Objective To investigate the effects of JNK inhibition on insulin signaling pathway. Methods HepG2 cells were treated with different kinds of JNK inhibitors for 12 h, and then the cells were treated with 10 nmol/L insulin for 5 min to stimulate insulin signaling pathway. Mitogen-activated protein kinases ( MAPK ) and insulin signaling pathways were analyzed by Western blot using the total cell lysates. Results JNK activity was significantly inhibited by JNK inhibitor JNKi-Ⅷand results showed that JNKi-Ⅷtreatment could reduce insulin signaling pathway in a dose-dependent manner. Furthermore, other JNK inhibitors including JNKi-Ⅴ, JNKi-Ⅲ, and SP600125 blocked JNK activity in HepG2 cells. Similar to JNKi-Ⅷ, these JNK inhibitors also impaired insulin signaling transduction in a dose-dependent manner. Conclusion In HepG2 hepatocytes, JNK activity inhibition blocks insulin signaling transduction.

Objective: To investigate the alteration of bcl- 2 gene family in the rat brain and the molecular mechanism of neuronal apoptosis following traumatic brain injury. Methods: Male Sprague -Dawley rats were subjected to lateral fluid percussion brain injury(FPI) of mo derate severity. Bcl-2, Bcl-x and Bax protein expression was detected by immun ohistochemistry. Results: (1) The immunoreactivity of Bcl-2 and Bcl-x protein decreased in the hippocampus ipsilateral impact site as early as 6 h post-injury, and this was the main cause of down-regulation of the ratio of Bcl-2+Bcl-x to Bax. (2) During 1-3 d after injury, the Bax protein express i on increased significantly, while the Bcl-2 and Bcl-x protein expression decre ased relatively slow. The decreased ratio of Bcl-2+Bcl-x to Bax was mainly due to the Bax up-regulation. Conclusion: The bcl-2 gene family is involved in neuronal apoptosis after FBI, and the protein expression alteration of the family members leads the neuronal cell to apoptosis.

Objective To study the relationship between serum cystatin C(Cys C)level and the development of insulin resistance(IR)in elderly patients with type 2 diabetes mellitus (T2DM).Methods A cross-sectional survey research involving 757 elderly patients with T2DM was performed and patients were divided into groups according to Cys C and IR levels.Serum 1evels of fasting insulin (FBI),fasting blood glucose(FPG),fasting C-peptide (FCP),glycosylated hemoglobin A1 c (H bA1 c),homeostasis model assessment of insulin resistance (IR) (HOMA2 IR-C),homeostasis model assessment of insulin secretion,homeostasis model assessment of insulin sensitivity(HOMA2-％ S-C),micro-albuminuria(mALB)and serum lipid were measured and compared among the groups.Possible influence factors were adjusted,and the correlation between Cys C levels and IR was analyzed.The influencing factors on IR were also analyzed.Results There were statistically significant differences in ages,course of T2 DM,FBI,FCP,HbA1c,urinary mALB,IR,insulin secretion,insulin sensitivity,morbidity rate of coronary heart disease,hypertension and diabetic nephropathy among the three groups(allP＜0.05).Insulin sensitivity was decreased with the increase of Cys C level,while others were increased.Among 757 patients,the level of serum Cys C was positively correlated with FCP,HOMA2-IR C levels,and was negatively correlated with HOMA2-％ S-C levels(P＜0.05).Multiple logistic regression analysis showed that the higher level of Cys C was independent risk factors for IR in elderly T2DM patients(OR=3.41,95％CI:2.22-5.22,P＜0.05).Conclusions Serum Cys C level is closely related with IR in elderly T2DM,and the elevated level of serum Cys C is one of independent risk factors for elderly T2DM.

Objective: To investigate the alterations of bcl-2 gene family in the rat brain and the molecular mechanism of neuronal apoptosis follow ing traumatic brain injury (TBI). Methods: Male Sprague-Dawley rats were subjected to lateral fluid percussion brain injury(FPBI) of moderate severity. bax and bcl-xL mRNA and protein expression was detected by RT-PCR an d immunohistochemistry. In addition to morphological evidence of apoptosis, TUNE L histochemistry was used to identify DNA fragmentation in situ under both l ight and electron microscope, whereas characteristic internucleosomal DN A fragm entation of apoptosis was demonstrated by DNA gel electrophoresis. Resul ts: bcl-xL mRNA and protein decreased in the ipsilateral hemisphere t o the impact site as early as 6 h post-injury［(67.42±7.54)% and (85.85±5.72)% r espectively］. The decrease in bcl-xL mRNA and protein preceded apoptosis was observed 12 h post-injury. And this was the main cause of up-regulation of the ratio of bax to bcl-xL in the acute period(minutes-hours) followin g FPBI. bax mRNA and protein were observed to rise slowly, doubled 3 d post- injury, returned to sham level slowly. The delayed cell death (days-weeks) migh t associated with the up-regulation of pro-apoptotic gene bax. Conclusio n: The expression of bcl-xL and bax coincide with apoptosis following TBI. The reg ulation of bax and bcl-xL by TBI occur before transcription. The balance of bax/bcl-xL ratio determines the neurocytes to survive or die following FPBI.

