Objective To construct 2 recombinant plasmids carrying cassette chromosome recombinase C(ccrC)and ccrAB respectively and introduce the plasmids into methicillin-resistant Staphlococcus aureus(MRSA)strain 81/0432,and to observe the precise excision of Staphylococcal cassette chromosome mec(SCCmec)island from bacterial chromosome.Methods ccrC and ccrAB genes were amplified with chromosomal DNAs isolated from MRSA strains 81/0342 and N31S as PCR templates.We constructed recombinant plasmids pSR5C and pSR2AB by cloning ccrC and ccrAB genes into temperature-sensitive plasmid pYI3,after introducing them into MRSA strains 81/0432 and N315 by electroporation.PCR was performed to identify SCCmec excision from the bacterial chromosome.The transformants were serial passaged for 10 days,and then the drug resistance of these rransformants was detected by replica experiment.Results The fragment length of ccrC gene was 1.9 kb,smaller than the fragment length of ccrAB from N315.The recombinant plasmids of pSR5C and pSR2AB were successfully constructed.After these 2 recombinant plasmids were introduced into MRSA strain 81/0342,type-V SCCmec island was excised from the chromosome and formed a closed circular structure in the bacteria.However,type-Ⅱ SCCmec island could be excised only in N31S strain after the expression of pSR2AB.Replica experiment verified that transformed strains of 0342(pSR2AB),0342(pSR5C),and N315 (pSR2AB)were mostly susceptible to ceftizoxim or tobramycin after the excision of SCCmec island.Conclusion cciC could serve as a recombinase as ccrAB,which could mediate precise excision of SCCmec island from the chromosome of type-V MRSA strain.This study shows that type-V SCCmec island is widely disseminated between Staphylococcus aweus strains in community setting.

Objective To observe the efficacy of modified Tianma?Sanqi Decoction on hypertension patients with anxiety. Methods Totally 64 hypertension patients with anxiety patients were randomly divid?ed into control group (32 cases) and treatment group (32 cases).The control group with standard medica?tions for hypertension and treatment group with addition of Tianma?Sanqi Decoction, and both groups were continuously treated for 8 weeks. The changes of 24 hours ambulatory blood pressure and SAS scores were compared.Results ( 1) After treatment in the control group and treatment group of 24 h mean systolic blood pressure (24 hSBP) ((130.13±3.81)mmHg,(123.56±4.37)mmHg) (1 mmHg=0.133 kPa) were lower than before treatment((145.36±7.17) mmHg,(145.16±6.77) mmHg) and the difference was statisti?cally significant( P<0.01).(2) After treatment in the control group and treatment group 24 h systolic pres?sure standard deviation,diurnal systolic pressure standard deviation and nocturnal systolic pressure standard deviation((12.19±3.21) mmHg,(11.16±2.26) mmHg,(10.49±3.76) mmHg and (10.95±2.35) mmHg, (10.08±2.67) mmHg,(10.12±3.78)mmHg) were lower than before treatment ((14.53±3.65) mmHg, (13.81±4.04) mmHg,(12.47±4.23) mmHg and (16.73±7.90)mmHg,(16.06±4.14)mmHg,(13.65± 4.92)mmHg),and the difference was statistically significant( P<0.01 or P<0.05).After treatment in the treatment group, 24 h systolic pressure standard deviation, nocturnal systolic pressure standard deviation ((10.95±2.35) mmHg,(10.08±2.67) mmHg) were lower than control group((12.19±3.21) mmHg, (11.16±2.26) mmHg) and the difference was statistically significant.( P<0.01).(3) After treatment in the treatment group,SAS scores (36.68±3.88) points were lower than before treatment (58.66±3.13) points and the difference was statistically significant(P<0.01).After treatment the SAS score ((36.68±3.88) points) in the treatment group was lower than control group((51.86+2.62) points).Conclusion The hypertension patients with anxiety treated with Tianma?Sanqi Decoction and benazapril can improve blood pressure variability and the symp?toms of anxiety.

Objective To evaluate the effect of surgical treatment for primary hepatocellular carcinoma(HCC) with bile duct thrombi(BDT).Methods Eleven cases were diagnosed as HCC with BDT by B ultrasound,CT,MRCP or ERCP.Hepatectomy and choledochotomy were performed,resecting the primary tumor and cleaning the BDT completely,followed with TACE.Results No patients died during operation.The survival time of 9 patients who received hepatectomy and BDT clearance ranged from 3 to 38(average 18.5) months,and that of the other two patients received TACE and BDT clearance survived 6 and 11 months respectively.Conclusion Accurate diagnosis and radical hepatectomy with complete BDT clearance are effective treatment for HCC with BDT.

