To explore the expressional information of PIWIL4 in human ovarian cancer, semi-quantitative reverse transcription polymerase chain reaction(semi-qRT-PCR) was used to detect the mRNA expression level of PIWIL1, PIWIL2, PIWIL3 and PIWIL4 in human ovarian clear cell carcinoma ES-2 cells.Meanwhile, in human ovarian cancer and adjacent normal tissues, the mRNA expression of PIWIL4 was determined.Then, PIWIL4-siRNA, which was designed to target PIWIL4 and chemical synthesized, was transfected into ES-2 cells with lipofectamine.MTT assay and clone formation assay were used to investigate the effects of PIWIL4-siRNA on the cell growth activity and proliferation capacity of ES-2 cells.Experimental results showed that the mRNA expression level of PIWIL4 was the highest compared to PIWIL1, PIWIL2 and PIWIL3 in ES-2 cells(P

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30% was considered sensitive and the rate ≤30% was considered resistant;and the 6 specimens were divided into 2 group according to the above standard.cDNA microassay combined with clustering analysis was used to screen for resistance-related genes.Semi-quantitative RT-PCR was used for verification of HDAC1 gene expression.Results:Three of the 6 glioma specimens belonged to the drug resistance group and the other 3 to the drug sensitive group.cDNA microarray analysis combined with cluster analysis screened out 21 genes,with 6 up-regulated and 15 down-regulated.High expression of gene HDAC1 was noticed in all the 6 specimens by semi-quantitative RT-PCR,and the trend was similar to that by microassay.Conclusion:The primary drug resistance of glioma may be associated with the 21 genes screened by cDNA microarray;the detailed mechanisms for drug controlling still need to discussed in the future.

To study the B cell linear epitopes of rabies virus CVS-11 nucleoprotein, peptides were synthesized according to the amino acid sequences of B cell linear epitopes. Linear epitopes predicted by bioinformatics analysis were evaluated with immunological techniques. Indirect enzyme-linked immunosorbent assay showed that titers of antibodies to peptides (355-369 and 385-400 residues of rabies virus CVS-11 nucleoprotein) were above 1:12 800 in mouse sera. The antibodies recognized denatured rabies virus CVS-11 nucleoprotein in Western blot analysis. Purified anti-peptide antibodies recognized natural rabies virus CVS-11 nucleoprotein in BHK-21 cells in indirect fluorescent antibody test. The 355-369 and 385-400 residues of rabies virus CVS-11 nucleoprotein were validated as B cell linear epitopes.
Amino Acid Sequence ; Animals ; Antibodies, Viral ; immunology ; Epitope Mapping ; Epitopes, B-Lymphocyte ; chemistry ; genetics ; immunology ; Female ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Molecular Sequence Data ; Nucleoproteins ; chemistry ; genetics ; immunology ; Rabies ; immunology ; virology ; Rabies virus ; chemistry ; genetics ; immunology

Growth patterns of trichome and contaminative bacteria in Spirulina sp. liquid culture were observed, and it was found that the number of neutral and alkalophilic bacteria was always 105～106 times of that of Spirulina sp. trichome. It would be very difficult to get real pure Spirulina sp. strain by classical methods of dilution plate, capillary and single trichome selecting methods. A great deal of contaminative bacteria was washed out by two pretreatment processes. Low speed centrifugation was designed to wash the strains which usually deposit at bottom, and filtration method was designed to treat the strains usually floating at surface. Sandwich plate and dilution plate were designed for the purification of the mobile strains and non-mobile strains, respectively. A lot of strains were purified by the above processes and pure single trichome formed pure colonies on plates.

