Yamato Suemitsu, Yusuke Ono, Yusuke Mizukami, Juanjuan Ye, Keiko Yamakawa, Takeshi Takamoto, Yuko Narusawa, Yuri Mukai, Manabu Takamatsu, Atsuko Nakazawa, Mari Mino-Kenudson, Toshio Kumasaka and Yoko Matsuda : Mutation and Genetic Heterogeneity., Frontiers in Oncology, 11, 2021.
(要約)
In conclusion, the clear and basophilic cells of the tumor were supposedly derived from the same clone and subsequently acquired additional mutations. This is the first report of clonal evolution in pancreatoblastoma.
Yuko Narusawa, Masanao Yokohira, Keiko Yamakawa, Juanjuan Ye, Misa Tanimoto, Linxuan Wu, Yuri Mukai, Katsumi Imaida and Yoko Matsuda : Relationship between Lung Carcinogenesis and Chronic Inflammation in Rodents., Cancers, 13, 12, 2021.
(要約)
mutations) have been widely studied. Animal studies have revealed that chronic inflammation significantly enhances lung carcinogenesis, and inhibition of inflammation suppresses cancer progression. Moreover, the reduction in tumor size by suppression of inflammation in animal experiments suggests that chronic inflammation influences the promotion of tumorigenesis. Here, we review rodent lung tumor models induced by various chemical carcinogens, including tobacco-related carcinogens, and transgenics, and discuss the roles of chronic inflammation in lung carcinogenesis.
Haiyan Xu, Keizo Hiraishi, Lin-Hai Kurahara, Yuko Narusawa, Xiaodong Li, Yaopeng Hu, Yoko Matsuda, Heping Zhang and Katsuya Hirano : Probio-M9 on Colitis-Associated Carcinogenesis by Restoration of the Gut Microbiota in a Mouse Model., Nutrients, 13, 4, 2021.
(要約)
/day) was orally administered for 7 days. Animals were sacrificed at 20 weeks after AOM administration and immunohistochemical staining and Western blotting was performed. The α-diversity of microflora (Shannon index), principal coordinate analysis, and distribution of intestinal bacterium genera and metabolic pathways were compared. The AOM/DSS group showed weight loss, diarrhea, intestinal shortening, increased number of colon tumors, proliferating tumorigenesis, increased inflammation score, fibrosis, increased CD68+, or CD163+ macrophage cells in the subserosal layer of non-tumor areas. Inflammation and tumorigenesis ameliorated after Probio-M9 treatment. Fecal microbial functions were altered by AOM/DSS treatment. Probio-M9 significantly upregulated the fecal microbial diversity and reversed fecal microbial functions. Thus, Probio-M9 could suppress tumor formation in the large intestine by regulating the intestinal environment and ameliorating inflammation, suggesting its therapeutic potential for treatment of inflammation and colitis-associated tumorigenesis.
Masanao Yokohira, Minoru Oshima, Keiko Yamakawa, Juanjuan Ye, Yuko Narusawa, Reiji Haba, Yuki Fukumura, Kenichi Hirabayashi, Hiroshi Yamaguchi, Motohiro Kojima, Keiichi Okano, Yasuyuki Suzuki and Yoko Matsuda : Adequate tissue sampling for the assessment of pathological tumor regression in pancreatic cancer., Scientific Reports, 11, 1, 2021.
(要約)
Standardized pathological evaluation of the regression assessment of neoadjuvant pancreatic cancer is necessary to improve prognostication and compare treatment outcomes in clinical trials. However, appropriate tissue sampling from surgically resected pancreatic cancer after neoadjuvant therapy has not been elucidated. We compared the tumor regression scores in the largest cancer slide determined macroscopically or histologically. We reviewed all slides and macroscopic photos of cut surfaces from resected pancreas of patients treated with neoadjuvant chemotherapy (n = 137; chemoradiotherapy or chemotherapy). The tumor regression scores (the Evans, College of American Pathologists, Japanese Pancreas Society grading systems, and Area of Residual Tumor [ART] score) were evaluated for the largest tumor slide determined by macroscopy or histologically as well as all slides from the resected pancreas. The largest cancer slides determined macroscopically and histologically were discrepant in 26% of the cases. Cancer cells were not detected in the largest macroscopically defined cut slides in 3%. Only ART scores assessed in the largest histological slides displayed significant difference in overall survival. We recommend obtaining the largest histological slides to provide adequate assessment for regression of neoadjuvant-treated pancreatic cancer. Sufficient sampling to detect the largest histological slides would be mandatory.
