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BACKGROUND:Estrogen/progestins replacement therapy prevents excess bone loss in postmenopausal women.Recently osteoprotegerin (OPG) has been identified in osteoblast and displayed to inhibit bone resorption.OBJECTIVE: To compare the action between 17β-estradiol (E2) and progesterone on OPG expression in cultured normal human osteoblast-like cells (hOB).DESIGN: A comparative investigation.SETTING: Institute of Metabolic Endocrinology, the Second Xiangya Hospital of Central South University.MATERIALS: α-MEM (Sigma Chemical Corp., St. Louis, MO, USA); Type Ⅳ collagenase (Sigma); Fetal bovine serum (Gibco-BRL Corp., Grand Island, NY, USA); Osteocalcin radioimmunoassay kit (DiaSorin Corp., Stillwater, MN, USA).METHODS: The experiments were carried out in the Institute of Metabolic Endocrinology, Second Xiangya Hospital of Central South University from January 2003 to March 2006. The osteoblasts were extracted from the cancelous bone of anterior superior iliac spine of normal people, then cultured. The hOB were treated with E2 and progesterone, and the expressions of OPG mRNA and OPG protein were determined by Northern blot analysis and enzyme-linked immunoabsorbent assay (ELISA) respectively.MAIN OUTCOME MEASURES: ①Characterization of human osteoblast-like cells; ②Effect of E2 and progesterone on OPG mRNA levels by Northern blot analysis; ③ Effect of E2 and progesterone on OPG protein levels in the conditioned medium by ELISA.RESULTS: ① Characterization of hOB in vitro The ALP levels in normal human osteoblasts were (74.3±4.7) U/g protein,and the detectable osteocalcin levels was (3.84±0.39) μg/L protein], which suggested that osteoblasts were the primary cell type found in our bone-derived cell cultures from donors. ② Effects of E2 and progesterone on the levels of OPG mRNA by Northern blot analysis: The OPG mRNA band was week in the control group [(12.3±3.5)%], treatment with 1 × 10-10, 1 ×10-9 1 ×10-8 mol/L E2 caused an increase in the levels of OPG mRNA. The expression of OPG mRNA in the 1×10-8 mol/L E2 group was gradually increased at 12, 24 and 48 hours. Progesterone had no influence on OPG mRNA expression. ③ Effects of E2 and progesterone on OPG protein production in conditioned medium determined with ELISA:ELISA revealed that treatment with 1 ×10-10, 1 ×10-9, 1 ×10-8 mol/L E2 induced obvious increase in the levels of OPG protein in cells media as compared with that in the control group [(1.27±0.26), (2.34±0.35), (3.62±0.23), (0.64±0.14)μg/L, P ＜ 0.01]. In the presence of 1×10-8 mol/L E2, OPG protein production in cells media at 12, 24 and 48 hours were significantly higher than that in the control group [(1.30±0.30), (3.07±0.14), (3.50±0.33), (0.62±0.12) μg/L, P ＜ 0.01]. 1 × 10-10, 1 ×10-9 1 × 10-8 mol/L progesterone had no influence on the OPG protein production after 12-24 hours (P ＞ 0.05).CONCLUSION: The different regulation of OPG production in osteoblasts by E2 and progesterone may contribute to the mechanisms by which estrogen or progestins exerts its different action on bone resorption.

Aim To investigate the induction of endothelial cell apoptosis and the suppression of VEGF expression in cancer cells by sodium caffeate (SCA). Methods Apoptosis of transformed human umbilical vein endothelial cells (ECV304 cell line) was detected by flow cytometry, DNA electrophoresis assay and morphological assessment. Western blotting analysis was applied for determination of VEGF expression in cancer cells. Substrate degradation by type Ⅳ collagenase was measured by zymography.ELISA was used to detect the binding of type Ⅳ collagenase with relevant monoclonal antibody. Results SCA induced ECV304 cell apoptosis in a time- and dose-dependent manner. After treatment with 100 and fluorescence and distinct changes of nuclear morphology, such as pyknosis and the occurrence of apoptotic bodies. VEGF expression in hepatoma HepG-2 cells and prostate carcinoma DU145 cells was reduced after SCA treatment. The degradation activity of type Ⅳ collagenase including MMP-2 and MMP-9 secreted by giant cell pulmonary carcinoma PG cells was inhibited by SCA in a dose-dependent manner. SCA also reduced the binding of mAb 3D6, a relevant monoclonal antibody, to type Ⅳ collagenase. Conclusion SCA can induce endothelial cell apoptosis and inhibit VEGF expression as well as type Ⅳ collagenase activity in cancer cells. SCA might be active in modulating tumor angiogenesis and the microenvironment.

