AIM:To investigate the effect of insulin and gliclazide therapies on the liver fat accumulation in type 2 diabetic rats .METHODS:A high-fat diet plus low-dose streptozotocin was implemented to establish a type 2 dia-betic rat model, and the rats were randomly divided into diabetes mellitus (DM) group, diabetic rats treated with insulin ( INS) group, diabetic rats treated with gliclazide per os ( PO) group, and normal control ( NC) group.The diabetic rats in INS group and PO group were given insulin and gliclazide for 3 weeks, respectively.The changes of the liver fatty were evaluated with oil red O staining .Fasting plasma adiponectin concentration was measured by ELISA .The expression of adi-ponectin receptor 1 ( AdipoR1 ) was detected by real-time PCR.The protein levels of AMP-activated protein kinase (AMPK), phosphorylated AMPK on threonine 172 ( Thr172p-AMPK), sterol regulatory element-binding protein 1c (SREBP-1c), phosphorylated SREBP-1c on serine 372 (Ser372p-SREBP-1c), acetyl-CoA carboxylase (ACC), phospho-rylated ACC on serine79 (Ser79p-ACC) and immunoglobulin-binding protein (BiP) in the liver homogenate were deter-mined by Western blotting .RESULTS:Compared with the normal rats , in DM group, the presence of cytoplasmic lipid deposits was confirmed by oil red O staining .In INS group, these changes were significantly lower than those in DM group . Similar results were obtained in PO group .Insulin therapy significantly increased the plasma concentration of diponectin and liver tissue levels of AdipoR1 compared with DM group.At the same time, these 2 indicators returned to normal levels after gliclazide therapy .Thr172p-AMPK/AMPK, Ser372p-SREBP-1c/SREBP-1c and Ser79p-ACC/ACC expression ratios were significantly reduced in DM group compared with control values .The expression of BiP was increased on the contrary . After insulin therapy, Thr172p-AMPK/AMPK and Ser372p-SREBP-1c/SREBP-1c were significantly increased, and Ser79p-ACC/ACC and BiP returned to the normal levels .After gliclazide treatment, Thr172p-AMPK/AMPK and Ser372p-SREBP-1c/SREBP-1c returned to the normal levels , the expression ratio of Ser79p-ACC/ACC had no significant improve-ment compared with DM group , and the expression of BiP significantly declined .CONCLUSION: Both the insulin and gliclazide therapies reduce the lipid deposition in the liver of rats with type 2 diabetes by activating AMPK , but the extent and mechanism are not the same.In insulin therapy, AMPK restrains the expression of SREBP-1c directly, increases the phosphorylation of SREBP-1c, and affects SREBP-1c by inhibiting the endoplasmic reticulum stress .Gliclazide treatment, which has no effect on the lipid oxidation , reduces lipid deposition in the liver only through the phosphorylation of SREBP-1c and the suppression of the endoplasmic reticulum stress .

Objective To analyze the expression of interleukin (IL)-31 in tuberculous pleural effusion,and to evaluate its diagnostic value of tuberculous effusion.Methods Seventy-one patients with pleural effusion were enrolled,including 40 cases of tuberculous pleural effusion and 31 cases of malignant pleural effusion.Luminex method was applied to detect the IL-31 expression in pleural effusion.IL-31 levels were compared using non-parametric Mann-WhitneyU test,and the receiver operator characteristic (ROC)curve was used to elvaluate the diagnostic value of IL-31 .Results IL-31 expression in tuberculous pleural effusion was significantly higher than that in malignant pleural effusion with statistical significance (529.4 ng/L vs 13.8 ng/L,U =62,P <0.01 ).Based on the level of IL-31 expression,area under the ROC curve was 0.95 with the optimum cut-off value of 67.5 ng/L.Thus,the sensitivity and specificity of IL-31 ≥67.5 ng/L for diagnosis of tuberculous pleurisy were 82.5 % (95 %CI :73.3% - 94.2%)and 100.0% (95 %CI :91 .4%-100.0%),respectively.Conclusion IL-31 is highly sensitive and specific for the diagnosis of tuberculous pleural effusion, which favors the differentiation of tuberculosis from malignance.

OBJECTIVETo investigate the additive effects of uncoupling protein 2 (UCP2) gene Ala55Val variation and ADR beta(3) gene Trp64Arg variation on the obesity in Chinese Han population.

METHODSThe UCP2 gene Ala55Val variation and ADR beta(3) gene Trp64Arg variation were examined by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP) in 119 obese subject with mean BMI (27.9+/-2.98)kg/m(2) and in 177 control subjects with mean BMI(21.9+/-1.9)kg/m(2). The additive effects of the two gene mutations were analyzed.

