Abstract:Objective:To investigate inhibitory effect of RNPC1a short hairpin RNA (shRNA) on the expression of RNPC1a in cell line SUM1315 and determine the effect of RNPC1a down-expression on the proliferation,migration and invasion of SUM1315 cells. Methods:The RNPC1a shRNA was transfected into SUM1315 cells with lipofectine. Puromycin was used for selecting stable cell line. Real-time PCR and Western blot were performed to analyze the mRNA and protein expression of RNPC1a in SUM1315 cells. Cell proliferation,migration and invasion were assessed by methyl thiazol tetrazolium(MTT),wound-healing experiment and Matrigel invasion assay. Results:The mRNA and protein expressions of RNPC1a gene in the shRNA-RNPC1a-SUM1315 group were significantly lower than those in the control group,confirmed by RT-PCR and Western blot,respectively(P < 0.05). The proliferation of SUM1315 cells was markedly inhibited by RNPC1a shRNA with the inhibition rate at 26.5% (P < 0.05),while scratch repair time extended. Meantime transwell cell invasion assay showed that the invasive ability of SUM1315 cell was also inhibited in the shRNA-RNPC1a-SUM1315 group(23 ± 7),compared with the SUM1315 group(213 ± 12)(P < 0.01). Conclusion:Down-regulation of RNPC1a by RNPC1a shRNA can inhibit the proliferation,migration and invasion of SUM1315 cells.

Abstract:Objective:To establish an NIH3T3 cell line to stable express human Trop-2 gene,and analysis the affection of Trop-2 on proliferation,migration and aggressiveness of NIH3T3-Trop-2 cell. Methods:The human Trop-2 gene was cloned into eukaryotic expression system pcDNA3.1 and transfected into NIH3T3 cells. The Trop-2 stable expression cells were selected by G418 and confirmed by RT-PCR. The cell proliferation,migration and aggressiveness were detected by MTS and wound healing assay,MMP-2/MMP-9 was analyzed. Results:The proliferation of NIN3T3-Trop-2 was higher than NIN3T3 cell. The wound healing assay results showed and that Trop-2 improved the cell migration,increased the number of foci generated(P < 0.05) and increased MMP-9 expression(P < 0.05) when compared to the NIH3T3 control group. Conclusion:These show that human Trop-2 is sufficient to improve the cell proliferation and induce the transformation of NIH3T3 cells.

Abstract:Objective:To investigate the expression changes of miR-27a in human lung adenocarcinoma cell line SPC-A1 after cisplatin chemotherapy in virto,and to discuss its relationship with cell apoptosis and clinical significance. Methods:SPC-A1 cells were cultured in vitro and treated with DDP at 2.5 μg/ml for 12,24,48 and 72 h. Cell morphology was observed by phase contrast microscope,as well as cell apoptosis was detected by flow cytometry;expression changes of miR-27a both in cultural supernatants and SPC-A1 cells were analyzed by real-time PCR. Results:SPC-A1 cells showed morphological changes of apoptosis in the DDP (2.5 μg/ml) group. The apoptotic rates of SPC-A1 cells at 48 and 72 h in the DDP (2.5 μg/ml) group[(59.3 ± 2.5)% and (76.4 ± 3.1)%,respectively] were higher than that in the control group (P < 0.05). PCR analysis demonstrated that the levels of miR-27a were significantly increased in the DDP (2.5 μg/ml) group both in cultural supernatants and SPC-A1 cells compared to the level of the control group(P < 0.05). Conclusion:The level of miR-27a raised in human lung adenocarcinoma cell line SPC-A1 after cisplatin chemotherapy in virto. Quantitative analysis of miR-27a may have important value for chemotherapy monitoring of lung adenocarcinoma.

Abstract:Objective:To investigate the expression of miR-125b and vascular endothelial growth factor-A (VEGF-A) in hepatocellular carcinoma(HCC) and to analyze their correlation. Methods:Real-time PCR was performed to detect miR-125b expression in HCC,tumor-adjacent tissue and Huh-7,HepG2,L02. Real-time PCR and Western blot were employed to detect the expression of VEGF-A mRNA and protein in HCC,tumor-adjacent tissue. miR-125b mimics was transfected into HepG2 cells and the expression of VEGF-A mRNA and protein were tested with real-time PCR and Western blot,respectively. Results:The expression of miR-125b was down-rengulated in HCC compared to tumor-adjacent tissue(P < 0.05). The expression of miR-125b mRNA in HepG2 cells was also less than to L02 cells(P < 0.05). VEGF-A mRNA and protein were up-regulated HCC and tumor-adjacent tissue compared to normal liver tissue(P < 0.05),which was down-regulated in HepG2 cells after transfection of an miR-125b mimic(P < 0.05). Conclusion:miR-125b is down-regulated,but VEGF-A is up-regulated in HCC,suggesting that the depressed expression of miR-125b promotes the carcinogenesis of hepatocellular carcinoma by upregulating VEGF-A expression.

