兔脂肪来源干细胞表面标记分析及向心肌样细胞分化

目的分析兔脂肪来源干细胞（adipose-derived stern cells，ADSCs）表面标记并且对其进行体外诱导，观察其向心肌细胞分化的潜能，探讨脂肪来源细胞治疗缺血性心脏病的可能性。方法应用酶消化法获得兔ADSCs，并在DMEM培养基中进行体外培养。取第二代细胞以流式细胞术鉴定其表面标记。第二代细胞经5-AZA（浓渡为10μmol／L）处理24h，第2天更换新鲜培养液，以后每3d换液1次，诱导3周后，应用RT-PCR检测心肌细胞特异性α肌球蛋白重链（α—MHC）基因及免疫荧光检测心肌肌钙蛋白（cTNT）。结果流式细胞术检测显示Sca1、CD44、CD117高表达，CD34和CD45低表达。诱导后细胞表达cTNT。RT—PCR显示与正常心肌组织＊MHC阳性条带对照，诱导后ADSC也在相同位置出现α-MHC的阳性条带，而未诱导ADSC没有阳性条带出现。结论ADSC具有间质干细胞的特征，ADSC在一定诱导条件下向心肌样细胞分化，是一种有潜力的治疗缺血性心脏病的移植细胞。     

成人脂肪间充质干细胞体外定向诱导分化为心肌样细胞的实验观察

目的:探索成人脂肪组织来源的间充质干细胞(ADMSCs)体外分离、培养方法,并化学诱导使其向心肌细胞分化,为心肌再生医学提供理想的种子细胞来源. 方法:体外分离成人ADMSCs并进行传代培养,5-氮胞苷(5-aza)诱导其向心肌细胞分化. 分别于诱导后14,21,28 d时用免疫细胞化学染色、RT-PCR法进行心肌细胞鉴定. 结果:ADMSCs经5-aza诱导后14 d时细胞形态趋向于心肌细胞,诱导28 d免疫细胞化学显示心肌特异性肌钙蛋白I(cTN-I)、结蛋白(Desmin)表达阳性,RT-PCR检测心肌β-肌球蛋白重链(Cardiac β-MHC)基因表达阳性. 结论:成人脂肪组织存在丰富的间充质干细胞且易于分离扩增,体外5-aza诱导可使其向心肌细胞分化,可做为心肌再生医学的理想种子细胞.     

肝细胞生长因子基因转染5-aza诱导后的骨髓基质干细胞及其表达、分化的研究

目的 将重组腺病毒(Ad-HGF)转染5-氮胞苷(5-aza)诱导后的骨髓基质干细胞(BMSCs),检测其心肌样细胞分化和HGF基因表达,探讨其用于缺血性心脏病治疗的可行性.方法 以5-aza诱导比格犬BMSCs向心肌样细胞分化,形态学观察、免疫组化检测心肌样细胞标志--β-肌球重链蛋白(β-MHC)和(α-横纹肌肌动蛋白的表达;以Ad-HGF转染5-aza诱导后的BMSCs,RT-PCR、ELISA检测肝细胞生长因子(HGF)的表达.结果 形态学观察与免疫组化结果表明,5-aza诱导BMSCs向心肌样细胞分化,Ad-HGF转染5-aza诱导后的BMSCs在mRNA和蛋白水平均有HGF表达,转染后48h表达量最高,达103ng/ml.结论 Ad-HGF可高效转染5-aza诱导后的BMSCs,为其应用于缺血性心脏病的生物治疗奠定了基础.     

脂肪干细胞的分离培养及成心肌诱导

目的 探讨脂肪干细胞的分离培养以及成心肌细胞分化的可能性,为脂肪干细胞治疗缺血性心脏病的应用提供理论依据。方法自SD大鼠提取脂肪组织,用酶消化法分离脂肪干细胞,培养于a-MEM培养基中。显微镜下观察细胞生长情况及形态特征,流式细胞仪检测脂肪干细胞表面分子的表达情况。用5-氮胞苷对脂肪干细胞进行诱导,显微镜下观察细胞生长情况及形态变化,免疫组织化学方法检测心肌特异性抗体-αActin的表达。结果分离培养的脂肪干细胞形态近似梭形,流式细胞仪检测结果显示CD90及CD105高表达,而CD34及CD45极低表达。经5-氮胞苷诱导后的脂肪干细胞呈类似心肌细胞的形态学特征,免疫组织化学检测结果见心肌特异性标志物-αActin表达阳性。结论脂肪干细胞具有较强的分化为心肌样细胞的能力,可以应用于缺血性心脏病细胞治疗的研究。     

