AIM: To optimize the process of extracting effective constituents from Radix Polygalae by central composite design/response surface methodology. METHODS: Independent variables were ethanol concentration,reflux time and solvent fold,dependent variable was extraction rate of senegenin in Radix Polygalae.Linear or nolinear mathematic models were used to estimate the relationship between independent and dependent variables.Response surface methodology was used to optimize the process of extraction.Prediction was carried out through comparing the observed and predicted values. RESULTS: Regression coefficients of binomial fitting complex model was as high as 0.979 0, the optimum conditions of extraction process were 60% ethanol,2.5 hours for reflux,10-fold solvent and 2 times for extraction.Bias between observed and predicted values was-5.93%. CONCLUSION: It shows that the optimum model is highly predictive.

Objective To explore the relation ship between the high-risk human papillomavirus(HPV) and prognosis of ovarian carcinomas.Methods In situ hybridization technique was used to examine HPV16/18 DNA in 32 cases of benign ovarian epithelial tumors,25 cases of borderline tumors and 45 cases of ovarian carcinoma.Immunohistochemistry was used to examine the expression of VEGF and ki-67 protein,which compared to usual ovarian tissues. Results (1)The positive rate of HPV16/18 was significantly different between ovarian carcinoma and normal epithelial ovarian tissues or benign epithelial ovarian tumors(P

To summarize the experiences of optimization of emergency medical care in Nanjing General Hospital.The principles and methods of optimizing emergency medical care were analyzed and summarized.The following methods were used to improve the emergency medical care: Applying modern conceptions of emergency medicine and establishing professional physician teams;Reinforcing the duty of each position and improving the quality of medical care;Modifying the mode and improving the working efficiency;Establishing clinical pathways for disasters and other accidents.The roles of emergency medicine are changed and modernized. Optimization of emergency care and popularization of emergency care skills may improve the quality and efficiency of emergency medical care.

OBJECTIVE To analyze the distribution,risk factors and preventive measures of nosocomial infection in patients with hemodialysis. METHODS Clinical data of 112 patients with hemodialysis were retrospectively reviewed.Statistic analysis was made on relation between the occurrence of nosocomial infection and patients′ age,adequacy of dialysis,dialysis duration,anemia,heart function,serum albumin,catheterization and reused dialyzer. RESULTS 67 infection cases were found.The main infection sites were blood vessel access,respiratory tract,and urinary tract.The infection rates increased significantly in the groups of age 60,inadequate dialysis,dialysis duration more than 1 year,serum albumin,heart failure and hypoalbuminemia compared with the correspondent controly group(P 0.05). CONCLUSIONS Patients with hemodialysis have higher infection rates,the main risk factors are old age,inadequate dialysis,long dialysis duration,severe anemia,heart failure,catheterization and hypoalbuminemia.Therefore the effective measures to reduce the nosocomial infection are strictly aseptic technology,adequate dialyzsis,reducing the invasive operation,ameliorating anemia and improving the nutrition.

Objective To explore the effects of TNF-α on the expression of IP3 R1 mRNA and protein in human mesangial cells (HMCs) and elucidate the role of protein kinase C (PKC) in this signal pathway.Methods Quantitative real-time polymerase chain reaction and immunoblot assay were used to examine the effects of TNF-α on IP3R1 mRNA and protein expression.Depletion PKC,the selective inhibitor of PKCα Safingol and inhibitor of PKCδ Rottlerin,overexpression of dominant negative mutant of PKC to examine the mechanism of signal transduction of TNF-α-regulated IP3 R1 in HMCs.PKCα activation was assayed by Western blot.Results TNF-α increased IP3R1 mRNA and protein expression in HMCs,effects that were blocked by prolonged incubted chronic PMA,Safingol and also by domain negative PKCα construct.TNF-α promoted PKCα activation with maximal PKCα phosphorylation that occurred 8 h post-stimulation.Conclusion TNF-α increased the expression of IP3 R1 and this was mediated through the PKCα activation signaling pathways in HMCs.

