Norimichi Nemoto

Increased Expression of Cyclooxygenase-2 in Local Lesions of Endometriosis Patients

PROBLEM: Human endometrial glands contain the highest levels of cyclooxygenase (COX), although whether it is COX‐1 and/or COX‐2 has not been previously determined. Overexpression of COX‐2 may result in the pathogenesis of endometriosis.

Expression and quantitative analysis of matrix metalloproteinase-2 and-9 in human gliomas

The matrix metalloproteinase (MMP) family members catalyze extracellular proteolysis. Recent reports have suggested that expression of MMP-2 and-9 might play a critical role in neoplastic tissue invasion or metastasis. In this study, the relationship between the expression of MMP-2 and-9 and the histological features of tissues from 21 cases of human glioma were investigated. MMP-2 and-9 proteins were detected by immnohistochemical studies. Amplification of MMP-2 and-9 mRNA was detected by reverse transcription-polymerase chain reaction (RT-PCR) assay. MMP-2 and-9 mRNA was measured quantitatively by the real-time RT-PCR method. Immunohistochemically, 38% of the cases were positive for MMP-2. Amplification of MMP-2 mRNA by RT-PCR was detected in. 62% of the cases. There was no significant relationship between the expression of MMP-2 protein or mRNA and the biological nature of the tumors, including aggressiveness and histologic classification. The quantity of MMP-2 mRNA was 0.035±0.113 (MMP-2/GAPDH%), which was significantly elevated in cases of neoplastic dissemination or recurrence (P<0.05). Tumor cells were immunohistochemically positive for MMP-9 in 81% of the samples. A positive reaction was found not only in neoplastic cells but also in endothelial cells, suggesting that the expression of MMP-9 protein might be associated with tumoral angiogenesis. The expression of mRNA in MMP-9 was detected in 91% of the cases, suggesting a close relationship between expression of MMP-9 and malignancy. The quantity of MMP-9 was 0.097±0.113 (MMP-9/GAPDH %) in all samples, which was significantly elevated in cases of glioblastoma (P<0.05). The average Ki-67 labeling index was 8.14±5.26 in samples from G2 glioma, 19.92±11.29 in samples from G3 glioma, and 23.52±10.14 in samples from glioblastoma. All of the cases with elevated indices had recurrence or dissemination. The results of our study suggest that quantity analyses of MMP-2 and-9 mRNA and Ki-67 labeling index should be useful for discerning tumoral behaviors such as invasion, dissemination, and recurrence.

Murine fetal resorption and experimental pre-eclampsia are induced by both excessive Th1 and Th2 activation

It has been proposed that immune responses in mammalian normal pregnancy are Th2-like, thereby protecting the fetus and placenta from being rejected. Administration of exogenous Th1 cytokines into pregnant mice is reported to induce feto-placental resorption. However, the effects of exogenous Th2 cytokines and Th2 directed responses in pregnant animals have not been well studied. In this study, we examined IL-4 and IL-12, which play decisive roles in the development of Th2 and Th1 responses, respectively, in the induction of fetal resorption and development of experimental pre-eclampsia. Transfer of either IL-4 and/or IL-12 stimulated splenocytes from BALB/C virgin female mice into BALB/C pregnant mice mated with either C57BL/6 or BALB/C male mice resulted in fetal resorption and glomerular nephritis associated with hypertension and proteinuria. In mice treated with IL-12 stimulated splenocytes, fatty liver degeneration associated with bile retention was observed. These results indicate that both excessive Th1 and Th2 activation contribute to the development of fetal resorption and pre-eclampsia, but that Th1 is critical to the development of liver degeneration.

Survivin as a prognostic factor for osteosarcoma patients.

Purpose: Survivin is one of the apoptosis inhibitor genes and is rarely expressed in adult tissues. However, survivin expression has been detected in various human cancers and correlations have been recognized between the level of expression of this gene in tumors and prognosis. In this study, we investigated the correlations between survivin mRNA expression in osteosarcoma tissues and clinicopathological parameters. Methods: There were 22 osteosarcoma patients in our hospital with paraffin-embedded tissues which could be extracted from biopsy specimens. We used the RT-PCR method after extracting total RNA and conducted a densitometric analysis to determine the ratio of survivin relative to h-GAPDH as an internal marker. Results: Expression of survivin mRNA was detected in all osteosarcoma samples. Patients with metastasis had high survivin mRNA levels in initial biopsy specimens (p<0.01). Moreover, there was a statistically significant difference in survivin mRNA expression between patients with and without metastasis (p<0.01). Conclusion: We concluded that high levels of survivin mRNA expression suggest poor prognosis for osteosarcoma patients.

Histopathological analysis of angiogenic factors in renal cell carcinoma

Aim: The present study was carried out to clarify whether a histopathological analysis of vascular endothelial growth factor (VEGF), transforming growth factor‐β1 (TGF‐β1) and matrix metalloproteinase 2 (MMP‐2) can help predict the outcome of renal cell carcinoma (RCC). We examined the expression of VEGF, TGF‐β1 and MMP‐2 in a large series of RCC with a long follow‐up, based on histopathological factors and survival.

