Biomedical subjects
Publications and source records attributed to Y Kamata.
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PURPOSE OF INVESTIGATION: Progestin is reported to suppress the growth of endometrial carcinomas, although its precise mechanism of action is not clear. This study aimed to transfect progesterone receptor-B (PRB) cDNA into endometrial carcinoma cells and investigate the effect of medroxyprogesterone acetate (MPA) on cell growth, and p21 and p27 expression in the transfectant. METHODS: Immunoblotting for p21 and p27 was performed at predetermined times after the administration of MPA. RESULTS: PR expression was maximally induced in Ishikawa cells at 24 hrs after the transfection. At 1 x 10(-6) M, MPA suppressed the growth of the transfectant by 34% on day 6 and stimulated p21 accumulation at 48 to 72 hrs and p27 accumulation at 48 to 96 hrs after its administration. PRB cDNA was effectively transfected and in the transfectant MPA at 1 x 10(-6) M, the dosage suppressing growth, induced p21 and p27expression. CONCLUSION: p21 and p27 may be related to progesterone-induced growth suppression in human endometrial adenocarcinoma.
We investigated Ang1, Ang2 and Tie2 expressions including balance and intratumoral vessels in the role of angiogenesis of endometrial adenocarcinoma. Immunohistochemical staining was performed on 133 patients with endometrial (endometrioid) adenocarcinoma, including 73 with G1, 34 with G2, and 26 with G3. The levels of Ang1, Ang2 and Tie2 expressions were expressed as staining score. Total vessel count (TVC), microvessel count (MVC) and mean vessel diameter (VD) in the CD34-stained tissues were measured in five hot spot areas at x 200 magnification by image cytometry. These results were compared with high and low vascular endothelial growth factor (VEGF) expressions. Ang1, Ang2, Tie2 and CD34 were expressed in the cytoplasm of tumor cells. A significant correlation was found among Ang1, Ang2 and Tie2 expressions. In high VEGF cases, Ang1 expression was correlated negatively with TVC and MVC, but positively with VD, and the Angl < Ang2 group was significantly higher in TVC and MVC and tended to be smaller in VD than the Ang1 > Ang2 group. VD was significantly larger in G3 than in G1. The Ang1 < Ang2 balance may be one of the key factors for angiogenesis of endometrial carcinoma in the presence of high VEGF expression.
A 70-year-old man presenting with a chief complaint of tongue swelling had been diagnosed with prostate cancer 1 year earlier. He had been on an oral angiotensin-converting enzyme inhibitor (ACE) inhibitor for hypertension for 20 years. Two months before the first of 4 episodes of tongue swelling within a period of 40 days, he had been prescribed oral estramustine phosphate (EMP) for the prostate cancer. He was admitted to our hospital for the evaluation after massive swelling of the tongue and epiglottis which necessitated tracheotomy. Food allergies, allergic reactions to environmental factors, and hereditary angioneurotic edema were excluded. Massive swelling of the tongue and epiglottis disappeared completely after EMP was discontinued. We concluded that angioedema was induced by EMP used concurrently with the ACE inhibitor.
Four hybridoma clones (ACV-1, -3, -4, and -5) were established for Chinemys reevesii (Reeves' turtle) vitellogenin (VTG) as a precursor protein of egg yolk and a biomarker of environmental pollution. Binding-inhibition experiments indicated that the epitopes of four mAbs were distinct. No binding of ACV-4 to C. reevesii VTG in the Western blot suggests that the epitope of ACV-4 would be dependent on the three-dimensional structure. ACV-1, -3, and -5 bound to C. reevesii VTG in the Western blot. The signal for ACV-1 and -5 disappeared by reduction of the VTG, suggesting that the construction of the epitopes for ACV-1 and -5 were dependent on the disulfide bridge in the VTG molecule. All four mAbs recognized Trachemys scripta and Mauremys japonica VTGs in the ELISA. The yolk proteins were tested for the binding of the mAbs in the Western blot. ACV-1 being capable to bind to the VTG in the reduced condition did not bind to any protein bands of the yolk. This indicates that ACV-1 recognizes a part of the VTG molecule that is not incorporated in the oocytes. Both ACV-3 and -5 bound to the 32- and 70-kDa yolk proteins. Since a mAb recognizes only one site (epitope) on a protein molecule, the 32-kDa protein originated from the 70-kDa one. An ELISA system using ACV-5 as the capture antibody and ACV-3 as the detecting antibody showed the lower detectable concentration (2 ng/mL) and a wide detectable range to 1000 ng/mL (R2=0.999). The system was used to determine serum VTG levels of juvenile turtles treated with estradiol-17beta or vehicle (corn oil). By the use of the mAbs described in this paper, basic and applied studies for turtle VTGs would be improved.
