Objective To establish a SYBR Green based real-time fluorescence quantitative PCR method for detecting human Annexin Ⅱ mRNA expression,and to detect the level of Annexin Ⅱ mRNA in human breast cancer cells MCF-7 and MDA-MB-231.Methods The specific primers were designed according to the conserved sequence of human Annexin Ⅱ gene.Total RNAs were extracted from human breast cancer cells(MCF-7,MDA-MB-231),then RNAs were transcribed reversely into cDNAs.The plasmid standards were constructed.The relative expression levels of human Annexin Ⅱ mRNA in human breast cancer cells were detected by this method.Results The square(r2 )of correlation coefficient of the standard curve in this method was 0.997,the melting curve analysis showed the single peak.The the intra-batch and inter-batch variable coefficients in the pGM-T Annexin Ⅱplasmid standard substance were 6.2%,7.8% and 9.1%,12.3% respectively.The further study indicated that AnnexinⅡ mRNA in MDA-MB-231 was higher than that in MCF-7(P<0.01).Conclusion The established SYBR Green real time fluorescence quan-titative PCR method for detecting human AnnexinⅡ is of good specificity and repeatability and can be used for quantitatively detec-ting AnnexinⅡ mRNA in breast cancer cells.

Objective Through studying change of serum level of IL-10 in patients of severe trauma with haemorrhagic shock,to investigate effect of severe trauma with haemorrhagic shock bypass on anti-infiammatory reaction in patients.Methods Menbers of experimental group are 43 patients of Severe trauma(ISS scope≥16).43 patients scheduled for attending with haemorrhagic shock are divided into simple severe trauma group(group B)and severe trauma with haemorrhagic shock(group C);and healthy control group(group A)is made up of randomly seleeted 20 healthy people of medical examination.Blood samples for cytokines and organ function were collected from the vetn of menbers of experimental group(B and C)with limosis at the following time:that day of injury,(T1),3th (T2),5th(T3),and 14th(T4)day after injury;blood samples of healthy control group(A)with limosis were collected at that day of examination;serum level of IL-10 was measured by radioimmunoassay.Results In experimental group(B and C),the serum peak level of IL-10 emerged on T3 and then stepped down;the serum peak level of them increased abviously compared with group A(P＜0.05 or P＜0.01).Furthermore,the serum level of them in C group was usually higher than that on the same time in group B.Conclusion It was longer and severer in time and degree that effects of severe trauma with haemorrhagic shock bypass on IL-10 of body compared with that of simple severe trauma.

Objective To explore a stable and feasible method for the primary culture of breast cancer-associated fibroblast (CAF), and to offer the theoretical basis for the further studies about the role of CAF in the processing of breast cancer by analyzing the biological property of CAF. Methods CAF was primary cultured with tissue block enzymolytic method. Flow cytometry (FCM) was used to detect the cell cycle. The expression of Vimentin, α-SMA and Ki-67 were detected by cell immunochemistry method. Transwell assay was used to detect the invasion and migration capacity of CAF. Results The CAF was successfullyculturedbyconditionoptimization.ImmunohistochemicalresultsshowedthatVimentin, α-SMA and Ki-67 were highly expressed in CAF.Comparedwithfibroblastsfrombreastcancersidestissues,the proliferating ability and the invasion and migration capacities of CAF were enhanced (number of invasive cells: 79.7 ± 3.5 vs. 66.3 ± 1.5, number of migrated cells: 242.3 ± 3.1 vs. 218.3 ± 2.1, both P<0.05). Conclusion CAF cultured from breast cancer has better proliferating ability and strong invasion and migration capacities.

Objective To observe the effect of silencing Snail gene on the invasion and proliferation ability of human pancreatic cancer cell line PANC1.Methods Lentiviral vectors that can express small hairpin RNA(shRNA) targeting human Snail gene(shRNA-Snail) or shRNA sequence that did not match any known mRNA(shRNA-NC) were constructed,and transfected into PANC1 cells.Untransfected cells served as control.mRNA and protein expression of Snail,α-smooth muscle actin (α-SMA) and E-cadherin was determined by real time quantitative PCR and Western blotting,respectively.In vitro invasion ability was tested by Transwell model.Proliferation ability was measured by CCK-8 assay.Results Compared with those in shRNA-NC group,Snail mRNA (0.27 ± 0.02 vs 0.92 ± 0.03) and protein level (0.26 ± 0.02 vs 0.80 ± 0.02),and α-SMA mRNA (0.33 ±0.04 vs 0.97 ±0.07) and protein level (0.31 ±0.04 vs 0.74 ±0.06) in shRNA-Snail group were obviously decreased,but E-cadherin mRNA (1.57 ± 0.45 vs 0.95 ± 0.08) and protein level (0.86 ± 0.03 vs 0.20 ± 0.03) were greatly increased.The number of cells permeating the septum of transwell [(6.80 ± 0.73)/400 magnification vs (26.80 ± 2.52)/400 magnification,P ＜0.01] was significantly decreased,and cell proliferation was inhibited (0.74 ± 0.05 vs 1.47 ± 0.04,P ＜ 0.01).All the differences above were statistically significant (all P ＜ 0.01).No significant differences were observed between shRNA-NC and normal control group.Conclusions Silencing Snail gene may restrain the invasion and proliferation ability of PANC1 cells to a certain degree.