Chemical constituents of 95% ethanol extract of the dried persistent calyx of Physalis pubescens were investigated. By chromatography on a silica gel column and reverse-phase preparative HPLC, 10 compounds were isolated from the dichloromethane fraction. Based on the MS and 1D/2D NMR data, these compounds were identified as 5-O-(E-feruloyl) blumenol (1), isovanillin (2), (E) -ethyl 3-(4-hydroxyphenyl) acrylate (3), 4-hydroxybenzaldehyde(4), 4-methylphenol (5), (E) -methyl cinnamate (6), 7,3',4' trimethoxyquercetin (7), 5,3', 5'-trihydroxy-3,7,4'-trimethoxyflavone(8), danielone (9), and 5,5'-diisobutoxy-2,2'-bifuran (10).
Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Molecular Structure ; Physalis ; chemistry ; Spectrometry, Mass, Electrospray Ionization

Objective To explore the clinical outcomes of different mild-stimulation in poor ovarian responders in vitro culture/intracytoplasmic injection (IVF/ICSI) to provide the evidence for clinical treatment,and investigate clinical parameters including pre-ovulation rate,rate of oocytes retrieved,fertilization rate,good quality embryo rate,and pregnancy rate per cycle.Methods The clinical date of a total 180 IVF cycles of infertile patients undergone from Jan 2013 to May 2016 in medical reprodution center of Xiang Tan Central Hospital.According to the different methods of controlled ovarian hyperstimulation (COH),the cycles were divided into 3 groups:group A with 60 cycles of early five days using clomiphene,group B with 60 cycles of persistent using clomiphene,and group C with 60 cycles using artificial luteal-phase ovarian stimulation protocol.Results The basic situations in three groups were no significant difference (P ＞0.05).Compared to groups B and C,the pre-ovalution rate was more in group A (P ＜ O.05),and the rate of oocytes retrieved,fertilization rate,good quality embryo rate,and preganacy rate per cycle were higher in groups B and C (P ＜ 0.05).The rate of oocytes retrieved,fertilization rate,good quality embryo rate,and preganacy rate per cycle were no significant difference among gourps B and C.The good quality embryo rate and preganacy rate per cycle in groups B and C were higher than group A,and group B was higher than group A (P ＜ 0.05).Conclusions Persistent using clomiphene or persistent using clomiphene in COH could decrease the rate of pre-ovalution,and get more rate of oocytes retrieved,good quality embryo rate,and preganacy rate per cycle.

Objective To approach the changes of cytosol phospholipase A2α(cPLA2α)and nitric oxide (NO)in patients with chronic obstructive pulmonary disease(COPD)and its significance. Methods One hundred patients with COPD admitted into Department of Critical Care Medicine of Affiliated Wuqing Traditional Chinese Medicine(TCM)Hospital of Tianjin University of TCM were enrolled,and according to the COPD severity grading standards,they were divided into mild group(25 cases),moderate group(25 cases),severe group(26 cases) and extremely severe group(24 cases);simultaneously,90 cases with normal pulmonary function who had taken health examination were chosen and assigned to the healthy control group. The cPLA2α level was detected by enzyme linked immunosorbent assay(ELISA),the level of uric acid(UA),total cholesterol(TC),triacylglycerol (TG)were detected by enzymatic method,and serum NO metabolites(NOx)level was detected by nitrate reductase method. Results Compared with the healthy control group,the serum levels of cPLA2α and UA in patients with different severity of COPD were significantly increased;along with the increase of patient's COPD grade of severity,the cPLA2α,UA levels were gradually increased,while NOx level was gradually decreased in mild, moderate, severe, extremely severe groups〔cPLA2α(ng/L):125.60±8.17, 155.20±6.42, 190.20±9.32, 255.80±11.28 vs. 88.50±7.99;UA(μmol/L):381.23±32.22,434.95±87.71,464.81±52.65,487.45±82.61 vs. 241.95±52.33;NOx(μmol/L):59.90±17.52,45.60±6.17,38.20±4.08,25.70±3.04 vs. 74.90±18.31,all P<0.05〕. The differences in blood cPLA2αand serum NOx level among groups with different severity of COPD were of statistical significance(P<0.05). The levels of TC,TG among these different severity groups had no statistical significance(all P>0.05). The cPLA2αand NOx levels presented significant negative correlation(rs=-0.798,P=0.013). Conclusion The combined examination of blood cPLA2αand serum NOx levels can evaluate the severity degree of COPD patients,and cPLA2αcan be used as a new target index for COPD grading.