血府逐瘀汤含药血清对MSC分化过程中α-MHC和β-MHC mRNA表达的影响

目的：探讨中药复方血府逐瘀汤含药血清体外诱导骨髓间充质干细胞（MSC）向心肌细胞分化的情况．为MSC移植治疗缺血性心脏病提供安全有效的种子细胞。方法：分离培养Wistar大鼠MSC,用血府逐瘀汤含药血清诱导后,RT-PCR技术检测诱导后的MSC心肌细胞标志物α-MHC、β-MHC的表达。结果：MSC经过血府逐瘀汤含药血清体外诱导后,细胞中有心肌细胞标志物α-MHC、β-MHC的表达。结论：中药复方血府逐瘀汤含药血清可以体外诱导大鼠MSC分化为心肌样细胞。     

体外诱导大鼠骨髓间充质干细胞分化为心肌样细胞

[目的] 探讨体外培养定向诱导大鼠骨髓间充质干细胞 ( MSCs )向心肌样细胞分化的潜能.[方法] 取大鼠下肢骨骨髓,分离培养 MSCs,用 5-氮胞苷 ( 5-aza )定向诱导 24 h,相差显微镜下观察其形态变化,免疫组化法检测肌钙蛋白 ( cTnT )和心肌细胞结蛋白 ( desmin )表达,并通过 RT-PCR对肌球蛋白重链 ( MHC )在细胞中的表达进行鉴定.[结果] MSCs经 5-aza诱导后,部分细胞呈梭形,并可见肌管样结构.免疫组化检测示诱导后 MSCs的 cTnT阳性细胞数为 15%, Desmin表达强阳性, RT-PCR示诱导后 MSCs有 MHC表达.[结论] MSCs在体外能在一定条件下被诱导分化为心肌样细胞,是心肌缺血干细胞移植的较理想的细胞来源.     

体外诱导骨髓间充质干细胞向心肌细胞的分化和鉴定

[目的]探讨大鼠骨髓间充质干细胞(MSC)体外分离、培养、纯化、扩增的方法和生物学特性,研究其在诱导因子诱导下向心肌样细胞定向分化的能力。[方法]取大鼠骨髓,采用贴壁培养筛选法分离MSC,进行培养扩增,观察其生物学特性;用5-杂氮胞苷(5-AZA)诱导P3代MSC向心肌样细胞定向分化,并通过细胞免疫化学法鉴定分化细胞。[结果]大鼠MSC可通过贴壁法成功分离并可在体外大量扩增;流式细胞仪分析所培养的P3代细胞为纯度超过95%的MSC;P3代MSC经5-AZA诱导后1周,相差鼹微镜下见细胞呈长梭形、多核;2周可见肌管样结构;细胞免疫化学染色肌钙蛋白I、结蛋白、α-肌动蛋白为阳性。[结论]MSC易分离和培养,体外培养条件下生长良好,可连续传代,在5-AZA的诱导下可定向分化为心肌样细胞。     

QY1骨髓多能间质干细胞系向心肌细胞和血管内皮细胞分化

目的:探索QY1骨髓多能间质干细胞系在体外向心肌细胞、血管内皮细胞分化的能力;优化骨髓多能间质干细胞向心肌细胞分化的体外适宜诱导条件;并初步探讨骨髓间质干细胞向成心肌成血管细胞分化的能力.方法:分别用5-氮胞苷、血管内皮生长因子对QY1骨髓多能间质干细胞进行定向诱导分化.利用免疫组化、RT-PCR鉴定分化后的细胞.结果:在传代培养至72 h时,加入10 μmol/L5-氮胞苷诱导24 h后换液培养,以后每7 d换液1次,培养14 d进行再次诱导的条件下,细胞能出现自发性搏动的心肌细胞和肌管样结构,诱导率达(39.47±0.56)%.心肌特异性的表面抗体α-横纹肌肌动蛋白、心肌连接蛋白-43表达阳性,RT-PCR检测有心肌特异性因子α-肌球蛋白重链表达.在用血管内皮生长因子诱导48 h后,出现呈直线排列的血管内皮细胞;7 d后,有部分区域细胞相互连接呈血管内皮细胞网状;14 d时免疫组化检测,发现诱导后的细胞血管内皮特异性表面抗体CD31和Ⅷ因子表达阳性.结论:QY1骨髓多能间质干细胞系细胞在体外特定条件下具有分化为心肌细胞和血管内皮细胞的能力.     

5-氮胞苷诱导骨髓间充质干细胞向心肌样细胞分化观察

目的：探讨5-氮胞苷（5-aza）对纯化培养的骨髓间充质干细胞（MSCs）向心肌样细胞分化的影响.方法：取雄性Wistar大鼠下肢长骨骨髓进行体外培养和连续传代,取第3代MSCs以10 μmol/L 5-aza诱导作用24 h后,用含体积分数为5%胎牛血清的完全培养基继续培养,观察细胞形态学变化,并在4周后行免疫细胞化学染色鉴定α-Actin,Desmin及心肌特异性肌钙蛋白T（cTnT）的表达.结果：5-aza作用MSCs 7d后细胞形态及排列发生变化,14～21 d左右细胞数量较诱导前明显减少,28 d后细胞稀落,周围伸出多个长的突起,细胞体积大小不等.经抗α-Actin抗体、抗Desmin抗体、抗cTnT抗体鉴定部分细胞出现阳性染色,阳性细胞率分别为20%,19%和10%.结论：骨髓间充质干细胞在体外特定的诱导条件下可以向心肌样细胞分化,有望成为缺血性心脏病细胞移植治疗的细胞材料.     