The high-copy-number plasmid pcDNA3.1+ is unstable within S almonella typhimurium. A novel plasmid pmcDNA3.1+ was constructed by removin g the promoter sequence of ampicillin resistance gene (bla gene) in plasmid pcDNA3.1+. In contrast to pcDNA3.1+, pmcDNA3.1+ was stable within Salmonel la typhimurium SL7207 in LB medium with or without ampicillin. Further experi ments showed the ?-lactamase activity of Salmonella typhimurium SL7207(pmc DNA3.1+) was apparently lowered than that of Salmonella typhimurium SL7207( pcDNA3.1+) and the high ampicillin concentration was maintained longer in LB me dium culturing Salmonella typhimurium SL7207(pmcDNA3.1+). When mice were a dministered with Salmonella typhimurium SL7207(pmcDNA3.1+) intraperitoneall y, more than 95% of Salmonella cells separated from the spleen still harbore d the plasmid pmcDNA3.1+ 7 days later; but 99% of Salmonella cells lost the plasmid pcDNA3.1+ at day 3 in mice innoculated with Salmonella typhimurium SL7207(pcDNA3.1+). By lowering the expression of bla gene, the rapid deco mposition of ampicillin in LB medium was avoided and the metabolic pressure was relieved for the host cells. This method offers a solution for the problem of t he instability of high-copy-number plasmid within Salmonella typhimurium.

Objective:To determine the difference of miRNA expression profiles in 6 types of human cancer cells by microarray technique.Methods:The microarray was prepared,with contained 210 oligonucleotides,including 206 probes complementary with human and mouse miRNA sequences and 4 positive control oligos.MiRNAs were extracted from HeLa(human cervical cancer epithelial cells),MCF-7(human breast cancer cells),A549(human lung adenocarcinoma cells),HT-29(human colonic cancer cells),ES-2(ovarian carcinoma cells),and K562(chronic myelogenous leukemia cells) cells and were labeled with Cy3 for hybridization to the miRNA microarray.The slides were scanned by ScanArrayTMExpress1.0 and images were analyzed by ScanArray3.0 and Cluster3.0;the results were confirmed by Northern blotting and RT-PCR.Results:Totally 115 miRNAs were found to be differentially expressed in the 6 cancer cell lines,with miR-21 expression up-regulated and miR-125b,let-7 expression down-regulated.The expression of miR-17-5p and miR-20a was in cluster and was more higher in ES-2 cells than in other cells.HeLa and MCF-7 cells were located on a single branch of the dendrogram in cluster analysis.Northern blotting showed that both pre-and miR-17-5p expressed in K562,pre-miR-17-5p was weakly expressed in A549 and ES-2 cells,and obvious pre-miRNA expression and weak miRNA expression were noticed in MCF-7,HeLa,and HT-29 cells.RT-PCR showed that expression of pre-miR-17-5p in K562 cells was higher than those in other cells.Expression of miR-21was high in all 6 cell lines and the highest expression was seen in A549 cells.Conclusion:Microarray can be used to detect miRNA expression files in cancer cells,which contributes to the study of the relation between miRNA and tumor.

OBJECTIVETo explain the essence of pungent-hot herb property express according to in vivo and in vitro studies on its effect on calmodulin on the base of the observation of the adjustment in hypothalamic-pituitary-gonad axis functions of Aconiti Lateralis Radix Praeparata, Curculiginis Rhizoma, Cinnamomi Cortex and bitter-cold herb Phellodendri Chinensis Cortex in rats under the state of yang deficiency.

METHODThe yang-deficient model was duplicated by intramuscularly injecting hydrocortisone sodium succinate powder injection. After the intervention with Aconiti Lateralis Radix Praeparata, Curculiginis Rhizoma, Cinnamomi Cortex and bitter-cold herb Phellodendri Chinensis Cortex for seven days, TSH, T3, T4, 17-OHCS, COR, T, E2 of hypothalamus-pituitary-target gland axis and other relevant indexes were detected. The calmodulin expression in livers and L02 cells cultured in vitro was detected by using Western blot.

RESULTPungent-hot herbs Aconiti Lateralis Radix Praeparata, Curculiginis Rhizoma, Cinnamomi Cortex can significantly correct indicators of hypothalamic-pituitary-gonad axis and calmodulin, whereas the bitter-cold herb Phellodendri Chinensis Cortex showed no obvious effect.

