Objective To analyse the mutation types of the NOTCH3 gene with cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy(CADASIL). Methods Including 4 probands of CADASIL,10 cases of CADASIL and 4 normal members from 3 CADASIL families and 100 healthy controls were recruited. Genomic DNA was extracted from white blood cell. The amplicons were analyzed by the denaturing high-performance liquid chromatography (DHPLC) technique. The positive samples which identified by the DHPLC were sequenced to determine the specific mutation or polymorphism, respectively. Results Three heterozygous missense mutations including Cys90Arg, Arg141Cys, Cys134Tyr located in the exon3 and exon4 were found in the 4 probands and 10 cases of CADASIL among the 3 families. 15 polymorphisms were also found. 2 members individual from family 1 and 2 were found to carry the same pathological mutations as in their proband but without clinical symptoms. They were identified as preclinical patients. Conclusions Mutation detection of NOTCH3 is the molecular genetic mechanical for CADASIL. The exon3 and exon4 are possible hot mutation spots in Chinese patients. The mutation of Cys134Tyr in exon4 is a novel mutation which has not been reported previously in China.

Objective:To perfect and improve the quality standard of Malloti Apeltae Radix. Methods:Microscopic identification was used to identify the transverse section and the powder. The water-soluble and the fat-soluble components were identified by TLC. Rutin was used as the reference in the characteristic chromatogram established by HPLC to determine the relative retention time and rel-ative peak area of each characteristic absorption band. A SPOLAR C18 chromatographic column ( 250 mm × 4. 6 mm,5μm) was used with acetonitrile-0. 4% phosphate solution as the mobile phase with gradient elution, the detection wavelength was set at 328 nm, the flow rate was 1. 0 ml·min-1, and the temperature was 35℃. Results: The microscopic characteristics were obvious. The spots of TLC were round and clear with good repeatability. Five characteristic absorption bands were shown in the fingerprints with the relative retention time within ± 5% of reference value of rutin, and the relative area of peak 3 and peak 4 was not lower than 0. 23 and 0. 66, respectively. Conclusion:The method is convenient, fast and repeatable, and the result is accurate and reliable, which can be used to control the quality of Malloti Apeltae Radix effectively, and as the main index of the quality standard.

To observe the effects of Fuhuang Shengji Yuchuang (FHSJYC) Ointment, a compound traditional Chinese herbal medicine, on the expressions of types I and III collagens in granulation tissue of wound in rats with diabetes.

Objective To explore the action mechanism of Yiqi Huayu Formula for postpartum subinvolution of uterus. Methods Fifty female SD rats were selected model with postpartum subinvolution of uterus was made by injecting escherichia coli in vagina. Rats were randomly divided into 5 groups according to pregnancy time:blank group, model group, Xinshenghua group, and Yiqi Huayu high and low dose groups. Blank group and model group received gavage with water;Yiqi Huayu low and high dose groups received gavage with Yiqi Huayu Formula;Xinshenghua group received gavage with Xinshenghua Granule, once a day for 7 days. Uterine endometrial morphology was observed by HE staining. The contents of NO and NOS were respectively detected by Nitrate reductase method. Results Uterus submucosal hyperemia, bleeding, and endometrial discontinuity with the infiltration of inflammatory cells of rats in the model group suggested that the models were successfully established. Compared with blank group, the contents of NO and NOS in model group were higher significantly. Compared with model group, the contents of NO and NOS significantly decreased in both Xinshenghua group and Yiqi Huayu high and low groups (P<0.01, P<0.01). Compared with Xinshenghua group, Yiqi Huayu high dose group was lower (P<0.05). Conclusion Yiqi Huayu Formula could promote reconstruction and repair of the endometrium by reducing the contents of NO and NOS and inhibiting abnormal uterine spiral arteries relaxation.