黄芪甲苷体外诱导骨髓间充质干细胞分化为心肌样细胞的实验研究

【目的】观察黄芪甲苷体外诱导骨髓间充质干细胞(MSCs)定向分化为心肌样细胞的作用。【方法】采用密度梯度离心和贴壁培养法分离大鼠MSCs,反复传代及纯化后,取第3代MSCs,采用流式细胞仪检测细胞表面抗原CD34和CD44;取第8代MSCs进行分组诱导:黄芪甲苷组(终浓度为250 mg/L)、5-氮胞苷(5-aza,终浓度为10μmol/L)、黄芪甲苷加5-aza组(终浓度分别为250 mg/L与10μmol/L),并设空白对照组,诱导后继续培养4周;计算心肌样细胞诱导率,采用免疫组化法鉴定诱导后MSCs中结蛋白(Desmin)和心肌特异性肌钙蛋白I(cTnI)的表达,采用逆转录聚合酶链反应(RT-PCR)法鉴定诱导后MSCs中心肌特异性蛋白α-心肌肌球蛋白重链(-αMHC)和β-心肌肌球蛋白重链(-βMHC)信使核糖核酸(mRNA)的表达。【结果】原代培养的MSCs首先形成集落,传代细胞体积变大,诱导后细胞呈梭形,并出现肌管;MSCs表面抗原CD44表达阳性,而骨髓造血干细胞表面抗原CD34表达阴性,表明本实验方法所得细胞为均一的MSCs细胞群;各组在不同时间的心肌样细胞诱导率无明显差异;免疫组化结果显示诱导后MSCs表达心肌特异性蛋白Desmin和cTnI;RT-PCR结果显示诱导后MSCs表达成熟的心肌特异性蛋白-αMHC和-βMHC。【结论】黄芪甲苷可在体外诱导大鼠MSCs定向分化为心肌样细胞,但诱导率仍有待提高。     

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.     

APPLICATION OF LORNOXICAM TO PATIENT-CONTROLLED ANALGESIA IN PATIENTS UNDERGOING ABDOMINAL SURGERIES

FOR anesthesiologis s ,treatingpostoperativepainhas alwaysbeen a problem.Althoughopioidshave been provedtobe effective,theirsideeffectscouldnotbeignored.With thedevelopmentofscienceand pharmacology,many drugs with aspectsof satisfactoryanalgesicefficacyand couldbe welltoleratedby patientshave been developed.And lornoxicamisone of them, which isa non-steroidalanti-inflammatorydrug (NSAID ), with analgesic, anti-infl-ammatory,andantipyreticproperties.Itseliminationhalf-time(3 to 5 hours) isle…     

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.     

Acupuncture Combined with Spinal Tui Na for Treatment of Primary Dysmenorrhea in 30 Cases

Objective: To observe the therapeutic effects in acupunture treatment of primary dysmenorrhea combined with spinal Tui Na, and study its mechanism. Methods: Thirty cases of the treatment group were treated by acupuncture combined with spinal Tui Na, and thirty cases in the control group were treated by routine acupuncture. Results: The total effective rate was 93.3% in the treatment group, and 73.3% in the control group, with a significant difference between the two groups (P<0.05). Conclusions: Acupuncture combined with spinal Tui Na has good prospects for treatment of primary dysmenorrhea.     

Luo Lingjie's Experience in Treating Chronic Nephropathy

In treating chronic nephropathy,Luo Lingjie,a chief physician,pays attention to regulating the balance between yin and yang,treating infection if present,and removing pathogenic factors.He prescribes gentle drugs and uses carefully strongly warming-tonifying ones,emphasizes the importance of persuading the patient to persist in treatment with medication and nurse one's health for recuperation,and is good at combined use of TCM and western medicine therapy and brings the merits of various therapies into full play,with obvious theraoeutic effects.     

Dr.Zhang Ren's Experience in the Acupuncture Treatment of Different Diseases with the Same Therapeutic Principle

Dr.Zhang Ren,the chief physician,is the chairman of Shanghai Acupuncture and Moxibustion Association.Having been engaged in medicine for about 40 years,he is experienced in treating various intractable diseases.In his long years of clinical practice,he advocates taking the TCM differentiation as the basis to seek for the acupuncture method for treatment of modern intractable diseases.The author of this essay had the fortune to follow Dr.Zhang in study.The following is a summary of Dr.Zhang's experience in the acupuncture treatment for different intractable diseases with the same therapeutic principle.     

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.