Abstract:Objective:To investigate the expression of miR-451 and MIF in colorectal cancer and colorectal adenoma and their correlation with clinicopathologic features in colorectal cancer. Methods:Real time polymerase chain reaction(RT-PCR) was performed to examine the expression level of miR-451 in 25 cases of colorectal cancer and matched non tumor adjacent tissue and 10 cases of colorectal adenoma tissue specimens. MIF protein was detected by immunohistochemistry(SP method). The relationships among the expression of miR-451, MIF and the clinicopathologic features of colorectal cancer were analyzed. Results:The expression level of miR-451 was significantly lower in the tumor tissues and adenoma tissues than that in the adjacent tissues (P < 0.01,P < 0.05). There were 6 cases up-regulated and 19 cases down-regulated in colorectal cancer,and the down-regulated rate was 76% (19/25). There were 3 cases up-regulated and 7 cases down-regulated in colorectal adenoma,and the down-regulated rate was 70% (7/10). The positive rate of MIF in colorectal cancer was 72%,the positive rate was 50% in colorectal adenoma and the positive rate was 32% in adjacent tissues. Down-regulated miR-451 expression and up-regulated MIF were associated with the cell differentiation (P < 0.05) in colorectal cancer patients. No significant association was found between the expression of miR-451 and MIF with the status of age,gender,Dukes staging system histological type, depth of tumor invasion and lymphatic metastasis. Conclusion:Up-regulated miR-451 expression may inhibit MIF expression,and then control colorectal carcinogenesis and the development process.

Abstract:Objective:To detect the clinicopathologic characters of micrometastasis and microvessel density(MVD) occur in non-small cell lung cancer(NSCLC),and to explore the correlation among the former factors and those cases with vascular invasion. Methods:Vascular invasion and MVD was recorded by immunochemistry staining to CD34. Blood samples from 47 patients were examined for CK19 mRNA expression by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Results:Serum CK19 mRNA positive was recorded in 13/18 cases with vascular invasion versus 9/29 in cases without invasion,with statistically difference. The average MVD in the invasion group was (29.3 ± 3.3),higher than the non-invasion group(25.2 ± 4.7). Conclusion:CK19 mRNA positive,pleura invasion,lymphnode state,metastasis and MVD are significantly correlated with vascular invasion in NSCLC patients,and these factors could be used to assess biological behavior of NSCLC.

Abstract:Objective:To detect the expression of chicken ovalbumin upstream promoter transcription factor Ⅱ(COUP-TFⅡ) protein in human gastric cancer(GC) tissues,and investigate the potential molecular mechanism of COUP-TFⅡ involved in the development and advancement of GC and its clinical significance. Methods:Paired tumorous and adjacent non-tumorous human gastric tissues were obtained from 66 patients with GC who underwent surgical resection. The expressions of COUP-TFⅡ and MMP-2 protein were detected using immunohistochemistry. The relationship between COUP-TFⅡ and the clinicopathologic factors as well as the correlation of COUP-TFⅡ and MMP-2 were analyzed. Results:COUP-TFⅡ proteins were positively expressed in 65.2%(43/66) and 19.7%(13/66) of GC tissues and paired non-GC tissues,respectively. The expression of COUP-TFⅡ was significantly correlated with the depth of invasion,differentiated grade,lymph node metastasis and tumor-node-metastatsis (TNM) stage (P < 0.05). However,the expression of COUP-TFⅡ was not correlated with the gender,age and volume of GC tissues. Patients with positive COUP-TFⅡ expression had lower mean survival time than those with negative COUP-TFⅡ expression (26.8 ± 2.6 months vs 36.4 ± 3.0 months, χ2 = 4.118,P = 0.042). MMP-2 proteins were positively expressed in 78.8% (52/66) and 34.8% (23/66) of GC tissues and paired non-GC tissues,respectively. Moreover,a positive correlation was found between COUP-TFⅡ and MMP-2 expression in GC tissues. Conclusion:COUP-TFⅡ protein has significant value in determining invasion and metastasis of gastric cancer and assessing prognosis in patients with gastric cancer,and MMP-2 may be involved in the above regulation progress.