RESULTS(1) The frequency of ADR beta(3) gene Trp64Arg variation in obese subjects was not significantly different from that in control subjects. In control subjects, the Trp64Arg variation carriers had higher fasting glucose level and 2-hour-post-prandial glucose level than did non-carriers. (2) The frequency of homozygote of UCP2 gene Ala55Val variation in obese subjects was higher than that in the control subjects (OR=3.71, P=0.001). In control subjects the Ala55Val variation carriers had higher BMI. (3) When there was only UCP2 gene or ADR beta(3) gene mutation, the frequency of gene mutation in obese subjects was not significantly different from that in control subjects (P>0.05). But when there were simultaneously two gene mutations, the frequency of gene mutations was higher in obese subjects than in control subjects (OR=2.57, P=0.009). (4) The genotype carriers with Val/Val+ Trp/Arg were the greatest relation to obese obesity (OR=8.58, P=0.002).

CONCLUSIONThe homozygote of UCP2 gene Ala55Val mutation increases the risk of obesity. Though the UCP2 gene mutation alone or the ADR beta(3) gene mutation alone is not associated with obesity, the possible additive effects of the two micro-genes increase the occurring of obesity.

Adult ; Aged ; Female ; Humans ; Ion Channels ; Male ; Membrane Transport Proteins ; genetics ; Middle Aged ; Mitochondrial Proteins ; genetics ; Mutation ; Obesity ; genetics ; Receptors, Adrenergic, beta-3 ; genetics ; Uncoupling Protein 2

Objective To investigate the influence of the individual genotype differences of DC-SIGN and DC-SIGNR on the mother-to-neonate intrauterine infection of HBV.Methods The genotypes of the gene DC-SIGN and DC-SIGNR in the pregnant women with HBV positive were detected by PCR and agarose gel electrophoresis.The significant difference of gene diversity of DC-SIGN and DC-SIGNR was analyzed by chi-square test.Results ①All of 29 cases in intrauterine infection group were 7/7 DC-SIGN genotype.In the non-intrauterine infection group,7/5 genotype were observed in 2 of 54 cases,and the other 52 cases were 7/7 genotype.The two groups was no significant difference ( P =0.54 ).②29 cases of intrauterine infection group was observed 4 genotypes of DC-SIGNR such as 7/7,7/5,9/7 and 6/5,the genotype frequencies were 0.3793,0.3448,0.2414 and 0.0345 respectively.54 cases of non-intrauterine infection group was found 6 genotypes such as 7/7,7/5,9/5,9/7,7/6 and 6/5,genotype frequencies were 0.5186,0.1481,0.0926,0.1852,0.0370 and 0.0185 respectively.The distribution of 7/5 genotype in the intrauterine infection group (29 cases) and the non-intrauterine infection group (54 cases ) was statistically significant ( P =0.038 ),and no significant difference was found in other genotypes between the two groups (P ＞ 0.05 ).Conclusion The gene DC-SIGN showed relatively little variation in the pregnant women infected with HBV.On the countrary,there were multiple genotypes of the gene DC-SIGNR in these women,and the genotype “7/5” of DC-SIGNR might be one of the susceptibility genes associated with intrauterine infection.

Objective:To explore the mechanism of herb-partitioned moxibustion in relieving rat intestinal inflammation by focusing on the neutrophil extracellular traps(NETs)in Crohn disease(CD)development. Methods:Rats were randomly divided into a normal group,a model group,a herb-partitioned moxibustion group,and a mesalazine group.The CD rat model was prepared with 2,4,6-trinitrobenzene sulfonic acid except for rats in the normal group.Rats in the normal group and model group did not receive any treatment but had the same fixation as the other groups.Rats in the herb-partitioned moxibustion group received herb-partitioned moxibustion at Qihai(CV6)and bilateral Tianshu(ST25).Rats in the mesalazine group received intragastric administration of mesalazine enteric-coated tablets.The general situation of rats in each group was recorded,and the histopathological changes in the colon were observed and scored by hematoxylin-eosin staining.The serum concentrations of NETs DNA(NETs-DNA),neutrophil elastase(NE)-DNA,and myeloperoxidase(MPO)-DNA were detected by ABC enzyme-linked immunosorbent assay,and the citrullinated histone 3(citH3),MPO,and NE protein and mRNA expression levels in rat colon tissue were observed by immunofluorescence and real-time quantitative polymerase chain reaction. Results:Compared with the normal group,the mucosal ulcer reached the muscularis,the epithelium was incomplete,the goblet cells decreased obviously with significant inflammatory cell infiltration in the colon;the colonic mucosa damage index(CMDI)score increased significantly(P<0.01);the serum NETs-DNA,NE-DNA,and MPO-DNA concentrations increased(P<0.05);the NE,citH3,and MPO protein and mRNA expression in the colonic tissue increased significantly in the model group(P<0.01 or P<0.05).Compared with the model group,the mucosal epithelium in the herb-partitioned moxibustion group and the mesalazine group was repaired and the goblet cells increased with a few infiltrating inflammatory cells in the colon;the CMDI score decreased(P<0.01);the serum NETs-DNA,NE-DNA,and MPO-DNA concentrations decreased(P<0.05);the NE,citH3,and MPO protein and mRNA expression in the colonic tissue was down-regulated(P<0.01 or P<0.05). Conclusion:Herb-partitioned moxibustion reduced the serum NETs complex and inhibited the protein and mRNA expression of NETs complex in the colon tissue,which may be one mechanism of herb-partitioned moxibustion in relieving colon mucosal inflammation in CD.