Objective To evaluate the effect of propofol preconditioning on endoplasmic reticulum stress induced by hypoxia-reoxygenation (H/R) in HEPG2 cells.Methods HEPG2 cells were randomly divided into 4 groups using a random number table:control group (group C),propofol group (group P),H/R group and H/R + propofol preconditioning group (group PP).In group C,the cells were cultured routinely for 42 h.In group H/R,after being cultured routinely for 6 h,the cells were exposed to 1％ O2 + 5％ CO2 + 94％ N2 for 12 h followed by 12 h reoxygenation.In group PP,the cells were cultured for 6 h in the culture medium containing propofol 10 μmol/L (final concentration),and then H/R was induced.The cell viability was detected by MTT assay.The expression of immunoglobulin heavy chain-binding protein (BIP),C/EBP homologous protein (CHOP) and activated caspase-3 was determined by Western blot.The expression of BIP,CHOP and caspase-3 mRNA was determined by RT-PCR.Results Compared with group C,the cell viability was significantly decreased,and the expression of BIP,CHOP and activated caspase-3 protein and mRNA was up-regnlated in H/R and PP groups,and no significant changes were found in the parameters mentioned above in group P.Compared with group H/R,the cell viability was significantly increased,and the expression of BIP,CHOP and activated caspase-3 protein and mRNA was down-regulated in PP group.Conclusion Propofol preconditioning can promote the cell proliferation and attenuate H/R injury to HEPG2 cells through inhibiting endoplasmic reticulum stress.

0.05).The prevalence of strains with resistance to rifampin was lower than that of strains with resistance to clarithromycin(P

ObjectiveTo investigate the popularity of the sedated gastrointestinal endoscopy in mainland China.MethodsThe survey on the popularity of the sedated gastrointestinal endoscopy was conducted with questionnaires sent by e-mails or telephones to the GI endoscopy units of three levels of hospitals.ResultsFrom May,2010 to November,2010,169 units from 28 provinces (90.3％ ) were enrolled,including 46 hospitals of grade Ⅲ,91 of grade Ⅱ and 32 of grade Ⅰ.Sedated gastrointestinal endoscopy were performed in 110 (65.1％,110/169) hospitals,i.e.93.5％ (43/46) grade Ⅲ hospitals,68.1％(62/91) grade Ⅱ hospitals and 15.6％ (5/32) grade Ⅰ hospitals.Significant difference was observed between the grade Ⅲ and the grade Ⅱ hospitals (P ＜ 0.05 ),so was between the grade Ⅱ and the grade Ⅰ hospitals ( P ＜ 0.05 ).Propofol was frequently used in sedation in 98 hospitals ( 89.1％ ).The only 9.1％( 10/110)hospitals,which had full-time anesthetists or anesthetic nurses,were all grade Ⅲ hospitals.ConclusionThere has been great progress in the popularity of sedated gastrointestinal endoscopy in mainland China during the past decade.However,it is less frequently pefformed in grade Ⅰ and Ⅱ hospitals.Shortage of full-time anesthetists is still a problem.

Aim To investigate the protective effect of pituitary adenylate cyclase activating polypeptide(PACAP)on focal cerebral ischemia reperfusion injury in rats.Methods The models of focal cerebral ischemia reperfusion in rats were established with suture-occluded method to simulate the middle cerebral artery ischemia reperfusion injury in clinic.The treatment groups were given three different dose PACAP(PACAP 10-11 mol,PACAP 10-10 mol and PACAP 10-9 mol) in 15 minutes before cerebral ischemia.PACAPs were infused into lateral cerebral ventricle by microinjector.The water content of brain,the activity of superoxide dismutase(SOD),the content of malondialdehyde(MDA) and the content of nitrogen monoxidum(NO) of the brain tissue were determined.Results Compared with NS group,the water content of brain and the concentration of MDA and NO were obviously lower in each PACAP treatment group,while the activity of SOD was elevated to different degrees.Conclusion PACAP has an obvious protective effect on ischemia-reperfusion injury of focal brain tissue in rats,in which edema-alleviating,free radicals-scavenging and lipid peroxidation-attenuating could be involved.The protective effect seems to be more clear in middle-and high-dose group than in low-dose group.

