0%) of twenty polyps that were recorded were reddish in color. Only two polyps revealed a yellowish tone. Microscopically, foamy cells were present in the lamina propria, but the submucosa did not contain foamy cells. Immunohistochemically, the foamy cells invariably expressed extensive positivity for CD68. The colonic glands showed a deformity in the case with moderate to intense density of the foamy cells. The surface epithelium showed a hyperplastic change in 22 (78.6%) xanthomas. The colonic glands in four xanthomas were also associated with hyperplastic changes. The basement membrane of the surface epithelium was often thickened. Cell debris and proliferation of the capillaries were observed just below the surface epithelium in 19 (67.9%) and 22 (78.6%) xanthomas, respectively. Previous mucosal minute injury was suggested as the pathogenesis of colorectal xanthomas. Colorectal xanthomas were not identical to gastric and esophageal xanthoma, endoscopically or microscopically. We prefer the term "xanthomatous polyp" rather than xanthoma in the colorectal region. They may be regarded as a novel type of colorectal non-neoplastic polyp.


Xanthogranulomatous hypophysitis mimicking a pituitary neoplasm

Hypophysitis is an inflammatory disease of the pituitary gland that clinically and radiologically mimics pituitary tumors. We report here a case of xanthogranulomatous hypophysitis mimicking a pituitary neoplasm.A 65-yr-old woman presented with weight loss, fatigue, and visual disturbance. Computed tomography demonstrated a round cystic low-density mass with calcification in the sella. A T1-weighted magnetic resonance imaging scan showed most of the mass as hyperintense. The capsule of the mass was strongly enhanced by gadolinium. Endocrinologic examination revealed hypocorticism and hypothyroidism. Diabetes insipidus (DI) developed after the administration of hydrocortisone. The patient also had hallucination and delusions of persecution. Transsphenoidal surgery was performed. Histologic examination of the removed tissue showed central necrosis surrounded by accumulation of foamy cells and epithelioid cells. Several multinucleated giant cells were also seen. The foamy cells and epithelioid cells were immunopositive for Kp-1, a marker of macrophages. The patient made an uneventful postoperative recovery. Although DI and hypofunction of adenohypophysis persisted, the visual disturbance and psychiatric disorder were resolved. We have described an unusual inflammatory lesion of the pituitary in the sellar region that was mimicking neoplasm. A high level of clinical suspicion of inflammatory disorders is necessary for correct diagnosis and optimal management.


"Honeycomb Golgi" in pituitary adenomas: not a marker of gonadotroph adenomas.

The vacuolar change in Golgi complexes known as "honeycomb Golgi" has been described as the ultrastructural hallmark of a specific tumor that has been called the "female gonadotroph" adenoma of the human pituitary. Recently, a few adenomas presenting with Cushing's disease have been reported to exhibit this feature. To clarify the significance of a "honeycomb Golgi" in the classification of pituitary adenomas, we studied clinically nonfunctioning adenomas with or without "honeycomb Golgi" using immunohistochemistry for adenohypophysial hormones and RT-PCR for the cell-specific transcription factors Tpit that identifies corticotrophs and SF-1 that identifies gonadotrophs. All adenomas were from women. Among 20 adenomas with complete "honeycomb Golgi" change, gonadotrophin subunits were totally immunonegative, but ACTH was positive in a few cells of 12 adenomas. Among eight adenomas with partial vacuolar change of the Golgi complex, five were positive for gonadotrophins and two were positive for ACTH. A subgroup of these lesions were examined by RT-PCR and among eight adenomas with typical "honeycomb Golgi" one case expressed both Tpit and SF-1, probably due to contamination with normal pituitary and another expressed neither Tpit nor SF-1. Of the remaining six cases, Tpit was expressed in two cases and SF-1 in four. These findings indicate that "honeycomb Golgi" change can been seen in corticotroph adenomas as well as gonadotroph adenomas. The reason why this vacuolar change occurs only in females remains to be clarified.


