诱导分化羊骨髓间充质干细胞作为组织工程瓣膜间质种子细胞的可行性

目的:对羊骨髓间充质干细胞加以诱导分化,并与大隐静脉来源的血管间质细胞进行比较,探讨其作为组织工程瓣膜种子细胞的可行性。方法:实验于2006-02/08在西京医院心血管病研究所完成。①家猪10只,屠宰后取主动脉瓣膜,尽量剔去血管外膜,浸入Hank’s液中洗涤,制备去细胞主动脉瓣,苏木精-伊红染色比较脱细胞前后的组织特点。②成年杂种绵羊10只,麻醉后分别于股骨大转子穿刺抽取肝素化骨髓10mL,稀释,离心,弃上清及脂肪层,余细胞用LG-DMEM无血清培养基重悬,铺于等体积的Percoll淋巴细胞分离液上,离心收集单个核细胞,消化传代培养。取第二、三代骨髓间充质干细胞,加入LG-DMEM条件培养基进行体外定向诱导分化。对诱导后细胞进行形态观察、免疫组化鉴定并绘制生长曲线。③上述10只绵羊抽取骨髓后,无菌条件下取大隐静脉,剥去静脉外膜,剪成1mm3,贴壁法培养,消化传代。④取第3代骨髓间充质干细胞与血管间质细胞,分别以5×104L-1密度接种于24孔培养板,常规孵育96h。每孔各吸取培养基0.5mL,测定两种细胞上清液中的羟脯氨酸含量。完毕后用胰蛋白酶消化处于对数生长期的细胞,加入冻存液,调整细胞密度为7×109L-1,置于液氮中,1周后复苏,计算两种细胞的存活率。⑤将去细胞猪主动脉瓣修剪成0.5cm×0.5cm×0.1cm大小,骨髓间充质干细胞、血管间质细胞均按105/cm2密度接种,置入37℃、体积分数为0.05的CO2孵箱中。反复种植3次,每次间隔24h,第4天各组均取出2份样本,扫描电镜观察。第7天向其余标本中加入四唑盐,采用酶联免疫检测仪490nm处骨髓间充质干细胞组、血管间质细胞组的吸光度值,比较两种细胞在去细胞主动脉瓣上的生长能力。结果:①新鲜的猪瓣叶中含有大量的成纤维细胞及内皮细胞。去细胞后,主动脉瓣中未见任何细胞及其碎片成分,纤维网架结构保持完整。②传代培养后的骨髓间充质干细胞和血管间质细胞形态相似,呈梭形或多角型,均于24h内贴壁,3～4d铺满瓶底;两种细胞对平滑肌肌动蛋白α、波形蛋白均呈部分阳性表达;且生长曲线相似,倍增时间分别为38h和36h(P>0.05)。③诱导分化后的骨髓间充质干细胞与血管间质细胞上清液羟脯氨酸含量基本相似[(1.52±0.21),(1.43±0.20)mg/L,P>0.05]。冻存复苏后,两种细胞的存活率亦基本相似[(85±3)%,(87±4)%,P>0.05]。④诱导分化后的骨髓间充质干细胞与血管间质细胞均能够种植在去细胞猪主动脉瓣上,两种细胞的吸光度值基本相似(0.50±0.04,0.48±0.03,P>0.05)。结论:诱导分化的骨髓间充质干细胞能够黏附在去细胞猪主动脉瓣膜支架上贴壁生长,与静脉来源的血管间质细胞在存活率、复苏后生长增殖情况、合成胶原功能等方面无明显差别,是合适的组织工程瓣膜间质种子细胞。     

