BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are in advantages of easy collection and amplification in vitro, but the culture and induction in vitro still need to be modified. OBJECTIVE: To investigate the differentiated potency of BMSCs into ostcoblasts by modified in vitro culture methods and inductor matching. DESIGN: Observational study. SETTING: Department of Medical Laboratory, General Hospital of Guangzhou Military Area Command of Chinese PLA. MATERIALS: This study was performed in the Stem Cell Tissue Engineering Laboratory, Department of Medical Laboratory, Genera Hospital of Guangzhou Military Area Command of Chinese PLA from July 2004 to June 2006. Twenty adult SD rats of clean grade, irrespective of gender, weighing 140-180 g were provided by Animal Experimental Center, General Hospital of Guangzhou Military Area Command of Chinese PLA. The experimental animals were disposed according to ethical criteria. Β-sodium glycerophosphate, dexamethasone, and vitamin C were provided by Sigma Company, USA; goat-anti-rat osteocalcin antibody by DSL Company, USA; S-P immunohistochemical kit by Maixin Biotechnology Developing Co., Ltd., Fujian. METHODS: Cells were cultured and induced into osteoblasts by modified methods. Separation and culture of BMSCs: By anesthesia, bone marrow of bilateral femur and tibia was separated to prepare single cell suspension; subsequently, the suspension was inoculated in culture bottle, and the culture media was semi-quantitatively changed after 48 and 96 hours in order to get rid of non-adherent hematopoietic cells. The liquid was changed every three days to further get rid of non-adherent cells. At 80% cell confluence, the medium was digested with 0.25% trypsin and cells were subcultured in the ratio of 1:2. MSCs in the second generation were inoculated in 6-well culture plate and glass fiat plate; after 48 hours, basic culture fluid was removed. Differentiation of BMSCs into osteoblasts: Subcultured BMSCs differentiated into osteoblasts induced by dexamethasone (10 mmol/L), β-sodium glycerophosphate (10-7 mol/L), and vitamin C (50 mg/L). MAIN OUTCOME MEASURES: Ten days after differentiation by modified culture methods and inductor matching, alkaline phosphatase activity was measured with Ca-Co technique. Twelve days after culture, osteocalcin secretion was detected with immunohistochemical method. Two weeks after culture, cell mineralization was detected with Von kossa staining. RESULTS: Alkaline phosphatase activity: Alkaline phosphatase staining of cells was apparent; gray-black particles or massive precipitations were observed in cytoplasm after positive reaction; regions expressing alkaline phosphatase activity were brown-black. Osteocalcin secretion: Osteocalcin was apparently positive; nucleus was blue; cytoplasm was brown. Cell mineralization: Induced cells grew layer by layer, and adiaphanous nodus was observed at the same time. Black mineralized nodus granules in various sizes were observed after Von kossa staining, and this suggested that mineralized apposition occurred. CONCLUSION: BMSCs may be successfully cultured and induced into osteoblasts by modified culture method.

BACKGROUND:With the potential of multi-directional differentiation and high proliferation, bone marrow mesenchymal stem cells (BMSCs) have wide application prospect in tissue engineering. OBJECTIVE:To investigate changes in cells and bioactivity in the differentiation from BMSCs into osteoblasts. DESIGN, TIME AND SETTING:A randomized control experiment was performed at the Laboratory of Stem Cells and Tissue Engineering, Department of Medical Experiment, General Hospital of Guangzhou Military Area Command of Chinese PLA from July 2004 to June 2006. MATERIALS:Twenty adult SD rats of both genders were used for bone marrow collection. METHODS:Rats were equally and randomly divided into an experimental group and a control group. BMSCs were isolated from adult rats by modified bone marrow culture method. BMSCs were inoculated in basic medium containing 10% fetal bovine serum, high glucose DMEM, 100 U/L penicillin, 100 U/L streptomycin and 2 mmol/L glutamine in the control group. BMSCs were inoculated in conditioned medium (basic medium, 10 mmol/L β-glycerophosphoric sodium, 10-7 mol/L dexamethasone, 50 mg/L vitamin C). Cell slide was prepared. MAIN OUTCOME MEASURES:Inverted microscope and transmission electron microscope were used to observe cell appearance. Gomori modified calcium-cobalt method was applied to do alkaline phosphatase staining. Immunocytochemistry was employed to measure osteocalcin expression. RESULTS:Forty-eight hours after inoculation, a mass of BMSCs adhered to a wall. Seventy-two hours later, BMSCs proliferated and well grew. Seven to nine days later, cells were grown to 80% confluency. Transmission electron microscope showed BMSCs with big cell nucleus and immature appearance. After in vitro osteoblast induction, BMSCs proliferated, aggregated, had node and mineralized. One week later, alkaline phosphatase activities were expressed in BMSCs. Two weeks later, osteocalcin expression was detected. CONCLUSION:After one week of in vitro osteogenic induction, BMSCs enter the process of osteoblast transformation, remain proliferative activities, and can be used as seed cells in bone tissue engineering