Expression of cell adhesion molecules in chordomas: an immunohistochemical study of 16 cases.

Chordomas are thought to be tumors originating from notochord remnants characterized histologically by cohesive cells with epithelial features and by immunohistochemical expression of epithelial markers. To investigate the expression and distribution of cell adhesion molecules in chordomas, we immunohistochemically studied the expression of representative cell adhesion molecules, E-cadherin, P-cadherin, N-cadherin, beta-catenin, CD44, ICAM-1 (CD54), NCAM (CD56), and VCAM-1 (CD106) in 16 tumors from 16 patients (skull base, n=5; cervical, n=2; sacral, n=9) and 3 cases of fetal notochord. Of 16 tumors, 12 (75.0%) expressed membranous immunoreactivity for NCAM, 10 (62.5%) for VCAM-1, 9 (56.3%) for CD44, 8 (50.0%) for N-cadherin, 6 (37.5%) for beta-catenin, 4 (25%) for ICAM-1, and 1 (6.3%) for P-cadherin. Nuclear staining for E-cadherin was recognized in 11 (68.8%) tumors, and membranous staining for E-cadherin in 3 (18.8%); none of the tumors showed both nuclear and membranous staining. Intranuclear accumulation of beta-catenin was not found in any chordoma. One fetal notochord case showed immunoreactivity for N-cadherin, E-cadherin (some cells showed staining in both cytoplasm and nuclei), CD44 and beta-catenin. These results indicate that chordomas frequently express immunoreactivity for multiple adhesion molecules including VCAM, CD44 and N-cadherin, as well as for NCAM and E-cadherin, as previously reported. These molecules may participate in producing the cellular cohesion evident in tumor morphological structure. Although the precise underlying mechanisms remain to be elucidated, the high frequency of nuclear expression of E-cadherin (11 of 16 cases) may be diagnostically useful.


Human Pituitary Adenomas Infrequently Contain Inactivation of Retinoblastoma 1 Gene and Activation of Cyclin Dependent Kinase 4 Gene.

Components of cyclinD1/cyclin-dependent kinase 4 (CDK4)/p16INK4a/pRb pathway are the frequent target of many tumor types. We examined the role of retinoblastoma susceptibility gene (RB1) and the CDK4 gene in human pituitary tumorigenesis. For the RB1 gene, pRb expression and loss of heterozygosity (LOH) on 13q in pituitary adenomas were analysed. Immunostaining of pRb revealed lack of expression in 1 of 29 pituitary adenomas. In 4 of 31 pituitary adenomas, allelic imbalances including LOH of RB1 on 13q14 were detected. Three of 4 pituitary adenomas, in which one adenoma lacked pRb expression, had a common LOH region at least from D13S219 on 13q12.3-q13 to D13S265 on 13q31-32. Interphase fluorescence in situ hybridization with a probe of RB1 showed 2 copies of RB1 gene suggesting that mitotic recombination events, not deletion or chromosome loss, led to LOH in the 3 pituitary adenomas analyzed. All 27 exons, intron-exon boundaries, and essential promoter region of RB1 gene were then sequenced in genomic DNA from 4 pituitary adenomas with allelic imbalance on 13q14 including one adenoma without pRb expression and 3 adenomas with pRb expression. Any somatic mutations, insertions, or microdeletions in the RB1 gene were not detected in 4 pituitary adenomas. Methylation sensitive (MS)-polymerase chain reaction (PCR) and bisulfite sequencing analysis revealed hypomethylated status of CpG islands in the promoter region of the RB1 genes of 4 pituitary adenomas. In addition, activating mutations of CDK4 gene, which is a component of cyclinD1/CDK4/p16INK4a/pRb pathway, were not detected in 31 pituitary adenomas. Based on these results, it is concluded that somatic mutations of the RB1 gene or CDK4 gene do not appear to play a major role in pituitary tumorigenesis. This supports the presence of potential tumor suppressor gene(s) on 13q12.3-q13 to 13q31-32 in pituitary adenomas.


