Background and purpose:Survivin,a member of the inhibiter of apoptosis gene family,has been detected in fetal tissue and in variety of human malignancies.In the current study,we not only investigated the expression of a novel inhibitor gene of apoptosis,survivin in Cervixneoplasms but also studied its relationship between the expression of P53 and HPV-16. Methods:Using streptavidin-bintin peroxidase(SP)method,We examinined the expression of the proteins survivin , p53 and HPV-16 in 21 normal cervical tissues ,33 CIN and 72 cervical cacinoma tissues.Results:Survivin was expressed in 52 of 72(72.2%) cases of cervical carcimomas ,however,it was not expressed in normal cervical tissues and it was expressed in6 of 33(18.18%)cases of CIN. Overexpression of survivn was related to the tumor grade and clinical stage.Survivin positive cases were strongly associated with p53 expression (P

Background and purpose:MMMT is known as a rare malignancy in gynecological tumor. Because of its difficulty in diagnosis and treatment, the prognosis is extremely poor. This study was to discuss the clinicopathologic feature and PRA,PRB expressions of uterine MMMT. Methods:We analysed clinicopathologic data and the expressions of PRA,PRB by immunohistochemical staining on 17 cases of uterine MMMT, and 11 patients were followed up. Results:①The patients usually presented with abnormal vaginal bleeding with no specifi city clinically. ②The morphological changes were various and complicated, including epithelial and mesenchymal components and a variety of inter-permeated and transitional tissue elements.③The positive rate of PRA in stageⅠ and stageⅡ were 66.7% and 40%, and PRB in stage Ⅰ and stageⅡwere 55.6% and 20%, respectively. ④The mean survival time in stageⅠ,Ⅱ and Ⅲ were 43.8 months (32-59), 34.25 months (19~41) and 5 months, respectively. Conclusion:The diagnosis of uterine MMMT was mainly based on tissue morphology; the development of uterine MMMT might be related with the loss of PRA and PRB ; the clinical stage and the expression of PRA and PRB might be the prognostic factors for uterine MMMT.

Objective A systematic review of implementation outcomes of the essential medicine system in China to identify scientific evidences for a better system.Methods A systematic review is made to extract data from the research papers on outcomes of the essential medicine system,followed by an analysis and description of such data.Results Of the 87 papers included,most of them focused on primary care institutions,while four of them on residents or patients,and one of them on pharmaceutical enterprises.The study found the medical institutions with rising availability of essential drugs,lowered medicine costs,rising or dropping business volume,and apparent drop of out-of-pocket expenses for patients.These have encouraged rational drug use.Evidences in hand indicate expected outcomes from the essential medicine system.Conclusion Current researches on the system focus on primary care institutions in developed areas in China,lacking rigorous design.Studies of broader scale,further depth and more rigorous designs of the implementations of the system are recommended for evaluation of the impacts and outcomes of the system on various stakeholders of the policy.

Background and purpose:The genesis and progression of cervical cancer are closly related to E6 and E7 oncogenes of HPV.Special ribozyme and antisense oligonucleotide could inhibit the expression of E6 or E7.but both of them were not the best methods.The aim of this study was to investigate the inhibitive effect of chemically synthetic siRNA on E6 gene in the cervical cancer cell line SiHa and the following biological changes of SiHa cells.Methods:Three specific siRNAs targeting to HPV16 E6 were synthesized and transfected into SiHa cells by lipofectamine.The level of E6 mRNA was tested by RT-PCR,proliferation activities,the cell cycle distribution,expression of p53 protein and the ultramicrostructure changes of SiHa cells were detected by MTT,FCM,immunochemistry and TEM respectively.Results:All the 3 siRNAs could inhibit the level of E6 mRNA singificantly,among which siRNA1 was most effective.Decreased cellular proliferation activity was observed by MTT,especially when the cells were transfected with siRNA1 at 50 nmol/L concentration.Flow cytometry revealed obvious G1 arrest in cell cycle.The expression of p53 protein in transfected cells(0.75?0.06)was increased compared to the control group(0.43?0.03),the difference was singificant.Cell-substance concentration and vacuolus were found in endochylema by TEM.Conclusions:Chemically synthetic siRNA can interfere in the expression of E6 mRNA in SiHa cells specifically and induce the biological changes of the target cells.

Objective To explore the value of peripheral blood samples in KRAS mutation testing of colorectal cancer patients and the correlation between the number of circulating tumor cells and KRAS mutation testing.Methods We detected KRAS mutation using amplification refractory mutation system PCR method in paraffin embedded tissues and matched peripheral blood samples obtained from 112 colorectal cancer patients and 10 proctitic peripheral blood samples in Affiliated Hospital of Jiangnan University between 2013 and 2014.Meanwhile,immunofluorescence in situ hybridization method was used to count the circulating tumor cells in peripheral blood samples and proctitic control samples.Results Among the 112 colorectal cancer samples tested,25 cases of peripheral blood samples found KRAS mutation (41.1％) and which was 46 in formalin fixed paraffin embedded tissues testing (22.3 ％),with a significant difference (x2 =40.12,P ＜ 0.001).One case with KRAS wild type in formalin fixed paraffin embedded tissues was mutation type in peripheral samples.In another case,mutation site was different in different kinds of samples.The sensibility of KRAS mutation testing was 73.3％,41.9％ and 16.7％ when the number of circulating tumor cells was more than 15,5 to 15,and 1 to 5,respectively,with significant differences (x2 =23.70,P ＜ 0.001).No KRAS mutation and no circulating tumor cells were found in 10 proctitic control samples.Conclusion We find high specificity in KRAS mutation testing of peripheral blood samples.but the accurate rate is not satisfying.KRAS mutation testing in peripheral blood samples may be an optional choice to test KRAS mutations for colorectal cancer patients who were not subjected to surgery.The sensibility of KRAS mutation testing in peripheral blood samples has a corretion with the number of circulating tumor cells.