Keiko Yamakawa, Juanjuan Ye, Yuko Narusawa and Yoko Matsuda : Pathological Changes in Pancreatic Carcinogenesis: A Review., Cancers, 13, 4, 2021.
(要約)
Despite advances in diagnostics and therapeutics, the prognosis of pancreatic cancer remains dismal. Because of a lack of early diagnostic methods, aggressive local progression, and high incidence of distant metastasis, most pancreatic cancers are inoperable; therefore, the characteristics of early pancreatic cancer have not been well understood. Autopsy studies revealed the characteristics of prediagnostic pancreatic malignancies, including precancerous lesions, early stage pancreatic cancer, and pancreatic cancer without clinical symptoms (occult cancers). Animal models using hamsters and genetically engineered mice have focused on mechanisms of carcinogenesis, thereby providing insights into risk factors and prevention and serving as a preclinical test for the development of novel diagnostic and treatment modalities. In this review, we have summarized pathological changes in the pancreas of humans and experimental animals during carcinogenesis.
Keiko Yamakawa, Masanao Yokohira, Yuko Narusawa, Sosuke Kishi, Shohei Kanie and Katsumi Imaida : Activation of MEK1/2-ERK1/2 signaling during NNK-induced lung carcinogenesis in female A/J mice., Cancer Medicine, 5, 5, 903-913, 2016.
(要約)
The extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway is activated by several growth factors and mitogens, and upregulation has been noted in many human cancers, including examples in the lung. In this study, to study the association of ERK1/2 activation with mutation of Kras encoding an upstream activator of ERK1/2 in lung premalignant lesions, we immunohistochemically examined expression of phosphorylated forms of ERK1/2 (pERK1/2) and MAP/ERK kinase 1/2 (pMEK1/2) proteins and correlation between ERK activation and mutation of Kras encoding an upstream activator of ERK1/2, in a mouse lung carcinogenesis model. Female 7-week-old A/J mice were administered a single dose of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), then maintained without additional treatment until sacrifice at week 52. Histopathologically, adenocarcinomas, adenomas and hyperplasias were observed in the lung. pMEK1/2 was expressed mostly in the cell cytoplasm in all three. In contrast, pERK1/2-positive cells were also relatively rare in any histological types as compared with level of pMEK1/2 expression. However, pERK1/2-positive cells in adenocarcinoma were still markedly more common than in hyperplasias and adenomas (~5-fold, ~4-fold; P < 0.01). Activating mutations of Kras gene at codons 12, 13 and 61 were detected in the majority of adenomas and adenocarcinomas, but without any significant relation to pERK1/2 expression. These results suggest that activation of ERK1/2 plays a key role in malignant transformation during lung carcinogenesis featuring Kras mutaion. Activation of ERK1/2 in lung premalignant lesions was little regardless of the mutation of Kras, and ERK1/2 activation in NNK-induced mouse lung carcinogenesis may be regulated not only by Kras mutation but also other signaling pathway or regulatory factor.
Masanao Yokohira, Nozomi Hashimoto, Toshitaka Nakagawa, Yuko Narusawa, Keiko Yamakawa, Sosuke Kishi, Shohei Kanie, Fumiko Ninomiya, Kousuke Saoo and Katsumi Imaida : Long-Term Chronic Toxicity and Mesothelial Cell Reactions Induced by Potassium Octatitanate Fibers (TISMO) in the Left Thoracic Cavity in A/J Female Mice., International Journal of Toxicology, 34, 4, 325-335, 2015.