BACKGROUND:ABO-incompatibility between donor and recipient is not a barrier for Successful allogeneic hematopoietic stem cell transplantation even though it is well established that major ABO incompatibility may lead to prolonged destruction of donor-derived erythrocytos and prolonged transfusioil requirements.OBJECTIVE:To explore the effect of ABO.incompatible on clinical characteristics in allogeneic-hematopoietic stem cell transplantation.DESIGN:A retrospective observation.SETTING:Department of Hematology.the Affiliated Drum Tower Hospital of Nanjing University Medical School.PARTICIPANTS:Fourteen patients(11 males and 3 feiliales,aged 15-60 years old)with malignant hematologic diseases who received ABO-incompatible allogeneic hematopoietic stem cell transplantation in the Affiliated Drum Tower Hospital of Nanjing University Medical School from May 2002 to September 2007 Were recruited for this study.Of the 14 patients,7 were human leukocyte antigen(HLA).matched,and the other 7 were HLA-half-matched.Controls were 11 patients who received ABO-compatibility bone marrow transplantation during the same period.Written informed consents for receiving allogeneic hematopoietic stem cell transplantation were obtained from each reciplent.The donors were sibling sister,sibling brother.son and mother,and they all agreed to provide marrow for transplantation.T1lis experiment was given an approval by the Ethics Committees of the hospital.METHODS:Regimen conditioning:HLA-matched transplantation regimen conditioning consisted of busulfan(Bu)and cyclophosphamide(Cy).HLA-half-matched transplantation regimen conditioning adopted GIAC program from Beijing People's Hospital.The GIAC program consisted of 4 parts:G:granulocyte colony-stimulating factors used for donors;I:stronger immunosuppressive regimen conditioning used for recipients;A: antihuman thymocyte globulin added:C: combined transplantation of bone marrow and peripheral blood;Perfusion of hematopoietic stem cells:The marrow from ABO-incompatible donor depleted erythrocytes by hydroxyethyl starch sedimentation.MAIN OUTCOME MEASURES:Adverse reaction.complication and hematologic recovery after ABO-incompatibility stem cell transplantation.RESULTS:One out of fourteen recipients developed pure red cell aplasia(PRCA)and dropped out of final analysis.Hematologic recovery:The median time of erythrocyte recovery after ABO-incompatible stem cell transplantation was delayed compared with ABO compatible stem cell transplantation (t=2.352.P＜0.05).There were no significant difieFences in the recovery of neutrophils and platelets between ABO-incompatible group and ABO-compatible group(P＞0.05).The median time of recovery of the erythrocyte and the blood type switching was delayed in HLA-mis-matched allogeneic hematopoietic stem cell transplantation compared with HLA-matched allogeneic hematopoietic stem cell transplantation,but without significant difference(P＞0.05).Complications:During the stem cell transfusion following transplantation.none of 14 Patieats had hemolytic complications or delayed haemolysis.CONCLUSION:There was no evidence of ABO-incompatibility between donor and recipient is a barrier for successful allogeneic hematopoietic stem cell transplantation.

Objective To evaluate the feasibility, methods and effectiveness of using a mechani-cal stapler for choledochojejunostomy.Methods The authors have operated on 118 patients in the management of carcinoma of head of pancreas, or periampullary tumor, or cholelithiasis.In the opera-tion, the bilio-enteric end-to-side, or end-to-end and side-to-side anastomosis was made by a circular stapler device, and then a Roux-en-Y or Brown's loop was formed for the preeedure.Results All the surgery of using stapler was done successfully.No postoperative complications such as stomal leak, bleeding and narrow were found.Meanwhile, no harmful consequences were observed through long-time follow-up.Conclusion Using mechanical stapler for bilio-intestinal anastomosis is time-saving, simple and reliable.It can be a choice for some diseases.

Objective The study was designed to observe influence of simvastatin on lung tissue angiogenesis and the gene expression of vascular endothelial growth factor(VEGF) and platelet factor 4(PF4) of rats with bleomycin (BLM)-induced pulmonary fibrosis. Methods Ninety-six healthy male SD rats were divided into four groups by random number table, including normal control group (A), bleomycin group (B), prednisone acetate treatment group (C) and simvastatin treatment group (D). Lung tissue of rats in each group was detected as specimens. HYP was detected by digestion method. Angiogenesis, VEGF and PF4 protein expression were determined by immunohistochemical method (SP). Expression of VEGF and PF4 mRNA were respectively detected by RT-PCR assay. Results (1)HYP content of group C, D was lower than the group B, which was statistical significance (P <0.01). (2)MVD and the expression of VEGF in group B, C and D was higher than that in group A. PF4 expression of group B, C and D were lower than that of group A (P < 0.01). MVD and the expression of VEGF of group D were lower than those of group B, the expression of PF4 of group D was higher than that in group B (P < 0.05). Conclusion Mechanism of simvastatin on pulmonary fibrosis may be related to regulate the expression of VEGF and PF4 in lung tissue, inhibit pathological angiogenesis.