Abstract:Objective:To investigate the androgenic activities of Microcos paniculata,licorice root,Angelica sinensis and the underlying molecular mechanisms. Methods:The effects of extracts on cell viability were examined by an assay based on 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium,inner salt(MTS). The expression of androgen receptor target gene was determined by real-time PCR. Results:Three extracts were screened for effects on viability of androgen-responsive 22RV1 cells. The extract of Microcos paniculata enhanced cell viability and the expression of PSA mRNA. But they were blocked by the androgen receptor(AR) antagonist flutamide and by the extracellular-signal-regulated kinase 1/2 (ERK1/2) inhibitor,U0126. Conclusion:Microcos paniculata has androgenic activities in an AR-dependent manner among the three plant extracts. And ERK1/2 signaling participates in Microcos paniculata-induced androgenic effects.

Abstract:Objective:To study the effects of aplasia rashomolog memberⅠ(ARHI ) on HOC SKOV3 cell lines. Methods:After transfection of the pIRES2-EGFP-ARHI plasmid to the HOC SKOV3 cells with low expression level of ARHI gene, absorbance A was measured and performed to calculate the inhibitory rate of cell growth using CCK-8 method. To analyze the cell cycle distribution and apoptosis rate by using flow cytometry and to test the change in expression level of LC3-Ⅱ protein by using Western blot approach in the pIRES2-EGFP-ARHI-SKOV3 (the test group), pIRES2-EGFP-SKOV3 (the plasmid control group) and SKOV3 (the negative control group) cell groups. Results:After the cells in the test group were cultured for 24, 48, 72, 96 and 120 h, we found that the inhibitory rates of cell growth were 64.69%, 70.17%, 67.01%, 66.87% and 67.70%, respectively. The inhibitory rates of cell growth in the test group were significantly higher than those in the plasmid control group(P < 0.01). The ratios of S phase cells were 64.18%, 38.43% and 15.15%, and the apoptosis rates were 47.97%, 26.53% and 9.33% respectively after culturing the cells for 48 h in the test, plasmid control and negative control group. The proportions of cells in S phase were 43.29%, 10.37% and 10.89%, and the apoptosis rates were 51.34%, 24.70% and 4.39% respectively after culturing the cells in the test, plasmid control and negative control group for 72 h. The significant difference remained between the test and control groups. The expression level of LC3-Ⅱ in test group increased with significant difference compared with the control group after cultured for 48 h. Conclusion:These findings suggest that ARHI gene inhibits the growth of SKOV3 cells, arrests the SKOV3 cells at S phase and induces apoptosis and autophagy.

Abstract:Objective:To study population distribution,ADR level,route of administration and the serious ADR reports of antineoplastic adverse drug reactions,in order to help related population to increase health awareness,pay attention to the prevention of cancer and minimize serious adverse drug reactions. Methods:Downloaded ADR reports occurred and reported in Affiliated Drum Tower Hospital of Nanjing University in 2011 from National Adverse Drug Reaction Monitoring Network database. We performed Excel to statistical analyze the medical records and data by category of age,gender,types of drugs,route of administration and ADR level. Results:Antineoplastic ADR had 422 cases,ranking the first place of all types of drugs. Among 1 503 cases of ADR reported in 2011,the mean age of 357 cases of adverse reactions was 50~59 years old,significantly higher than other age,in which there were 163 male cases and 194 female cases. Of the 357 cases,there were 151 cases of antineoplastic ADR,in which there were 90 male cases and 61 female cases. Of 422 cases of antineoplastic ADR,there were 349 cases (82.70%) in the level of severe or severe new,route of administration was mostly intravenous infusion (385 cases,91.23%). Severe antineoplastic adverse reactions were firstly caused by Oxaliplatin (50 cases). Conclusion:ADR is significantly reduced after the anti-infective standardized management,but is significant increased in the proportion of antineoplastic ADR. Antineoplastic ADR mostly occurred in the 50~59 years old male population. This particular population should pay special attention to cancer prevention and rational use of drugs to minimize the serious adverse reactions.