OBJECTIVETo explore the effect of Danlou Tablet (DT) on arrhythmia model rats induced by transient myocardial ischemia/reperfusion (I/R).

METHODSTotally 45 healthy Wistar rats were randomly divided into 3 groups, the sham-operation group, the model group, and the DT group, 15 in each group. Rats in the sham-operation group and the model group were administered with distilled water by gastrogavage at the daily dose of 0.1 mL/kg. Rats in the DT group was administered with 0.53 g/mL DT suspension by gastrogavage at the daily dose of 0.1 mL/kg. All medication was lasted for 10 successive days. The myocardial I/R experiment was performed at 1 h after the last gastrogavage. ECG was performed before ligation and at I/R. The jugular arterial blood pressure of all rats was measured during the whole course. ST segment changes were observed at each time point of I/R. The ventricular fibrillation, the premature ventricular, the number and the duration of ventricular tachycardia within 30 min reperfusion were also observed. Activities of Na(+)-K+ ATPase and Ca2+ ATPase in the myocardium homogenate were detected as well.

RESULTSThe jugular arterial blood pressure and the heart rate were slightly lower in the DT group than in the model group, but with no statistical difference (P > 0.05). Compared with the sham-operation group, the degree of ST segment was obviously elevated in the model group at 0, 5, and 7 min (P < 0.05). It was significantly lower in the DT group than in the model group (P < 0.01). ST seg ment was more elevated at 5 min than at 0 min in the model group, but the degree of ST segment elevation was still obviously lower in the DT group than in the model group (P < 0.05). There was no statistical difference in the degree of ST segment elevation at 7 min between the two groups (P > 0.05). At 0 min when the decrement of ST segment exceeded one half the ischemia, there was no statistical difference in the degree of myocardial ischemia between the model group and the DT group (P > 0.05). Compared with the model group, the incidence of fatal and nonfatal ventricular fibrillation, the frequency and duration of ventricular tachycardia and premature ventricular beats were obviously lessened, and activities of Na(+)-K+ ATPase and Ca(2+)-ATPase increased (all P < 0.05).

CONCLUSIONSDT could significantly protect arrhythmias induced by transient I/R. Its effect might be related to lowering the degree of myocardial ischemia, and increasing ion transport channel related enzyme activities.

Animals ; Arrhythmias, Cardiac ; drug therapy ; etiology ; Disease Models, Animal ; Drugs, Chinese Herbal ; therapeutic use ; Male ; Myocardial Reperfusion Injury ; complications ; Rats ; Rats, Wistar

