3T3-L1 adipocytes were cultured and differentiated into mature adipocytes in vitro. The adipocytes were intervened by ACAT inhibitor( 2 μg/ml) and ox-LDL with various concentrations (0,25,50,75,and 100 μg/ml)for 48 h,ACAT inhibitor( 2 μg/ml) and ox-LDL( 50 μg/ml) at the 0,6,18,36,and 48 h,or ACAT inhibitor( 2 μg/ml),ox-LDL( 50 μg/ml),and TUDCA with various concentrations(0,100,200,and 400 μ mol/L)for 48 h,respectively.The levels of visfatin in supernatant were examined by ELISA and the expressions of protein GRP78 and CHOP in adipocytes were detected by Western blot.After the adipocytes were treated with ACAT inhibitor and ox-LDL at different concentrations for 48 h,the cholesterol concentration and the expressions of GRP78 and CHOP protein in adipocytes and the visfatin levels in the supernatant fluid were increased with the increase of the ox-LDL concentration.The differences had statistical significance in the experimental groups compared with blank control group( all P＜0.05 ).After the intervention with ACAT inhibitor and ox-LDL for different durations,the expressions of GRP78 and CHOP protein in adipocytes and the visfatin levels in the supernatant fluid were up-regulated in a timedependent manner.The differences between experimental groups and blank control had statistical significance( all P＜0.05 ).After the intervention with ACAT inhibitor,ox-LDL,and different concentrations of TUDCA for 48 hours,the expressions of GRP78 and CHOP protein in adipocytes and visfatin levels in the supernatant fluid were down-regulated in a dose-dependent manner and as compared with blank control group the difference were statistically significant( all P＜ 0.05 ).The increase of cholesterol load in adipocyte may promote the visfatin secretion,denoting that the mechanism might be due to the enhancement of endoplasmic reticulum stress in aidpocytes.

Objective To investigate the role of intestinal immune dysfunction in the pathogenesis of irritable bowel syndrome(IBS) and to study the effects of Clostridium butylicum on the regulation of intestinal immune disorders.Methods A total of 50 male 6-week-old C57BL/6 mice were randomly divided into three groups,including the experimental group (n =20),the control group (n =20) and the Clostridium butylicum group(n =10).A mouse model of constipation-predominant IBS (C-IBS) was established by perfusing sodium butyrate solution(200 μl,concentration of 500 mmol/L) into the mouse colon twice a day for three consecutive days.The mice in control group were intrarectally perfused with normal saline enema (200 μl).Two hours before the perfusion of sodium butyrate into colon,the mice in Clostridium butylicum group were given Clostridium butylicum 500 μl(viable cell concentration of 1×109 CFU/ml) by oral gavage once a day for six days.The colorectal distention test(CRD) was carried out for evaluation of clinical parameters.HE staining of intestinal tissue section was performed for histopathological assessment of colonic mucosal inflammation.Intestinal intraepithelial lymphocytes (IELs) and lamina propria mononuclear cells (LPMCs) were isolated and analyzed by flow cytometry to evaluate the correlation between IBS and intestinal immune dysfunction/abnormal activation of intestinal immune cells in mouse model of C-IBS,and to assess the regulatory effects of Clostridium butylicum on the intestinal immune disorder.Results (1) Compared with the control group,the mice in experimental group showed a significant change in physiological parameters,histological structure of colon,inflammatory cells infiltration and low-grade inflammatory state.There was a significant increase in scores of CRD and a decrease in lowest sensory threshold (t=8.926 and t=6.103,both P＜0.001) ; (2) There was a decrease in the numbers of DC in IELs (t =2.878 and t =3.086,both P＜0.05),but an increase in the numbers of macrophage (t=3.191,P＜0.05) and the memory T cells in mice with IBS (t=3.071,P＜0.05) as compared with that in control group; (3)DCs were decreased (t=2.880 and t=2.664,both P＜0.05),but memory T cells were increased (t =3.732 and 2.682,P＜0.01 and P＜0.05) in the LPMCs of mice in experimental group; (4)There was no significant difference in the physiological index between the mice in control group and the Clostridium butylicum group.Levels of memory T cells,macrophages and DCs in the IELs were close to the normal level (6 d,t =1.103,0.0213,0.418,all P＞0.05),and levels of macrophages and DCs in the LPMCs of mice in the Clostridium butylicum group were also similar to that in the control group (6 d,t =0.782,0.347,both P＞0.05) ; (5) Compared with the mice in experimental group,the level of memory T cells in LPMCs of mice treated with Clostridium butylicum was dramatically declined (6 d,t=2.346,P=0.0470,P＜0.05),however,which was still higher than that of mice in control group (6 d,t =2.233,P =0.0476,P＜0.05).The intestinal immune function was restored to normal level with Clostridium butylicum intervention.Conclusion The pathophysiologic mechanism of IBS might be closely related to the abnormal activation of intestinal immune cellsand disordered functional state in the intestinal mucosa.Clostridium butylicum could regulate the intestinal immune homeostasis and restore the physiological function of gastrointestinal tract.