Abstract:Objective:To construct a human scFv phage antibody library against rabies virus G protein,selected the single chain antibody against rabies virus G protein for different epitopes,and identificated its neutralizing activity. Methods:The human scFv phage antibody library against rabies virus G protein was constructed by using peripheral blood lymphocytes from 130 Rabies vaccine immune volunteers. Screening by the purified rabies virus G protein,the positive clones were identified and expressed,identified neutralizing activity by AMMS. Results:Through the prokaryotic expression system and purification of rabies virus G protein,the human anti rabies virus G protein scFv phage antibody library was constructed with about 5.0×107 size. The nucleic acid sequence analysis confirmed the insertion of a fragment is scFv. After 5 rounds of screening,we randomly picked 140 clones by phage-ELISA,and obtained 4 nucleic acid sequences in different scFv antibodies,purified and identified activity. We finally obtained 1 scFv antibody with neutralizing activity. Conclusion:We have successfully constructed a whole human anti-rabies virus G protein scFv antibody library,and obtained a whole human anti-rabies virus G protein scFv antibody with neutralizing activity from the library. It is established foundation of the new type of rabies virus vaccine for human immunization.

Abstract:Objective:To explore the protective effects of interleukin-10 (IL-10) pretreatment on the behavioral and molecular biochemical changes of Alzheimer’disease(AD) rat models and its possible mechanism. Methods:Okadaic acid(OA) was microinjected into the hippocampus of rats to establish the AD rat models. The escape latency of rats was tested by water maze. Western-blot method was used to detect the expression of protein phosphatase 2A (PP2A) and amyloid precursor protein (APP) in the hippocampus. Simultaneously,the ratio of CD4+ cells to the CD8+ cells in the mesenteric lymph node cells was examined by flowcytometry. Results:The latent period in water maze of rats that were injected with OA in the hippocampus was significantly increased,and the latent period of the rats with high concentration of OA injection was longer than that of the rats with low concentration of OA injection. The PP2A protein expression in hippocampus after the OA injection was down-regulated,and the inhibitory effect of OA on PP2A protein expression was enhanced as the OA concentrations increased. Conversely,APP protein expression in hippocampus was increased with the elevated concentrations of OA injected in the hippocampus. After IL-10 and OA were injected into the rat hippocampus respectively,the latent period of water maze was significantly lower than that of the AD rats only with the OA injection. IL-10 reversed the inhibitory effect of OA on PP2A expression and alleviated the augument of APP;furthermore,IL-10 reversed the increasing of CD4+/CD8+ induced by OA and caused the CD4+/CD8+ return to the control level. Conclusion:AD-like changes can be induced by the microinjection of OA into the hippocampus. IL-10 can reverse the AD-like changes induced by OA,and the effects of IL-10 are dose-dependent and also closely related to its inhibition of inflammatory reaction.

Abstract:Objective:To observe the effect of berberine on high-fat diet induced mice chronic kidney injury and the macrophages in the kidney. Methods:A total of 30 C57BL/6 mice were randomly divided into the normal control group, high-fat diet group and berberine treatment group. At the end of experiment,the mice were sacrificed. Blood,urine and kidney tissue were collected. Blood was used for detecting total cholesterol,triglyceride,serum creatinine,urea nitrogen,cystamin C. Urine was used for detecting 24 h urinary albumin. Pathological changes were observed by light microscope by HE staining. The expression of IL-1β and iNOS were detected by Western blot. The expressions of F4/80 and iNOS were determined by immnofluorescence staining and immunohistochemistry staining,respectively. Results:Compared with the normal control group,total cholesterol,triglyceride,cystamine C,urine volume and 24 h urinary albumin were significantly higher in the high-fat diet group (P < 0.05). The expressions of IL-1β,F4/80 and iNOS increased significantly in the high-fat diet group (P < 0.05). Compared with the high-fat diet group,the berberine treatment group showed less aggravated kidney pathological changes. The expressions of IL-1β,F4/80 and iNOS in the berberine treatment group were decreased than in the high-fat diet group (P < 0.05). Conclusion:Berberine may decrease the macrophages,especially the M1 macrophages and then intervene the chronic kidney injury induced by high-fat diet.