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Explore the source record for details and available documents.
Explore the source record for details and available documents.
Mutant p53 protein is not degradated but accumulates in the nuclei. However, the relation between p53 gene mutation and quantity of p53 protein has not been clarified yet in endometrial carcinoma. We investigated the correlation between p53 gene mutation and its protein expression quantitatively using 11 cell lines of endometrial adenocarcinoma, endometrioid type and two serous-type cell lines. To examine p53 mutation, PCR-SSCP analysis in exon 5 to 8 and direct sequence were carried out. p53 expression was determined by immunocytochemistry and immunoblotting. The percentage of positive staining in the nuclei by immunocytochemistry was calculated as a labeling index (LI). The amount of p53 detected by immunoblotting was expressed as a comparative value of Ishikawa cells. Point mutation of p53 gene was detected in four of 11 (36.4%) cell lines of endometrioid adenocarcinoma, and all two of serous adenocarcinoma. There was a significant positive correlation between p53 LI and p53 values. The LI and the values of p53 were significantly higher in the mutant group than the wild one, thus showing a quantitative correlation between p53 protein expression and p53 gene mutation in endometrial carcinoma cell lines. It is plausible that immunohistochemical analysis of p53 could be qualified to be a convenient indicator of p53 gene mutation on clinical materials, if p53 LI is more than 40 (M-SD in mutant p53).
OBJECTIVE: Endometrial carcinomas are grouped into two types of carcinogenic entities. These two different types of cell lines designated as HEC-251 and HEC-265 were established from human endometrial adenocarcinoma. We report their characteristics. MATERIALS: HEC-251 was derived from moderately differentiated endometrioid adenocarcinoma and HEC-265 from well-differentiated adenocarcinoma. RESULTS: These cell lines grow well and serial passages can be successively carried out more than 100 times. The monolayer cultured cells reveal neoplastic and pleomorphic features, and grow in multilayers. HEC-251 cells are immuno-cytochemically positive for p53 and HEC-265 cells for PgR. Both cell lines are transplantable to nude mice and reflect the original histopathologic characteristics. CONCLUSION: The cell lines HEC-251 and HEC-265 will contribute to clarifying the characteristics of two different types of human endometrial carcinomas.
PURPOSE: PTEN is a tumor suppressor gene that inhibits cell proliferation by regulating intracellular signaling pathways, and this activity can be abolished by mutations of the PTEN gene. This study was designed to examine the correlation of PTEN expression with the expression of cell cycle regulators and with clinicopathological parameters in endometrioid adenocarcinoma of the uterine corpus. METHODS: Tissue samples of 117 endometrioid adenocarcinomas in addition to those of 19 normal endometria and 20 endometrial hyperplasias were used for the study. Immunohistochemical staining for PTEN protein was performed with the labeled streptavidin-biotin method on formalin-fixed and paraffin-embedded tissue samples. PTEN expression was represented as the staining score. RESULTS: Immunohistochemistry showed that the nuclei of cells were positive for PTEN. The PTEN staining score of normal endometrium was significantly higher in the proliferative phase than in the secretory phase. The scores of various endometrial hyperplasias were not significantly different from each other, regardless of the type of hyperplasia. The PTEN staining scores of endometrioid adenocarcinomas were 7.6+/-5.2 in G1, 9.6+/-5.2 in G2, and 11.9+/-3.7 in G3, and increased significantly as the histological grade increased. PTEN staining score was not significantly correlated with clinicopathological parameters such as FIGO stage, myometrial invasion, lymph-vascular space invasion (LVSI), lymph node metastasis or group, but was significantly correlated with labeling indices (LIs) of cell cycle regulators such as Ki-67, cdk2, cyclin A, cyclin D1, cyclin E, p27, and p53. The PTEN staining score of p53-wild cases was significantly lower than that of p53-mutant ones, but there was no significant difference of the score in cases with different PTEN gene status. PTEN expression was significantly lower in cases with both high levels of estrogen receptor and progesterone receptor. CONCLUSION: PTEN protein expression was decreased in well-differentiated and less growth-aggressive endometrial carcinoma with wild-type p53 gene and high levels of ER and PR. This suggests that disturbed PTEN expression occurs in an early phase of the tumorigenesis of well-differentiated endometrial carcinoma.
