Kazuko Shichijo

The mechanism of low susceptibility to stress in gastric lesions of spontaneously hypertensive rats

The mechanism of low susceptibility to stress in gastric lesion formation in spontaneously hypertensive rats (SHR) was investigated focusing on the sympathetic and parasympathetic nervous systems. In the gastric tissues of SHR, norepinephrine (NE) and dopamine (DA) contents were higher, while acetylcholine content and choline acetyltransferase activity were lower than those of Wistar-Kyoto rats (WKY). Water-immersion restraint induced gastric lesions frequently in WKY (ulcer indices : 52 +/- 7mm2) but less frequently in SHR (ulcer indices : 3 +/- 1mm2). Although NE content decreased in both SHR and WKY as a result of water-immersion restraint, it remained higher in SHR than in WKY. ACh content decreased by the procedure in WKY but not in SHR. DA content was increased by the procedure in all gastric regions of SHR. The gastric lesions induced in SHR were aggravated by pretreatment with 6-hydroxydopamine, an agent for chemical sympathectomy, following decreases of NE and DA contents. These results indicate that the relative sympathetic hyperfunction, parasympathetic hypofunction and dopaminergic mechanism in the stomach contribute to the prevention of gastric lesion formation in SHR.

The role of sympathetic neurons for low susceptibility to stress in gastric lesions

Stroke prone spontaneously hypertensive rats (SHRSP) were less susceptible to stress in gastric lesions than Wistar-Kyoto rats used as normotensive controls. The gastric lesions induced by water-immersion restraint (WIR) in SHRSP were aggravated by pretreatment with 6-hydroxydopamine (6-OHDA), an agent for chemical sympathectomy, following decreases in blood pressure and sympathetic nerve function. 6-OHDA treatment remarkably reduced norepinephrine content but caused increases in dopamine content and in choline acetyltransferase activity in the stomach. The mechanism of aggravation of gastric lesions in SHRSP was investigated with regard to gastric acid and motility. The pretreatment with 6-OHDA of SHRSP significantly increased the acid secretion stimulated by 2-deoxy-D-glucose indirectly acting on parietal cells via the vagus nerve, but did not change the acid secretions stimulated by carbachol, pentagastrin and histamine acting directly on parietal cells. Gastric (corpus) motility associated with WIR was completely blocked by atropine. The pretreatment with 6-OHDA in SHRSP decreased the gastric motility during WIR, which was facilitated by treatment with domperidone. These results indicate that the sympathetic hyperactivity of the stomach prevents WIR-induced gastric lesion formation mainly via the inhibition of gastric acid secretion.

Foci formation of P53-binding protein 1 in thyroid tumors: activation of genomic instability during thyroid carcinogenesis.

Defective DNA damage response (DDR) can result in genomic instability (GIN) and lead to the transformation into cancer. P53‐binding protein 1 (53BP1) belongs to a family of evolutionarily conserved DDR proteins. Because 53BP1 molecules localize at the sites of DNA double strand breaks (DSBs) and rapidly form nuclear foci, the presence of 53BP1 foci can be considered as a cytologic marker for endogenous DSBs reflecting GIN. Although it has been proposed that GIN has a crucial role in the progression of thyroid neoplasms, the significance of GIN during thyroid tumorigenesis remains unclear, particularly in patients. We analyzed, therefore, the level of GIN, as detected with immunofluorescence of 53BP1, in 40 cases of resected thyroid tissues. This study demonstrated a number of nuclear 53BP1 foci in thyroid cancers, suggesting a constitutive activation of DDR in thyroid cancer cells. Because follicular adenoma also showed a few 53BP1 nuclear foci, GIN might be induced at a precancerous stage of thyroid tumorigenesis. Furthermore, high‐grade thyroid cancers prominently exhibited an intense and heterogeneous nuclear staining of 53BP1 immunoreactivity, which was also observed in radiation‐associated cancers and in mouse colonic crypts as a delayed response to a high dose ionizing radiation, suggesting increased GIN with progression of cancer. Thus, the present study demonstrated a difference in the staining pattern of 53BP1 during thyroid carcinogenesis. We propose that immunofluorescence analysis of 53BP1 expression can be a useful tool to estimate the level of GIN and, simultaneously, the malignant potency of human thyroid tumors.

The effect of chemical sympathectomy on acute liver injury induced by carbon tetrachloride in spontaneously hypertensive rats

The effects of 6-hydroxydopamine (6-OHDA) on carbon tetrachloride (CCl4)-induced acute liver injury were examined in spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). Plasma glutamate-pyruvate transaminase (GPT) activity was increased in both strains after one dose of CCl4 administration, although the increase was more significant in SHR than in WKY. Pretreatment with 6-OHDA inhibited the increase in GPT activity after CCl4 administration in both strains and especially in SHR. 6-OHDA also reduced the noradrenaline (NA) content of the liver and increased the hepatic blood flow in both strains. It caused a greater decrease in blood pressure in SHR than in WKY. The results suggest that 6-OHDA blocks the sympathetic neural activity in the liver, resulting in vasodilatation and increase in hepatic blood flow and thereby alleviating the circulatory disturbance produced by CCl4 preventing acute liver damage, especially in SHR.