Objective To observe the effect of Tongqiao Huoxue decoction on the changes of motilin (MTL) and vasoactive intestinal peptide (VIP) and its protective effect on gastric mucosa in rats with cerebral infarction at acute stage.Methods According to random number table, 96 Wistar rats were divided into six groups: normal control, sham operation, cerebral infarction model, cimetidine, Tongqiao Huoxue decoction routine dose and low dose groups, eachn = 16. Cerebral infarction model group was established by using a suture to ligate the middle cerebral artery, resulting in its occlusion. The rats of cerebral infarction model group and sham operation group were lavaged with 2 mL of normal saline. Tongqiao Huoxue decoction routine dose group was lavaged with 2 mL of Tongqiao Huoxue decoction (1.3 g/mL), while Tongqiao Huoxue decoction low dose group was lavaged with 1 mL of Tongqiao Huoxue decoction (1.3 g/mL) plus 1 mL of normal saline. Cimetidine group was lavaged with cimetidine (0.1 g/kg), and nothing was done for the normal control group. On the 4th and 7th day after the delivery of treatment, inferior vena venous blood samples were collected to detect the plasma MTL and VIP concentrations. Then the gastric tissues were obtained and examined under an electron microscope.Results There were no statistically significant differences between the normal control group and sham operation group in plasma levels of MTL and VIP (bothP > 0.05). The plasma levels of MTL and VIP at 4 days and 7 days after the delivery of treatment in cerebral infarction model group were higher than those in the normal control group and sham operation group [4 days MTL (ng/L): 189.51±13.48 vs. 117.01±11.38, 117.67±12.73, VIP (ng/L): 94.86±4.14 vs. 60.98±5.42, 62.55±6.60, bothP < 0.05; 7 days: MTL (ng/L): 183.05±14.49 vs. 119.79±10.64, 120.27±11.48, VIP (ng/L): 91.92±3.77 vs. 59.63±4.29, 58.10±4.99, bothP < 0.05]. Plasma levels of MTL and VIP in cimetidine group, Tongqiao Huoxue decoction routine dose group and low dose group were significantly lower than those in cerebral infarction model group. Compared with Tongqiao Huoxue decoction low dose group and cimetidine group, the degrees of descent in plasma levels of MTL and VIP were apparently lower on the 7th day in Tongqiao Huoxue decoction routine dose group [MTL (ng/L): 138.72±8.02 vs. 152.16±12.66, 156.60±11.87, bothP < 0.05; VIP (ng/L):68.18±3.41 vs. 81.12±3.98, 78.89±2.13, bothP < 0.05]. The results of electron microscope: compared with normal control group and sham operation group, obvious pathological changes in gastric mucosa in model group were found. In Tongqiao Huoxue decoction routine group, Tongqiao Huoxue decoction low dose group and cimetidine group, the gastric mucosal lesions were milder, and the curative effects of Tongqiao Huoxue decoction routine group were much better.Conclusions During the acute stage of cerebral infarction, significant damage is seen in gastric mucosa in rats. Tongqiao Huoxue decoction not only can effectively regulate the expression levels of MTL and VIP, but also has certain protective effect on gastric mucosa, and its efficacy being directly proportional to the dosage.

We established methods to isolate human amniotic fluid-derived progenitor cells (hAFPCs), and analyze the ability of hAFPCs to secrete human coagulation factor IX (hFIX) after gene modification. The hAFPCs were manually isolated by selection for attachment to gelatin coated culture dish. hFIX cDNA was transfected into hAPFCs by using a lentiviral vector. The hFIX protein concentration and activity produced from hAFPCs were determined by enzyme-linked immunosorbent assay (ELISA) and clotting assay. The isolated spindle-shaped cells showed fibroblastoid morphology after three culture passages. The doubling time in culture was 39.05 hours. Immunocytochemistry staining of the fibroblast-like cells from amniotic fluid detected expression of stem cell markers such as SSEA4 and TRA1-60. Quantitative PCR analysis demonstrated the expression of NANOG, OCT4 and SOX2 mRNAs. Transfected hAFPCs could produce and secrete hFIX into the culture medium. The observed concentration of secreted hFIX was 20.37% +/- 2.77% two days after passage, with clotting activity of 16.42% +/- 1.78%. The amount of hFIX:Ag reached a plateau of 50.35% +/- 5.42%, with clotting activity 45.34% +/- 4.67%. In conclusion, this study established method to isolate and culture amniotic fluid progenitor cells. Transfected hAFPCs can produce hFIX at stable levels in vitro, and clotting activity increases with higher hFIX concentration. Genetically engineered hAFPC are a potential method for prenatal treatment of hemophilia B.
Amniotic Fluid ; cytology ; Blood Coagulation ; Cell Culture Techniques ; Cell Separation ; methods ; DNA, Complementary ; Factor IX ; biosynthesis ; Genetic Engineering ; Genetic Vectors ; Humans ; Stem Cells ; cytology ; metabolism ; Transfection