Objective To investigate the effects of sex hormone-binding globulin (SHBG)gene in the apoptosis of human trophoblastic cells.Methods The siRNA specific-targeting SHBG gene was transfected into human trophoblastic cells and they were divided into six groups:trophoblasts without transfection in normal control groups(group Ⅰ);transfect liposome in blank control groups(group Ⅱ);transfect nonspecific siRNA in negative control groups(group Ⅲ);transfect SHBG siRNA-Ⅰ,SHBG siRNA-Ⅱ,SHBG siRNA-Ⅲ respectively in trans-fection group(group Ⅳ,Ⅴ,Ⅵ).Hoechst 33258 dying method was used to detect cell apoptosis.SHBG and Caspase-3 mRNA profiling and the level of SHBG and caspase-3 protein were detected by real-time PCR and Western blot.Results There was no statistical significant difference in the gene expression and protein level of SHBG and caspase-3 in group Ⅰ,Ⅱ and Ⅲ (P >0.05).In Ⅳ,Ⅴ and Ⅵ group,there was no statistical significant difference in the expression level of SHBG and caspase 3 (P >0.05).Compared with group Ⅰ,Ⅱ and Ⅲ,the a-mount of SHBG gene expression decreased obviously,the caspase-3 mRNA and protein level increased obviously and the trophoblast cell ap-optosis increased markedly (P <0.05).Conclusion Through siRNA interference technology can reduce SHBG gene expression in human trophoblastic cells,and it can lead to excessive apoptosis of human trophoblasts cells.

Objective To explore the difference in the regulating effect of electroacupuncture on serum insulin (Ins) and fat consumption between male and female simple obesity rat and the possible mechanism of its action.Methods A rat model of simple obesity was made by induction with sodium glutamate. The rats were randomized into model and electroacupuncture groups (male and female), 10 rats each. A normal group of 10 rats (male and female) was established as a control. Points Quchi, Zhongwan, Guanyuan, Housanli, Fenglong and Sanyinjiao were selected in the electroacupuncture group. Stimulation with a low frequency of 2 Hz was provided after needle insertion. The treatment was given once daily for 28 consecutive days. Serum Ins content, and greater omentum, pericardiac and perirenal fat weights were measured in the male and female groups of rats before and after electroacupuncture intervention. The comparisons were made.Results Serum Ins content, and greater omentum, pericardiac and perirenal fat weights were significantly lower in the electroacupuncture male and female groups of rats than in the model male and female groups of rats (P<0.01) and also in the electroacupuncture male group of rats than in the electroacupuncture female group of rats (P<0.05).Conclusions Electroacupuncture has different degrees of weight-reducing effect in both male and female obesity rats. The reducing effect on serum Ins content, and greater omentum, pericardiac and perirenal fat weights is better in male obesity rats.

OBJECTIVE:To optimize extraction technology of polysaccharides from Stemona tuberosa so as to increase the ex-traction rate and efficiency. METHODS:Anthrone-sulfuric acid method was used for detection. With water and alkaline water (NaOH solution)as the solvent respectively,the effects of decocting method,ultrasound method and microwave method on the ex-traction rate of crude polysaccharide were observed respectively. By carrying out single factor and orthogonal tests,with the extrac-tion rate of crude polysaccharide as the index,the effects of the amount of water,NaOH solution concentration,extraction tempera-ture and extraction time on the extraction of crude polysaccharide by alkaline water ultrasound method were observed to optimize the technology parameters,and verification test was conducted. RESULTS:The alkaline water ultrasound method is superior to oth-er extraction methods,where the optimal technology parameters were as follows as 0.3 mol/L NaOH aqueous solution 30 times as much as the amount of crud drug,ultrasonic processing for 70 min at 50 ℃. The verification tests showed that the average extrac-tion rate of crude polysaccharide was 25.76%(RSD=3.51%,n=6). CONCLUSIONS:The optimal alkaline water ultrasound method can better extract crude polysaccharide from S. tuberosa.

