In order to improve the expression level of segment of GABAA receptor a1 subunit in E. coli, growth conditions of the recombatant, which influence the final yield of protein expression, including growth medium, inoculation ratio, temperature, pH, rotation speed, inducing time and concentration of IPTG and so on, were studied in shaking flasks. The results indicated that, with 3% inoculation ratio, cultured 3.5 hours at 37℃, and then induced 5 hours by IPTG at 32℃, the yield of GABAA receptor protein was 95mg/L and the biomass was 3.25 g/ L. In contrast, using a 16 L stirred fermentor instead of shaking flasks, the highest level of the protein expression, 136mg/L with 4.95g/L of biomass, was achieved after fermenting 5.5 hours.

Objective To observe the ultrastructure changes of detrusor of urinary bladder following partial outlet obstruction. Methods 14 New Zealand white male rabbits were divided into "A" and "B" group.The "A" group rabbits underwent operation to induce bladder outlet obstruction,"B" group rabbits served as controls.The ultrastructures including rough endoplactmic reticulum and mitochondrion of detrusor cell of each bladder were examined after 5 weeks. Results The average area of rough endoplactmic reticulum in the obstructed group was (5.377?2.318) ?m 2 in each unit cytoplasm area (115.28 ?m 2) versus (0.476?0.319) ?m 2 in the controls ( P

Objective To observe the effect of electroacupuncture on improvement of learning and memory ability and the expression of connective tissue growth factor(CTGF)mRNA and protein in hippocampus in diabetic rats with cognitive impairment.Methods The rat diabetes model was induced by injecting streptozotocin(20 g/L),and then the rats were randomly divided into three groups:electro-acupuncture group(EA),diabetes-mellitus-untreated group(DM)and control group(CN).After four weeks of electroacupuncture treatment,blood glucose level was determined and the effect of electroacupuncture on learning and memory was examined with the device of Morris water maze.RT-PCR was used to detect CTGF mRNA level,and immunohistochemistry was used to detect CTGF protein expression.Results Blood glucose level and the latency period in DM group were increased compared with those in EA and CN groups(P

Blood Coagulation ; Blood Viscosity ; China ; Diagnosis, Differential ; Humans ; Medicine, Chinese Traditional

Blood Coagulation ; Blood Viscosity ; Diagnosis, Differential ; Humans ; International Cooperation ; Medicine, Chinese Traditional

OBJECTIVE To investigate the immunoregulatory role of tolerogenic dendritic cells(TolDC) in allergic airway inflammation in mouse model of allergic rhinitis(AR).METHODS A total of 24 Balb/c mice were equally and randomly divided into 4 groups with 6 mice in each group. AR group was established with oval bumin(OVA) sensitization and challenge, meanwhile the physiological saline(PBS) sensitization and challenge as the control group. AR group were treated by adoptive transfer of TolDC as treatment group. And treatment group were injected intraperitoneally with TGF-β/IL-10R neutralizing antibody as blockade group. In each group, allergic nasal symptoms score, inflammatory cell infiltration in lung, the expression of Th1/Th2-derived cytokines in the bronchoalveolar lavage fluid(BALF) and nasal lavage fluid(NALF), the expression of OVA-specific serum IgE and the expression of Treg in lung were measured.RESULTS The mouse model of OVA-induced AR was successfully developed. Compared with AR group, treatment group exhibited lower allergic nasal symptoms score, a decrease in inflammatory cell infiltration in lung, lower expression of Th1/Th2-derived cytokines in BALF and NALF and OVA-specific serum IgE, as well as up-regulation of Treg in lung, which were abolished by TGF-β/IL-10 neutralizing antibody shown in blockade group.CONCLUSION TolDC suppress airway inflammation in AR by inducing regulatory T cells through TGF-β/IL-10-dependent mechanisms.

Increasing attention has been focused on the antitumor activity of artemisinin derivatives in recent years, for artemisinin had been reported to have cytotoxic effects against HL-60, P388 and MCF-7 tumor cells. We report here the synthesis and evaluation for antitumor activity of a series of artemisinin-ether derivatives bearing tetrahydropyrrole, morpholine, piperidine, substituted piperidines and azoles with various linkers. Sixteen 10-O-substituted dihydroartemisinin derivatives were designed and synthesized, all of which have never been reported in literatures and whose antiproliferative effects on human breast cancer MCF-7, MCF-7/Adr and HL-60 cells were determined by MTT assay or direct cell counting. Each of these artemisinin derivatives possessed better effects than dihydroartemisinin evidently against HL-60 and MCF-7 cells growth, while less potent than doxorubicin. All target compounds exhibited significantly improved potency compared to DHA and doxorubicin on the doxorubicin-resistant MCF-7/Adr cells, so did they in their sensitive counterparts MCF-7 cells. Among them, compounds GF02, GH04 and ZH04 showed strong activity against these three cell lines growth. Further research is undergoing.
Antineoplastic Agents ; chemical synthesis ; chemistry ; Artemisinins ; chemical synthesis ; chemistry ; Breast Neoplasms ; pathology ; Cell Proliferation ; Doxorubicin ; Drug Design ; HL-60 Cells ; drug effects ; Humans ; MCF-7 Cells ; drug effects

