外胚间充质细胞向神经胶质/雪旺细胞的诱导分化及在BAM生物材料的存活研究

目的 探讨体外诱导外胚间充质细胞向神经胶质/雪旺细胞分化的潜能,观察细胞在不同材料上的生长状况,利用分化细胞观察在BAM生物材料的复合情况,为进一步构建组织工程神经提供理论依据.方法 取妊娠10.5d胎鼠颌突外胚间充质细胞,纯化后分别用8.5mg/L牛脑垂体提取物BPE、50μmol/L弗司扣林(forskolin)及联合应用诱导细胞分化.相差显微镜下观察细胞的形态变化,免疫组化鉴定细胞性质,将诱导后的细胞种植于胶原膜及胶原凝胶,并于培养3、6d后分别进行光镜、HE染色及扫描电镜观察细胞的生长情况.结果 培养3d后外胚间充质细胞形态发生变化,细胞形态为梭形,突起变长.而对照组细胞形态未见明显改变;免疫组化显示外胚间充质细胞表达S-100蛋白,但无GFAP的表达.而诱导分化后出现GFAP的表达,其中BPE及Forskolin联合应用后分化效果最好,约为79.2%.而在BAM凝胶和膜上细胞都可伸展并增殖.结论 BPE和Forskolin可不同程度诱导外胚间充质细胞向神经胶质细胞分化.在BAM膜上分化细胞生长状态良好,为该细胞与BAM生物材料构建组织工程神经导管提供了理论依据.     

兔血管内皮细胞的培养及生物学特性

目的 为构建组织工程血管提供种子细胞.方法 兔麻醉取主动脉,采用酶消化法培养血管内皮细胞并传代,倒置显微镜、扫描电镜、透射电镜观察和免疫组织化学鉴定.结果 培养出血管内皮细胞并传代,倒置显微镜下血管内皮细胞呈“鹅卵石”形态,扫描电镜显示“铺路石”样生长,透射电镜可见内皮细胞特征性W-P小体,免疫组织化学鉴定Ⅷ因子（＋）.结论 兔血管内皮细胞培养成功,本法可为组织工程血管的研究提供细胞来源.     

细菌纤维素与雪旺细胞体外生物相容性研究

目的 探讨雪旺细胞与细菌纤维素体外共培养时的生物相容性,为细菌纤维素制备成组织工程化神经提供实验依据.方法 将背根神经节与细菌纤维素膜体外共培养,分别在培养的第1、2、5天通过扫描电镜和光镜观察雪旺细胞的形态及增殖、迁移情况,免疫细胞化学染色检测S-100蛋白表达情况;将坐骨神经来源的纯化雪旺细胞在细菌纤维素浸出液中培养,分别在24、48、72 h采用倒置显微镜和免疫细胞化学染色观察雪旺细胞形态及S-100蛋白的表达情况,并通过MTT检测细胞活力,流式细胞仪检测细胞周期来综合评价雪旺细胞与细菌纤维素的生物相容性.结果 雪旺细胞能在细菌纤维素膜表面正常生长、迁移;与普通培养基相比,细菌纤维素浸出液对雪旺细胞的生长、增殖及S-100蛋白的表达无明显影响.结论 雪旺细胞能够在细菌纤维素膜表面及细菌纤维素浸出液中良好生长,体外与细菌纤维素具有较好的生物相容性.     

成年大鼠周围神经损伤后远侧端雪旺细胞变化的体外研究

目的:体外观察大鼠坐骨神经损伤不同时间段远侧端雪旺细胞的变化。方法:切断成年SD大鼠坐骨神经,以远侧端为研究对象,术后1、2周和1个月取材,进行雪旺细胞体外培养,倒置相差显微镜,S-100免疫荧光染色及激光扫描共聚焦显微镜观察雪旺细胞的形态,MTT检测细胞活力,流式细胞仪检测细胞周期。结果:神经被切断后,雪旺细胞形态发生改变,1月组最明显;细胞活力和增殖能力也发生变化,切断1周组活力和增殖能力最强,随后逐渐变弱。结论:周围神经损伤后雪旺细胞形态发生变化,细胞的活力和增殖能力随着时间的推移逐渐下降但仍未停止增殖。     

GDNF基因修饰的雪旺细胞对大鼠坐骨神经缺损的修复作用

目的应用携带胶质细胞源性神经营养因子基因的逆转录病毒载体质粒(pLXSN-GDNF)对体外培养的雪旺细胞进行基因修饰,并结合细胞外基质凝胶及生物可降解聚乳酸-聚羟基乙酸共聚物(PLGA)管构建神经移植复合体,用于大鼠坐骨神经缺损的修复,同时对其促轴突再生及神经元保护作用予以检测评价.方法 40只成年Wistar大鼠随机分为4组:A组(n=10) 细胞外基质凝胶-PLGA管桥接组;B组(n=10)雪旺细胞-细胞外基质凝胶-PLGA管桥接组;C组(n=10) GDNF基因修饰的雪旺细胞-细胞外基质凝胶-PLGA管桥接组;D组(n=10)自体神经桥接组.损伤各组12周时应用神经电生理及甲苯胺蓝染色和透射电镜观测神经再生情况,辣根过氧化物酶神经逆行示踪技术用于脊髓前角运动神经元再生评价.结果 12周时再生神经运动传导速度检测、甲苯胺蓝染色轴突形态计量学分析、透射电镜超微结构观察以及脊髓前角运动神经元再生评价结果提示:C组优于A、B组,而与D组相比无明显差异.结论雪旺细胞的转基因处理可能弥补单纯细胞移植神经营养因子含量的不足,而可能达到与自体神经移植相似的效果.     