(要約)
The present study was conducted to examine the chronic effects of potassium octatitanate fibers (trade name TISMO; chemical formula K2O·6TiO2) on the mouse lung and thoracic cavity. This method of infusion was employed to examine the direct effects of the fibers to the pleura. In the present study, 52- and 65-week experiments were employed to examine the long-term chronic effects after infusion of fiber-shaped TISMO into the thoracic cavities of A/J mice. Following this infusion, TISMO fibers were observed in the alveoli, indicating penetration through the visceral pleura. The additional histopathological detection of TISMO fibers in the liver, spleen, kidneys, ovary, heart, bone marrow, and brain of TISMO-infused mice indicated migration of the fibers out from the thoracic cavity. Atypical mesothelial cells with severe pleural proliferation were observed, but malignant mesotheliomas were not detected. This study demonstrated that intrathoracic infusion of TISMO fiber did not cause malignant mesothelioma but did cause severe chronic inflammation and proliferation of pleural mesothelial cells.
Yuko Narusawa, Masanao Yokohira, Nozomi Hashimoto, Keiko Yamakawa, Sosuke Kishi, Fumiko Ninomiya, Shohei Kanie, Kousuke Saoo and Katsumi Imaida : Rat strain differences in levels and effects of chronic inflammation due to intratracheal instillation of quartz on lung tumorigenesis induced by DHPN., Experimental and Toxicologic Pathology, 66, 8, 391-401, 2014.
(要約)
Chronic inflammatory effects of single intratracheal instillation (i.t.) of quartz on rat lung tumorigenesis were examined using 4 different animal models. At first, in order to determine an appropriate dose of quartz i.t. to promote lung tumorigenesis, F344 male rats were administrated single 0, 0.5, 1, 2 or 4 mg quartz/rat after initiation by N-bis(2-hydroxypropyl) nitrosamine (DHPN). Further studies were performed to examine strain differences of the effects of chronic inflammation caused by quartz i.t. in 3 strains of rat, i.e. F344, Wistar-Hannover and SD. Each was instilled with 2mg quartz/rat after DHPN administration and sacrificed in week 24. In addition, strain differences in generation of inflammation were determined at days 1 and 28. Finally, for determination of long-term effects period, F344 and Wistar-Hannover rats were similarly treated, but the experiment was terminated at week 52. In F344 rats, the tumor areas in DHPN treated groups showed a tendency to increase along with the dose of quartz. F344 rats demonstrated the highest and Wistar-Hannover rats the lowest sensitivity to quartz in acute and chronic phases in the 3 strains. In 52 week, in F344 rats, the multiplicity of tumors and the serum concentration of IL-6 in the group treated with DHPN and quartz were significantly increased. The present experiments indicated that chronic inflammation due to quartz instillation exerted promoting effects on lung carcinogenesis in F344, SD and Wistar-Hannover rats. The strain differences in tumor promotion appeared to correlate with inflammatory reactions to quartz and increase of IL-6.
Masanao Yokohira, Keiko Yamakawa, Yuko Narusawa, Takamasa Numano, Fumio Furukawa, Sosuke Kishi, Fumiko Ninomiya, Shohei Kanie, Hiroko Hitotsumachi, Kousuke Saoo and Katsumi Imaida : Immunohistochemical characteristics of surfactant proteins a, B, C and d in inflammatory and tumorigenic lung lesions of f344 rats., Journal of Toxicologic Pathology, 27, 3-4, 175-182, 2014.
(要約)
Surfactant proteins (SPs), originally known as human lung surfactants, are essential to respiratory structure and function. There are 4 subtypes, SP-A, SP-B, SP-C and SP-D, with SP-A and SP-D having immunological functions, and SP-B and SP-C having physicochemical properties that reduce the surface tension at biological interfaces. In this experiment, the expressions of SP-A, SP-B, SP-C and SP-D in lung neoplastic lesions induced by N-bis (2-hydroxypropyl) nitrosamine (DHPN) and inflammatory lesions due to quartz instillation were examined and compared immunohistochemically. Formalin fixed paraffin embedded (FFPE) lung samples featuring inflammation were obtained with a rat quartz instillation model, and neoplastic lesions, hyperplasias and adenomas, were obtained with the rat DHPN-induced lung carcinogenesis model. In the rat quartz instillation model, male 10-week old F344 rats were exposed by intratracheal instillation (IT) to quartz at a dose of 2 mg/rat suspended in saline (0.2 ml) on day 0, and sacrificed on day 28. Lung tumorigenesis in F344 male rats was initiated by DHPN in drinking water for 2 weeks, and the animals were then sacrificed in week 30. Lung proliferative lesions, hyperplasias and adenomas, were observed with DHPN, and inflammation was observed with quartz. The expressions of SP-A, SP-B, SP-C and SP-D were examined immunohistochemically. SP-B and SP-C showed strong expression in lung hyperplasias and adenomas, while SP-A and SP-D were observed in mucus or exudates in inflammatory alveoli. These results suggest the possibility that SP-B and SP-C are related to lung tumorigenesis.