Background and purpose:Loss or altered expression of Ras association domain family 1A gene (RASSF1A) through DNA methylation has been associated with the pathogenesis of a variety of cancers, which suggests the tumor suppressor function of this gene. This study aimed to explore the effect of DNA methyltransferase inhibitor 5-Aza-2’deoxycytidine (5-Aza-dc) on demethylation and expression of RASSF1A in cervical cancer cell lines. Methods:HPV positive cervical cancer cell lines HeLa and Caski, HPV negative cell lines HT-3 and C-33A were treated with two different concentration of 5-Aza-dc (5 μmol/L, 10 μmol/L). MSP (methylation-specific PCR) and Bisulfite genomic sequencing PCR (BGS) combined with TA clone were used to investigate methylation status of RASSF1A gene promoter and exon 1 before and after treatment of 5-Aza-dc. RASSF1A gene mRNA expression was detected by RT-PCR. Results:Two HPV positive cell lines showed hypomethylated RASSF1A promoters and expressed RASSF1A mRNA, and after treatment with 5-Aza-dc, the mRNA expression of RASSF1A did not change significantly (FHeLa=3.003, P=0.125; FCaski=0.045, P=0.956). Two HPV negative cervical cancer cell lines showed hypermethylation status of RASSF1A promoter and silenced RASSF1A. After treatment with 5-Aza-dc, demethylation occurred in the promoter region of RASSF1A gene, which subsequently induced re-expression of this gene in HT-3 and C-33A. The F test (FHT-3=18.002, P=0.03;FC-33A=17.179, P=0.03) and LSD-t test (P<0.05) demonstrated that significant difference in the expression of RASSF1A was found upon two different concentrations drug treatment.Conclusion:The methylation status of promoter and exon 1 of RASSFIA gene in HPV positive and HPV negative cervical cancer cell lines are different. The promoter hypermethylation is correlated with RASSF1A gene expression in HPV negative cervical cancer cell line HT-3 and C-33A, and plays a key role in RASSF1A silencing. 5-Aza-dc may effectively reverse the methylation status of RASSFIA gene promoter in cervical cancer HT-3 and C-33A cells and reactivate gene expression silenced by aberrant hypermethylation in a dose-dependent manner within certain extent.

Background and purpose:Studies have proved that the serine/threonine kinases 31 (STK31) gene plays important roles in human cancers. TheSTK31 gene expression was demonstrated to be regulated by the methylation status of its promoter/exon1 region. Viral infection was revealed to be associated with the hypermethylation of some tumor suppressor genes in some tumor samples. The purposes of this paper were to study the roles ofHPV16E6,E7, orE6/E7 oncogenes in methylation status and expression of theSTK31 gene, and potential effects of DNA methyltransferases (DNMTs) onSTK31 gene methylation status.Methods:Ectopically-expressed HPV16 E6, E7, or E6/E7 cells were estab-lished by transfectingHPV16E6,E7, orE6/E7 oncogenes with lentivirus vectors into HPV-negative cervical cancer cell lines HT-3 and C33A. Bisulfite genomic sequencing PCR (BGS) combined with TA clone and MSP (methylation-specific PCR) were used to analyze methylation status of theSTK31 gene promoter/exon1 region in HPV-positive cervical cancer cell lines (HeLa, SiHa, CaSki), HPV-negative cervical carcinoma cell lines (C33A, HT-3) and the transfected cells. The mRNA and protein expression of STK31, DNMT1, DNMT2, DNMT3a, DNMT3b and DNMT3L were detected by RT-PCR and Western blot.Results:Transfection efficiency was tested by Western blot, which showed that the transfected cells successfully expressed E6, E7, or E6/E7 proteins, respectively. TheSTK31 gene promoter/exon1 was hypomethylated in HPV-positive cell lines HeLa, SiHa and CasKi resulting in detection of mRNA and protein expression.STK31 gene promoter/exon1 showed hypermethylation leading to silenced expression in the two HPV-negative cervical cancer cells HT-3 and C33A. Compared with primary HT-3 and C33A cells, the methylation status ofSTK31 promoter/exon1 was down-regulated that led to expression of STK31 in the ectopically-expressed HPV16 E7 and E6/E7 cells. Expressions of DNMT1,DNMT3a andDNMT3b genes at the level of transcription were higher in C33AE6/E7 and HT-3E6/E7 cells than those in C33A-vector and HT-3 vector cells, respectively (P<0.001). mRNA levels of DNMT1, DNMT3a and DNMT3b were higher in HPV16-positive cervical cancer tissues than those in HPV-negative cervical cancer tissues, respectively (t=5.997,P<0.001;t=6.743,P<0.001;t=7.926,P<0.001). DNMT2 mRNA level was lower in C33AE6/E7 and HT-3E6/E7 cells than those in C33A-vector and HT-3 vector cells, respectively (t=7.451,P<0.001;t=2.451,P<0.05). mRNA level of DNMT2 gene was lower in HPV16-positive cervical cancer tissues than in HPV-negative cervical cancer tissues (t=9.134, P<0.001). There was no statistically significant difference in expression levels of DNMT3L mRNA between cervical cancer cell lines before and after transfection, or HPV16-positive and HPV-negative cervical cancer tissues, respectively (P>0.05, data not shown).Conclusion:HPV infection leads to the down-regulated methylation status ofSTK31 promoter/exon1 that results in the expression of STK31.STK31 gene expression is regulated by methylation status of its promoter/exon1 region. HPV16E7 andE6/E7 oncogenes may influence the methylation status ofSTK31 gene promoter/exon1 region by regulating the expression of DNMT2.

AIM: To explore the structural changes of brain microvasculature and mechanism in microvascular lesion after focal cerebral ischemia with reperfusion. METHODS: Using the techniques of immunohistochemical staining, in situ hybridization,optical microscopy and transmission electron microscopy, the expression of uPA, uPA mRNA, and changes in miocrovascular structure were examined in ischemic focus and perifocal areas after focal cerebral ischemia 2 hours with various time points of reperfusion in stroke-prone renovascular hypertensive rats (RHRSP). RESULTS: The brain edema and hemorrhage were severe 12 hours to 3 days after reperfusion. Ultrastructural change showed that the damage characterizations of the basement membrane were degradation, defection, and exfoliated of basement membrane, while uPA, which attack the basememt membrane around cerebral capillaries and extra-cellular matrix, and uPA mRNA expression increased significantly in ischemic and perifocal areas 12 hour to 3 day after reperfusion. CONCLUSION: The main pathologic mechanism of brain edema and hemorrhage after cerebral ischemia with reperfusion may result from the basement membrane lesion of brain microvasculature. The increase in the expression of uPA in reperfusion area may be the main cause of the basement membrane lesion .

OBJECTIVE: Cervical cancer is one of the most common malignant tumors. Our previous results showed that long non-coding RNA (lncRNA) XLOC_006390 plays an important role in cervical cancer. In this study, we have explored the mechanism of action of lncRNA XLOC_006390. METHODS: LncRNA XLOC_006390 was proposed to exercise its function as a competing endogenous RNA (ceRNA), and its potential targeted miRNAs was predicted through the database LncBase Predicted v.2. Two miRNAs, miR-331-3p, and miR-338-3p, were chosen for the study. Expression of miRNAs and lncRNA in cervical cancer cells and tissues was detected by reverse transcription polymerase chain reaction. To determine the correlation, silencing of XLOC_006390, over-expression of miR-331-3p, and miR-338-3p was performed in SiHa and Caski cell lines, respectively. RESULTS: Based on the interactive effect between miRNA and lncRNA, miR-331-3p and miR-338-3p were significantly downregulated in cervical cancer cells and tissues, and their expression levels were negatively related to that of lncRNA. Our results also showed that the expression of miR-331-3p target gene NRP2, miR-338-3p target genes PKM2, EYA2 was significantly downregulated when the XLOC_006390 was knocked down. Further, XLOC_006390 was found to facilitate cervical cancer tumorigenesis and metastasis by downregulating miR-331-3p and miR-338-3p expression. CONCLUSION: Taken together, our study demonstrated that XLOC_006390 may serve as a ceRNA and reversely regulates the expression of miR-331-3p and miR-338-3p, thus facilitating cervical cancer tumorigenesis and metastasis.
Carcinogenesis* ; Cell Line ; MicroRNAs ; Neoplasm Metastasis* ; Polymerase Chain Reaction ; Reverse Transcription ; RNA ; RNA, Long Noncoding* ; Uterine Cervical Neoplasms*

Objective To investigate the changes of the levels of TNF and IL 8 in blood and in gastric mucosa and their mRNA expression after liver ischemia reperfusion (I/R). Methods 130 rats were randomly divided into 3 groups: (1) sham operation group(control group, n=10);(2)liver ischemia reperfusion group (n=60), including ischemia for 20min and 40min, and reperfusion for 1h, 24h and 72h respectively; (3) liver I/R rats treated with famotidine group (n=60). ALT and AST were detected after operation, pathomorphological changes of gastric mucosa were studied by light microscope and transmission electronic microscope, the concentrations of TNF and IL 8 in blood and gastric mucosa were measured by radioimmunoassay (RIA), TNF mRNA and IL 8 mRNA were detected by in situ hybridization. Results After liver I/R,inflammatory damage appeared in gastric mucosa, TNF and IL 8 concertrations increased significantly(P