[Role of minocycline in an immature rat model of hypoxic-ischemic brain damage].

OBJECTIVE: To establish a model of immature rat hypoxic-ischemic brain damage (HIBD) which was expected to be similar to periventricular leukomalacia in human preterm infants pathologically and neuroethologically, and to investigate the role of minocycline (MN) in this model. METHOD: Totally 192 Sprague-Dawley rats (postnatal day 2, P(2)), of either sex, were randomly divided into 4 groups: normal-group, sham operation group, HIBD-group, HIBD + MN group, each group had 48 rats. HIBD group and HIBD + MN group survived the left common carotid artery (CCA) ligation followed by 4h exposure to 8% O(2). Rats in sham operation group only survived the left CCA isolation. Rats in normal group were not treated with anything. In HIBD + MN group, the rats were treated with intraperitoneal injection of minocycline 45 mg/kg, immediately after HI and every 24 h for 2 days. Brain tissues were collected on day 3, 1 week, 2 weeks, 4 weeks after HI, for hematoxylin-eosin staining and histological scoring. Frozen sections of the brains were stained with anti-O4, anti-O1 immunohistochemistry on day 3 after HI, and MBP immunohistochemistry 2 weeks after HI. Rats in the four groups underwent neuroethologic examination 4 weeks after HI. RESULT: In the HIBD group, there were pathological changes in the periventricular white matter. The pathological changes were milder in HIBD + MN group; There was no statistically significant difference between the normal group and HIBD + MN group in the number of positively stained O4 cell (P > 0.05). The number of positively stained O4 cell in the HIBD group was significantly reduced, compared with that of normal group, sham operation group, and HIBD + MN group (23.67 ± 12.00 vs. 52.89 ± 10.68, 39.28 ± 11.78, 41.63 ± 8.41, P < 0.05). The differences in the number of positively stained O1 cell among the normal group, sham operation group, HIBD group and HIBD + MN group had no statistical significance (P = 0.093). The numbers of myelin basic protein (MBP) positively immunostained fiber bundles in the HIBD + MN group were significantly less than that of the normal group and sham operation group (P < 0.05). The numbers of MBP positively immunostained fiber bundles in the HIBD group were significantly less than that of the normal group, sham operation group, and HIBD + MN group (14.71 ± 7.42 vs. 36.67 ± 6.50, 35.50 ± 3.24, 26.33 ± 5.92, P < 0.05). The HIBD group had long-term neuroethologic abnormality. There was no statistically significant difference in the inclined plane test, hanging test and cylinder test among the HIBD + MN group, normal group, and sham operation group (P > 0.05). The scores of the HIBD group had statistical significantly among the normal group, sham operation group and HIBD + MN group (P < 0.05). In the open field test, there was no statistically significant difference between the HIBD group and HIBD + MN group (P = 0.772), but there was significant difference between these two groups and the normal group, sham operation group (P < 0.05). CONCLUSION: Minocycline protects the pre-oligodendrocyte and has protective effects in terms of long-term neuroethology.

Prevalent expression of MHC class I chain-related molecule A in human osteosarcoma.

MHC class I chain-related molecule A (MICA) is one of the major ligands for activating immune-receptor NKG2D which is expressed on NK cells and cytotoxic T lymphocytes. The release and sustained expression of MICA protein can impair NKG2D-mediated cytotoxic activity by reducing NKG2D receptor on immune effector cells. The aim of the study was to investigate the expression and release of MICA in human osteosarcoma. RT-PCR, immunohistochemistry, western blotting and flow cytometry were used in analyzing the expression of MICA. Serum level of soluble MICA was quantitated by ELISA. Our data showed that MICA is prevalently expressed in osteosarcoma both in mRNA and protein level. Upregulation of MICA was found in osteosarcoma compared with benign tumors and normal bone tissues. Higher level of soluble MICA in serum can be detected in osteosarcoma patients. In conclusion, prevalent expression of MICA and higher serum level of soluble MICA may suggest a deficiency of MICA-NKG2D mediated immunosurveillance in osteosarcoma patients. Restoring the expression of NKG2D receptor on immune effector cells may contribute to a therapeutic strategy for human osteosarcoma.

[Expression of membrane/soluble MHC class I chain-related molecule A and NKG2D receptor in osteosarcoma and its clinical significance].