Masanao Yokohira, Sosuke Kishi, Keiko Yamakawa, Yuko Narusawa, Fumiko Ninomiya, Shigemi Kinouch, Junko Tanizawa, Kousuke Saoo and Katsumi Imaida : Napsin A is possibly useful marker to predict the tumorigenic potential of lung bronchiolo-alveolar hyperplasia in F344 rats., Experimental and Toxicologic Pathology, 66, 2-3, 117-123, 2013.
(要約)
There are 2 types of bronchiolo-alveolar hyperplasia found in rat lungs. One is 'inflammatory hyperplasia' with a potential to recover in future with removal of the stimulating insult and the other is 'latent tumorigenic hyperplasia' as an independent preneoplastic lesion for adenocarcinoma. In the present experiment, we focused on rat lung bronchiolo-alveolar hyperplasia induced by 4-(methyl-nitrosamino)-1-(3-pyridyl)-1-butanone (NNK), which decreases with time after induction and reverts to normal, or by N-bis(2-hydroxypropyl)nitrosamine (DHPN), with tumorigenic potential to progress to adenoma and adenocarcinoma. Though NNK is a typical carcinogen inducing lung adenocarcinoma in female A/J mice, the tumorigenic potential by NNK in rats is weak. Differences between hyperplasias induced by DHPN and by NNK were here examined immunohistochemically. Formalin fixed paraffin embedded lung samples with hyperplastic and inflammatory lesions were obtained from rats exposed to DHPN or NNK and from lung inflammation models induced with fine particles like CuO, NiO and quartz. The 19 markers were examined immunohistochemically. Napsin A, in the inflammatory lesions and hyperplasia induced by NNK, was positive for macrophages and secretions in the alveoli spaces but less so in the walls of the alveoli. In the proliferative lesions including hyperplasia induced by DHPN, strong positive staining for napsin A was observed in the walls of the alveoli. Thus high expression was suggested to be possibly useful for detecting tumorigenic potential of rat lung hyperplasia.
Fumiko Ninomiya, Masanao Yokohira, Sosuke Kishi, Yuko Narusawa, Keiko Yamakawa, Tatsushi Inoue, Toshiya Kuno and Katsumi Imaida : Gender-dependent effects of gonadectomy on lung carcinogenesis by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in female and male A/J mice., Oncology Reports, 30, 6, 2632-2638, 2013.
(要約)
The present study was conducted to investigate the effects of gonadectomy on lung carcinogenesis in female and male mice, and to determine an association between sex hormone and lung carcinogenesis. Female and male A/J mice were divided into gonadectomized and unoperated control groups and all animals were treated intraperitoneally with 1 or 2 injections of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) at the dose of 2 mg/mouse. The mice were sacrificed 18 or 56 weeks after surgery. Serum levels of estradiol in females and testosterone in males were confirmed to be decreased by gonadectomy. Lung white nodules were detected in all mice of all groups. In the control groups of 18- and 56-week studies, the multiplicities of lung nodules in females were significantly greater than in males. In males in the 56-week study, the multiplicity of macroscopical lung nodules, bronchiolo-alveolar hyperplasias, adenomas and tumors (adenomas and adenocarcinomas) showed significant increase with castration. In females in the 18-week study, the multiplicity of adenomas decreased significantly by ovariectomy. Based on the results of the present study, female A/J mice were confirmed to be more susceptible to NNK-induced lung carcinogenesis than males. Furthermore, it was suggested that the process is inhibited by testosterone and accelerated by estradiol. These findings indicate the possibility that sex hormones play important roles in determining sex differences in lung carcinogenesis in the A/J mice initiated by NNK.
Masanao Yokohira, Yuko Narusawa, Keiko Yamakawa, Sosuke Kishi, Fumiko Ninomiya, Kousuke Saoo and Katsumi Imaida : Strain differences in pleural mesothelial cell reactions induced by potassium octatitanate fibers (TISMO) infused directly into the thoracic cavity., Experimental and Toxicologic Pathology, 65, 6, 925-932, 2013.
(要約)
Although we have previously reported that the fiber-shaped TISMO, morphologically similar to asbestos, can induce a severe mesothelial reaction in A/J mice, it is important to clarify any strain differences. In the present study, female A/J, C3H/HeN, ICR and C57BL/6 mice were therefore employed as test strains. At the beginning of the experiment, all mice underwent a left thoracotomy and direct administration of 3mg of TISMO particles suspended in 0.2 ml saline into the left thorax. The experiment was terminated after 21 weeks and all groups were sacrificed and the mesothelium and main organs were examined histopathologically. To contribute to mechanistic analysis, iron staining with Berlin blue and Turnbull's blue, and immunostaining for calretinin were also performed. The present experiment demonstrated only minor strain differences in the degree of pleural reaction to TISMO. However, there was clear variation in the iron and lymphocyte accumulation in the pleura and in the liver. This difference in response to TISMO fibers in vivo is important information when considering the development of mesothelioma as an animal model and the extrapolation to human risk from such animal studies.
Masanao Yokohira, Yuko Narusawa, Nozomi Hashimoto, Keiko Yamakawa, Fumiko Ninomiya, Sosuke Kishi, Kousuke Saoo and Katsumi Imaida : Toxicity of nicotine by repeated intratracheal instillation to f344 rats., Journal of Toxicologic Pathology, 25, 4, 257-263, 2012.
(要約)
In vivo, nicotine in cigarette smoke induces various effects not only on the respiratory system but also the central and peripheral nerve systems, circulatory organs and digestive organs, and there is a possibility of promotion of lung tumorigenesis. The present experiment was conducted to examine histopathological changes caused by nicotine in the lung with repeated intratracheal instillation (i.t.). Six-week-old male F344 rats were administered nicotine by i.t. at doses of 0.05, 0.1 and 0.2 mg nicotine/rat every 3 weeks beginning at week 4, for up to a total of 9 times and were then sacrificed at week 30. The total number of administrations, total dose of nicotine and effective number of rats were 9 times, 0.45 mg and 5 rats and 4 times, 0.20 mg and 5 rats for the 0.05 mg nicotine/rat group; 3 times, 0.30 mg and 5 rats and 4 times, 0.40 mg and 3 rats for the 0.1 mg group; and 3 times, 0.60 mg and 3 rats for the 0.2 mg group, respectively. As a control group, 5 rats were administered 0.2 ml saline/rat 9 times. Some rats administered 0.1 and 0.2 mg nicotine suffered convulsions just after administration. Histopathologically, though proliferative changes were not observed, neutrophil infiltration, edema and fibrosis in the lung were induced by nicotine. In conclusion, repeated treatment of nicotine promoted neurologic symptoms in the acute phase, and strong inflammation in the lungs in the chronic phase, even at a low dose. Toxicity of nicotine is suggested to depend not on total dose of nicotine in the experiment but rather on repeated injury with consecutive administration.
Satoshi Suzuki, Masanao Yokohira, Nozomi Hashimoto, Kousuke Saoo, Yoko Matsuda, Keiko Yamakawa, Yuko Narusawa, Toshiya Kuno and Katsumi Imaida : Different threshold levels for 2-amino-3,8 dimethylimidazo[4,5-f]quinoxaline (MelQx) initiation of lung and colon carcinogenesis and the effects of an additional initiation by 4 (methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in A/J mice., Molecular Medicine Reports, 3, 2, 301-307, 2010.