To establish the quality standard for Zhike Xiaoyan Zhitong lotion. Methods: Phellodendri chinensis Cor-tex, Angelicae sinensis Radix and Astragali Radix in the preparation were identified by TLC. The content of berberine hydrochloride, the effective component in Phellodendri chinensis Cortex, was determined by HPLC with the chromatographic conditions as follows: an Agilent 5 TC-C18 column (250 mm × 4. 6 mm, 5 μm) was used, the mobile phase was acetonitrile-0. 05 mol·L-1 potassium dihydro-gen phosphate (30∶70) with the flow rate of 1. 0 ml·min-1 , the detection wavelength was 265 nm, and the column temperature was at 30 ℃. Results:The spots of Phellodendri chinensis Cortex, Angelicae sinensis Radix and Astragali Radix in Zhike Xiaoyan Zhitong lotion on TLC were clear with strong specificity and without any interference from the negative controls. There was a good linear rela-tionship for berberine hydrochloride within the range of 0. 029-3. 628 μg(r=0. 999 9), and the average recovery was 97. 83% (RSD=2. 05%, n=6). Conclusion:The qualitative and quantitative methods are accurate, reliable and repeatable, which can effectively control the inherent quality of Zhike Xiaoyan Zhitong lotion.

Objective: To observe the effects of moxibustion at different times on prostaglandin E2 (PGE2), prostaglandin F2α (PGF2α) and arginine vasopressin (AVP), in the uterine tissues of rats with dysmenorrhea due to cold-dampness retention, and to explore the differences and possible mechanisms of moxibustion at different times in easing pain in dysmenorrhea due to cold-dampness retention. Methods: Forty-three female Wistar rats were randomly divided into a blank control group (n=7), a model group (n=9), a pre-moxibustion group (n=9), an immediate-moxibustion group (n=9) and a pre-moxibustion plus immediate-moxibustion group (n=9). Rat models of primary dysmenorrhea due to cold-dampness retention were established using (0±1) ℃ ice water immersion method combined with injection of estradiol benzoate for 10 d, followed by injection of oxytocin on the 11th day. Rats in the 3 intervention groups received moxibustion to Shenque (CV 8) and Guanyuan (CV 4), 10 min for each acupoint, once a day. Rats in pre-moxibustion group were given mild moxibustion, beginning on the 8th day during modeling, for 3 continuous days; rats in immediate-moxibustion group were given one time mild moxibustion, immediately after injection of oxytocin on the 11th day during modeling; rats in pre-moxibustion plus immediate-moxibustion group were given mild moxibustion, beginning on the 8th day during modeling till immediately after injection of oxytocin on the 11th day during modeling, for 4 continuous days. The level of PGF2α in the rat uterine tissues was measured by enzyme-linked immunosorbent assay (ELISA), and the levels of PGE2 and AVP in rat uterine tissues were measured by radioimmunoassay. Results: Compared with the blank control group, the levels of PGF2α and AVP, the PGF2α/PGE2 ratio in the model group were significantly increased (P<0.01), and the PGE2 level was significantly decreased (P<0.01) in the rat uterine tissues in the model group. Compared with the model group, the writhing latency was significantly prolonged, the writhing number and the total writhing score were all decreased in the pre-moxibustion group, the immediate-moxibustion group and the pre-moxibustion plus immediate-moxibustion group (all P<0.01); the levels of PGF2α and AVP, and the PGF2α/PGE2 ratio were all significantly decreased (P<0.05, P<0.01), and the PGE2 level was significantly increased (P<0.01) in the rat uterine tissues of the 3 treatment groups. Compared with the pre-moxibustion group, the writhing number and the total writhing score were all decreased in the immediate-moxibustion group and the pre-moxibustion plus immediate-moxibustion group (all P<0.01), the writhing latency was significantly prolonged in the pre-moxibustion plus immediate-moxibustion group(P<0.01); the levels of PGF2α and PGF2α/PGE2 ratio were significantly decreased (P<0.05, P<0.01), and the PGE2 level was significantly increased (P<0.01) in rat uterine tissues in the immediate-moxibustion group and the pre-moxibustion plus immediate-moxibustion group. Compared with the immediate-moxibustion group, the writhing latency was significantly prolonged and the writhing number was decreased (all P<0.05), and the total writhing score was decreased (P<0.01) in the pre-moxibustion plus immediate-moxibustion group; the PGF2α level and the PGF2α/PGE2 ratio were significantly decreased (P<0.01), and the PGE2 level was significantly increased (P<0.01) in rat uterine tissues in the pre-moxibustion plus immediate-moxibustion group. Conclusion: Moxibustion at different times all can produce obvious analgesic effects on dysmenorrhea due to cold-dampness retention in rats, and pre-moxibustion plus immediate-moxibustion ranks the top. The mechanism of this analgesic effect may be via the regulation of abnormal PGF2α, PGE2 and AVP levels, to effectively inhibit the spastic contraction of uterine smooth muscle in dysmenorrhea rat, thereby improving the ischemia and hypoxia in uterus.