OBJECTIVE: To study the expression of membrane MICA (mMICA), soluble MICA (sMICA) and NKG2D receptor in cases of osteosarcoma and to analyze its clinical significance. METHODS: Expression of mMICA in osteosarcoma tissue of 43 cases was detected with immunohistochemistry. Expression of NKG2D in peripheral blood lymphocytes of 16 cases was analyzed by flow cytometry. Serum level of soluble MICA (sMICA) was measured by ELISA. RESULTS: mMICA was widely expressed in osteosarcoma tissue (37/43). Expression of NKG2D in peripheral blood lymphocytes was significantly decreased. High levels of mMICA and NKG2D expression were associated with better differentiation and earlier tumor stage of osteosarcoma (P < 0.05). A significant increase in serum level of sMICA was demonstrated in patients with metastasis and advanced tumor. CONCLUSIONS: The mMICA expression in tumor tissue, NKG2D expression in peripheral lymphocytes and serum sMICA level correlate with the differentiation and stage of osteosarcoma. These parameters may thus represent potential diagnostic and prognostic markers in patients with osteosarcoma. Manipulation of the MICA-NKG2D pathway may become a target of immunotherapy for osteosarcoma.

[Evolutive characters of oval cells in experimental hepatocarcinogenesis].

BACKGROUND & OBJECTIVE: The role of oval cells in hepatocarcinogenesis is unclear yet. This study was to explore the correlation of oval cells to hepatocarcinoma through dynamic observation on evolutive characters of oval cells in experimental hepatocarcinogenesis. METHODS: Male SD rats were fed with 3o-me-DAB to establish an animal model of experimental hepatocarcinoma. Evolutive characters of oval cells in liver tissue during experimental hepatocarcinogenesis was dynamically observed with routine HE staining, Alcian blue staining, and immunohistochemistry. RESULTS: Oval cells (OV-6-positive) appeared sparsely around the portal tract in the 4th week of tumor-induction. In the 8th and the 14th weeks, OV-6-positive cells were increased gradually and expanded into hepatic lobules; the hepatic tissue was divided as pseudo-lobule-like. Till the 17th and the 24th weeks, carcinoma foci were formed, meanwhile, the total amount of oval cells were decreased, and OV-6-positive cells were observed in carcinoma foci. On Alcian blue-stained preparations, two distinct histologocal types of carcinoma foci could be seen: cholangioepithelial carcinoma foci were positive and hepatocellular carcinoma foci were negative. CONCLUSION: Oval cells, as intrahepatic stem cells, might play an important role in hepatocarcinogenesis.

[Induced differentiation of rat hepatic oval cells in-vitro by combined hepatocyte growth factor and epidermal growth factor treatment].

OBJECTIVE: To characterize the biologic featrues of hepatic oval cells and their protein expression profiles during induced differentiation in vitro. METHODS: Rat hepatic oval cells were treated with epidermal growth factor (EGF) and hepatocyte growth factor (HGF) in vitro, followed by morphological and molecular marker assessment by electromicroscopy, immunocytochemistry, RT-PCR and protein expression chip technology. RESULTS: Ten weeks after induction, the levels of GST-P mRNA and M2-PK mRNA were significantly reduced, whereas those of ALB and CK18 were elevated. Significant variations of expression was seen in 8 protein species during the course of the induced differentiation. CONCLUSION: Combined EGF and HGF treatment in vitro induces cell differentiation of hepatic oval cells, a process in which 8 protein species may play some regulatory roles.

Correlation of AIB1 overexpression with advanced clinical stage of human colorectal carcinoma.

AIB1, a member of the steroid receptor coactivator 1 family, has been cloned on 20q12 and is a candidate oncogene in human breast cancer. It is commonly amplified and overexpressed in several types of human cancers. In this study, we examined the expression of AIB1, as related to clinicopathologic features, in 85 human colorectal cancers (CRCs). The status of the number of AIB1 copies, p53 expression, and DNA ploidy was also analyzed. The overexpression of AIB1 was detected in 35% of CRCs. Amplification of AIB1 was observed in 10% of CRCs. In addition, the overexpression of AIB1 was observed more frequently in CRCs in later clinical stages (T3 N1 M0/T3 N0 2M1), compared with that in T3 N0 M0 stage (P < .05). These results suggest that overexpression of AIB1 might provide a selective advantage for the developmental growth and/or progression of subsets of CRCs. In addition, a significant correlation (P < .05) of overexpression of AIB1 with p53 overexpression as well as with aneuploid DNA content was observed in these CRCs. The overexpression of p53 was also correlated significantly with CRC DNA ploidy (P < .05). Furthermore, there was a substantial population of CRCs showing overexpression of both AIB1 and p53 protein and all had aneuploid DNA content; most of these were in the later clinical stage. These findings suggest a possible convergence of AIB1 with a pathway involving p53, which might induce chromosomal instability and affect the clinical phenotype of a subset of CRCs.

Oncogenic role of clusterin overexpression in multistage colorectal tumorigenesis and progression.