(要約)
The existence of possible threshold dose levels of genotoxic carcinogens for carcinogenesis is of particular interest for human risk assessment. Recently, no observed effect levels (NOELs) for various hepatocarcinogens have been reported. However, reports on threshold levels for lung carcinogenesis have hitherto been lacking. In the present study, we first investigated low dose response lung and colon carcinogenesis with a food-derived genotoxic carcinogen, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) (0, 0.01, 0.1, 1, 10 and 100 ppm in the diet) alone for 32 weeks using female A/J mice. The endpoints were histopathologically diagnosed hyperplasias and adenomas in the lung, and aberrant crypt foci (ACF) in the colon. The results showed NOELs of 100 and 1 ppm, respectively. We next investigated the effect of additional pre-treatment with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) (2 mg/mouse, single dose, intraperitoneal injection) prior to the low-dose application of MeIQx (0, 0.01, 0.1, 1, 10, 100 and 600 ppm in the diet) for 32 weeks. Lung lesions were significantly increased in the NNK + MeIQx (1 ppm) group, but not in the NNK + MeIQx (10 ppm) groups. Since the dose-response curve was not of so-called 'hockey stick type', it was not possible to determine a NOEL for lung tumorigenesis. Significant increase in the mRNA expression of CYP2A5, a major metabolic enzyme for NNK, was also observed in the NNK + MeIQx (1 ppm) group, and a similar pattern was noted for O6-methylguanine DNA methyltransferase (MGMT). By contrast, the formation of colon ACF showed a dose-dependent increase. The NOEL for the formation of colon ACF was considered to be 10 ppm MeIQx with NNK. These results suggest that MeIQx may have different threshold dose levels for the induction of lung tumorigenic lesions and ACF formation in the colon. Pre-treatment with NNK, a potent lung carcinogen, concealed the effects of MeIQx in the lung, but exerted minimal influence in the colon. CYP2A5 and MGMT expression may be of importance, particularly in the lung. The present study provides critical suggestions for the human risk assessment of genotoxic carcinogens.
Keiko Yamakawa, Toshiya Kuno, Nozomi Hashimoto, Masanao Yokohira, Satoshi Suzuki, Yuko Narusawa, Kousuke Saoo and Katsumi Imaida : Molecular analysis of carcinogen-induced rodent lung tumors: Involvement of microRNA expression and Krαs or Egfr mutations., Molecular Medicine Reports, 3, 1, 141-147, 2010.
(要約)
Genetic and epigenetic alterations play a key role in lung carcinogenesis, and a high frequency of KRAS and epidermal growth factor receptor (EGFR) mutations have been observed in human lung cancers. Recent evidence indicates that the expression of specific microRNAs (miRNAs) may be involved. In rodent lung carcinogenesis models, Kras mutations are frequently observed, whereas genetic alteration of the Egfr gene is generally rare. Since little is known regarding the involvement of miRNAs in rodent lung carcinogenesis, the present study of miRNA expression levels in the liver and lung during 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)- and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx)-induced lung tumorigenesis in A/J mice was conducted. In addition, incidences of Egfr and Kras gene mutations in rat and mouse lung tumors induced by the chemical carcinogens NNK, MeIQx and N-bis(2?hydroxypropyl)nitrosamine (DHPN) were examined. Three miRNAs, let-7a, miR-34c and miR-125a-5p, were selected for attention. In rat lung tumors, one silent mutation was detected in the Egfr gene exon 20 (AAC↷AAT; N772). Activating mutations of the Krαs gene at codon 12 were detected in neoplastic lesions induced by NNK (5/6, 83%), MeIQx (1/1, 100%) and DHPN (7/15, 47%), all resulting in G/C↷A/T transitions. NNK or MeIQx administration reduced the expression of miR-125a-5p (MeIQx alone group, 86.3%; MeIQx + NNK group, 83.6%; p<0.05, at day 15) and let-7a (MeIQx + NNK group, 56.3%; p<0.001, at day 22) in the liver. miR-34c was up-regulated 3.5-fold with NNK treatment as compared to the control group (p<0.001). These findings raise the possibility that aberrant expression of miRNA is involved in lung tumorigenesis, at least in its early stages.