Background and purpose:miR-124 is considered to be a tumor suppressor in multiple tumors, including lung cancer, prostate cancer, bladder cancer, and breast cancer. However, its function is unclear in gastric cancer. This study aimed to explore the expression of miR-124 in human gastric mucosal epithelium cells and different gastric cancer cells, as well as gastric cancer tissues and matched para-cancerous tissues. The correlations of miR-124 expression with gender, age, histological grade, T stage, TNM stage, lymph node metastasis and prognosis of gastric cancer patients were analyzed.Methods:A real-time lfuorescent quantitative polymerase chain reaction (RTFQ-PCR) was employed for detecting the expression of miR-124 in human gastric mucosal epithelium cells and gastric cancer cells. The expression of miR-124 in gastric cancer tissues and matched para-cancerous tissues was detected by in situ hybridization.Results:The RTFQ-PCR results indicated that the expression of miR-124 was down-regulated in MKN-74, MKN-28, MKN-45, MGC-803, SGC-7901 and AGS cells compared to GES-1 cells.In situ hybridization showed that miR-124 was strongly expressed in normal gastric mucosa. However, low expression, focal positive expression or lack of miR-124 expression were observed in gastric cancer tissues. Statistical analysis showed that miR-124 was close-ly correlated to histological stage, TNM stage and node metastasis of gastric cancer patients, but not the age, gender and tumor size. The OS and DFS of the patients with low expression of miR-124 were shorter than that of the patients with high expression of miR-124. Multivariate analysis suggested that miR-124 down-regulation was an independent prog-nostic factor for survival in patients with gastric cancer.Conclusion:miR-124 is down-regulated in gastric cancer cells and tissues. The expression of miR-124 is correlated to histological stage, TNM stage, node metastasis and prognosis.

Objective Analysis and investigate the suitable method and results of auxiliary diagnostic test applicated in newly di‐agnosed patients with multiple myeloma(MM) .Methods The auxiliary diagnostic test results of the 171 patients newly diagnosed with MM were analyzed retrospectively .Results In bone marrow smear examination ,the percentages of plasma cell in all the 171 patients increased and were 5 .0% -88 .0% ,while the morphology of plasma cells was abnormal .The levels of immunoglobulin in 112 patients increased in different degrees .81 patients had different degrees of renal function damage .110 patients had different de‐grees of anemia and reduction of white blood cells and platelets .Conclusion Comprehensive considerate all the results of auxiliary diagnostic tests and the clinical manifestations could improve the detection rate of MM.

The construction of characteristic talent training program is crucial to strengthening the development of independent college.During the construction process,efforts should be made in the following aspects:the training aim being the inheritance of its matrix with the emphasis of application ; the curriculum system emphasizing skill training as well as improvement; the training model design making full use of resources with sufficient practice; the quality improvement highlighting regional characteristics and inheriting culture.Besides,special attention should be paid to understanding the government's relevant policies,to studying relevant theories about higher education and to grasping the generality of the education aimed at undergraduates and individuality of talents in independent college.

Objective To study the antidepressant effect of total timosaponin(TT)and its mechanism.Methods The antidepressant effect of TT was examined by mice forced swimming test(FST),learned helplessness(LH)experiment,chronic mild stress(CMS)model,yohimbine induced lethality test and 5-HTP induced head-twitches test.Results TT(25 or 50 mg?kg-1,ig,qd?14 d)markedly shortened the immobility time in the FST,but didn't affect the autonomic activity.TT(12.5,25 or 50 mg?kg-1,ig,qd?14 d)significantly decreased the number of escape deficits in the LH mice.TT(25 or 50 mg?kg-1,ig,qd?21 d)markedly enhanced the locomotor activity and increased consumption of sucrose solution in CMS mice.TT(50 mg?kg-1,ig,qd?14 d)enhanced the mortality of mice after administration of yohimbine for 4 h,and distinctly increased the head-twitch number in the 5-HTP induced head-twitches test.Conclusion TT has antidepressant effects in various depression mouse models.Its mechanism may be related to the reinforcement of NE and 5-HT nerves system.

Objective To evaluate the performance of UW2000 automatic urine system .Methods Using UW2000 automatic u‐rine system ,to evaluate intra ,inter batch precision ,linear ,carryover ,and the accuracy of the urine visible components(RBC ,WBC);at the same time intra ,inter batch precision and accuracy of of urine dry chemistry component .Results The within‐run coefficients of variation(CV% ) of RBC in high ,middle and low‐value samples were 15 .6% ,6 .8% ,20 .3% ,WBC were 16 .4% ,10 .5% ,24 .6% , respectively .Inter‐run precision of variation(CV% ) of RBC in middle and low‐value were 12 .8% ,13 .2% ,WBC were 15 .3% , 22 .8% ,respectively .The correlation coefficient of RBC was 0 .991 and the linear range was 88-19 513/μL .The correlation coeffi‐cient of WBC was 0 .997 and the linear range was 7-8 254/μL .And carryover rate was 0 .00% for RBC and 0 .06% for WBC ;The coefficient of UW2000 and microscopy on RBC and WBC were 0 .992 ,0 .995 .Within‐run precision ,inter‐run precision and accuracy for urine dry chemical composition of high ,low controls were all within the requirements of standards .Conclusion UW2000 auto‐matic urine analyzer workstation was not only save manpower ,but also with high detection speed and save cost .Importantly ,the re‐sults are reliability .In addition ,it could meet the working requirement in the general hospital ,and has great application value .