Abstract:Objective:To observe the effects of endotoxin tolerance on the production of inflammatory cytokine TNF-α and anti-inflammatory cytokine IL-10 in THP-1 cells. Methods:THP-1 cells were pretreated with 1 μg/ml Porphyromonas gingivalis (P.gingivalis) lipopolysaccharides (LPS) or 1μg/ml Escherichia coli (E.coli) LPS for 24 h. Then,the cells were washed and stimulated with the same LPS for additional 24 h. The levels of TNF-α and IL-10 in supernatants were detected by ELISA. Results:The amounts of TNF-α and IL-10 secreted by THP-1 cells stimulated with P.gingivalis LPS or E.coli LPS were increased significantly after 24 h (P < 0.05). After restimulations for another 24 h,TNF-α production was decreased significantly compared with that secreted by the cells stimulated with the same LPS only once(P < 0.05),while the level of IL-10 was increased significantly(P < 0.05). Conclusion:Repeated LPS stimulations triggered endotoxin tolerance and led to a decrease in TNF-α production and an increase in IL-10 production,which might have effects on periodontal inflammation.

Abstract:Objective:To investigate the effects of acrylamide on apoptosis and miR-21 expression in SH-SY5Y neuroblastoma cells. Methods:Cell viability of SH-SY5Y cells treated with different concentrations of acrylamide for 24 hours was measured by MTT. Hoechst 33258 staining was used for the determination of cell apoptosis;miR-21 expression was detected by real-time PCR; the expression levels of PTEN,p-AKT,AKT,Bcl-2,Bax,caspase 9 and caspase 3 were examined by Western blot. Results:Acrylamide exposure resulted in decreased viability and induction of apoptosis of SH-SY5Y cells in a dose-dependent manner. miR-21 expression was downregulated by acrylamide. Meanwhile,acrylamide increased the expression levels of PTEN,Bax,caspase 9 and caspase 3,and decreased p-AKT and Bcl-2 expression in SH-SY5Y cells. Conclusion:Acrylamide induces apoptosis of SH-SY5Y cells via mitochondrion pathway,and down-regulation of miR-21 may be involved in the regulation of apoptotic process induced by acrylamde.

Abstract:Objective:To investigate the activity of medial prefrontal cortex (mPFC) pyramidal neurons in rats and their response to 5-hydroxytryptamine-7(5-HT7) receptor stimulation. Methods:The change of the spontaneous firing of pyramidal neurons in mPFC was observed by extracellular recording in vivo. Results:In this study,we reported that systemic and local administration of 5-HT7 receptor agonist AS19 produced excitation,inhibition and no change in the firing rate of pyramidal neurons in mPFC of rats. The mean response of the pyramidal neurons to AS19 (0.08 μg/100 nl) by systemic and local administration in mPFC was excitatory. The inhibitory effect by systemic administration of AS 19 was reversed by -酌-aminobutyricacid A receptor antagonist picrotoxinin(2 mg/kg). Systemic administration of picrotoxinin excited all the neurons examined in rats. After treatment with picrotoxinin,the local administration of AS19 increased the firing rate of the neurons. Conclusion:These results indicate that the activity of mPFC pyramidal neurons is regulated through activation of 5-HT7 receptor by direct or indirect action.

Abstract:Objective:To investigate the association between vitamin D receptor(VDR) gene polymorphism and the bone mineral density of the chronic renal failure patients in the northern China. Methods:A total of 211 patients with chronic renal failure coming from unrelated families in the northern China were selected. The subjects were divided into the chronic renal failure group (n=110) and the uremia group(n=101) by the level of serum creatinine. Polymerase chain reaction and restriction fragment length polymorphism(PCR-RFLP) was used to detect VDR BsmⅠpolymorphisms. Information on environmental-related risk factors was collected using a pre-tested standard questionnaire. Bone mineral density was measured by dual energy X-way absorptiometry. Concentration of serum intact parathyroid hormone was measured by radioimmunoassay. Results:According to the χ2 test,there was no significant difference between the chronic renal failure group and the uremia group (χ2 = 0.088,P = 0.591). Age had significant negative correlation with bone mineral density(P < 0.01) while body mass index had significant positive correlation with bone mineral density(P < 0.01). Statistical differences of bone mineral density in lumbar were showed among different VDR genotypes(P < 0.05). People with aa genotype had significant lower bone mineral density in lumbar spine (P < 0.05) after correcting the body mass index and age. Conclusion:The results show that there is a possible correlation between the polymorphisms of VDR gene BsmⅠand bone mineral density in chronic renal failure patients in the northern China. It indicates that the restriction site polymorphisms of BsmⅠgene may be used as a genetic markers.