Objective To understand the levels of plasma endothelin (ET) and calcitonin gene-related peptide (CGRP) in cough variant asthma(CVA) patients, and to investigate their correlation and clinical implications. Methods Thirty CVA patients,30 typical bronchial asthma patients and 25 healthy controls were recruited. Three milliliter limosis venous blood were drawn from each patient to measure the levels of plasma ET and CGRP by radioimmanity. Results ①The levels of ET in the CVA group and typical bronchial asthma group were higher than that in healthy controls (P < 0.01). Their values were (103.58±28.66) ng/L, (129.37±27.28) ng/L and (72.63±21.52)ng/L, respectively. The levels of CGRP in the CVA group and typical bronchial asthma group were (7.62±2.56) ng/L and(6.63±2.09)ng/L The level of CGRP in the healthy controls group was (21.60±3.29) ng/L. The first two groups were lower than the latter(P < 0.01). However, there was no significant difference between the CVA group and typical bronchial asthma group(P >0.05). ②ET and CGRP were negatively correlated in both CVA group(r= -0.819,P<0.05) and typical bronchial asthma group(r= -0.738,P<0.05). Conclusions ET and CGRP were negatively correlated in both CVA group and typical bronchial asthma group, which means that ET and CGRP were a couple of antagonistic factors participated in the regulation of CVA, and may play an important role in the process of CVA.

Objective To investigate the effects of carboxymethyl-chitosan (CM-CH) on keloid fibroblasts (KFB) proliferation and collagen synthesis. Methods Keloid fibroblasts were isolated from fresh keloid tissue and cultured. The effect of CM-CH on proliferation was examined by MTT assay. The synthesis of collagen was evaluated by hydroxy proline (HP) colorimetric analysis. Results The fibroblasts were treated with CM-CH at concentration of 10 ?g/ml, 50 ?g/ml, and 100 ?g/ml 48h and 72h after treatment, and all of the three concentrations showed inhibitory effects significantly (P0.05) at 200 ?g/ml concentration after 24h, 48h, and 72h. CM-CH at concentration of 10 ?g/ml, 50 ?g/ml, and 100 ?g/ml after treatment for 48h on KFB could markedly decrease the synthesis of collagen (P

Objective:To explore the methods and condition in which calcium ionorphore(CI) induces the CML cells to differentiate into dendritic cells(DCs).Methods:Mononuclear cells were separated from peripheral blood or bone marrow of CML patients whose WBC counts were more than 30?30~9 L~ -1 when samples were collected,then lymphocytes and monocytes were discarded by pouring out supernatant twice at different culture time point. Slightly adherent cells were cultured in RPMI1640 containing 10%FCS(Fetal calf serum),with or without CI(375 ng/ml) and GM-CSF(200 ng/ml) at 37℃,5%CO_2 humidified atmosphere for 96 h. To evaluate the effect of CI on inducing CML cells to differentiate into DCs,the phenotype of these cells were analyzed by flow cytometry and the morphology change was observed under inverted microscope and electron microscope. Better condition was also explored for DCs differentiation from CML cells under the effect of CI.Results:After 96 h of culture with CI and GM-CSF,the CML cells acquired morphology of mature DCs and significantly up-regulation of CD80,CD86,CD40,CD86 and HLA-DR.Conclusion:CML cells might acquire typical morphology and immunological phenotype of mature DCs when being cultured with CI and GM-CSF.

BACKGROUND: The involvement of genetic factors in migraine has been under close investigation, and geneiic epidemiological study and segregation analysis have confirmed genetic disposition as an important risk factor for migraine.OBJECTIVE: To analyze the connection between mutations in CACNA1A gene and familial hemiplegic migraine (FHM) in southern Chinese Han patients by examining the three most frequently mutated sites in CACN1A gene.DESIGN: Sampled survey.SETTING: First Affiliated Hospital of Zhongshan University and Baoan Xixiang People's Hospital of Shenzhen City.PARTICIPANTS: All the participants were selected from patients in the above two hospitals and their relatives, including 10 patients with FHM, 12 relatives of the patients in 2 pedigrees, 53 migraine patients with aura without family history, and 10 healthy control subjects.METHODS: The exons 13, 16 and 17 of CACNL1A4 gene were amplified by PCR. Single-strand conformation polymorphism technique was employed to detect the most frequent mutations in the 3 exons (T666M, R583Q and D715E) in these subjects.MAIN OUTCOME MEASURES: ① Results of PCR amplification of the 3 exons of CACNL1A4 gene; ② Results of SSCP for mutation analysis of the 3 exons.RESULTS: Participants completed the study. The target fragment length of exons 13, 16 and 17 were 247 bp, 268 bp and 204 bp, respectively.None of mutations of T666M, R583Q and D715E were detected in the subjects, including FHM patients and their relatives, migraine patients without family history and the healthy control subjects.CONCLUSION: None of the 3 most frequent mutations (T666M, R583Q and D715E) can be detected in southern Chinese FHM pedigrees or migraine patients without family history.