CONCLUSIONThe pungent-hot herb property expression may be closely related to calmodulin.

Animals ; Calmodulin ; metabolism ; Drugs, Chinese Herbal ; administration & dosage ; chemistry ; Gonads ; drug effects ; metabolism ; Hypothalamo-Hypophyseal System ; drug effects ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Yang Deficiency ; drug therapy ; metabolism

Objective To investigate the relationship between serum secreted frizzled-related protein 4 (SFRP4) and the first-phase of glucose-stimulated insulin secretion from pancreatic β cell under different glucose tolerance statuses. Methods Fifty-six patients with newly diagnosed type 2 diabetes mellitus ( T2DM group), 52 patients with impaired glucose tolerance (IGT group), and 42 subjects with normal glucose tolerance (NGT group) underwent intravenous glucose tolerance test. Fasting serum SFRP4 and interleukin ( IL)-1β were assayed by ELISA. Acute insulin response ( AIR), the area under the curve of the first-phase (0-10 min) insulin secretion (AUC), glucose disposition index(GDI), homeostasis model assessment for β cell function index(HOMA-β), and insulin resistance index(HOMA-IR) were calculated. Results (1) The levels of SFRP4 and IL-1β in T2DM group and IGT group were significantly higher than that in NGT group [(184. 38 ± 61. 34 or 141. 64 ± 40. 46 or 95. 46 ± 20. 13)ng/ ml, P<0. 01]. AIR, AUC, and GDI in T2DM group and IGT group were significantly lower than those in NGT group(P<0. 01), and these results were more significantly reduced in T2DM group compared with those in IGT group. (2) SFRP4 was negatively correlated with AIR, AUC, GDI, HOMA-β (P<0. 01), and positively correlated with fasting plasma glucose, 2 h plasma glucose after glucose loading, HbA1C , IL-1β, and high sensitive C-reactive protein(P<0. 01). (3) Multiple stepwise regression analysis showed that AUC, HOMA-IR, and serum IL-1β level were independently associated with SFRP4. Conclusion The concentration of serum SFRP4 is closely correlated with the glycolipid metabolic disorder, the first-phase of glucose-stimulated insulin secretion, and chronic low-grade inflammation. SFRP4 may be involved in the mechanism of β cell dysfunction in type 2 diabetes mellitus.

Objective To investigate the endoscopic findings and treatment of gastrointestinal neu-roendocrine neoplasms.Methods Endoscopic manifestation and treatment were analyzed retrospectively in 72 patients who were diagnosed as having gastrointestinal neuroendocrine neoplasms by endoscopy and pathol-ogy from May 2011 to December 2014.Results In 72 patients of gastrointestinal NEN,rectum was most commonly involved (48 patients,66.7%),followed by the stomach (16 patients,22.2%),duodenum, esophagus and the ileocecal valve.There were certain features in rectal neuroendocrine neoplasms under endoscopy,mostly manifested by submucosal tumors.But gastric,esophageal,and small intestine NEN man-ifestations showed various forms,without specific typical features,and easy to be misdiagnosed.Some patients diagnosed as having NEN G1 or G2 underwent EMR,ESD or laparoscopy combined with endoscopy resection.Patients diagnosed as having NEN G2 or neuroendocrine carcinoma by pathology received surgical resection,chemotherapy or palliative treatment.All 50 patients underwent endoscopic treatment successful-ly.Perforation occurred in one duodenal bulbar G1 patient during ESD.No bleeding occurred during and after the operation.All patients after treatment were followed up for 15.6 +13.2 months on average with no recurrence or metastasis.Conclusion Manifestations of gastrointestinal neuroendocrine neoplasms vary under endoscopy.Some tumors that locate in mucosa or submucosa with diameter less than 1cm can be resec-ted through EMR or ESD.