ObjectiveTo investigate the expression of G protein-coupled ER (GPER) and ER in the activation of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) induced by 17β-estradiol (17β-E2 )in endometrial carcinoma cells,Ishikawa and HEC-1A.Methods Expressions of GPER,Erα and Erβ protein in Ishikawa and HEC-1A cells were detected by immunohistochemical SP method.Levels of GPER,Erα and Erβ were examined by western blot in Ishikawa and HEC-1A cells after treated with 1 ×10-6 mol/L 17β-E2 at different time (0,15,30,60,120 minutes).ResultsGPER was positive expressed in Ishikawa and HEC-1A cells.Erα and Erβ were both positive expressed in Ishikawa cells.While,Erα was weakly expressed and Erβ was almost negatively expressed in HEC-1A cells.Western blot analysis showed that 1× 10-6 mol/L 17β-E2 treatment,the Ishikawa and HEC-1A cells GPER protein level for 15 minutes markedly increased (P ＜ 0.05 ),which Ishikawa 30 minutes,when cells reached the highest level (0.192 ± 0.004),HEC-1A cells for 15 minutes and reached the highest level (0.184 ±0.006) ; Ishikawa and HEC-1A cells,Akt,activation of 15 minutes from the treatment start was significantly increased (P ＜ 0.05 ),which Ishikawa cells for 30 minutes and reached the highest level (0.666 ± 0.021 ),HEC-1A cells for 15 minutes and reached maximum (0.788 ± 0.035); Ishikawa and HEC-1 A cells,Erα and Erβ protein expression did not change significantly ( P ＞ 0.05 ).Conclusion GPER likely involved in non-nuclear activation of PI3K/Akt signaling pathways in endometrial carcinoma cells,Ishikawa and HEC-1A.

ObjectiveTo investigate the effect of SalB on bone marrow-derived mesenchymal stem cells (BMSCs) apoptosis induced by hypoxia and serum deprivation (hypoxia/SD) in the vitro. Methods BMSCs were cultured in the vitro and randomly divided into control group, hypoxia/SD group and SalB group.SalB group was composed by four groups and were pretreated by complete medium with 0.1、 1、 10、 100 mg/L SalB for 1 hour. And after that they were washed with phosphate buffer for 2 times, added by IMDM with 0.1、1、 10、 100 mg/L SalB and cultured with hypoxia/SD group together in the same condition of hypoxia/SD for 6hours. The control group was cultured for 6 hours in the condition of aerobic and enough serum. Apoptosis was detected by Hoechst33342 staining with inverted phase contrast, fluorescence microscope and Annexin V/PI dual-color flow cytometry. Results Significant apoptosis of BMSCs was induced by hypoxia/SD in the vitro.The early apoptosis of BMSCs induced by hypoxia/SD was significantly decreased by SalB of 0.1、 1、 10 mg/L(P＜0.05) . Conclusion0.1、 1、 10 mg/L SalB can decrease the early apoptosis of BMSCs induced by hypoxia/SD.

Objective To investigate the effects of signal pathway inhibitors PD98059 and LY294002 on cell proliferation, apoptosis, expressions of phosphorylated extracellular signal-regulared kinase (p-ERK) and phosphorylated protein kinase B ( p-Akt) in endometrial carcinoma xenografts. Methods Human endometrial carcinoma Ishikawa cells were cultured in vitro. The effects of PD98059 and LY294002 on proliferation, apoptosis, and cell cycle distribution of endometrial cancer cells were detected by monotetrazolium ( MTT) assay and fluorescence-activated cell sorting technique. The models of xenografted tumor were established by the subcutaneous inoculation in 24 nude mice, and then they were randomly divided into 4 groups ( n = 6) , normal saline group, PD98059 group (PD group) , LY294002 group ( LY group) or PD98059 + LY294002 group ( PD + LY group) by intraperitoneal injections, respectively. The anti-tumor efficacy was evaluated by measuring tumor volume and tumor growth status. The histopathological change of tumor specimens was observed using HE staining and terminal deoxynucleotidyl transferasemediated dUTP-digoxigen in nick and labeling method (TUNEL) testing and the expression levels of p-ERK and p-Akt were detected by immunohistochemistry method. Results ( 1) The proliferation of Ishikawa cells were suppressed after treated by PD98059 and ( or) Y294002, in which A570 values of cells decreased showing both time-dependent and concentration-dependent manner ( LY294002: Fgroup = 9. 801, P = 0. 002; Ftime = 10. 398, P = 0. 001. PD98059: Fgroup= 8. 213, P = 0. 015; Ftime = 6. 839, P = 0. 036). Cell cycle distribution analysis revealed that percentage of Ishikawa cells at G0/G1 phase(Ftime =35.049, P= 0.004; Fgroup = 32. 024, P ＜0. 01) increased and percentage of S phase cells (Ftime = 7. 789, P = 0. 049; Fgroup = 30. 132, P ＜0. 01) decreased significantly. The percentage of apoptotic cells increased significantly among PD group, LY group and PD + LY group, in which there were significant difference [(63. 3 ±0.5)% vs (30. 7 ± 20. 1) % vs(40. 8 ± 1. 3) % ; F = 621. 059, P ＜ 0. 01]. (2) Compared with the control group, the increasing of transplanting tumor volume in the treated groups were obviously ( F = 23. 545 , P ＜ 0. 01) , and the inhibited rate of the tumor was higher in PD + LY group than that in PD group or LY group [(68 ± 9 ) % vs ( 32 ± 16 ) % or ( 38 ± 17 ) % ; F = 10. 283 , P ＜ 0. 05]. ( 3 ) HE staining shown that there were different degrees of necrosis for endometrial carcinoma cell in different groups. The apoptosis of tumor cells were significantly increased in treated groups by TUNEL testing [(13. 7 ± 1. 5)% , ( 14. 1 ± 1. 2)% , (29. 0 ± 1. 8 ) % ; F = 320. 344, P ＜ 0. 01]. Immunohistochemistry results demonstrated that the expressions of p-ERK and p-Akt in treated groups were lower than that in control group, of which LY + PD group was the lowest one. Conclusion The signal pathway inhibitors PD98059 and LY294002 could inhibit the growth of human endometrial carcinoma in vivo and in vitro, in which may induce cell apoptosis.