GH-Secreting Pituitary Adenomas Infrequently Contain Inactivating Mutations of PRKAR1A and LOH of 17q23-24.

The molecular events leading to the development of GH-secreting pituitary tumours remain largely unknown. Gsalpha (GNAS1) mutations are found in 27-43% of sporadic GH-secreting adenomas in the Caucasian population, but the frequency of GNAS1 mutations in Japanese and Korean acromegalic patients was reported to be lower, 4-9% and 16%, respectively. Other genes responsible for the tumourigenesis of GH-secreting pituitary adenomas have not been detected yet. PRKAR1A, which codes for the RIalpha regulatory subunit of cyclic AMP-dependent protein kinase A (PKA) on 17q23-24, was recently reported to contain inactivating mutations in some Carney complex families, which involved GH-secreting adenomas in about 10%. We re-evaluated the frequency of GNAS1 mutations and investigated PRKAR1A on the hypothesis that it might play a role in the tumourigenesis of GH-secreting adenomas. We analysed exons 8 and 9 of GNAS1 and all exons and the exon-intron boundaries of PRKAR1A with the PCR and by direct sequencing using genomic DNA extracted from 32 GH-secreting pituitary adenomas (30 GH-secreting adenomas, two GH and PRL-secreting adenomas) and 28 corresponding peripheral blood samples, and performed loss of heterozygosity (LOH) analysis of 17q23-24 with four microsatellite markers and intragenic markers of PRKAR1A. Seventeen of 32 (53.1%) tumours showed somatic-activating mutations of GNAS1: 16 (53.3%) of 30 GH-secreting adenomas and one of two GH and PRL-secreting adenomas. Neither inactivating somatic mutations of PRKAR1A nor LOH of 17q23-24 were detected in any of the tumours examined. We reconfirm the important role of activating mutations of GNAS1 in GH-secreting adenomas, and conclude that PRKAR1A does not play a significant role in the tumourigenesis.


A Predominant Increase of APC Gene Isoform with Exon 9a in a Case of Attenuated Familial Adenomatous Polyposis.


Cribriform-Morular Variant of Papillary Thyroid Carcinoma: A Pathological and Molecular Genetics Study with Evidence of Frequent Somatic Mutations in exon 3 of the b-catenin Gene.

The cribriform-morular variant (C-MV), an unusual and peculiar subtype of papillary thyroid carcinoma (PTC), has been observed frequently in familial adenomatous polyposis (FAP)-associated thyroid carcinoma and also in sporadic thyroid carcinoma. In this paper, five young women with the C-MV of PTC, aged 22-34 years at cancer diagnosis, are reported; two of them had attenuated FAP. Grossly, one FAP-associated tumour and one sporadic tumour were multicentric and the others were solitary. Histologically, the tumours were encapsulated and exhibited a combination of cribriform, follicular, trabecular, solid, and papillary patterns of growth, with morular areas. Immunohistochemically, the tumour cells showed cytoplasmic expression of thyroglobulin, neuron-specific enolase, epithelial membrane antigen, high- and low-molecular-weight cytokeratins, vimentin, and bcl-2 protein; nuclear expression of oestrogen and progesterone receptors, and retinoblastoma protein; and cytoplasmic and nuclear accumulation of beta-catenin. Germline mutations of the adenomatous polyposis coli (APC) gene were investigated using the protein truncation test in four subjects, including two FAP individuals. Germline APC mutation was identified in only one FAP patient with the multicentric C-MV of PTC, who had a thymidine deletion at codon 512, resulting in a frameshift leading to a premature stop codon. No loss of heterozygosity of loci close to the APC gene was detected in tumour tissues from these four patients. Somatic mutation analysis of exon 3 of the beta-catenin gene (CTNNB1) revealed alterations in seven tumours from all five individuals: one at a serine residue (codon 29), three at amino acids adjacent to serine or threonine residues (codons 22, 39, and 44), and three at other amino acids (codons 49, 54, and 56). Moreover, each of two different tumours examined from two patients with the multicentric C-MV of PTC, had different somatic mutations of the CTNNB1 gene. Taken together, these data suggest that accumulation of mutant beta-catenin contributes to the development of the C-MV of PTC.