Objective To establish appropriate correction models of serum marker α -fetoprotein (AFP) multiple of the median (MOM) for prenatal screening on patients with neural tube defect (NTD) at the second trimester to replace the foreign data currendy used in Beijing with a higher sensitivity.Methods A sample of 5815 mid-gestation pregnant women with normal pregnancy outcome were selected including 5557 cases of age ＜ 35 years and 258 cases of age ≥35 years.The regression relationship between AFP median and gestational age was builded by exponential model and the regression relationship between MOM and weight was builded by power model in age ＜ 35 years.The regression relationship between AFP median and gestational age was builded by S model and the regression relation ship between MOM and weight was builded by quadratic model in age ≥35 years.The normal reference values in two age sections were estimated approximately by percentiles method.Results Corrected AFP MOM values with mean value (age ＜ 35 years:1.001; age ≥35 years:1.113) and median value (age ＜ 35 years:0.934;age ≥35 years:0.990) obtained by regression analysis were more close to 1.00 than the data from the software[mean value(age ＜ 35 years:1.164; age ≥35 years:1.254) and median value (age ＜35 years:1.093; age ≥35 years:1.149)].There were significant differences (age ＜35 years:t =65.120,P =0.000;age ≥35 years:t =9.812,P =0.000).Conclusion Correction model for maternal serum marker is more applicable for Beijing population at the second-trimester prenatal screening.

OBJECTIVE To discuss the best scheme for fungemia detection by analyzing and comparing the ability and performance characteristics of the BACTEC 9120 automated blood culture system and 4 kinds of blood culture bottles in the detection of simulated fungemia.METHODS Simulated blood culture was produced using 65 fungi isolates from clinical specimens and BACTEC Plus Aerobic/F,Plus Anaerobic/F,Peds Plus/F and Myco/F Lytic blood culture bottles and detected by BACTEC 9120 automated blood culture system.The final inoculum densities were 1-5 CFU/ml blood.The(time to detection TTD) of simulated blood culture with different concentrations of suspension produced using 2 kinds of standard strains and 4 kinds of blood culture bottles was compared.RESULTS From the 260 bottles in this study 216 had growth detected by the BACTEC 9120 blood culture system.The positive rates of BACTEC Plus Aerobic/F and Anaerobic/F,which were 90.77%and 41.54%,respectively,were significantly(P

Objective To investigate the relationship between the resistance of the Klebsiella pneumoni-ae and the Klebsiella pneumoniae plasmid pNDM-LJ carrying blaNDM-1 by high-throughput DNA sequencing. Methods High-throughput DNA sequencing was carried out by the Illumina Miseq platform , and sequencing data were assembled by Edena software. Contigs were annotated by the RAST server and analyzed by the BLAST server. Results The plasmid pNDM-LJ was 54-kb in size with a GC content of 49%. The plasmid encoded 52 putative functional genes and belonged to the IncX3 group in incompatible classifications. Analysis of the plasmid sequence revealed high similarity with other IncX3 plasmids. The blaNDM-1 gene was located in a complicated gene environment possibly constructed by several transposition events. The 5′ and 3′ ends of the blaNDM-1 gene were adjacent to the ISAba125 and IS 26 respectively , forming a 10.8-kb transposon-like structure. Conclusion The plasmid pNDM-LJ carried the blaNDM-1 gene being resistant to carbapenems and played a possibly impor-tant role in transmission of blaNDM-1 in China.

Objective To explore the establishment methods of animal models of adult growth hormone deficiency, and to provide a good model for experimental research and treatment for abnormal bone metabolism caused by growth hor-mone deficiency.Methods The methods of establishment of animal models of adult growth hormone deficiency were re-viewed and evaluated refering to literature.Results There were three methods including spontaneous lack-of, pituitecto-mized and gene knockout can establish animal models of adult growth hormone deficiency.Conclusions Hypophysecto-mized animal models are inexpensive and easy to create, but not suitable for studying the relationship between growth hor-mone and bone metabolism.Spontaneous lack-of and gene knockout models are specific growth hormone deficiency and of great research significance in exploring the relationship between growth hormone and bone metabolism.