Background:Reduced expression of tripartite motif-containing 3 (TRIM3) has been reported to be involved in the pathogenesis of human glioblastoma.In our previous research,we found that TRIM3 expression was markedly reduced in human primary hepatocellular carcinoma (HCC) tissues and that low TRIM3 expression was associated with short survival of HCC patients.However,the role of TRIM3 in liver cancer remains unknown.This study aimed to investigate the function of TRIM3 in liver cancer cells.Methods:The protein levels of TRIM3 in five liver cancer cell lines (SK-Hep1,Hep3B,Huh7,HepG2,Bel-7402) and one normal liver cell line (L02) were detected with Western blotting.HepG2 and Bel-7402 cells with IowTRIM3 expression were infected with recombinant lentiviruses overexpressing TRIM3 (LV-TRIM3),whereas Huh7 and Hep3B cells with high TRIM3 expression were transfected with TRIM3-targeted small interfering RNA (siTRIM3).The functions of TRIM3 in the proliferation,colony formation,cell cycle,migration,invasion,and apoptosis of the above cell lines were examined.The effect of TRIM3 on tumor growth and metastases in nude mice was also investigated.Results:TRIM3 was overexpressed in HepG2 and Bel-7402 cells with LV-TRIM3 infection,which further reduced proliferation,colony formation,migration,and invasion of both cell lines.Cell cycle analysis showed thatTRIM3 overexpression induced G0/G1 phase arrest in HepG2 and Bel-7402 cells.Moreover,apoptosis was not increased in HepG2 or Bel-7402 cells overexpressing TRIM3.Contrarily,silencing TRIM3 expression in Huh7 and Hep3B cells by siTRIM3 led to significantly decreased percentages of both cells in the G0/G1 phase and promoted cell proliferation,colony formation,migration,and invasion.In vivo experiment results confirmed thatTRIM3 overexpression suppressed tumor growth and metastasis.Conclusions:TRIM3 plays a tumor-suppressing role in the regulation of liver cancer development by reducing cell proliferation through cell cycle arrest at the G0/G1 phase.

Objective: To systemically investigate the volatile organic compounds (VOCs) of Viola yedoensis, and to compare the VOCs differences of V. yedoensis obtained by the needle trap, static headspace and hydrostillation methods. Method: The needle trap, static headspace and hydrostillation methods coupled with gas chromatography-mass spectrometer (GC-MS) were developed for isolation and identification of the VOCs in V. yedoensis. The relative content of each component was obtained by peak area normalization with a triple-bed needle packed with Tenax, Carbopack X and Carboxen 1000 sorbents. Result: The 112 compounds were trapped by using needle trap, mainly moderate volatile components, including aldehydes, ketones, alcohols, monoterpenes, sesquiterpenoids and aromatic compounds. The static headspace and hydrodistillation methods were allowed to obtain 37 (mainly the high-volatile components) and 78 compounds (mainly low-volatile components), respectively. Only 13 common volatile components were detected in all these three methods. Conclusion: The results clearly demonstrated that the needle trap method is an alternative method for sampling VOCs of herbs, characterized by fast analysis, simple operation, good enrichment effect and high sensitivity.These three methods for VOCs analysis are complementary for each other.

OBJECTIVETo clone the gene encoding nucleocapsid protein (NP) of hantavirus strain Z10 (HV-Z10), to construct its prokaryotic expression system as well as to establish a rNP-IgM direct capture ELISA based on HRP-labeled recombinant NP (rNP), in order to detect serum samples of patients suffering from hemorrhagic fever with renal syndrome (HFRS) and to evaluate the effects of detection.

METHODSGene encoding NP of strain HV-Z10 was amplified by PCR and then its prokaryotic expression system pET28a-Z10N-E. coli BL21DE3 was constructed, using routine genetic engineering method. SDS-PAGE was applied to measure the expression of rNP and ion-exchange plus Ni-NTA-affinity chromatography was performed to purify the recombinant product. Western blot assay was used to determine the specific immuno-reactivity of rNP while HRP-labeled rNP-IgM direct capture ELISA was established to detect the serum samples from 95 cases of confirmed HFRS patients. The detection effect was compared with that by routine HV-IgM indirect capture ELISA method.