Spindle Epithelial Tumor with Thymus-like Differentiation of the Thyroid: A Case Report with Pathological and Molecular Genetics Study.

We report an unusual case of spindle epithelial tumor with thymus-like differentiation (SETTLE) of the thyroid present in a 6-year-old boy. The tumor, located at both the left lobe and isthmus, was a circumscribed mass with slightly gritty whorled appearance. Microscopically, the lobulated, highly cellular, spindle cell neoplasm was arranged in intersecting bundles and fascicles separated by fibrous bands. Benign-appearing glands entrapped within fibrous bands and foci of squamous differentiation within spindle cells were observed. Immunohistochemically, the spindle cells were diffusely positive for cytokeratins, vimentin, and alpha-smooth muscle actin and patchily reactive for muscle-specific actin and epithelial membrane antigen, exhibiting myoepithelial differentiation. The spindle cells were also patchily immunopositive for p53 protein. Molecular genetic analysis revealed Ki-ras gene mutations at codons 13 (GGC(gly) to AGC(ser)) and 15 (GGC(gly) to AGC(ser)) on the same allele. Mutation of the p53 gene was not detected. This is the first report on Ki-ras oncogene mutations in a case of SETTLE.


空腹時の血漿インスリン濃度は常に低値で，グルカゴンにより著明なインスリン分泌反応が誘導されたインスリノーマの1例


Role of E-cadherin, alpha-, beta-, and gamma-catenins, and p120 (cell adhesion molecules) in prolactinoma behavior.

E-cadherin/catenin complex regulates cellular adhesion and motility and is believed to function as an invasion suppressor system. In a number of cancers, abnormal and reduced expression of E-cadherin/catenin complex is associated with tumor invasion and metastasis. Prolactinomas show frequent invasion on the surrounding structures, despite their histologically benign nature. Furthermore, gender-based differences in endocrine and surgical findings are found in patients with prolactinoma. To understand biological factors governing prolactinoma behavior, this study analyzed the expression of E-cadherin; alpha-, beta-, and gamma-catenins; p120; and cell proliferation marker MIB-1 labeling index in 13 invasive tumors (9 in men, 4 in women), 26 noninvasive tumors (4 in men, 22 in women), and 8 normal anterior pituitaries by immunohistochemistry. Immunostaining of E-cadherin; alpha-, beta-, and gamma-catenins; and p120 showed a membranous pattern of reactivity and generally stronger in normal pituitaries than in prolactinomas. Expression of E-cadherin and beta-catenin was significantly lower in invasive than in noninvasive prolactinomas (P <.002 and P <.005, respectively), and reduced expression of E-cadherin and beta-catenin was more frequent in invasive than in noninvasive prolactinomas (P <.001 and P <.05, respectively); in contrast, gamma-catenin expression showed higher in invasive than in noninvasive prolactinomas (P <.05). Expression of E-cadherin was significantly lower in macroprolactinomas than in microprolactinomas (P <.01), and decreased expression of E-cadherin and beta-catenin predicted high MIB-1 expression (P <.05). Moreover, the expression of E-cadherin and beta-catenin was significantly lower in macroprolactinomas in men than in those in women (P <.01 and P <.02, respectively). No statistical correlations were observed between expression of alpha-catenin, p120, and clinicopathologic features. In conclusion, the reduction of E-cadherin and beta-catenin expression was related to invasiveness and proliferative status of prolactinomas and correlated with the more aggressive behavior of prolactinomas in men compared with in women.