Aim To develop an in vitro high throughput drug screening system based on reporter gene assay for identification of novel compounds with PXR, FXR and LXRα agonist activity. Methods The expressions of exogenous PXR, FXR and LXRαgene in HEK293, HepG2 and LS174T cells were examined by Real-Time quantity PCR. pSG5-hPXR and pGL3-XREM-CYP3A4, pEGFP-N3-hFXR and EcRE-TK-Luc, pCMX-FLAG-hLXRα and pGL3-XREM-CYP3A4 were cotransfected into cells and the optimal ratio of three plasmids was determined. The dose-response relationship between the positive drug and the fold induction was determined. The specificity of the model was ex-amined, and the repeatability was also determined by Z′ value. Results ① The PXR, FXR and LXRα mRNA expression in HEK293 cell is low among three different cells. ②reporter gene vector and expression plasmid ratio of 1∶ 1, 2∶ 1 and 2∶ 1 were proved to be suitable for highest relative luciferase activity for PXR, FXR or LXRα agonist screening model. ③ The relative luciferase activity was induced by Rif, CDCA or T0901317 in a dose-dependent manner. ④Only Rif, CDCA or T0901317 could significantly increase the relative luciferase activity in PXR,FXR or LXRα agonist screening model, no effect of other nuclear re-ceptors agonist was observed, and the values of Z′-factor for PXR, FXR and LXRαagonist screening model were 0. 58, 0. 66 and 0. 63, respectively. Conclusion An in vitro PXR, FXR and LXRα agonist high-throughput screening models are devel-oped with acceptable specificity and repeatability, and the mod-els can be used to screen PXR, FXR and LXRα agonist.

Objective To analyze the clinical characteristics , diagnosis, therapy and prognosis of new diagnosed pri-mary gastric diffuse large B cell lymphoma ( PGDLBCL) and to discuss the efficacy of rituximab .Methods Between Jan 2005 and May 2012 , twenty-six new-diagnosed PGDLBCL patients were reviewed retrospectively .The clinical characteris-tics, diagnosis, therapy, results and prognostic factors were analyzed .Results There were 14 males and 12 females.Their age ranged from 25 to 82 (median, 50.1) years old.The most common symptom was stomachache .Treatment strategies were chemotherapy alone ( n=9) [ scheduled as cyclophosphamide , doxorubicin , vincristine and prednisone ( CHOP) and CHOP-like] and chemotherapy combined with rituximab (n=17), followed by radiotherapy of the stomach with or without regional nodes .All clinical and pathological features were similar between the two groups .The median follow-up time was 40 months.The overall response rate was 100%(9/9)in CHOP group, including 55.56%(5/9) CR, and 93.75%(15/16) in RCHOP group including 50%(8/16) CR (P>0.05).The total PFS and OS of 5 years were 60.3%and 74.4%respectively.The PFS in CHOP group and RCHOP group was 66.7% and 58.9%, respectively,and the OS was 66.7%and 84.6%, respectively.Although the OS of RCHOP group was much better than that of CHOP group , there was no sta-tistically significant difference.Univariate analysis showed that IPI (P<0.05) and Lugano staging (P<0.05) were inde-pendent factors of survival in patients with PGDLBCL .Conclusion Chemotherapy could be the first-line therapy of PGDL-BCL.The overall survival rate might be increased by adding rituximab to chemotherapy .The Lugano stage and IPI are im-portant prognostic factors .

Background:Fenretinide,which is capable of generating reactive oxygen species( ROS ),has emerged as a promising antineoplastic agent based on numerous in vitro and in vivo studies and clinical chemoprevention trials. Preliminary studies showed that fenretinide could induce apoptosis in human hepatocellular carcinoma( HCC)cells in vitro, however,the precise mechanism was not clarified. Aims:To elucidate the effect of ROS on apoptosis of human HCC cells induced by fenretinide and the underlying mechanism. Methods:Human HCC cell line Huh-7 was treated with antioxidant vitamin E,fenretinide or their combination,respectively. ROS in live cells was evaluated by confocal microscopy and flow cytometry;cell viability and apoptosis were assessed by CellTiter-Glo Luminescent Cell Viability Assay Kit and Caspase-Glo3/7 Assay Kit;expression and intracellular localization of nuclear receptor Nur77,as well as expression of stress-induced transcription factor GADD153 were measured by immunofluorescence staining and Western blotting,respectively. Results:Vitamin E pretreatment fully blocked the fenretinide-induced ROS production. In Huh-7 cells pretreated with vitamin E,cell apoptosis induced by fenretinide was significantly reduced(P<0. 05). Furthermore,effect of vitamin E pretreatment was noteworthy on reducing fenretinide-induced GADD153 expression, while no significant impact on fenretinide-induced Nur77 expression and translocation was observed. Conclusions:Elimination of ROS by vitamin E can abrogate the pro-apoptotic effect of fenretinide on Huh-7 cells,which indicates the participation of ROS in fenretinide-induced apoptosis of human HCC cells. Its mechanism might be associated with induction of GADD153 protein expression.