Mechanism of the antiulcerogenic effect of IL-11 on acetic acid-induced gastric ulcer in rats

The purpose of this study was to evaluate the healing effect of interleukin-11 (IL-11) on acetic acid-induced gastric ulcer in rats. Gastric ulcers were induced in male Wistar rats by applying acetic acid to the fundus of the stomach. Recombinant human interleukin-11 (rhIL-11 100 microg/kg/twice daily, subcutaneously) was administered starting on the 2nd day before ulcer induction up through the 7th day after ulcer induction. Control rats were injected with bovine serum albumin. At 12 hours and 7 days after ulcer induction, the animals were sacrificed, and the ulcer index, proliferating cell nuclear antigen (PCNA) expression, and IL-11alpha receptor expression in the gastric tissues were studied. The ulcer index of the rhIL-11-treated rats was significantly lower than that of the control rats at the 7th day. The expression of PCNA as evaluated by Western blotting and immunohistochemistry, was enhanced in both the mucosal proliferative zone and proper muscle layer of the rhIL-11-treated rats in comparison with that in the control rats. IL-11alpha receptor expression was observed in the mucosal neck cells of the rhIL-11-treated rats and control rats. These findings suggest that IL-11 accelerates ulcer healing by inducing the proliferation of mucosal and muscular cells.

Altered Expression of β-Catenin during Radiation-induced Colonic Carcinogenesis

Summary Radiotherapy for malignant pelvic disease is commonly accompanied by treatment-induced proctitis, and rarely by colorectal cancer. Translocation of the b-catenin protein, which is a key downstream effector of the Wnt signal transduction pathway, is frequently found in colorectal cancer. Nuclear β-catenin enhances transcriptional activity of the cyclin D1 gene in cancer cells. Here, we evaluate the involvement of the Wnt pathway in radiation-induced colon carcinogenesis with rats (n = 36). β-catenin, APC, and cyclin D1 expression profiles were analyzed by immunohistochemistry in radiation-induced chronic colon injury including cancers and ulcerative lesions in rats (n = 12 in treated group, n = 12 in control group). In total, 3 cases of invasive adenocarcinomas were developed in the irradiated portion 50 weeks after a single dose of 36 Gy irradiation.Nuclear translocation of β-catenin was observed in all radiation-induced colon cancers, whereas this protein was also found in the cytoplasm and/or nucleus of 9 cases of non-neoplastic irradiated colonocytes. Nuclear translocation of β-catenin correlated with loss of APC and gain of cyclin D1 expression, suggesting activation of the Wnt pathway during radiation-induced colorectal carcinogenesis. A single dose of 10 Gy was also given for acute injury (n = 12: 3 each in days 0, 3, 5, and 7, respectively). β-catenin expression was distributed in the cytoplasm of degenerating glands at day 3 and 5, and was observed in the cell membrane of those glands with histological normalization at day 7 after irradiation. Because translocation of β-catenin was found in irradiated-colonic mucosa as well as colon cancer, disruption of β-catenin expression might be one of the early events in radiation-induced colonic carcinogenesis.

Alteration of p53-binding protein 1 expression during skin carcinogenesis: association with genomic instability.

Epidermal cells are the first cells to be exposed to environmental genotoxic agents such as ultraviolet and ionizing radiations, which induce DNA double strand breaks (DSB) and activate DNA damage response (DDR) to maintain genomic integrity. Defective DDR can result in genomic instability (GIN) which is considered to be a central aspect of any carcinogenic process. P53‐binding protein 1 (53BP1) belongs to a family of evolutionarily conserved DDR proteins. Because 53BP1 molecules localize at the sites of DSB and rapidly form nuclear foci, the presence of 53BP1 nuclear foci can be considered as a cytological marker for endogenous DSB reflecting GIN. The levels of GIN were analyzed by immunofluorescence studies of 53BP1 in 56 skin tumors that included 20 seborrheic keratosis, eight actinic keratosis, nine Bowens disease, nine squamous cell carcinoma, and 10 basal cell carcinoma. This study demonstrated a number of nuclear 53BP1 foci in human skin tumorigenesis, suggesting a constitutive activation of DDR in skin cancer cells. Because actinic keratosis showed a high DDR type of 53BP1 immunoreactivity, GIN seems to be induced at the precancerous stage. Furthermore, invasive cancers exhibited a high level of intense, abnormal 53BP1 nuclear staining with nuclear accumulation of p53, suggesting a disruption of DDR leading to a high level of GIN in cancer cells. The results of this study suggest that GIN has a crucial role in the progression of skin carcinogenesis. The detection of 53BP1 expression by immunofluorescence can be a useful histological marker to estimate the malignant potential of human skin tumors. (Cancer Sci 2008; 99: 946–951)