Morules with optically clear nuclei in ovarian borderline endometetrioid tumor

Optically clear nuclei (OCN) have been observed in morules of some neoplasms and in some conditions unrelated to the development of the morules. We first report a case of ovarian borderline endometrioid tumor (BET) showing the morules associated with OCN. The patient was a 47-year-old premenopausal woman with a left ovarian cystic tumor, atypical endometrial hyperplasia, and elevated serum levels of FSH, LH, estradiol, and CA 125. The resected ovarian tumor measured 6 cm in diameter, and showed a papillary growth. Histologically, the ovarian tumor was consistent with BET, and the morules with OCN were scattered. Immunohistochemically, OCN were proven to be rich in biotin. An aberrant nuclear expression of beta-catenin was observed in both the tumor cells and the morular cells. Our case may suggest the possibility that the appearance of OCN with or without morules in ovarian tumors is related to endometrioid differentiation of the tumor cells, and should be recognized as a diagnostic clue of ovarian endometrioid tumors. Although female sex hormones have been reported to play a role in the occurrence of OCN, the participation of beta-catenin mutation has also been suggested.

卵巣の境界悪性腫瘍の類内膜腫瘍で，淡明な核を有し扁平上皮性桑実様細胞巣を呈するはじめての症例報告をする．47歳，閉経前の女性で，左卵巣に嚢胞性の腫瘤があり，子宮内膜に異型的過形成を認め，血清中のFSH，LH，エストラジオール，CA125の上昇があった．抽出卵巣は直径6cmで乳頭状発育をしていた．組織学的には淡明な核を有する扁平上皮性桑実様細胞巣が散見された．免疫組織化学的に淡明核はビオチンに富んでおり，腫瘍細胞，桑実様細胞巣の細胞質と核にベータカテニンの発現が認められた．淡明核は腫瘍の類内膜への分化と関係があることが示唆された．


Gastric inverted hyperplastic polyp. Report of four cases and relation to gastritis cystica profunda

Gastric inverted hyperplastic polyp (IHP) is a rare type of gastric polyp, and is characterized by downward growth of the hyperplastic mucosal components into the submucosa. To the best ofour knowledge, 16 gastric IHP cases have been described in the English literature, but the pathogenesis has not been established. We report the clinical and pathological findings of four gastric IHP cases. The lesions were mainly composed of hyperplastic foveolar-type glands with focal cystic dilatation. Pyloric type glands, endocrine cells, acinic cell metaplasia, and smooth muscle bundles were also seen as components of the polyp. Two cases (cases 1 and 4) coexisted with multifocal gastritis cystica profunda (GCP) and gastric adenocarcinoma. Case 4 furthermore exhibited an intermediate form between IHP and GCP. We suggest that IHP may be GCP associated with exaggeratedly hyperplastic and metaplastic changes. In case 4, the coexisting gastric carcinoma was mainly located in the submucosa, whilst the mucosal component was minimal. Five out of twenty reported gastric IHP cases, including our cases, coexisted with gastric adenocarcinoma. These facts would lead us to further investigate the relation between gastric IHP and carcinoma.


Expression of E-cadherin, b-catenin, and Ki-67 in goblet cell carcinoids of the appendix: an immunohistochemical study with clinical correlation.

Goblet cell carcinoid (GCC) of the appendix is a rare entity, of which both the histogenesis and biologic behavior remain controversial, and prognostic tools and therapeutic strategies for this unusual tumor have yet to be defined. The aim of this study was to analyze expression of E-cadherin and b-catenin in GCCs of the appendix with long-term follow-up data as related to the expression of Ki-67 proliferation marker to provide a rationale for treatment guidelines. We analyzed the expression of E-cadherin, b-catenin, and Ki-67 in 11 GCCs of the appendix and control groups of typical carcinoids of the large intestine (n = 29), well to moderately differentiated adenocarcinomas of the colon (n = 10), poorly differentiated adenocarcinomas of the colon (n = 12), and normal appendiceal tissues (n = 10). There was no significant difference between the GCCs and normal appendiceal tissues regarding the expression of E-cadherin or b-catenin (p = 0.297 and 0.103, respectively). The percentage of positive GCC cells ranged between 0.52 and 10.35% (4.27 +/- 0.80), and only one case had a score >10%. Metastatic tumor spread and death were found in high MIB-1 labeling index (LI) cases of GCC (>3%). Our findings suggest that the behavior of the majority of GCCs might be indolent and different from adenocarcinomas because of the preserved expression of E-cadherin and b-catenin and relatively low MIB-1 LI. However, some of these tumors act aggressively and MIB-1 LI might be a good parameter to determine the therapeutic procedure.