Abstract:Objective:To investigate the relationship between gene polymorphisms of IGFⅠ,IGFⅡ and birth weight. Methods:In the case-control study,88 small for gestational age infants(SGA),477 appropriate for gestational age infants (AGA) and 265 large for gestational age infants(LGA) were chosen in Nanjing. All the neonates were genotyped for genetic polymorphisms in the IGFⅠ rs35767 and IGFⅡ rs3741205 by using a polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP) assay. Linear regression models were used to estimate the association of the genetic polymorphism with LGA and SGA. Results:TT and CT/TT genotype of IGFⅠ rs35767 reduced the risk of SGA significantly(P = 0.024,P = 0.031),the polymorphism in IGFⅡ rs3741205 had no relationship with SGA;CT genotype of IGFⅠ rs35767 and TT homozygote of IGFⅡ rs3741205 increased the risk of LGA significantly(P = 0.045,P = 0.029). When mother’s educational background beyond the college,the IGFⅡ rs3741205 GT/TT genotype reduced the risk of LGA. Conclusion:IGFⅠ rs35767 is contribute to SGA. IGFⅠ rs35767,IGFⅡ rs3741205 are contribute to LGA.

Abstract:Objective:To study the variation of skinfold thicknesses of Tunbu adult with increased age,and the differences with other ethnic groups in China. Methods:Using the random sampling method, we investigated 6 items of skinfold thickness(including facial,triceps,biceps,subscapular,suprailiac and calf skinfold thickness) of 506 cases of Tunbu adults(251 males and 255 females) in area of Anshun Sichuan, and analyzed the variation features that the skinfold thickness varies with increased age. Results:Male Tunbu subscapular skinfold was thickest,biceps skinfold was thinnest,subcutaneous fat thickness of body was thicker than the limbs. Female Tunbu triceps skinfold was thickest,biceps skinfold was thinnest. All of the 6 items of skinfold thickness were greater in female than male,and had no clear correlation with age in male (P > 0.05),only facial skinfold had positively correlated with age in female (P < 0.01). Variance analysis of 6 items of skinfold thickness in 5 age groups of male and female showed suprailiac skinfold was statistically significant within age groups only in male(P < 0.01). All of the 6 items of skinfold thickness of Tunbu were greater than ethnic groups in southern China. Conclusion:Tunbu subcutaneous fat development has a big difference with South Asian ethnic group,while relatively close to other Han ethinc groups,particularly close to Han(Sichuan),Han(Jiangsu),Han(Anhui) and Buriats.

Abstract:Objective:In order to guide clinical doctor to select antimicrobial agents rationally,we sought to analyze the pathogen distribution and resistance pattern of positive clinical specimens from the department of gerontology of our hospital in 2011. Methods: The bacteria and fungi were identified by API or Vitek 2 compact system. The susceptibility of antimicrobial and antifungal agents was tested by K-B and the data were analyzed by WHONET5.6 software. Results:A total of 993 strains were isolated and collected in the department of gerontology,including 657 strains (66.2%) of Gram-negative bacilli,124 strains (12.5%) of Gram-positive cocci and 119 fungi(12.0%). The pathogen of the highest isolating rate was Pseudomonas aeruginosa,which accounted for 25.6%,the next was Acinetobacter species(19.0%) as well as Klebsiella(12.7%). Resistance analysis showed that among nonfermenters,Pseudomonas aeruginosa was resistant to the most of the antimicrobial agents,and Acinetobacter for most antimicrobial resistant was more than 70.0%,however for the resistance pattern of imipenem and meropenem were 88.7% and 91.1% respectively. Meanwhile,Escherichia coli were resistant to the most of cephalosporins,but for the resistance pattern of imipenem and meropenem were 0.0% and 1.5%,respectively. The resistance pattern of Klebsiella was higher than Escherichia coli,and for the resistance pattern of imipenem and meropenem were 35.2% and 42.3%,respectively. As for Staphylococcus species,none was found to resistant to vancomycin. However,they were resistant to many other antimicrobial agents. Conclusion:The pathogens in our department of gerontology have generated diversity,and the resistance pattern of the pathogens is rising. Especially,the resistance to carbapenems among Acinetobacter species and Enterobacteriaceae were serious,and we need to pay more attention on this issue.