Objective To characterize the growth curve of Brucella in Bact/Alert blood culture system which may be helpful to predict the growth of Brucella strain for earlier clinical diagnosis of brucellosis.Methods The epidemiological,clinical and labo-ratory data including growth curve of Brucella were reviewed and analyzed retrospectively for 15 cases of brucellosis as con-firmed by positive serological test and brucellosis agglutination test.Results All the 15 patients had irregular fever and history of animal exposure,even though their clinical feature and signs varied greatly.The growth curve of Brucella in blood culture showed the same chracteristics in the 15 patients,including:the time to positive alarm was about 72 hours,a longer lag phase, shorter logarithmic phase,a shorter vertical axis corresponding to the logarithmic phase,and a flat stable phase.Conclusions The clinical manifestation of brucellosis is variable and non-specific.Lack of awareness of this disease makes the clinicians mis-diagnose it easily.Therefore,blood culture is critical for clarifying the etiology in febrile patients.The growth curve of bacteria during blood culture is useful for early diagnosis of brucellosis and prevention of laboratory infections.

Objective To observe the expression of cysteine rich-protein 61 (Cyr61) on renal tubular cells,to explore its effects against hypoxic induced kidney injury and the underlying mechanisms.Methods A stably Cyr61 expressed tubular cell line Cyr61-HK2 was established based on HK2 cells and recombinant Cyr61-lentivirus.BrdU incorporation assay was used for cell proliferation.The apoptosis of cells was analyzed by flow cytometry with Annexin V and propidiumiodide staining.Western bloting was used to detect the protein expression of BAD,Akt and ERK.Results (1) Cyr61-HK2 cells displayed more proliferation ability than HK2 cells.(2) Under hypoxia condition,the apoptosis of both HK2 and Cyr61-HK2 cells increased,but the apoptosis of Cyr61-HK2 cells was lesser than HK2 cells.(3) The expression of Cyr61 led to the phosphorylation of BAD,Akt and ERK on 0 h,0.5 h,1 h (all P ＜ 0.05).Conclusion The expression of Cyr61 can promote cell proliferation and dampen cell apoptosis induced by hypoxia,which may be involved in the Akt/ERK signal pathway.

OBJECTIVE To investigate the distribution of clinical commonly encountered pathogens and their drug resistance in our hospital,and provide reference for reasonable choices of the clinical antibiotics.METHODS The K-B method was used to test the sensitivity to antibiotics of 859 strains pathogens isolated from all kinds of infected samples during from Jan to Dec 2006 in our hospital,at the same time the ESBLs of Escherichia coli and Klebsiella and the MRS were detected.RESULTS The more pathogens isolated from our hospital were E.coli,Staphylococcus,Pseudomonas aeruginosa,Klebsiella,Acinetobacter and Enterococcus.The ESBLs isolating rate was 32.8% in E.coli and 29.4% in Klebsiella,and the sensitivity to antibiotics was degraded obviously in those ESBLs producing strains.Imipenem was the most effective antibiotic to Gram-negative bacilli,cefoperazone/sulbactam and piperacillin/tazobactam also had better antibacterial activity.The isolating rate of MRSA and MRCNS was 32.6% and 40.7% in S.aureus and MRCNS.Gram-positive cocci had the best sensitivity to vancomycin.There was no drug-resistant Enterococcus strain to vancomycin being found.CONCLUSIONS We should think highly of the bacterial drug resistance and use antibiotics reasonably.

