牛胎儿成纤维细胞分离与体外培养

本研究以 4 5d牛胎儿为材料 ,使用 0 .2 5 %胰酶 + 0 .0 4 %EDTA消化液 ,分离得到牛胎儿成纤维细胞 ,体外培养传代至第 2 8代。本实验描绘出牛胎儿成纤维细胞生长曲线 ,并发现α MEM是一种适合牛胎儿成纤维细胞生长的培养基。分离得到的牛胎儿成纤维细胞传至第 1 8代时核型仍然正常 ,冷冻、解冻后细胞可继续传代     

Echocardiography and conventional Doppler examination in clinically healthy adult Cavalier King Charles Spaniels: Effect of body weight,age, and gender,and establishment of reference intervals

ObjectivesThe objectives of this study were (1) to assess the potential effect of body weight (BW), age, and gender on the most commonly used echocardiographic and conventional Doppler variables in a large population of healthy Cavalier King Charles Spaniels (CKCS), and (2) to establish the corresponding reference intervals (RI).Animals134 healthy adult CKCS.MethodsUltrasound examinations were performed by trained observers in awake dogs. M-mode variables included left ventricular (LV) end-diastolic and end-systolic diameters, LV free wall and interventricular septal thicknesses at end-diastole and end-systole, and LV fractional shortening (FS%). The left atrium (LA) and aortic (Ao) diameters were measured using a 2D method, and the LA/Ao was calculated. Pulsed-wave Doppler variables included peak systolic aortic and pulmonary flow velocities, mitral E and A waves, and E/A ratio. Effects of BW, age, and gender on these 15 variables were tested using a general linear model, and RIs were determined by applying the statistical procedures recommended by the Clinical and Laboratory Standards Institute.ResultsA significant BW effect was observed for all variables, except LA/Ao, FS%, and mitral E/A ratio. A significant but negligible effect of gender and age was also observed for 5/15 and 4/15 of the tested variables, respectively. Only the BW effect on M-mode variables was considered as clinically relevant and the corresponding regression-based RIs were calculated.ConclusionsBody weight should be taken into account when interpreting echocardiographic values in CKCS, except for LA/Ao, FS%, and mitral E/A ratio.     

药用真菌双向固体发酵草乌后菌质对小鼠的急性毒性及致大鼠心律失常作用

[目的]研究药用真菌灵芝、茯苓、红栓菌和扇菇菌运用双向固体发酵草乌后菌质(分别称为灵乌菌质、茯乌菌质、栓乌菌质和扇乌菌质)对小鼠的急性毒性作用和对大鼠心脏毒性的影响.[方法]运用双向发酵技术获得菌质,根据急性毒性预试验结果进行最大耐受量试验,采用Powerlab生理信号记录系统,通过观察大鼠Ⅱ导联心电图变化研究不同药物对大鼠的心脏毒性.[结果]相同剂量下灌胃给药,在给药7d后各处理组小鼠体重增长明显低于正常对照组;给药14 d后,灵乌菌质组体重增长很快,接近于正常对照组;与炮制品组相比,灵乌菌质组、扇乌菌质组和栓乌菌质组对大鼠心律失常发生次数有显著性降低作用(P ＜0.05或P＜0.01),而茯乌菌质组对大鼠心律失常的影响和炮制品组没有显著差异;各发酵组间没有显著差异.[结论]灵乌菌质、扇乌菌质和栓乌菌质具有降低心律失常发生率的作用.     

Effect Comparison of Adenovirus Vector Transfer into Different Buffalo Cells

Buffalo mammary epithelial cell,cumulus cell and fibroblasts were transfected by adenovirus vectors and compared their transfection efficiency.293 cells were transfected with pBHGloxdelE13cre and pDC316-eGFP by liposome,the virus was collected and titer was detected.Buffalo mammary epithelial cell,cumulus cell and fibroblasts were exposed to different multiplicity of infection (MOI) of adenovirus vectors.After 72 h,the cells were observed with inverted fluorescence microscope,and transfection efficiency was calculated.When the MOI was 25,50,100,200 and 400,the transfection efficiency of fibroblasts were 0.7%,7.0%,9.0%,12.5% and 34.0%,the transfection efficiency of cumulus cells were 42.5%,55.3%,57.4%,76.0% and 80.0%,the transfection efficiency of mammary epithelial cells were 88.7%,100%,100%,100% and 100%.The results showed that the transfection efficiency of mammary epithelial cell was the best,followed by cumulus cell,and fibroblast was poor.     

Papillary muscle measurements in cats with normal echocardiograms and cats with concentric left ventricular hypertrophy

BACKGROUND: Papillary muscle hypertrophy can occur in conjunction with, or as the only indication of, hypertrophic cardiomyopathy or other diseases that result in left ventricular concentric hypertrophy (LVCH). Assessment of papillary muscle size is usually subjective because objective measures have not been reported. HYPOTHESIS: The study hypothesis was that papillary muscle dimensions are different between normal cats and cats with LVCH. ANIMALS: Echocardiograms from 44 normal cats and 40 cats with LVCH were included in the study. METHODS: All measurements were taken from the right parasternal short-axis view at the level of the papillary muscles at end-diastole. Three methods were used to assess papillary muscle size: the area subtraction method, the direct area trace method, and the diameter method. Measurements were compared between cat groups and method comparisons were made among methods for area determination. RESULTS: Cats with LVCH were older and had significantly greater left ventricular septal and free wall thicknesses and larger left atrial measurements than normal cats (P < .0006). Papillary muscle measurements were significantly greater by all measurement methods in cats with LVCH than in cats with normal echocardiograms (P < .0001). The area subtraction method and direct area trace method showed moderate agreement. CONCLUSIONS AND CLINICAL IMPORTANCE: Papillary muscle measurements were larger for LVCH cats than normal cats; however, some overlap was present. The establishment of these objective measures adds to the echocardiographic examination of cats.     

Cardiovascular effects of acute pulmonary obstruction in horses with recurrent airway obstruction

BACKGROUND: Recurrent airway obstruction (RAO) is common in horses. Although pulmonary artery (PA) pressure increases during RAO, cardiac function in horses with RAO has received limited attention. HYPOTHESIS: The purpose of this study was to noninvasively determine the cardiovascular effects of acute pulmonary obstruction (APO) in horses with RAO and their reversibility. ANIMALS: Five geldings with RAO, inducible by exposure to moldy hay, were studied. METHODS: Pulmonary mechanics, echocardiography, serum troponin I concentrations, arterial blood gases, and hematocrit were obtained before and after 7 days of APO. Heart rate, PA diameter and flow characteristics, right and left ventricular luminal dimensions and wall thicknesses, global cardiac performance, and evidence of myocardial damage were evaluated. Pulmonary mechanics and echocardiography were reevaluated during remission. RRESULTS: Severe, transient APO did not induce chronic cor pulmonale in horses, because cardiac anatomy and function were normal between episodes. An acute episode of APO produced anatomical and functional cardiac changes in both the right and left heart (including increased PA diameter, abnormal septal motion, and decreased left ventricular diameter and estimated stroke volume), possibly because of the development of pulmonary hypertension, without apparent myocardial damage. The decrease in stroke volume was offset by the increase in heart rate. CONCLUSIONS AND CLINICAL IMPORTANCE: With APO of 7 days' duration, cardiovascular abnormalities and the functional airway changes that produce them are reversible when the offending allergens are removed.     

p300/p53/Smad3 pathway involved in aging-related atrial fibrosis in human atrial fibroblasts

AIM To investigate the role of p300 in aging-related atrial fibrosis in human atrial fibroblasts （HAFs） and its potential mechanism. METHODS HAFs were obtained from human left atrial tissue, and the senescence model was established by cell passage. Senescence-associated β-galactosidase （SA-β-Gal） staining was used to detect the cell senescence, and Western blot was used to determine the protein levels of p300, p53, Smad3 and other senescence and fibrosis associated proteins in HAFs. RESULTS Compared to passage 3 HAFs, the proportion of senescent cells, and the protein levels of p300, p53, p-Smad3 and other senescence and fibrosis associated proteins were increased in HAFs at passage 7 and 11 （P<0.05）. After treated with curcumin （a p300 inhibitor） or transfection with p300 small-hairpin （sh） RNA plasmid, the protein levels of p300, and the senescence and fibrosis associated proteins were decreased in HAFs at passage 7（P<0.05）. Up-regulation of p300 by transfection with p300 over-expression plasmid increased the protein levels of p53, Smad3 and MMP-2 in HAFs at passage 3 （P<0.05）. CONCLUSION p300/p53/Smad3 signaling pathway plays an important role in aging-related atrial fibrosis in HAFs.     

Mouse melanoma cell exosomes promote expression of Rac1 protein in fibroblasts

AIM To investigate the effect of exosomes secreted by mouse melanoma cells on the expression of Ras-related C3 botulinum toxin substrate 1 (Rac1) protein in fibroblasts. METHODS Ultracentrifugation was adopted to separete exosomes secreted by mouse melanoma B16-F10 cells. The morphological structure of exosomes was observed by negative-staining electron microscopy. The size distribution of exosomes was determined by nanoparticle tracking analysis (NTA). The exosomal markers, tumor susceptibility gene 101 (Tsg101) and tyrosinase-related protein 2 (Tyrp2), were identified by Western blot. Laser confocal microscopy was used to observe the process that mouse embryonic fibroblasts (MEF) took in exosomes during co-culture. Immunocytochemical staining and Western blot were used to detect the expression of Rac1 protein in MEF. RESULTS B16-F10 cell exosomes showed a typical tea tray-like structure, with a size range of 141~255 nm, and expressed protein markers Tsg101 and Tyrp2. The results of laser confocal microscopy showed that compared with co-culture at 0 h, a small number of exosomes appeared in the MEF at 12 h, and a large number of exosomes accumulated in the MEF after co-cultured for 24 and 36 h. Western blot analysis showed that compared with co-culture at 0 h, the expression of Rac1 protein in the MEF was significantly increased at 24 h and 36 h of co-culture (P<0.01). The results of immunocytochemical staining showed that compared with co-culture at 0 h, the positive expression level of Rac1 in the MEF cells was significantly increased at 12 h, 24 h and 36 h of co-culture (P<0.05 or P<0.01). CONCLUSION Intake of exosomes secreted by mouse melanoma cells promotes the expression of Rac1 protein in fibroblasts.