In order to establish an UPLC-MS method for determination of twelve active compounds in Qili Qiangxin capsules including astragaloside, calycosin-7-0-glucoside, ginsenoside Rb1, ginsenoside Re, ginsenoside Rd, ginsenoside Rg1, ginsenoside Rf, periplocin, periplocoside H1, hesperidin, narirutin, isoquercitrin, the chromatographic separations were performedon a Phenomenex UPLC Kinetex C18 column (2.1 mm x 100 mm, 2.6 microm) with gradient elution of acetonitrile and 0.1% aqueous formic acidat a flow rate of 0.4 mL x min(-1). The temperature was set as 40 degrees C and injection volume was 5 microL. The monitoring of all analytes was achieved under the negative ionization mode with TOF-MS and TOF-MS/MS method. The twelve analytes showed good linearity (R2 > 0.9990) within the test ranges, the average recoveries were 98.0%-102%, respectively, and the RSD were less than 3.9%, respectively. The established method is simple, rapid, and sensitive, and can be used for quality control of Qili Qiangxin capsules.
Capsules ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Quality Control ; Spectrometry, Mass, Electrospray Ionization ; methods ; Tandem Mass Spectrometry ; methods

Copolyesters consisting of 3-hydroxybutyrate (3HB) and 3-hydroxyhexanoate (3HHx) (PHBHHx), a new type of biodegradable material, are receiving considerable attentions recently. The material properties are strongly related to the 3HHx fraction of PHBHHx. As the 3HHx fraction increase, crystallinity and melting point of PHBHHx decrease, flexibility and tractility increase. PHBHHx of different 3HHx fraction can meet different demands of commercial application and research. Aeromonas are the best studied PHBHHx-producing strains. Recent studies have been focused on optimizations of fermentative culture media and culture conditions for low-cost and efficient fermentative production. Aliphatic substrates such as long-chain fatty acid and soybean oil were used in the PHBHHx fermentation as the sole carbon source and energy source. Two-stage fermentation method was also developed for more efficient PHBHHx production. While studies on Aeromonas hydrophila revealed that the monomer composition of PHBHHx could not easily be regulated by fermentative process engineering methods such as changing substrates and fermentative conditions because precursors involved in the PHBHHx synthesis were all from the beta-oxidation pathway. In this study, phbA gene encoding beta-ketothiolase and phbB gene encoding acetoacetyl-CoA reductase were introduced into a PHBHHx-producing strain Aeromonas hydrophila 4AK4 so as to provide a new 3HB precursors synthesis way. phbA gene encodes beta-ketothiolase which can catalyze two acetyl-CoA to form acetoacetyl-CoA; phbB gene encodes acetoacetyl-CoA reductase catalyzing acetoacetly-CoA into 3HB-CoA which is the precursor of 3HB. The introduced novel 3-hydroxybutyrate precursor synthesis pathway allowed the recombinant strain to use unrelated carbon source such as gluconate to provide 3HB precursors for PHBHHx synthesis. Shake-flask experiments were carried out to produce PHBHHx of controllable monomer composition and fermentations in 5 L fermentor were also proceeded for confirmation of these result in large-scale culture. In flask culture, it was possible to reduce the 3HHx mol fraction in PHBHHx from 15 % in the wild type to 3% - 12% in the recombinant by simply changing the ratio of gluconate to lauric acid in the culture media. When lauric acid was used as the sole carbon source, 51.5 g/L Cell Dry Weight (CDW) containing 62 % PHBHHx with 9.7 % 3HHx mol fraction was obtained in 56 hours of fermentation in a 5 liter fermentor. When co-substrates of sodium gluconate and lauric acid (1:1) were used as carbon sources, 32.8 g/L CDW containing 52 % PHBHHx with 6.7% 3HHx mol fraction was obtained in 48 hours of fermentation. These results showed the possibility for fermentative production of PHBHHx with controllable monomer composition.
3-Hydroxybutyric Acid ; metabolism ; Acetyl-CoA C-Acyltransferase ; genetics ; metabolism ; Aeromonas hydrophila ; enzymology ; genetics ; metabolism ; Alcohol Oxidoreductases ; genetics ; metabolism ; Bacterial Proteins ; genetics ; metabolism ; Biotechnology ; methods ; Caproates ; metabolism ; Fermentation ; genetics ; physiology ; Lauric Acids ; metabolism

OBJECTIVETo explore effects of electroacupuncture of different frequencies on stroke.

METHODSForty-seven cases of stroke were treated with electroacupuncture at the motor region of the scalp and divided into 2 Hz, 2/15 Hz and 100 Hz groups according to the used frequency. Cerebral blood perfusion and brain functions before and during electroacupuncture in the 3 groups were investigated with single photon emission computed tomography (SPECT).

RESULTSLocal cerebral blood perfusion and brain cell functions could be improved by electroacupuncture of the 3 frequencies, and the actions of 2/15 Hz and 100 Hz were better.

CONCLUSIONElectroacupuncture of 2/15 Hz and 100 Hz has a better therapeutic effect on stroke when the stimulating intensity is fixed.