BACKGROUND: Color Doppler flow imaging can exam the early myocardial disorder in type 2 diabetic patients. What does exercise tolerance test work for the examination of such disorder in combination with color Doppler flow imaging (CDFI) ?OBJECTIVE: To analyze in comparison the early evaluation on reduced diastolic function in left ventricle in patients with type 2 diabetes between the examinations of exercise tolerance test combined with color Doppler flow image and simple color Doppler flow imaging.DESIGN: Cases-controlled comparison and self-comparison.SETTING: Department of electrodiagnosis and department of Endocrinology in a municipal hospital.PARTICIPANTS: Thirty-six cases of inpatients with type 2 diabetes were selected from Department of Endocrinology of Shenyang Red Cross Hospital from March to December in 2004, of which, 25 cases were males and 11cases females. The diabetic patients included had no cardiac vascular complications and participated in the study in volunteer. Thirty-two patients who received annual routine health check at the same period were selected as the control, of which, 20 cases were males and 12 cases females.METHODS: Metronics treadmill exercise test equipment was used for exercise tolerance in two groups. Before exercise(at quiescent state) and after exercise tolerance, Vivid 4 CDFI was used to determine flow velocity at E and A peak values respectively, ratio between flow velocities at E peak value and A peak value as well as isovolumtric relaxation time (IRT).MAIN OUTCOME MEASURES: Comparison of cardiac functional indexes before and after exercise tolerance in two groups.RESULTS: Thirty-four diabetic patients accomplished exercise tolerance test, of which, 1 case presented frequent ventricular extrasystole, another one presented precordial pain and stopped the test. In the control, all of 32 cases ratio between flow rates of E peak value and A peak value was remarkably lower than that in the control (0. 90 ± 0. 25, 1.40 ± 0.30, P ＜ 0.05 ); IRT was remarkably longer than that in the control [ (112. 07 ± 20. 16),imental group, the ratio between flow rates of E peak value and A peak value was remarkably lower than that in the control (0.62 ±0. 12, 1.28 ±0.87, P＜ 0.01 ); IRT was remarkably longer than that in the control[ (138. 10± 19.21), (97.37±9.61) ms, P ＜0.01].CONCLUSION: CDFI can supervise and evaluate at early stage the cardiac functional changes in type 2 diabetic patients. Due to the induction of exercise tolerance, the combination of exercise tolerance test and CDFI provides more accurate, objective and valuable conclusions at early stage.

OBJECTIVETo investigate the relationship between hormone sensitive lipase (HSL) gene polymorphism and aerobic endurance.

METHODSThe (CA)n repeats polymorphism genotypes in HSL intro 6 of 123 outstanding long distance runners and 127 controls of Han nationality in northern China were analyzed by PCR and Fluorescence labeled STR-genescan. Repeat polymorphisms were grouped according to segmentation point and alleles were divided into long or short chains. Chi-square test was used to analyze the frequency difference of allelic and genotypic between athlete and control groups.

RESULTS(CA) n repeats polymorphism in HSL gene was total of 9 different repeat number of alleles, which composed of 25 different genotypes. The chi-square test result showed that when compared short and long chain alleles split by 4, there was a significant difference (P <0.05) of genotype distribution in 5/10 km group compared with control. Compared the rest groups with control, there was no significant difference.

CONCLUSIONCompared short and long chain alleles split by 4, the LL genotype of (CA)n of HSL was associated with aerobic endurance and it might be a molecular marker of elite 5/10 km long distance runners.

Alleles ; China ; Ethnic Groups ; Genotype ; Humans ; Physical Endurance ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Sterol Esterase ; genetics

Fourteen compounds were obtained from Glechoma longituba by the chromatographic methods of silica gel, ODS, Sephadex LH-20 and preparative of HPLC. According to physicochemical properties and spectral data, these compounds were identified as stilbostemin B (1), trilepisiumic acid (2), 3, 4-dihydroxyphenyl ethanol ketone (3), bergeninmonohydrate (4), oresbiusin A (5), norbergenin (6), stilbostemin D (7), ehretioside B (8), ethyl ferulate (9), E-p-hydroxy-cinnamic acid (10), methyl gallate (11), protocatechuic acid (12), 4'-Hydroxyacetophenone (13), and E-3-2,4-dihydroxyphenyl-2-acrylic acid (14). Among them, compounds 1-10, 13 and 14 were isolated from this plant for the first time.
Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Lamiaceae ; chemistry ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Spectrometry, Mass, Electrospray Ionization

Objective To investigate plasma levels of total carnitine (TC) and free camitine (FC) in patients undergoing hemodialysis and peritoneal dialysis.Methods 200 cases of normal group came from physical examination in this hospital,all testing cases were the in-hospital patients in the department of nephropathy.TC and FC were determined by use of an enzymatic cycle assay on Hitachi 7170 automatic biochemical analyzer.Results In 200 cases of normal group,TC level was (56.52?9.61) ?mol/L,and FC was (46.60?8.23) ?mol/L.In 37 hemodialysis patients,TC and FC levels were (41.47?13.22) ?mol/L and (24.58?8.91)?mol/L before dialysis,a statistic difference was observed against the control group (P0.05).Conclusions Carnitine deficiency was seen in most patients undergoing hemodialysis and peritoneal dialysis.Furthermore,the deficiency status got worse along with the dialysis course in hemodialysis patients.Carnitine infusion can effectively improve the status of these patients.