Quantitative analysis of human tissue-specific differences in methylation

Tissue-specific differentially methylated regions (tDMRs) have been identified and implicated for their indispensable involvement in mammalian development and tissue differentiation. In this report, a quantitative DNA methylation analysis was performed for 13 human orthologous regions of recently confirmed mouse tDMRs by using Sequenom Mass Array, by which bisulfite-treated fragments are quantitatively detected using time of flight mass spectroscopy analysis. Eight regions were shown as tDMRs in various tissues from three independent individuals. Testis DNA samples from eight individuals were also analyzed for methylation. Interestingly, there is evidence that the DNA methylation level is divergent among individuals. DNA methylation levels of five testis-specific DMRs were significantly inversely correlated with the number of spermatocytes. However, a positive correlation was seen at tDMRs located near the TRIM38 and CASZ1 genes. Our results indicate that tDMRs are conserved between mouse and human and may have an important role in regulating tissue function, differentiation, and aging.

Expression of Somatostatin Receptor (SSTR) Subtypes (SSTR-1, 2A, 3, 4 and 5) in Neuroendocrine Tumors Using Real-time RT-PCR Method and Immunohistochemistry.

Molecule targeting therapy using somatostatin (SS) analogues has become a widely accepted modality to treat neuroendocrine tumors (NETs), particularly gastrointestinal (GI) and pancreatic endocrine tumors. On the other hand, little is known about the expression of somatostatin receptor (SSTR) subtypes in neuroendocrine carcinomas (NECs). We investigated the expression of SSTR subtypes (SSTR-1, 2A, 3, 4 and 5) using real-time reverse transcription polymerase chain reaction (RT-PCR) method and immunohistochemistry in 32 neuroendocrine neoplasms (9 NET G1, 2 NET G2, 18 NECs G3 and 3 mixed NEC G3) of various primary sites. Expression of more than two SSTR subtypes was detected in all neuroendocrine neoplasms examined. Expression of SSTR-2A mRNA was significantly higher than other subtypes. In addition, mRNA expression of SSTR-3 and SSTR-5 was significantly low or below the detection level except for gastroduodenal NET G1. No significant difference of the expression of SSTR subtypes was observed between the NET and NEC groups. The expression of protein and mRNA was generally well correlated. In conclusion, NECs would be a good candidate for molecule targeting therapy using SS analogues, and the expression of SSTR-2A can be useful as a biomarker of neuroendocrine differentiation. We have demonstrated that NEC G3 small cell type shows a different expression profile of SSTR subtypes compared with NET and NEC non-small cell type.

Collision Tumor of the Stomach A Rare Case of an Adenocarcinoma and Carcinoid Tumor

We report a rare case of gastric collision tumor composed of moderately differentiated tubular adenocarcinoma and carcinoid in an 84-year-old woman. On endoscopic examination, an invasive tumor was noted at the cardia of the stomach, and a pathologic examination of the biopsy specimen revealed adenocarcinoma. After total gastrectomy, a thorough histopathologic examination of the resected tumor revealed the concurrent presence of moderately differentiated adenocarcinoma and a typical carcinoid tumor, which had a colliding pattern of tissue proliferation. There was no intermixing or transition area between the 2 components. The final pathologic diagnosis was collision tumor of the adenocarcinoma and carcinoid tumor. The presence of either tumor individually would not be especially noteworthy, but this collision-type tumor of both histopathologic types in the stomach is, to our knowledge, only the sixth such case in the literature.

Application of liquid-based preparation to fine needle aspiration cytology in breast cancer.

OBJECTIVE To examine the performance of liquid-based cytology (LBC) in breast cytology to confirm the diagnosis of carcinoma. STUDY DESIGN Using cell clusters directly scratched from surgically removed tumor masses, we examined the immunocytochemistry, molecular biology and cytomorphology of the specimens. RESULTS LBC was very useful for gene analysis and evaluating the immunocytochemistry. The cytologic features of LBC were slightly different from those ofa conventional aspiration cytology smear. CONCLUSION LBC is a promising method for improving the standardization ofpreparations in breast cytology, although care should be taken to account for its characteristic cytologic features. The quantitative analysis of HER-2 mRNA correlated with the results of immunohistochemistry.

The Expression of Murine Double Minute 2 (MDM2) on Helicobacter pylori-Infected Intestinal Metaplasia and Gastric Cancer

The overexpression of murine double minute 2 (MDM2) is found in several human tumors, and increased expression of MDM2 inactivates the apoptotic and cell cycle arrest function of p53. Interleukin-16 (IL-16) is a pleiotrophic cytokine and the properties of IL-16 suggest that it involve in the pathophysiological process of chronic inflammatory diseases. In this study, we investigated the expression of MDM2 in intestinal metaplasia and gastric cancer as well as the effect of H. pylori infection and IL-16 on epithelial cell proliferation and MDM2 expression in gastric cells in vitro. The expression of MDM2 on gastric biopsies was studied immunohistochemistry. AGS cells were incubated with a combination of IL-16 and Helicobacter pylori (H. pylori). Gastric epithelial cell proliferation was studied by BrdU uptake and the expressions of MDM2 were studied by ELISA. There was no significant difference on the expression of MDM2 between with and without H. pylori infected chronic gastritis. In H. pylori infected gastric mucosa; the MDM2 expression was higher on intestinal metaplasia and gastric cancer than chronic gastritis. IL-16 administration was increased MDM2 expression and cell proliferation on AGS cells, which was decreased by H. pylori infection. In conclusion, the expression of MDM2 in long-term H. pylori infected gastric mucosa may indicate a risk for carcinogenesis. IL-16 secretion in H. pylori infected mucosa is one of the factors for gastric cancer. The expression of MDM2 on mucosa can be a mediator for gastric cancer.