Downregulation of E-cadherin and its Undercoat Proteins in Pituitary Growth Hormone Cell Adenomas with Prominent Fibrous Bodies.

The cadherin-catenin complex regulates cellular adhesion and motility, and genetic alterations in these molecules play a critical role in multistage tumorigenesis. In this study, the expression of three major type I classic cadherins E-, N-, and P-cadherin and their undercoat proteins alpha-, beta-, and gamma-catenin, and pp120 was investigated in 127 pituitary adenomas and 10 normal adenohypophyseal glands using an immunohistochemical technique with highly specific monoclonal antibodies. In normal pituitary glands, E-cadherin, catenins, and pp120 were strongly expressed on almost all hormone-producing cell-cell boundaries, N-cadherin was weakly immunoreactive on a few cell-cell boundaries, and P-cadherin was negative. In pituitary adenomas, a correlation was not identified among expression of E-cadherin, catenins, or pp120 with patient age, sex, hormone level, tumor size, and/or invasiveness, respectively. Expression of E-cadherin, catenins, and pp120 was significantly reduced in 24 growth hormone (GH) cell adenomas with prominent fibrous bodies compared with the other subtypes of pituitary adenomas and normal pituitary glands (p < 0.0001, respectively). Methylation-specific polymerase chain reaction analysis revealed that the E-cadherin gene promoter region was methylated in 6 of 16 (37.5%) GH cell adenomas with prominent fibrous bodies examined, 2 of which displayed total methylation, but not in 10 GH cell adenomas without fibrous bodies. No mutation of exon 3 of the beta-catenin gene was found in 16 GH cell adenomas with prominent fibrous bodies or in 10 other subtypes of pituitary adenomas that showed unremarkable intracellular presence of beta-catenin protein. In conclusion, the decreased expression of the E-cadherin catenin complex and methylation of the E-cadherin gene promoter region only in GH cell adenomas with prominent fibrous bodies may be an event associated with the formation of fibrous bodies.


Alterations of E-cadherin, a-catenin and b-catenin expression in neuroendocrine tumors of the gastrointestinal tract.

Neuroendocrine tumors (NETs) of the gastrointestinal tract comprise a heterogeneous group of neoplasms arising from the diffuse neuroendocrine system. These tumors strongly differ from each other on the basis of different pathogenetic, clinical, functional, histological, and prognostic patterns. Previous studies have shown that abnormal and reduced expression of the E-cadherin/catenin complex in several human cancers is associated with tumor dedifferentiation, advanced clinical stages, and poor survival rate. We assessed correlations between the expression of E-cadherin and catenins, Ki-67, and the following clinicopathological factors: age, embryological site of origin, size, histological growth pattern, the depth of penetration into the intestinal wall, and the presence of metastasis. In this study, reduction of membranous E-cadherin expression to a variable degree was detected in more than two-thirds (42 of 51) of gastrointestinal NETs (19 foregut, 8 midgut, and 24 hindgut) belonging to the complete neuroendocrine neoplastic spectrum [18 well-differentiated NETs, 22 well-differentiated neuroendocrine carcinomas (NECs), and 11 poorly differentiated NECs]. The reduction of E-cadherin expression was concomitant with the reduction of alpha-catenin (44 of 51) and beta-catenin (35 of 51) expression. Our immunohistochemical analysis demonstrated significant differences of percentage of membranous positive cells of E-cadherin, alpha-catenin, or beta-catenin between normal tissues and well-differentiated NETs (P=0.0038, P=0.004, and P=0.0329, respectively), well-differentiated NECs (P<0.001, P<0.001, and P<0.001, respectively) and poorly differentiated NECs (P=0.0002, P<0.0002, and P=0.0002, respectively). Among the gastrointestinal NETs, there were significantly more positive cells of E-cadherin, alpha-catenin, or beta-catenin in well-differentiated NETs than well-differentiated NECs (P=0.0006, P=0.0065, and P=0.0001, respectively) or poorly differentiated NECs (P=0.0053, P=0.0041, and P<0.001, respectively). MIB-1 labeling index generally showed a low proliferative activity in well-differentiated NETs (0.49+/-0.37) and well-differentiated NECs (0.662+/-0.66). A high proliferation rate was observed in poorly differentiated NECs (41.518+/-16.59). MIB-1 labeling index was significantly higher in poorly differentiated NECs than well-differentiated NETs and well-differentiated NECs (P<0.0001 and P<0.0001, respectively). E-cadherin, alpha-catenin, and beta-catenin expression were correlated significantly with transmural tumor invasion (P<0.0001, P=0.0001, and P<0.0001, respectively) and with size (P=0.0013, P=0.0001, and P<0.0001, respectively). These results indicate that the alteration in the E-cadherin/catenin expression may be involved in the growth and progression of gastrointestinal NETs.