吡咯喹啉醌对大鼠雪旺细胞形态及增殖细胞核抗原表达的影响

目的:观察吡咯喹啉醌(PQQ)对体外培养雪旺细胞的形态、细胞数量及雪旺细胞内增殖细胞核抗原(PCNA)表达的影响。方法:从出生3-5dSD大鼠坐骨神经分离培养雪旺细胞,纯化并经S-100免疫荧光标记鉴定;设立对照组和实验组,分别加入100nmol/LPQQ和等量PBS培养48h后倒置显微镜下观察细胞形态并进行细胞计数;将不同浓度的PQQ(0,10,50,100,500nmol/L)加入雪旺细胞培养基中,培养48h后提取总mRNA及总蛋白进行PCNA的RT-PCR和Western blotting检测。结果:100nmol/LPQQ干预雪旺细胞48h可以使细胞的形态发生变化并使细胞数量增加,细胞数量较对照组增加13.6%;RT-PCR及Western blotting检测结果表明:PQQ浓度在10-500nmol/L浓度范围内可增加雪旺细胞内PCNA的表达,且当PQQ浓度为100nmol/L时该上调效应最明显,分别是对照组的2.0倍和3.4倍。结论:PQQ可增加体外培养雪旺细胞的数量;并可使体外培养雪旺细胞内PCNA表达上调。     

体外构建可注射性组织工程软骨的初步研究

目的利用关节软骨细胞复合壳聚糖-磷酸甘油(chitosan/glycerophosphate, C/GP)凝胶,体外构建可注射性组织工程软骨,为微创方式治疗关节软骨缺损提供依据.方法以体外培养扩增的兔关节软骨细胞为种子细胞,以自制温固化可注射C/GP凝胶为支架,按5×107/ml细胞浓度复合体外培养.于倒置显微镜下观察细胞形态与分布,四甲基偶氮唑盐(MTT)法计算培养1周时种子细胞存活率,培养4周时样本进行组织学及扫描电镜观察.结果软骨细胞在7:1(体积分数)C/GP凝胶中保持球形,细胞存活率95%以上;HE染色可见软骨较为典型的陷窝样结构、软骨囊及同源细胞群等现象;甲苯胺蓝异染及免疫组化Ⅱ型胶原染色呈强阳性.结论软骨细胞在C/GP凝胶中可存活并保持分泌软骨基质的功能,形成类软骨组织.温固化可注射性C/GP凝胶能作为软骨组织工程的载体材料.     

Biocompatibility research of hADSCs cultivated in PLGA/OAg hydrogel in vitro

天然高分子聚合物聚L-谷氨酸(PLGA)与氧化海藻酸钠(OAg)共价交联合成PLGA/OAg水凝胶,将人脂肪来源干细胞(hADSCs)接种于水凝胶内,倒置显微镜、扫描电镜观察细胞在支架中的存活、增殖、黏附情况,DNA定量法检测细胞的增殖.体外证实该水凝胶材料生物相容性良好,具有性能可控性和微创性等优势,对于进一步开发脂肪组织工程材料具有指导性意义.     

人早孕胎盘绒毛膜滋养层细胞的培养

目的: 培养纯度较高的人绒毛膜滋养层细胞, 为胎盘绒毛在妊娠期间的作用及其机制研究提供细胞学基础.方法: 胰蛋白酶-DNase消化法培养人绒毛膜滋养层细胞,间接免疫染色进行细胞纯度检测,透射电镜观察细胞内部结构.结果: 倒置显微镜下,可见细胞为上皮样细胞形态,呈片状铺展生长.细胞角蛋白染色阳性,波型蛋白染色阴性,表明不含污染细胞.透射电镜观察示所获细胞有典型滋养层细胞结构.结论: 该法简便易行,可获得合乎试验要求的人绒毛膜滋养层细胞.     

beagle犬骨髓间质干细胞与β-磷酸三钙多孔陶瓷复合培养的实验研究

目的:研究beagle犬骨髓间质干细胞(BMSCs)与β-磷酸三钙多孔陶瓷(β-TCP)的生物相容性.方法:体外诱导培养beagle犬BMSCs作为种子细胞,β-TCP作为组织工程支架,制备β-TCP-BMSCs复合物,相差倒置显微镜及扫描电镜观察细胞在材料孔隙区的贴附及生长情况.结果:相差倒置显微镜观察:自原代、2代、3代及4代培养,贴壁细胞由成纤维细胞样向三角形、棱形、多边形和多角形转化,最终重叠生长形成结节状;将细胞接种于材料上培养2～4 d,孔隙间大量生长增殖,呈细胞交叠覆盖.扫描电镜下观察:材料表面细胞呈梭形,相互融合成片状,表面有丝状突起,孔隙边缘有大量胶原分泌及细胞外基质分泌.结论:β-TCP与细胞的亲和力好,能够为基质细胞增殖、分化和成骨提供自然的三维空间,是极具潜力的组织工程支架之一.     

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.