Aim To investigate the effect of gelsemium alkaloids on chloride channels and cell volume in he-patic carcinoma cells. Methods The time-lapse live cell imaging and whole-cell patch clamp techniques were used respectively to detect the volume changes and currents induced by gelsemium alkaloids in HepG2 cells. Results It was found that the cell volume was decreased by (12. 48 ± 2. 2) % (P<0. 01) when ex-posed to gelsemium alkaloids for 50 min and this phe-nomenon could be inhibited by the chloride channel blocker tamoxifen. It was shown by whole-cell patch clamping that a chloride current could be evoked by extracellular application of gelsemium alkaloids ( 2μmol·L-1 ) . The current was outward-rectified with-out obvious voltage- and time-dependent inactivation. The reversal potential of the current was ( -3. 21 ± 0. 67) mV ,which was close to the equilibrium poten-tial of chloride. The extracellular application of the chloride blockers, tamoxifen and 5-notro-2-(3-phenyl-propylamino)benzoic acid (NPPB), and 47% hyper-tonic solution inhibited the current significantly ( P <0. 01 ) . Conclusion Gelsemium alkaloids could acti-vate chloride channels and induce a volume decrease ( named apoptotic volume decrease, AVD) , and these effect could be inhibited by chloride channel blockers. The results suggest that the chloride channel can be one of the targets of gelsemium alkaloids in their anti-cancer action.

Objective To analyze the efficacy of gefitinib in the first-line and the second-line treatment of advanced non-small cell lung cancer (NSCLC) with different mutations of epidermal growth factor receptor (EGFR). Methods The clinical data of 70 patients with advanced NSCLC harboring different EGFR mutations and taking gefitinib as the first-line or the second-line treatment in Shanxi Provincial Cancer Hospital from January 2013 to December 2014 was analyzed retrospectively. According to the treatment method, the patients were divided into the first-line treatment group (36 cases) and the second-line treatment group (34 cases); according to the type of gene mutations, the patients were divided into exon 19 deletion group (EGFR gene exon 19 LREA deletion, 46 cases) and exon 21 mutation group (exon 21 L858R mutation, 24 cases). Progression-free survival (PFS), effective rate (ORR), and disease control rate (DCR) were observed in each group. Results Of the 70 evaluable patients, the median PFS time in patients with exon 19 LREA deletion was 8.88 months (95％ CI 7.72-10.04), and the median PFS time in patients with exon 21 L858R mutation was 8.67 months (95％ CI 7.17-10.17), the difference between the two groups was not statistically significant (P = 0.959). The ORR in patients with exon 19 LREA deletion was 69.6％ (32/46), and the ORR inpatients with exon 21 L858R mutation was 54.2％ (13/24), the difference was not statistically significant (χ 2= 1.629, P = 0.202). The DCR in patients with exon 19 LREA deletion was 84.8％ (39/46), and the DCR in patients with exon 21 L858R mutation was 91.7％ (22/24), the difference was not statistically significant (χ 2= 0.194, P = 0.659). The median PFS time in patients with the first-line treatment was 9.22 months (95％ CI 7.92-10.52), and the median PFS time in patients with the second-line treatment was 8.37 months (95％ CI 7.08-9.65), the difference was not statistically significant (P = 0.507). The ORR in patients with the first-line treatment was 63.9％ (23/36), and the ORR in patients with the second-line treatment was 58.8％ (20/34), the difference was not statistically significant (χ 2 = 1.460, P = 0.227). The DCR in patients with the first-line treatment was 88.9％ (32/36), and the DCR in patients with the second-line treatment was 88.2％ (30/34), the difference was not statistical significant (χ 2 = 0.060, P = 0.940). Conclusion The short-term efficacy and PFS are similar between NSCLC patients with different mutations of EGFR or with the first-line and the second-line treatment with gefitinib.