AIM: To study the effects of exogenous metallothionein (MT) and ZnCl 2-induced MT production on biological action of homocysteine(HCY)in vascular fibroblasts METHODS: -TdR, -Pro incorporation and LDH leakage were measured, the cellular viabilities were calculated by trypan blue exclusion test and the intracellular contents of MT were assayed by [ 109 Cd]-hemoglobin saturation method in cultured rat vascular fibroblasts. RESULTS: Proliferation, collagen production of vascular fibroblasts in HCY-treated group were significantly increased compared with control group in a concentration-depedant manner. HCY (500 ?mol/L) increased LDH leakage and decreased the cellular viabilities ( P

Objective To investigate the diagnostic technique of Alport syndrome(AS)by immunohistochemical staining of type Ⅳ collagen ? chains on paraffin-embedded renal sections.Methods Renal biopsies were obtained from 2 patients with autosomal recessive form of AS,2 female patients and 2 male patients with X-linked dominant form of AS and 2 patients with hematuria(1male and 1 female).AS was diagnosed according to symptoms,family history,pathology,immunofluorescence staining of type Ⅳ collagen ? chains on renal and skin biopsies and gene analysis.Normal portions of nephrectomized kidneys from 2 patients with renal tumor were used as controls.Type Ⅳ collagen ? chains were stained by two-step immunohistochemistry staining method on paraffin-embedded renal sections.Three antigen retrieval methods including autoclave heating,pepsin digestion and proteinase were investigated to find the best antigen retrieval method for type Ⅳ collagen ? chains.The findings were compared with those examined by immunofluorescence staining on fresh frozen sections.Results By immunohistochemistry staining,type Ⅳ collagen ?3 and ?5 chains showed continuous linear pattern along glomerular basement membrane on sections from the controls and the hematuria patients,intermittent linear pattern for X-linked dominant female AS patients,negative for X-linked dominant male AS patient.For patients with autosomal recessive AS,the staining of type Ⅳ collagen ?3 and ?5 chains were negative on glomerular basement membrane,but ?5 chain was positive on glomerular capsules and partial tubular basement membrane.The results were the same as those examined by immunofluorescence staining.Conclusion AS can be diagnosed by immunohistochemistry staining of type Ⅳ collagen on paraffin-embedded renal sections,which is a new technique for diagnosis of AS in China.

AIM: To discuss the single application on iris localization technology in excimer laser for the treatment of astigmatism.
METHODS:Totally 203 cases (406 eyes) of laser in situ keratomileusis ( LASIK ) in the treatment of compound myopic astigmatism patients were operated from November 2011 to November 2012 in our hospital. They were divided into two groups. One was observation group using iris localization and the other was control group using routine operation. Patients in the observation group of 100 cases ( 200 eyes ) , aged 18-43 years old, spherical diopter was - 1. 25 to - 8. 75D, astigmatism was -1. 0 to -3. 25D. In control group, 103 patients ( 206 eyes ) , aged 19-44 years old, spherical diopter was -1. 75-9. 50D, astigmatism was -1. 0 to -3. 25D. The patients in the observation group before the application of WaveScan aberrometer check for iris image, spherical lens, cylindrical lens and astigmatism axis data operation, only single application of iris location, without using wavefront aberration guided technology, laser cutting patterns for conventional LASIK model, spherical, cylindrical mirror and astigmatism axis  RESULTS: Postoperative residual astigmatism, the observation group was significantly better than the control group. Astigmatism axial measurement after operation, the observation group was significantly less than that of the control group. Postoperative visual acuity at 6mo, the observation group was better than that of the control group. The difference was statistically significant.
data source to preoperative wavefront aberration results. The control group received routine LASIK. It was applicated comprehensive optometry optometry respectively to examine astigmatism and axial, based on the computer analysis during the preoperative, 1wk after the operation, and 6mo. Analysis of using SPSS 17 statistical software, it was independent-sample t test between the two groups of residual astigmatism and astigmatism axis.
CONCLUSION: For patients who cannot complete the wavefront aberration guided treatment of astigmatism, can separate the application of wavefront aberration analyzer automatic iris recognition technology to improve precision of astigmatism treatment, give full play to advanced technical performance of equipment, which has good application value.