Experimental stress ulcer and gastric catecholamine contents in spontaneously hypertensive rats

SummaryTo determine the role of the sympathetic nervous system in the development of stress ulcer, we carried out experiments on spontaneously hypertensive rats (SHR) exposed to restraint and water immersion stress for 7 h. Normotensive Wistar-Kyoto rats (WKY) were used as controls. Catecholamine (CA) contents in gastric tissues, divided into mucosal and muscular layers of both antrum and corpus, were quantitated by high-performance liquid Chromatographic electrochemical detection (HPLC-EC). The stress ulcer formation was much less frequently induced in SHR than in WKY. The noradrenaline (NA) contents in all regions of gastric tissues were higher in SHR than in WKY. The contents of adrenaline (A) and dopamine (DA), present in small quantities in gastric tissues showed no difference between SHR and WKY. After exposure to stress, the NA contents in mucosal and muscular layers of the gastric corpus decreased significantly in both SHR and WKY, whereas the value remained higher in the former. On the contrary, a remarkable increase of A contents (probably released from the adrenal medulla by the stress) was observed in all gastric tissues, of both SHR and WKY. Increase of the A contents in the mucosal layer was remarkable in the SHR. The DA contents increased in both strains. These results suggest that the peripheral sympathetic hyperfunction in the stomach in the SHR may have an inhibitory role in stress ulcer formation.

X Radiation Up-regulates the Occurrence and the Multiplicity of Invasive Carcinomas in the Intestinal Tract of Apcmin/+ Mice

Abstract Nakayama, T., Yamazumi, K., Uemura, T., Yoshizaki, A., Yakata, Y., Matsuu-Matsuyama, M., Shichijo, K. and Sekine, I. X Radiation Up-regulates the Occurrence and the Multiplicity of Invasive Carcinomas in the Intestinal Tract of Apcmin/+ Mice. Radiat. Res. 168, 433–439 (2007). X rays are well known to cause genetic damage and to induce many types of carcinomas in humans. The Apcmin/+ mouse, an animal model for human familial adenomatous polyposis (FAP), contains a truncating mutation in the APC gene and spontaneously develops intestinal adenomas. To elucidate the role of X rays in the development of intestinal tumors, we examined the promotion of carcinogenesis in X-irradiated Apcmin/+ mice. Forty out of 77 (52%) X-irradiated Apcmin/+ mice developed adenocarcinomas that invaded the proprial muscle layer of the small intestine; 24 of 44 (55%) were in males, and 16 of 33 (49%) were in females. In contrast, invasive carcinomas were detected in the small intestines of only 13 of 64 (20%) nonirradiated Apcmin/+ mice; nine of 32 (28%) were in males and four of 32 (13%) were in females. These differences between X-irradiated and nonirradiated Apcmin/+ mice in the occurrence of invasive intestinal carcinomas were statistically significant (P < 0.05 for males, P < 0.005 for females). In wild-type mice, invasive carcinomas were not detected in either X-irradiated or nonirradiated mice. Apcmin/+ mice had many polyps in the large intestine with or without X irradiation; there was no difference in the number of polyps between the two groups. Also, invasive carcinomas were not detected in the large intestine with or without irradiation. The occurrence of mammary tumors, which was observed in Apcmin/+ mice, was found to be increased in irradiated Apcmin/+ mice (P < 0.01). Apcmin/+ mice had many polyps in the small and large intestines with or without X irradiation. X-irradiated Apcmin/+ mice had highly invasive carcinomas in the small intestine with multiplicities associated with invasiveness. Our results suggest that X radiation may promote the invasive activity of intestinal tumors in Apcmin/+ mice.

Neuronal release of endogenous dopamine from corpus of guinea pig stomach

Neuronal release of endogenous dopamine was identified in mucosa-free preparations (muscle layer including intramural plexus) from guinea pig stomach corpus by measuring tissue dopamine content and dopamine release and by immunohistochemical methods using a dopamine antiserum. Dopamine content in mucosa-free preparations of guinea pig gastric corpus was one-tenth of norepinephrine content. Electrical transmural stimulation of mucosa-free preparations of gastric corpus increased the release of endogenous dopamine in a frequency-dependent (3-20 Hz) manner. The stimulated release of dopamine was prevented by either removal of external Ca2+ or treatment with tetrodotoxin. Dopamine-immunopositive nerve fibers surrounding choline acetyltransferase-immunopositive ganglion cells were seen in the myenteric plexus of whole mount preparations of gastric corpus even after bilateral transection of the splanchnic nerve proximal to the junction with the vagal nerve (section of nerves between the celiac ganglion and stomach). Domperidone and sulpiride potentiated the stimulated release of acetylcholine and reversed the dopamine-induced inhibition of acetylcholine release from mucosa-free preparations. These results indicate that dopamine is physiologically released from neurons and from possible dopaminergic nerve terminals and regulates cholinergic neuronal activity in the corpus of guinea pig stomach.