Objective To explore the effects of TNF-α on the expression of IP33R1 mRNA and protein in human mesangial cells (HMCs) and elucidate the mechnism of TNF-α indnces the IP3R1 expression in the occurrence of hepatorenal syndrome (HRS).Methods HMCs was stimulated by tumor (TNF-α) with 100 ng/mL for different hours (2,4,8,24 hours).The expression change of IP3R1 mRNA and protein were detected by quantitative real-time polymerase chain reaction and immunoblot assay.Several inhibitors including D609,U73122,PP1,Safingol,Rottlerin and non-radioactive PKC assay to examine the mechanism of signal transduction of TNF-α-regulated IP3R1 in HMCs.Results The levels of IP3R1 mRNA at 2 h post-TNF-α exposure were significantly enhanced and reached peak at 8 h in HMCs (P ＜ 0.01),then descened at 24 h (P ＜ 0.01).The levels of IP3R1 protein at 4 h post-TNF-α exposure were obviously increased and reached peak at 24 h post-TNF-α exposure (P ＜ 0.01).Compared with the control group,safingol (PKC-α inhibitor) and D609 (PC-PLC inhibitor) each significantly suppressed TNF-α-induced expression of IP3R1 mRNA (3.30 ± 0.81) vs.(1.95 ± 0.130,P ＜ 0.05 ; (2.10 ± 0.49),P ＜ 0.01 andIP3R1 protein (3.09±0.13) vs.(1.86+0.39),P＜0.01; (1.98±0.02),P＜0.01.TNF-αpromoted autophosphorylation,and hence the activation,of PKC-α with maximal phosphorylation that occurred 8 h post-stimulation measured by non-radioactive PKC assay,and the effect was marked attenuated by pretreated with D609 or safingol.Conclusions TNF-α increased the expression of IP3R1 and this was mediated,at least in part,through the PC-PLC/PKC-α signaling pathways in HMCs.

Objective To investigate the effects of specific protein 1 ( Sp1 ) on the TNF-αin-duced expression of inositol 1, 4, 5 trisphosphate receptor type 1 ( IP3R1 ) in human mesangial cells ( HMCs) and to further elucidate the molecular mechanism regarding the decreased glomerular filtration rate ( GFR ) during hepatorenal syndrome .Methods Quantitative real-time polymerase chain reaction and Western blot assay were used to analyze the effects of TNF-αon the expression of IP3R1 at mRNA level and the expression of IP3R1 and Sp1 at protein level in HMCs , respectively.HMCs were transfected with a re-combinant plasmid PGL3-IP3R1 promoter to determine the effects of TNF-αon the activity of IP3R1 promot-er.HMCs were treated with Mithramycin A , an inhibitor of Sp1 binding, and transfected with Sp1-siRNA plasmid respectively to evaluate the expression of IP 3R1 regulated by TNF-α.The role of TNFR1 and TNFR2 in the TNF-αinduced expression of Sp 1 and IP3R1 proteins were detected by Western blot .Results TNF-αincreased the expression of IP3R1 at mRNA level and the expression of IP3R1 and Sp1 at protein lev-el in HMCs.Moreover, the activity of IP3R1 promoter in HMCs was remarkably increased by TNF-αas well.TNF-αinduced expression of IP3R1 was inhibited by Mithramycin A in a concentration dependent manner.HMCs transfected with Sp1-siRNA plasmid showed a significantly decreased expression of IP 3R1 protein.Both anti-TNFR1 and anti-TNFR2 antibodies blocked the TNF-αinduced IP3R1 expression, while only anti-TNFR1 antibodies inhibited the TNF-αinduced Sp1 expression.Conclusion TNF-αmight in-crease the expression of IP3R1 through the TNFR1/Sp1 signaling pathways in HMCs .

Objective To investigate the relationship between renin (REN) gene G10631A, angiotensinogen (AGT) gene T704C mononucleotide polymorphisms and cerebral infarction and to investigate the mechanisms and characteristics of cerebral infarction from molecular level. Methods REN gene G1063A and AGT gene T704C polymorphisms in 82 patients with cerebral infarction and 89 controls were detected with polymerase chain reactionrestriction fragment length polymorphism. The differences of the genotypes and allele frequencies were compared between the patient group and the control group. Results The frequency of REN 10631AA genotype (31. 7% vs. 10. 1%,χ2 =12. 816, P = 0. 002) and the frequency of A genotype (49. 4% vs. 30. 3% χ2 = 12. 969, P =0. 000), as well as the frequency of AGT 704 CC genotype (63. 4% vs. 34. 8% χ2 = 15. 029, P = 0. 001) and the frequency of A genotype (79. 9% vs. 61. 2% χ2 = 14. 173, P = 0. 000) in the cerebral infarction group were all significantly higher than those in the control group; the frequency of haplotype 704C 10631A was also significantly higher than that in the control group (P=0. 000). Conclusions REN 10631AA genetype and A allele as well as AGT 704 CC genetype and C allele may be the susceptible factors of cerebral infarction. Haplotype 704C-10631 A may be a genetic risk factor for the occurrence of cerebral infarction.