Objective To investigate the optimal condition for transforming alkanna tinctoria pigment into shikonin. Methods Transformation rate of shikonin served as index. Transformation temperature, time, ratio of 2% NaOH to alkanna tinctoria pigment (v/w) was optimized. Results With ratio of 2% NaOH to alkanna tinctoria pigment being 4.5 mL·mg-1, temperature 35℃ and the reaction time 4 h, the transformation rate reached the highest, and the average transformation rate was 64.86%. Conclusion This method is easy and simple, and suitable for industrialized production.

Objective Cellular response to estradiol is mediated both by estrogen receptor (ER) binding to estrogen response element (ERE) and by non-nuclear actions like activation of signal transduction pathways such as mitogen activated protein kinase (MAPK) pathway. However, the signal transduction of estrogen involving phosphatidylinositol 3-kinase-protein kinase B (PI3K -PKB) is not clear in endometrial carcinoma. Our purpose was to study if PI3K-PKB signaling pathway could be activated rapidly by 17?-E 2 through non-nuclear action and also, whether PI3K inhibitor, LY294002, could inhibit such non-nuclear action of 17?-E 2 in endometrial carcinoma cell line Ishikawa. Methods Levels of phosphorylated PKB(Ser473 site, p-PKB) and total PKB were examined by western blotting in Ishikawa cells after stimulation with 17?-E 2 at 1?10 -6 mol/L for different time periods and at varied doses for 30 min. Optimal time and appropriate dose for 17?-E 2 to activate PKB in Ishikawa cells were observed. Inhibitory effect of LY294002 on activation of PKB induced by 17?-E 2 was also studied. p-PKB/PKB ratio was used to indicate levels of activation of PKB. Results p-PKB/PKB at 15 min (0.533?0.029) was significantly higher than the control (0.361?0.029, P 0.05, 0.05,

Objective To investigate the influence of pertussis toxin(PTX)on G protein-coupled estrogen receptor(GPER)-mediated activation of phosphatidylinositol 3-kinase(PI3K)/protein kinase B (Akt)signaling activated by 17 β-estradiol(17β-E2)in endometrial carcinoma cells.Methods Expressions of GPER protein were detected by immunohistochemical SP method in Ishikawa and HEC-1A cells.Changes of levels of GPER,ERα and ERβ protein and the activation of Akt protein were observed by western blot in the two cells after they were treated by PTX for 30 minutes at different concentrations(0,0.1,0.5,1.0 μg/ml),and then co-stimulated with with 1 × 10-6 mol/L 17β-E2 respectively at different time (Ishikawa 30 minutes,HEC-1A 15 minutes).Results(1)Immunohistochemical SP method showed that GPER was positive stained in cell cytoplasm of Ishikawa and HEC-1A cell.(2)After co-treated with PTX at different concentrations(0,O.1,0.5,1.0 μg/ml)and 10-6 mol/L 17β-E2,in Ishikawa cell,the ratio of pAkt/Akt was 0.74 ±0.54,0.34 ±0.06,0.18 ±0.03,0.07 ±0.15,the gray values of GPER was 0.872 ± 0.490,0.395 ± 0.054,0.145 ± 0.014,0.034 ± 0.008,and with increasing concentration of PTX,the ratio of p-Akt/Akt and the expression of GPER decreased gradually(P ＜ 0.05),which was most obviously when the concentration was 1.0 μg/ml(F =63.729,P =0.0001;F =160.284,P =0.0001);ERα and ERβ protein had no significant change among different groups(P ＞0.05).In HEC-1A cell,the ratio of pAkt/Akt was 0.73 ±0.09,0.26 ±0.14,0.11 ±0.03,0,the Gray values of GPER is 0.927 ±0.134,0.485 ± 0.022,0.194 ± 0.004,0,and with increasing concentration of PTX,the ratio of p-Akt/Akt and the expression of GPER decreased gradually(P ＜ 0.05),which were also completely inhibited when the concentration was 1 μg/ml(F =1039.321,P =0.0001;F =109.646,P =0.0001),ERα protein had no significant differences(P ＞ 0.05)among different groups.ERβ was negatively expressed.Conclusion The results proposed that the activation of PI3K/Akt signaling in Ishikawa and HEC-1A cells could be inhibited after blocking the role of GPER by PTX.

Objective To investigate the impact of preload fasting and meal replacement in patients with metabolic syndrome.Methods A total of 92 subjects with metabolic syndrome were enrolled in the study.They were assigned into the preload fasting group (PFG),the meal replacement group (MRG),and the control group (CG) for 12-weeks intervention.Special dietary with 100 kcal was provided 30 min before each meal in the PFG,and while in the MRG the same dietary was taken just before each meal and the amount of meal was reduced appropriately.The subjects in CG took meals as usual.Body mass index,waist circumference,and insulin resistance were assessed.Satiety situation was investigated by the scale.Results After 12 weeks,improvement were found in fasting insulin(-3.29 mU/L) and waist circumference (-4.04 cm) in the PFG and significant difference was shown compared to the CG (P＜0.05).Satiety index in the PFG was the most significant among the three group.Conclusion Preload fasting is helpful in improving insulin resistance,reducing waist circumference,and enhancing satiety.