A quantitative immunological method was developed for measuring serum vitellogenin levels of Reeves' pond turtles (Chinemys reevesii) to investigate the effects of endocrine disruptors on the freshwater ecosystem. Vitellogenin was induced by injecting estradiol-17beta into C. reevesii turtles (adult females, juvenile females, and males) and was purified from the turtle serum by EDTA-MgCl2 precipitation followed by gel filtration. Using a polyclonal antibody raised against C. reevesii vitellogenin, an enzyme-linked immunosorbant assay was established. The detectable range, recovery of vitellogenin, and coefficient of variation in this assay were 0.0040-1.0 microg.ml(-1), 85.3-109% and 3.4-11.5%, respectively. This assay was also applicable for measurement of the concentrations of vitellogenins from other species, Japanese pond turtles (Mauremys japonica) and red-eared turtles (Trachemys scripta). The serum vitellogenin concentration of 131 C. reevesii turtles captured at a Japanese local river was measured by the assay. In females, vitellogenin ranged from 0.10 microg.ml(-1) to 15,000 microg.ml(-1) with two peaks, 0.10-1.0 microg.ml(-1) (juveniles) and 1,000-10,000 microg.ml(-1) (adults). However, in males, it ranged from 0.10 microg.ml(-1) to 0.60 microg.ml(-1), showing one peak, 0.10-0.20 microg.ml(-1). Therefore, if relatively high concentrations of vitellogenin are detected in males or juvenile females, it is suggested that they would have been exposed to xenobiotic estrogens.
PURPOSE: Cyclin E is known as a G1-S phase regulatory protein and its abnormal expression has been implicated in cellular proliferation. This study aimed to investigate the correlation of cyclin E expression with tumorigenesis of the endometrium, proliferative activity, and clinicopathological features of endometrial adenocarcinoma. METHODS: Immunohistochemical staining for cyclin E in addition to cyclin-dependent kinase 2 (cdk2), Ki67, p27, and p53 was performed by the labeled streptavidin-biotin method on formalin-fixed, paraffin-embedded tissues of normal endometria (20 cases), endometrial hyperplasias (20 cases), and endometrial adenocarcinomas (endometrioid type) (127 cases). Positive staining was expressed as a labeling index (LI) based on percentages of positive nuclei in tumor cells. RESULTS: Immunohistochemistry showed that the nuclei of the cells were positive for cyclin E. Both proliferative and secretory endometria, and endometrial hyperplasia regardless of type were negligible for cyclin E expression. The expression in normal endometrium and hyperplasia was significantly less than that in endometrial adenocarcinomas (P<0.0001). LIs of cyclin E in well-differentiated, moderately differentiated, and poorly differentiated endometrial adenocarcinomas were 31.5+/-33.3%, 37.8+/-31.9%, and 51.1+/-30.8%, respectively. Cyclin E expression increased significantly more in histological grades. The LI of cyclin E in carcinoma was positively correlated with that of cdk2, Ki67, and p53 but not with p27. The cyclin E expression was correlated with myometrial invasion and lymph-vascular space involvement, but not with FIGO stage, lymph node metastasis, coexisting endometrial hyperplasia, estrogen receptor, progesterone receptor, and menopause. CONCLUSION: Cyclin E as a complex with cdk2 is associated with carcinogenesis and disease progression in endometrial adenocarcinoma, and might be a prognostic indicator of endometrial adenocarcinoma.
It has been reported that tumor suppressor gene p53 regulates vascular endothelial growth factor (VEGF) expression, but the relation between them in endometrial carcinoma remains unclear. We investigated VEGF expression in 11 endometrial carcinoma cell lines and the effect of p53 gene transfection on VEGF expression in the p53-mutated endometrial carcinoma cell line, HEC-50B. Immunoblotting for detecting VEGF, p53, and beta-actin was performed. Wild type p53 gene was transfected using the SuperFect method. The mean VEGF value of 0.8 +/- 0.3 (n = 6) in p53 wild-type group was significantly lower than the 1.6 +/- 0.8 (n = 5) that was found in the p53 mutant group (P < 0.05). Levels of VEGF in the culture medium were measured by enzyme immunoassay (EIA). VEGF levels in the p53 gene-transfected HEC-50B cells and the conditioned medium were decreased at 48 h after p53 gene transfection. VEGF expression was downregulated by p53 in endometrial carcinoma cells.
Systemic injection of an adenovirus vector into adult mice resulted in pathological improvements in multiple visceral organs of mice with mucopolysaccharidosis VII; however, no therapeutic efficacy was observed for mental retardation, skeletal deformities, corneal clouding, and retinal degeneration. In this study, an adenovirus vector expressing human beta-glucuronidase was injected into mice with mucopolysaccharidosis VII within 24 h of birth, and therapeutic efficacy was evaluated. In the brains of the mice, more than 20% of GUSB activity was maintained for at least 20 weeks after birth, and histopathological analysis showed no obvious lysosomal storage. Furthermore, no vacuolated cells were detected in corneal stroma and retinal pigment epithelium in the eyes of the mice treated in the neonatal period, while pathological improvement was not observed in adult MPSVII mice that received similar treatments. The treated mice also lacked characteristic facial skeletal deformities, and radiographic analysis demonstrated that their facial and cranial bones were morphologically normal. These results indicate that a single systemic adenovirus injection in the neonatal period could prevent the progression of mental retardation, corneal clouding, retinal degeneration, and skeletal deformities, all of which are frequently observed clinical manifestations and difficult to treat in adulthood.