Background Tumstatin is the most active endogenous angiogenesis inhibitor,which has a marked inhibitory effect on pathological neovascularization,and Tum5 is an angiogenesis inhibitors fragment of fulllength tumstatin.Objective This study was to investigate the effects of adenovirus-mediated overexpression of recombinant Tum5 gene on the proliferation,migration and tube formation of human umbilical vein endothelial cells (HUVECs) in physiological status.Methods The empty adenoviral vector expressing green fluorescent protein (rAd-GFP) and the viral vector expressing recombinant Tum5 gene were constructed.The HUVECs cultured in RPMI1640 medium were divided into normal control group,empty vector group (rAd-GFP group) and Tum5 gene infection group (rAd-GFP-Tum5 group).The rAd-GFP and rAd-GFP-Tum5 adenoviral particles at the density of 1 × 1010/ml were added into the medium to infect the cells for 48 hours.The proliferation of the cells was assayed at 24,48 and 72 hours by cell counting kit-8 (CCK-8) to evaluate the proliferative rate;the migration number of the cells was detected at 48 hours after infection by Transwell chamber;the tube formation number of the cells were detected by Matrigel method.The concentration of vascular endothelial growth factor (VEGF) in cell supernatants was assayed by ELISA at 24,48,and 72 hours following adenoviral infection.Results The cultured cells showed green fluorescence in the rAd-GFP group and rAd-GFP-Tum5 group under the inverted fluorescence microscope,and the infection efficiency of rAd-GFP and rAd-GFP-Tum5 was 55.13％ and 50.31％,respectively.No significant difference was found in cell proliferative rate among normal control group,rAd-GFP group and rAd-GFP-Tum5 group both at 24 and 48 hours after infection (both at P＞0.05),and the cell proliferative rate was significantly lower in the rAd-GFP-Tum5 group than that in the normal control group and rAd-GFP group at 72 hours after infection (both at P＜0.01).The migration number of the cells at 48 hours after infection was 2 260.25-±930.44,2 370.00±441.06 and 723.75± 363.80 in the normal control group,rAd-GFP group and rAd-GFP-Tum5 group,showing a significant difference among the groups (F =8.524,P =0.008),and the migrated cells were evidently decreased in the rAd-GFP-Tum5 group compared with the rAd-GFP group and the normal control group (both at P＜ 0.01).The tube number at 48 hours after infection was 95.67±5.86,88.00±4.58 and 20.67±3.51 in the normal control group,rAd-GFP group and rAdGFP-Tum5 group,showing a significant difference among the groups (F=226.498,P＜0.01),and the tube number in the rAd-GFP-Tum5 group was significantly reduced in comparison with the normal control group and rAd-GFP group (both at P＜ 0.01).The considerably differences in VEGF concentration in the cell supernatants were found in different groups and various time points (Fgroup =73.260,P＜0.01;Ftime =73.477,P＜0.01),and VEGF concentration in the cell supernatants was significantly decreased in the rAd-GFP-Tum5 group compared with the rAd-GFP group at both 48 hours and 72 hours (both at P＜0.01).Conclusions The overexpression of the recombinant Tum5 can inhibit the proliferation,migration and tube formation of the HUVECs in physiological status,which may be associated with Tum-5-mediated down-regulation of VEGF protein in the cell supernatant.

A rapid method for the simultaneous determination of daidzein, genistein and formonetin in solanum Lyratum Thunb by high performance liquid chromatography (HPLC) was developed. Separation was achieved on a Diamonsil C18 column (250 mm×4.6 mm, 5 μm) with isocratic elution, using a mobile phase of methanol-tetrahydrofuran-water (44∶3∶53, v/v). The wavelength was set at 260 nm and column was maintained at 35 ℃. The linear ranges of daidzein, genistein and formonetin were 1.0-40.0, 0.1-4.0 and 0.1-4.0 μg/mL, respectively. The average recoveries were between 98.4% and 101.3%. This method could be used for the quality control of Solanum lyratum Thunb due to its simplification, reliability, rapidity and excellent precision.