Objective　To observe some biological features of bone marrow mesenchymal stem cells and explore the best conditions for isolatin g and culturing in vitro. Methods　Common cell culture techn ique, light and electron microscopy were used to study the effects of the growth , proliferation, morphology of the bone marrow mesenchymal stem cells in differe nt adherent time, concentration of serum and cell density. Results　The best culture condition in vitro for growth was 4-24 hours adherent time, 5%-10% fetal bovine serum, (4-8)×104/ml cell density. The cells were sp indle in shape and had a strong ability of proliferation. The time for cell duplication was 3 to 4 days. The cells showed the characteristics of stem cell s in electron microscope. Conclusion　The best condition for iso lation and culture of bone marrow mesemchymal stem cells was successfully establ ished and some biological features were obserred. It found a base for further in vestigation and using of mesenchymal stem cells.

OBJECTIVETo study the feasibility and safety of transumbilical single incision laparoscopic cholecystectomy.

METHODSEighty-two cases with gallbladder diseases were underwent with transumbilical single incision laparoscopic cholecystectomy. Some difficulties and countermeasure in operations were analyzed.

RESULTSNine in all patients were converted because of the surrounding inflammation of gallbladder, difficult to dissect in Calot's triangulation and variation of gallbladder artery. Of them, 6 cases converted to two-hole laparoscopic cholecystectomy and 3 cases to open operation. Other 73 cases were successfully operated with transumbilical single incision laparoscopic cholecystectomy. The success rate was 89.0%. The average operative duration was 48.4 min. The average operative blood loss was 20.8 ml. Patients returned to liquid food on the first day after operation. Hospital duration was 3 ~ 6 d. During 13.7 months of follow-up, there was no bile duct injury, large bleeding, incision infection, bile fistula and umbilical hernia. The incision healed well. The scar in umbilicus was concealing and difficult to be observed.

CONCLUSIONSTransumbilical single incision laparoscopic cholecystectomy is safe and feasible. The difficulties in technique are easy to be broken through.

Adolescent ; Adult ; Aged ; Cholecystectomy, Laparoscopic ; methods ; Female ; Follow-Up Studies ; Gallbladder Diseases ; surgery ; Humans ; Middle Aged ; Retrospective Studies ; Treatment Outcome ; Umbilicus ; surgery ; Young Adult

OBJECTIVETo construct a retrovirus-mediated expression system carrying human SHP-1 gene to transfer SHP-1 gene in human breast cancer MDA-MB-231 cells.

METHODSThe full-length SHP-1 gene fragment was amplified by RT-PCR from the total RNA extracted from human breast cancer cell line MCF-7 over-expressing SHP-1 protein. The gene fragment was inserted into the vector pLNCX2 to construct the recombinant retroviral plasmid, which was transfected into the packaging cell PT67 via Lipofectamine2000. A cell line stably producing the virus was selected with G418. MDA-MB-231 cells was infected with the virus, and the expression of SHP-1 gene in the positive cell clone was detected with Western blotting.

RESULTSA 1.8 kb cDNA fragment of SHP-1 gene was obtained from MCF-7 cells and successfully inserted into the pLNCX2. A stable cell clone PT67/SHP-1 and virus supernatant were obtained. Expression of SHP-1 protein was detected in the cells infected with the virus.

CONCLUSIONThe recombinant retroviral vector carrying SHP-1 gene has been successfully constructed and MDA- MB-231/SHP-1 cell line expressing SHP-1 has been obtained to allow further functional study of SHP-1 in breast cancer.

Breast Neoplasms ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Cloning, Molecular ; Genetic Vectors ; genetics ; Humans ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Retroviridae ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection