不同浓度羊自体血清对骨髓间充质干细胞增殖能力影响的研究

目的 观察不同浓度羊自体血清对骨髓间充质干细胞(bone marrow-derived mesenchymal stem cells,bMSCs)增殖活力的影响,并筛选出最佳血清培养浓度.方法 以12月龄青山羊为实验动物,首先制备自体血清,分离获取bMSCs;然后选DMEM/F12为基础培养基,采用不同体积分数(分别为5%、10%、15%)自体血清及10%体积分数胎牛血清培养基分别对bMSCs进行培养;通过显微镜下细胞形态学观察、活细胞计数及MTT等方法对细胞进行检测,并对检测数据做统计学分析.结果 不同浓度自体血清培养培养条件下bMSCs形态正常.与10%胎牛血清组进行比较,5%自体血清组bMSCs增殖较慢(P＜0.01),10%、15%自体血清组增殖较好,但组间无统计学差异(P＞0.05).结论 羊自体血清培养bMSCs是可行的,10%浓度自体血清是最佳培养浓度.     

不同浓度兔自体血清对脂肪源性干细胞增殖能力的影响

目的：观察不同浓度兔自体血清对脂肪源性干细胞（adipose-derived stem cells,ADSC）增殖活力的影响,并筛选出最佳血清培养浓度。方法：以大鼠为实验动物,首先制备自体血清,分离获取ADSC;然后选DMEM为基础培养基,采用不同体积分数（分别为5%、10%、20%）自体血清及10%体积分数胎牛血清培养基分别对ADSC进行培养;通过显微镜下细胞形态学观察、活细胞计数及MTT等方法对细胞进行检测,并对检测数据做统计学分析。结果：不同浓度自体血清培养培养条件下ADSC形态正常。与10%胎牛血清组进行比较,5%自体血清组ADSC增殖较慢（P〈0.01）,10%、15%自体血清组增殖较好,但组间无统计学差异（P〉0.05）。结论：兔自体血清培养ADSC是可行的,10%浓度自体血清是最佳培养浓度。     

补肾行气活血法对骨髓基质干细胞诱导成软骨细胞作用的影响

[目的]观察补肾行气活血法对诱导骨髓基质干细胞(BMSCs)成软骨细胞作用的影响.[方法]将传至第3代的BMSCs分为阴性对照组和治疗组,分别采用软骨细胞诱导培养基加体积分数为10%的空白对照血清和诱导培养基加体积分数10%的骨炎定含药血清培养诱导成软骨细胞,观察软骨细胞的生长状况及细胞表型特征.[结果]治疗组在细胞贴壁、生长形态及成软骨特性方面明显优于对照组,细胞表型的免疫组化染色均为阳性.[结论]以补肾行气活血法为组方依据的骨炎定含药血清既可促进BMSCs的增殖,又可促进诱导的软骨细胞分化.     

补肾活血中药对激素性股骨头坏死家兔骨髓基质干细胞增殖的影响

目的探讨补肾活血中药对激素性股骨头坏死家兔的骨髓基质干细胞(BMSCs)增殖的影响。方法对正常家兔及激素性股骨头坏死家兔骨髓基质干细胞分离、筛选、培养后,选择第3代家兔BMSCs,将5%、10%、20%体积分数补肾活血方含药血清对上述第3代家兔BMSCs进行干预,通过MTT法观察不同时间BMSCs增殖情况。结果 (1)经过含药血清的培养基培养的第3代细胞增殖速度明显的优于前者;(2)补肾活血方含药血清组A490值均高于同体积分数正常血清组(P<0.05),同时,补肾活血方含药血清组A490值随着含药血清体积分数增加而上升,差异有统计学意义(P<0.05)。结论补肾活血生骨法对激素性股骨头坏死家兔BMSCs增殖和分化有明显促进作用,其作用强度随浓度、时间而改变。     

细胞密度和血清对小鼠骨髓基质细胞体外生长的影响

目的:研究不同细胞接种密度、血清浓度及种类对小鼠骨髓基质细胞体外生长的影响.方法:设置不同细胞密度、不同种类血清及血清量的培养体系,观察细胞贴壁情况,细胞80%贴壁时间,用台盼蓝排斥试验检测活细胞率,MTT法检测细胞增殖情况,HE染色光镜下观察细胞形态.结果:原代全血培养的小鼠骨髓基质细胞适宜接种密度为1～4×106/ml;细胞最适生长的血清浓度为10%～20%;与胎牛血清相比马血清对小鼠骨髓基质细胞的促增殖作用更好(P＜0.05);不同种类血清培养的小鼠骨髓基质细胞类型有一定差异(P＜0.05).结论:不同细胞接种密度及血清浓度和血清种类对小鼠骨髓基质细胞体外生长有一定影响.     

骨髓间质干细胞体外增殖及诱导分化成肌样细胞实验研究

目的比较SD大鼠、Wistar大鼠和C57BL/6J小鼠骨髓间质干细胞在体外的增殖及在体外将骨髓间质干细胞诱导分化成肌样细胞的情况。方法体外分离成年SD大鼠、Wistar大鼠和C57BL/6J小鼠的骨髓干细胞,培养增殖,用5-氮杂胞苷诱导分化,使其向肌细胞定向分化。结果SD大鼠和Wistar大鼠的原代骨髓间质干细胞约10 d可达90%以上融合,平均3 d左右传代,C57BL/6J小鼠的骨髓间质干细胞在1周内可迅速增殖,1周后基本处于停滞状态,且细胞集落很少,难传代。SD大鼠的骨髓间质干细胞在含5-氮杂胞苷、两性霉素B和5%马血清的分化培养基作用下,可以分化为肌肉特异性抗原desmin和-αsarcomeric actin染色阳性反应的肌样细胞。结论不同种系的骨髓干细胞在体外用相同的培养基培养,可以具有截然不同的增殖能力;5-氮杂胞苷可以促使骨髓干细胞定向分化为肌样细胞。     

小鼠骨髓间质干细胞向胰岛样细胞的转分化及其鉴定

目的:培养小鼠骨髓间质干细胞,使其转分化为胰岛素样细胞,为细胞移植治疗糖尿病提供细胞来源.方法:分离小鼠骨髓间质干细胞,以添加诱导剂DMSO的DMEM培养基培养,于光镜下观察细胞形态,RT-PCR及葡萄糖刺激的胰岛素释放试验检验诱导形成的胰岛的生理功能.结果:分离初大部分细胞呈圆形,10 d后细胞快速增殖成胰岛样细胞团,RT-PCR显示培养后细胞内胰岛素和胰高血糖素表达增强(P均＜0.01),对高糖刺激的胰岛素释放较低糖时增加(P＜0.01).结论:小鼠骨髓间质干细胞在体外培养条件下可增殖,并可转分化为有内分泌功能的胰岛样细胞.     

低氧有助于小鼠骨髓源间充质干细胞的定向分化

目的本研究在测量小鼠体内股骨骨髓正常氧分压的基础上设定相应的体外低氧浓度(3%),研究低氧环境对小鼠骨髓源间充质干细胞向内皮细胞系定向分化功能的影响。方法用微电极测量正常小鼠股骨骨髓的氧分压,并以此为依据设定体外培养的低氧浓度。将小鼠骨髓源单个核细胞置于3%(低氧)和20%(常氧)氧浓度下用内皮细胞专用培养基(EGM-2)进行定向诱导分化培养,培养至第2代进行Dil-Ac-LDL摄取实验和FITC-UEA结合实验双染色鉴定。对3%和20%氧浓度下培养的内皮样细胞用MTT法进行增生能力的鉴定;用荧光染色法对其进行黏附能力的比较。结果小鼠股骨骨髓生理性氧分压浓度为(21.55±3.40)mm Hg(1 mm Hg=0.133 k Pa),相当于(2.8±0.45)%氧体积分数(约为3%)。在不同氧浓度下培养的细胞,DilAc-LDL摄取实验和FITC-UEA结合实验均为阳性。。MTT细胞增生实验显示低氧(3%)培养的细胞吸光度为0.31±0.05,常氧(20%)培养的细胞吸光度为0.16±0.01(P<0.01,n=5)。低氧培养的细胞,其黏附率为(8.72±2.95)%,常氧培养的内皮样细胞,其黏附率为(3.91±0.72)%(P<0.05,n=5)。结论 3%的氧浓度是骨髓生理状态下的氧浓度。在此浓度下,骨髓源间充质干细胞可被诱导分化为内皮样细胞。与常氧相比,低氧培养的内皮样细胞具有较高的增生和黏附能力。     

犬骨髓间充质干细胞的培养及生物学特性

目的 研究探讨犬骨髓间充质干细胞的分离、培养方法以及生长增殖特性.方法 无菌技术抽取比格犬骨髓,密度梯度离心法获取细胞.加入含胎牛血清体积分数为0.1的低糖DMEM培养基中进行原代培养;待细胞生长至80%～90%融合时,1:3传代培养.倒置显微镜观察细胞形态.MTT法测定3、5和10代细胞生长曲线.流式细胞仪检测第5代细胞的表面抗原分子.结果 原代培养24h后看见微少贴壁细胞,初起为小圆形,3 d后可见梭形类成纤维细胞呈漩涡样生长.10代以后细胞逐渐有宽大梭形样变,增殖逐渐减缓,偶有细胞飘零.流式细胞仪检测传5代细胞有96%表达CD44,95%表达CD29,而只有5%表达CD34.生长曲线显示,传3、5代骨髓间充质干细胞均有较强的增殖能力,其中传3代细胞更为明显,而传10代细胞的增殖能力较前有所减弱.结论 犬骨髓间充质干分离提取操作简便易行,且有较强的生长增殖能力,能够长期培养,符合组织工程种子细胞的基本条件.     

杜仲对大鼠骨髓间充质干细胞增殖的影响

【目的】探讨杜仲对大鼠骨髓间充质干细胞(BMSCs)增殖的影响。【方法】分别取10日龄SD大鼠胫骨、股骨的骨髓,过滤离心后将所得BMSCs进行体外单层培养、扩增,采用DMEM培养基,37℃、体积分数5%CO2条件培养。在细胞达到80%～90%融合时,使用2.5 g/L胰酶消化传代。将传代细胞分别置于空白血清培养液(对照组)、杜仲水提取物含药血清及杜仲醇提取物含药血清培养液进行培养,观察细胞在3种条件下的形态、贴壁生长、增殖、集落等情况,采用四甲基偶氮唑盐(MTT)法绘制生长曲线,比较3种培养条件下达到100%融合的时间。【结果】杜仲水提取物组、杜仲醇提取物组对大鼠BMSCs增殖的影响显著高于空白血清对照组(P<0.05),且杜仲醇提取物组作用优于杜仲水提取物组(P<0.05)。【结论】杜仲对大鼠BMSCs增殖具有一定促进作用。     

Value of D-Dimers in patients with acute aortic dissection

Objective: To evaluatel the value of D-dimers in patients with acute aortic dissection (AAD). Methods: This study consisted of 16 patients with AAD and 27 non-AAD patients. Serum D-dimets were measured by Sta-Liatest D-DI immunoturbidimetric assay. Results: D-dimer level was higher (P ＜ 0.001) in patients with AAD(7.91 ± 5.52 μg/ml) than that in non- AAD group(1.57±1.24 μg/ml). D-dimer was positive (＞0.4 μg/ml) in all patients with AAD and in 10 control group patients (37%). Among patients with acute AAD, D-dimers tended to be higher in Stanford A than in Stanford B (8.67 ± 4.31 μg/ml vs. 3.24±1.27 μg/ml, P ＜0.01). D-dimer values tended to be higher in more extended disease(3.84 ± 1.65 μg/ml, 8.57 ± 3.58 μg/ml and 11.87 ± 5.69 μg/ml in thoracic aorta, thoracic and abdominal aorta, thoracic and abdominal aorta and iliacal arteries, respectively, P ＜ 0.05 for both 8.57 ± 3.58 and 11.87 ± 5.69 vs. 3.84 ± 1.65 ). Including the control group into the analysis, we found a sensitivity of 100%, a negative predictive value of 100%, and a specificity of 66% and a positive predictive value of 64% for D-dimer in diagnosis of AAD in our patients with suspected AAD. Conclusion: D-dimer was elevated in patients with AAD. A negative D-dimer test result could be useful in excluding AAD.     

Research of combined liver-kidney transplantation model in rats

Objective： To set up a simple and reliable rat model of combined liver-kidney transplantation. Methods： SD rats served as both donors and recipients. 4℃ sodium lactate Ringer＇s was infused from portal veins to donated livers,and from abdominal aorta to donated kidneys, respectively. Anastomosis of the portal vein and the inferior vena cava （IVC） inferior to the right kidney between the graft and the recipient was performed by a double cuff method, then the superior hepatic vena cava with suture. A patch of donated renal artery was anastomosed to the recipient abdominal aorta. The urethra and bile duct were reconstructed with a simple inside bracket. Results： Among 65 cases of combined liver-kidney transplantation, the success rate in the late 40 cases was 77.5%. The function of the grafted liver and kidney remained normal. Conclusion： This rat model of combined liver-kidney transplantation can be established in common laboratory conditions with high success rate and meet the needs of renal transplantation experiment.     

BLOOD PRESSURE CHANGE WITH AGE IN SALT-SENSITIVE TEENAGERS

Objective To observe blood pressure change with age in salt-sensitive teenagers whose salt sensitivity were determined by repeated testing.Methods Salt sensitivity was determined through intravenous infusion of normal saline combined with volume-depletion by oral diuretic furosemide in 55 teenagers. After five years, salt sensitivity was re-examined and subject blood pressure was followed up. Blood pressure changes in salt-sensitive teenagers were compared to that of non-salt sensitive teenagers over five years.Results After 5 years, the repetition rate of salt sensitivity determined by intravenous saline loading is 92.7%. In teenagers with salt sensitivity on the baseline, both the systolic blood pressure increments and increment rates were much higher than non-salt sensitive teenagers (12.7±12.1 mmHg vs. 2.8±5.2 mmHg, P＜ 0.01; 12.2%± 12.0% vs. 2.5% ±4.4%, P＜ 0.001,respectively). There was a similar trend for diastolic blood pressure (8.4 ± 6.4 mmHg vs. 3.7 ± 6.4 mmHg, P = 0.052; 13.2% ±10.6 % vs. 6.8%± 10.1%, P = 0.053, respectively).Conclusions Salt sensitivity determined by intravenous saline loading showed good reproducibility. Blood pressure increments with age were much higher in salt-sensitive teenagers than non-salt sensitive teenagers, especially in terms of systolic blood pressure.     

安心颗粒对急诊经皮冠状动脉介入术后生活质量影响的研究

目的：评价使用安心颗粒对急诊经皮冠状动脉介入术（PPCI）术后生活质量的影响.方法：将160例接受PPCI的急性ST段抬高型心肌梗死患者随机分为安心颗粒组（术前顿服安心颗粒8.8g,术后安心颗粒4.4 g/次,每日2次）和对照组（仅接受基础药物治疗）.所有患者均服用阿司匹林、氯吡格雷和阿托伐他汀.分别在入院时、出院前1d、出院后180 d时,应用心肌梗死多维度量表（MIDAS）、中文版SF-36评价量表对患者生活质量评分.并观察术后30 d以内的出血并发症、血小板减少症发生情况.结果：入院时和出院前1d,两组患者的心肌梗死MIDAS、SF-36量表评分比较无差异（P＞0.05）;出院后180 d时,与对照组比较,安心颗粒组MIDAS、SF-36评分明显减低（P＜0.05）;组内与入院时比较,两组出院前1d、出院后180 d时,MIDAS、SF-36评分均降低（P＜0.05）.两组患者在随访期间均无大量出血、少量出血、重度和极重度血小板减少症发生,安心颗粒组有4例、对照组有7例发生不明显出血（P＞0.05）.两组发生轻度血小板减少症的患者数比较无差异（P＞0.05）.结论：PPCI使用安心颗粒,能改善急性ST段抬高型心肌梗死患者的生活质量,且不增加出血风险.     

The comparative studies of the influences of Urapidil and Nicardipine on sino-atrial node function, atrio-ventricular node function and hemodynamics

Objective:To investigate the influences of urapidil and nicardipine on rabbit sinus function,atrio-ventricular node function and hemodynamics.Methods:Thirty-two Angora's rabbits were selected and randomly divided into four groups.U1 group:urapidil 0.25 mg/kg;U2 group:urapidil 0.5 mg/kg;N1 group:nicardipine 10 μg/kg;N2 group:nicardipine 20 μg/kg.All these medicine were administrated within 30 seconds.Measurements were taken before and after the administration of urapidil or nicardipine for the following data:mean blood pressure(MAP),heart rate(HR),sino-atrial conduction time(SACT),maximal sinoatrial recovery time(SNRTmax)corrected sinus node recovery time(CSNRT),index of sinus node recovery time(SNRTI),Wenckebach A-V conduction frequency (WB),and P-R interval.Results:Significant MAP and HR changes were identified in all of the four groups before and after administration of both urapidil and nicardipine.No significant changes could be found in the rest of the parameters.Intergroup analysis showed that SACT and CSNRT of N1 and N2 groups were shorter than those of the U2 group(P＜0.01);the MAP decreased(P＜0.01)and the HR increased drastically(P＜0.01).Conclusions:Neither urapidil(0.25 mg/kg,0.5 mg/kg)nor nicardipine(10μg/kg,20μg/kg)has any significant influence on rabbit sinus function or rabbit atrio-ventricular node function.Nicardipine could be a better choice than urapidil for parafunctional sinus node patients.     

Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.     

The clinicopathological and immuohistochemical analysis of solid-pseudopapillary tumor of the pancreas： report of 9 cases

Objective：To investigate the clinical features, pathological characteristics and immunophenotype of solid-pseudopapillary tumor of the pancreas（SPTP）. Methods：Nine surgically treated cases of SPTP were retrospectively reviewed. Hematoxylin and Eosin（HE） staining and immunohistochemical staining were used to analyze all cases, and the general clinical data was collected. Results：Six patients were asymptomatic except for a palpable mass. Two patients complained of vague-epigastric pain. One patient appeared jaundice. The tumor was encapsulated and solid tissues alternately with cystic tissues. Histologically, the histological structure of solid portion was pseudopapillary with a fibrovascular core. Tumor cells were uniform and medium-sized which were arranged in sheets ets or nests or pseudopapillary patterns. Immunohistochemical studies demonstrated that SPTP proved positive in vimentin（9/9 cases）, AAT（9/9 cases）, NSE（9/9 cases）, ACT（7/9 cases）, CK20（2/9 cases）, CgA（1/9 cases）, S-100（3/gcases）, PR（4/gcases）, Syn（3/9 cases） and CD56（5/9cases）, negative in CEA and ER. Conclusion：SPTP is a tumor predominantly occurring in young women frequently without special symptoms. This tumor has various characteristical histological patterns with different immunophenotype.     

Dynamic Changes in Serum Estradiol and Progesterone levels in Patients of Premenstrual Syndrome with Adverse Flow of Liver-qi

Objective:To probe into the influence of changes of ovarian hormones on the pathogenesis of the specific sub-type premenstrual syndrome(PMS)and reveal partial microcosmic mechanisms of adverse flow of liver-qi.Methods:Estradiol(E2)and progesterone(P)levels in serum were determined at different phases of menstrual cycle by radioimmunoassay.Results:In the group of PMS with adverse flow of liver-qi.the secretive peak value Of E2 and P at the follicular phase significantly decreased,and the secretive peak value at the luteal phase did not come into being.Conclusions:Low E2 and P secretive peak at the follicular phase and absence of secretive peak at the luteal phase is one of the microcosmic mechanisms of PMS with adverse flow of liver-qi.One of the pathophysiologic mechanisms of specific sub-type PMS is probably the continuous low level of E2and P.     

Real-time Three-dimensional Echocardiography in Assessment of Left Ventricular and Right Ventricular Volumes

Real-time three-dimensional echocardiography (RT3DE)is a new ultrasound technique that enables dynamic threedimensional visualization and quantification of the heart in real time. Investigation of feasibility and methodology of RT3DE in determining left ventricular (LV) and right ventricular (RV) volumes, RT3DE was performed in 35 normal adults using Philips SONOS 7500 system with a 2-4 MHz matrix array transducer. The 60°×60° "pyramid" volume database was obtained and analyzed on a TomTec echo workstation. Both LV and RV volumes were calculated with four 3DE methods (i.e. apical 2, 4, 8, and 16-plane) through manually tracing ventricular endocardial borders in end diastole and end systole. Stroke volumes were then calculated. LV volume was also measured by 2DE Simpson's rule using GE VIVID 7 ultrasound machine.     

SCREENING FOR A 21-CHROMOSOME ABNORMALITY IN PREIMPLANTED EMBRYOS OF ELDERLY WOMEN

Increasing maternal age is the only etiological factor unequivocally linked to Down's syndrome in humans. The occurrence rate of newborns with Down's syndrome is about 1/220 in women over 35 years old. However, the occurrence rate in embryos fertilized in vitro, of the elder woman is unclear. Using FISH we screened the number of chromosome 21 in preimplanted embryos of 5 elderly women (average age, 38.4 years) to study the feasibility and necessity of screening trisomy 21 in embryos in patients over 35 years old at the in vitro fertilization (IVF) center.