RESULTSpET28a-Z10N-E. coli BL21DE3 was able to express rNP with high efficiency. The purified rNP only showed a single protein fragment in the gel after SDS-PAGE. HV-IgG could efficiently recognize rNP and hybridize with the recombinant protein. 94.73% (90/95) of HFRS patients' serum samples were positively confirmed by rNP-IgM direct capture ELISA, while a positive rate of 92.63% (88/95) in the same samples was confirmed by HV-IgM indirect capture ELISA. The distributions of A450 values of the serum samples detected by the two IgM capture ELISAs as well as the changes of the A450 mean values from several serum samples with different dilutions were similar.

CONCLUSIONWe successfully constructed a high efficient prokaryotic expression system of NP encoding gene of hantavirus strain HV-Z10. The rNP-IgM direct capture ELISA that established in this study could be used as a new serological test for HFRS diagnosis because of its simplicity, safety, with high sensitivity and specificity.

Blotting, Western ; Capsid Proteins ; genetics ; immunology ; metabolism ; Enzyme-Linked Immunosorbent Assay ; methods ; Humans ; Immunoglobulin M ; immunology ; Recombinant Proteins ; genetics ; immunology ; metabolism ; Viral Core Proteins ; genetics ; immunology ; metabolism

OBJECTIVETo investigate the effect of hepatitis B virus(HBV) X gene on the expression of SPG21.

METHODSThe expressions of SPG21 mRNA and protein in HepG2 and HepG2.2.15 cells were tested by RT-PCR and western blot. HepG2 cells were co-transfected with reporter plasmid pGL3-SPG21 and plasmids carrying individual genes of HBV, the luciferase activity was measured and the expressions of SPG21 were detected by RT-PCR and western blot.

RESULTSThe expressions of SPG21 mRNA and protein were higher in HepG2.2.15 cells than in HepG2 cells (0.36+/-0.06 vs 0.21+/-0.05, P value is less than 0.05). The activity of SPG21 in HepG2 cells transfected with pCMV-X was higher (875+/-27 vs 67+/-12, P value is less than 0.01) as compared to blank control group (transfected with pCMV-tag2B). HBV X gene enhanced SPG21 gene promoter activity, SPG21 mRNA expression and SPG21 protein production in HepG2 cells in a dose-dependent manner.

CONCLUSIONHBV X gene can specially activate SPG21 expression.

Adaptor Proteins, Signal Transducing ; genetics ; metabolism ; DNA, Viral ; genetics ; Hep G2 Cells ; Hepatitis B virus ; genetics ; Humans ; RNA, Messenger ; genetics ; Trans-Activators ; genetics ; Transfection

BACKGROUND: Immune eye diseases such as hyperthyroidism exophthalmos and uveitis seriously endanger the eye health of patients, which are common and difficult eye diseases. Current treatments for these diseases include oral administration of hormones and immunosuppressive agents, with poor efficacy, recurrent attacks and poor prognosis. Meanwhile, these treatments can induce systemic adverse reactions. Lymphocytes are directly or indirectly involved in these diseases. Therefore, we try to make papua eye patch carrying immunosuppressant, and deliver the drug through the topical use. Cyclosporin A microemulsion targeting lymphocytes can treat or control palpebral lymph nodes involved in the immune eye diseases. It is a topical method rather than the systemic medication, which is targeted and has small doses of drugs. If possible, this treatment can effectively treat immune eye diseases and avoid systemic drug adverse reactions and long-term adverse reactions induced by original drugs. OBJECTIVE: To study the preparation of cyclosporin A microemulsion papua cream eye patch, and its transdermal absorption characteristics in vitro. METHODS: Cyclosporine A microemulsion was fully mixed with water-soluble polymer materials at a ratio of 1 mg:1 mL, including sodium polyacrylate, polyvinyl alcohol, polyvinylpyrrolidone, gelatin, peach gum, sodium carboxymethylcellulose, hydroxypropylcellulose, and then coated onto the non-woven fabric to prepare Babu cream. Permeability of the Babu cream on the abdominal skin of ICR mice was determined by Franz diffusion cell method. High-performance liquid chromatography was used to detect the concentration of cyclosporine A, and skin irritation and anaphylaxis were also measured. RESULTS AND CONCLUSION: Cyclosporin A microemulsion papua cream eye patch was successfully prepared with appropriate viscosity, good permeability, good permeability, comfortable application, no skin irritation and allergic reaction. The content of cyclosporine A was 10 mg/tablet, and the concentration was 1 g/L. The concentration of cyclosporine A microemulsion increased with the increase of time, and it had good transdermal effect. This study proved that it is feasible to prepare cyclosporine A microemulsion into papua patch. It has good performance in skin permeability, adhesion and skin comfort.