AIM: To investigate the expression pattern of clusterin in colorectal adenoma-carcinoma-metastasis series, and to explore the potential role of clusterin in multistage colorectal tumorigenesis and progression. METHODS: A colorectal carcinoma (CRC)-tissue microarray (TMA), which contained 85 advanced CRCs including 43 cases of Dukes B, 21 of Dukes C and 21 of Dukes D tumors, were used for assessing the expression of clusterin (clone 41D) and tumor cell apoptotic index (AI) by immunohistochemistry and TUNEL assay, respectively. Moreover the potential correlation of clusterin expression with the patient's clinical-pathological features were also examined. RESULTS: The positive staining of clusterin in different colorectal tissues was primarily a cytoplasmic pattern. Cytoplasmic overexpression of clusterin was detected in none of the normal colorectal mucosa, 17% of the adenomas, 46% of the primary CRCs, and 57% of the CRC metastatic lesions. In addition, a significant positive correlation between overexpression of clusterin and advanced clinical (Dukes) stage was observed (P<0.01). Overexpression of cytoplasmic clusterin in CRCs was inversely correlated with tumor apoptotic index (P<0.01), indicating the anti-apoptotic function of cytoplasmic clusterin in CRCs. CONCLUSION: These data suggests that overexpression of cytoplasmic clusterin might be involved in the tumorigenesis and/or progression of CRCs. The anti-apoptotic function of cytoplasmic clusterin may be responsible, at least in part, for the development and biologically aggressive behavior of CRC.

Up-regulated expression of cytoplasmic clusterin in human ovarian carcinoma.

BACKGROUND: Recently, tumorigenic roles of the clusterin gene in several human malignancies have been suggested, but its potential role in the development and progression of ovarian carcinoma is unclear. METHODS: In the current study, immunohistochemistry was used to examine the expression status of clusterin in 10 normal ovaries, 20 ovarian cystadenomas, 15 borderline ovarian tumors, and 240 ovarian carcinomas (nonmetastatic and metastatic) by tissue microarray. In addition, the apoptotic index of each tumor was assessed with a terminal deoxyuridine triphosphate nick-end labeling assay. RESULTS: Positive staining for clusterin in different ovarian tissues was observed primarily a cytoplasmic pattern. Cytoplasmic overexpression of clusterin was detected in none of the normal ovaries, in 17% of cystadenomas, in 38% of borderline tumors, and in 58% of invasive ovarian carcinomas. A significant association was observed (P < 0.001) between the overexpression of clusterin and late clinical stage according to the International Federation of Gynecology and Obstetrics staging system. In addition, the overexpression of clusterin was detected more frequently in metastatic lesions than that in their matched primary tumors. The current results also provided evidence that the overexpression of cytoplasmic clusterin in carcinomas was correlated inversely with the tumors' apoptotic index, demonstrating an antiapoptotic function of cytoplasmic clusterin in ovarian carcinomas. CONCLUSIONS: The current results suggested that the overexpression of cytoplasmic clusterin may represent an acquired malignant phenotypic feature of ovarian carcinoma and may be one of the important factors in determining the aggressive nature of ovarian carcinoma.

Heterogeneous expression and association of beta-catenin, p16 and c-myc in multistage colorectal tumorigenesis and progression detected by tissue microarray.

Most colorectal carcinomas (CRCs) arise from adenomas through an archetypal pathogenic pathway, the adenoma-carcinoma-metastasis sequence. Aberrant expression of beta-catenin, p16, E-cadherin and c-myc appears to have played important roles in the development and/or progression of CRC, but their precise distribution pattern and associations in different pathologic loci along CRC's pathogenic pathway have not been thoroughly examined. In this study, a tissue microarray (TMA) containing 85 advanced CRCs in different Dukes stages was constructed. In each of 85 cases, tissue specimens from normal mucosa and primary carcinomas in different layers of the bowel wall were included in the TMA. Tissue specimens from matched adenoma, lymph node metastases and distant metastases were obtained from 22, 21 and 21 cases, respectively. Expression patterns of beta-catenin, p16, E-cadherin and c-myc were evaluated by immunohistochemistry. The results revealed that nuclear expression of beta-catenin, p16 and c-myc was quantitatively increased from normal mucosa to premalignant adenoma, primary carcinoma and lymph node metastatic carcinoma; the frequency of nuclear overexpression of beta-catenin and p16 in lymph node metastases was significantly higher than that in distant metastases (p < 0.05). These results suggest an association between nuclear overexpression of beta-catenin and/or p16 and CRC lymph node metastasis but not distant metastasis. The results also showed that correlative high nuclear expression of beta-catenin and c-myc was observed in primary carcinomas involving the serosa and lymph node metastases (p < 0.05) but not in other pathologic regions of CRCs, suggesting that the tumor microenvironment in different pathologic loci of colorectal tumorigenesis and progression may influence c-myc responsiveness to beta-catenin/Tcf activation.