Vascular endothelial growth factor (VEGF) that activates endothelial cell growth induces angiogenesis, which is indispensable to tumor igenesis and tumor progression. On the other hand, tumor suppressor gene p53 has been considered to regulate VEGF expression, but the detailed relationship between them remains unclear. In this study, we aimed to study VEGF expression in endometrial carcinoma cells and the effect of p53 gene transfection on VEGF expression using p53-mutated endometrial carcinoma cell line, HEC-50B. Immunoblotting for detecting VEGF protein, p53 protein and beta-actin was performed using 11 endometrial carcinoma cell lines. Levels of VEGF in the cultured media were measured by Enzyme immunoassay(EIA). Transfection of wild p53 gene was carried out by SuperFect method in HEC-50B cells, which had mutant p53 gene and did not express p53 protein. The results of immunoblotting were analyzed by NIH image and expressed as values. The results of EIA were expressed as the relative value. The VEGF value was 0.8 +/- 0.3 (n = 6) in p53-wild group, whereas in p53-mutant group it was 1.6 +/- 0.8 (n = 5). VEGF expression was correlated significantly with p53 status (P < 0.05). VEGF levels in p53 gene-transfected cells and the conditioned medium were decreased in 48 hours after p53 gene transfection. VEGF expression was down-regulated by p53 in endometrial carcinoma cells.
p27 is regarded as a cyclin-dependent kinase inhibitor of the G1-to-S cell cycle progression by suppressing the kinase activity of cyclin/cyclin-dependent kinase complex. This study aimed to investigate p27 expression in the normal endometrium and endometrioid adenocarcinoma of the uterine corpus and the correlation of its expression with cell proliferation and clinicopathological parameters. Tissue samples of 127 endometrioid adenocarcinomas and 15 normal endometria were used in the study. Immunohistochemical staining for detecting p27 and Ki-67 was performed by the labelled streptavidin-biotin method on formalin-fixed and paraffin-embedded tissue samples. The expression was given as the labelling index, which indicates the percentage of positive nuclei. p27 staining was observed in the nuclei of the glandular cells in the functional layer of the secretory phase endometrium, whereas it was negative in those of the proliferative phase. In endometrioid adenocarcinomas, the labelling index of p27 expression paradoxically increased more significantly in the higher histological grades and was correlated with that of Ki-67. The high level of p27 expression was associated with clinicopathological parameters such as FIGO stage, lymph node metastasis, lymphovascular space involvement and myometrial invasion. High p27 expression was linked to higher grades of endometrioid adenocarcinoma, cell proliferation and some clinical prognostic factors. These results indicate that p27 might be an indicator of poor prognosis.
In an outbreak of gastroenteritis on 23 July 1996, in Osaka, Japan, 54 of 91 persons who had attended a meeting the previous day became ill. Escherichia coli O166:H15 was isolated from stool specimens of patients (29/33, 88%). Laboratory tests for other bacterial pathogens and viruses were negative. The E. coli 0166 organisms did not adhere to HEp-2 cells in a localized, diffuse, or enteroaggregative manner. The organisms did not express known enterotoxigenic E. coli (ETEC) colonization factors. In polymerase chain reaction tests, the bacteria did not have coding genes for shigatoxin of enterohemorrhagic E. coli (EHEC), heat-labile, or heat-stable enterotoxin of ETEC, attachment and effacement (eaeA) of EPEC, or invasion (invE) of enteroinvasive E. coli (EIEC). Consequently, they could not be assigned to any of the recognized diarrhoeagenic groups of E. coli: EPEC, ETEC, EHEC, EIEC, enteroaggregative E. coli (EAggEC), or diffusely adhering E. coli. However, the organisms possessed the EAggEC heat-stable enterotoxin (EAST1) gene. To our knowledge, this is the first report of an outbreak caused by E. coli that did not have well-characterized virulence genes other than EAST1. The isolates showed the same DNA banding pattern in pulsed-field gel electrophoresis after digestion with the restriction enzymes XbaI or NotI. Three O166:H15 strains isolated from two sporadic cases and another outbreak during 1997-8 were distinct, indicating that multiple clones have spread already. We propose that diarrhoeal specimens should be examined for E. coli possessing the EAST1 gene.