Abstract:Objective:To establish an optimal method of template preparation for sequencing avian influenza virus (H7N9) genome. Methods:Sequencing templates were prepared by using random hexamers primers to synthesize single strand cDNA and double strands cDNA;by using influenza virus specific-U12 primer to synthesize single strand cDNA and using random hexamers primers to obtain double strands cDNA;or by using influenza virus specific-U12 primer to synthesize single strand cDNA and using influenza virus gene-specific primer to obtain double strands cDNA. Sequencing library was constructed by using above template and sequencing. After analysis of the data,the results were compared with Sanger sequencing. The best method for template preparation was selected for high throughput sequencing. Results:The cluster pass filter,coverage,single nucleotide polymorphisms (SNPs),and median length were the best in using influenza virus specific-U12 primer to synthesize single strand cDNA and using influenza virus gene-specific primer to obtain double strands cDNA compared with other two methods. Conclusion:High accuracy,short turnaround times,relatively low cost and high throughput make this method becoming possible for surveillance of H7N9 viruses evolution.

Abstract:Objective: To assess the reliability and validity of randomized response technique(RRT) model of cluster sampling survey on multichotomous sensitive question investigation method and statistical formulae. Methods:A total of 404 female sex workers were selected randomly by cluster sampling in Xichang during May 2011 to July 2011. RRT was applied to obtain their sensitive characters. With the sensitive characters taken as parameter of simulated overall,simulated direct questioning results taken as criterion,results got from multichotomous RRT model were compared with ones from simulated direct questioning by using chi-square test with Monte Carlo simulation to access validity and reliability. Results:There were 99 results have no statistically significant different from criterion (α=0.05). Conclusion:The method and corresponding formulae have high validity and reliability,and can be used for sample survey of large-scale sensitive issues.

Abstract:Objective:To design and synthesize the pueraria isoflavonoid-imprinted monolithic column by molecular modeling,and theoretically explore the mechanism of its specific molecular recognition. Methods:Genistein was selected as template,4-vinylpyridine,acrylamide and methacrylate were selected as candidate monomers. Density functional theory(DFT) was adopted to accurately predict the hydrogen bonding energy between template and different functional monomers,respectively. And then the optimal functional monomer and the appropriate ratio of monomer to template were screened out. As a result,the pueraria isoflavonoid-imprinted monolithic column was prepared by in situ polymerization method. Results:4-vinylpyridine was the optimum functional monomer and ideal ratio of monomer to template was 2∶1. Meanwhile,pueraria isoflavonoid-imprinted monolithic column was successfully prepared. Conclusion:These results provide the theoretical reference for the design and synthesis of pueraria isoflavonoid-imprinted monolithic column,and can be applied to the other non-covalent imprinting system.

Abstract:Objective:To evaluate the immunity effect of H1N1 influenza vaccine prepared by film-dispersion and freeze-thawing method,and to provide expermental foundation for attenuated live vaccine liposome preparation. Methods:Experimental mice were divided into the influenza vaccine non-liposome group,the film-dispersion prepared H1N1 influenza vaccine liposome group,the freeze-thawing lyophilized prepared H1N1 influenza vaccine liposome group,the positive control group and the negative control group (n=5). 4 μg and 6 μg hemagglutinin of H1N1 subtype per mouse were tracheally delivered to the mice for the non-liposome group and the lyophilized liposome groups,with the same dose intraperitoneally delivered groups as the positive control,and the PBS intraperotoneal injection group as the negative control. After 7,14 and 28 d of immunization,serum levels of HA antibodies were measured by the hemagglutination-inhibition method,and serum cytokine levels were measured by the ELISA method. Results:By pulmonary injection of the two liposome groups,the antibody titers of 6 μg dose groups were higher than those of 4 μg dose groups (P < 0.05). HA antibody titers of the two pulmonarily delivered liposome groups were higher than those of the pulmonarily (P < 0.05) and intraperitoneally (P < 0.01) injected non-liposome group. Antibody titers of the film-dispersion liposome group for both doses were higher than those of the freeze-thawing lyophilized liposome group. IL-2 and IFN--酌 produced by the pulmonary injected liposome groups were higher than those of the intraperitoneally delivered non-liposome group. Conclusion:The humeral and cellular immunities could be effectively induced by pulmonarily delivered vaccine liposomes;The film-dispersion method group is better than the freeze-thawing lyophilized group in terms of immunity strength. For the freeze-thawing lyophilized prepared H1N1 influenza vaccine liposome group,the immune response occurs at 14 d after vaccination,while the titer ratio is only 1.8 for normal vaccinated flu vaccine. The freeze-thawing lyophilized liposome vaccine also stimulates higher levels of cytokines.