二次的変性を伴った側頭骨軟骨芽細胞腫の1例

二次的変性を伴った側頭骨軟骨芽細胞腫の1例


Spectrum of different types of hypophysitis : A clinicopathologic study of hypophysitis in 31 cases

下垂体炎31例を臨床病理学的に，リンパ球性下垂体前葉炎19例，リンパ球性下垂体後葉炎3例，肉芽腫性下垂体炎5例，黄色肉芽腫性下垂体炎2例の4つに分類した．それぞれの下垂体炎で，下垂体にリンパ球の浸潤と種々の程度の繊維化が見られた．リンパ球性下垂体前葉炎の2例ではリンパ球形質細胞浸潤は軽度ながら，副腎皮質刺激ホルモン陽性細胞が選択的に消失していた．このことは下垂体炎における下垂体機能低下は高度な浸潤細胞によって非特異的な前葉細胞の破壊によるものと，選択的な前葉細胞の破壊によるものの少なくとも二つの機序が考えられた．


Superantigens and autoantigens may be involved in the pathogenesis of gastric mucosa-associated lymphoid tissue lymphoma

To clarify the origin of tumor cells and the possible role of antigens in the pathogenesis of mucosa-associated lymphoid tissue lymphoma (MALTL) of the stomach, we analyzed the DNA sequences of the immunoglobulin (Ig) variable region gene from tumor cells of 4 patients with low-grade and 2 patients with high-grade MALTL associated with Helicobacter pylori infection. There were few somatic mutations in the Ig variable region gene, but intraclonal variations were observed in 2 of the 4 low-grade MALTL cases. In the remaining 2 low-grade MALTL and 1 of the 2 high-grade MALTL cases, somatic mutations and intraclonal variations were evident. In contrast, somatic mutations in the Ig variable region gene were prominent, but intraclonal variation was absent in the other high-grade MALTL cases. The deduced amino acid sequences of the antigen-binding fragments (Fab) from 2 MALTL cases revealed homology with anti-thyroglobulin antibodies, 3 MALTL cases with lupus anti-DNA antibodies, and 1 MALTL case with a rheumatoid factor. Furthermore, the heavy-chain variable region 3 (V(H)3) family genes were used in 5 of the 6 MALTL cases and had conserved amino acid residues for binding to staphylococcal protein A (SpA), a superantigen of B cells. Considering that another superantigen, protein Fv, competes for binding to Fab with SpA and has been shown to play a major role in immune defenses against gut pathogens, SpA and possibly protein Fv may contribute to the development of MALTL. Thus, these observations suggest that most gastric MALTLs arise from memory B cells that are preliminarily activated by superantigens and autoantigens.