更换髓内钉治疗股骨干骨折髓内钉内固定术后骨不连：12例资料回顾性分析

目的:通过对12例股骨干骨折髓内钉内固定术后骨不连患者进行更换髓内钉治疗的回顾性分析,观察股骨干骨折髓内钉内固定术后骨不连更换髓内钉治疗的疗效。方法:选择吉林大学第一临床医院2004-03/2006-09住院的股骨干骨折髓内钉内固定术后骨不连患者12例。其中男9例,女3例。更换髓内钉治疗:手术取出原有髓内钉,扩大骨髓腔,更换比原有髓内钉长2～4cm且直径粗1～3mm的髓内钉,其中3例骨缺损致接触面积<50%的病例取同侧髂骨行植骨治疗。12例患者获得随访,随访时间为6～24个月,平均13.75个月。分别于术后1,3,6个月拍X射线片检查,观察材料及宿主反应。按Neer评定标准进行功能评估。结果:①6个月时12例患者均参加随访,8～15个月时每月各1例,21,22,24个月时各1例。②12例中9例术后6～9个月X射线片显示达到骨性愈合,平均7.33个月。3例骨移植病例于术后8～11个月X射线片显示达到骨性愈合,平均9.67个月;骨性愈合率100%。③按Neer评分标准评估疗效结果:85分以上1例,70分以上11例。④所用髓内钉等植入物未发现有腐蚀、断裂等情况,未发现急慢性全身及局部反应。结论:更换髓内钉治疗具有促进骨不连愈合的生物学和力学效应,是治疗股骨干骨折髓内钉内固定术后骨不连的有效方法。     

中学生网络成瘾与父母养育方式的关系

目的:分析中学生网络成瘾与父母养育方式的关系。方法:①于2006-09在被试所在学校采用分年级简单随机抽样方法,共选取新乡市初一至高三的12班622名中学生为调查对象。②采用父母养育方式评定量表评价父母教养态度和行为,有4个等级,记1～4分,1为从不,4为总是(此量表包含11个因子,父亲教养方式的6个分量表包含情感温暖、理解,惩罚、严厉,过分干涉,偏爱被试,拒绝、否认,过度保护等58个题目;母亲教养方式的5个分量表包含情感温暖、理解,过干涉、过保护,拒绝、否认,惩罚、严厉,偏爱被试等57个题目)。采用修订的中文网络成瘾量表界定出网络成瘾群体,高分代表成瘾群体,低分代表非成瘾群体。③计量资料间差异比较采用t检验,相关性分析采用直线相关分析。结果:获得有效答卷612份,有效应答率为98.4%。男生304名,女生308名,平均年龄(16±2)岁。①网络成瘾与非成瘾中学生的父母养育方式的比较:网络成瘾中学生的父亲养育方式的惩罚严厉、过分干涉、拒绝否认和过度干涉四因子得分明显高于非网络成瘾中学生(t=2.186～2.518,P<0.05),而在情感温暖理解和偏爱被试因子上差异不明显(P>0.05);网络成瘾中学生在母亲养育方式的过分干涉保护、拒绝否认、惩罚严厉和偏爱被试因子得分明显高于非网络成瘾中学生(t=2.039～2.789,P<0.05～0.01),而在情感温暖理解因子上的得分差异不明显(P>0.05)。②中学生网络成瘾与父亲养育方式的相关性:网络成瘾总分与父亲养育方式的惩罚严厉、过分干涉、拒绝否认和过度保护因子呈明显正相关(r=0.172,0.112,0.204,0.149,P<0.01),与情感温暖理解因子呈明显负相关(r=-0.081,P<0.05),而与偏爱被试因子无明显相关(r=0.058,P>0.05)。③中学生网络成瘾与母亲养育方式的相关性:网络成瘾总分与母亲养育方式的过分干涉保护、拒绝否认、惩罚严厉和偏爱被试因子呈明显正相关(r=0.208,0.218,0.216,0.109,P<0.05～0.01),而与情感温暖理解因子无明显相关(r=-0.052,P>0.05)。结论:网络成瘾中学生父母的教养方式存在问题,网络成瘾与父母养育方式存有一定的相关性。     

Risk factors for coronary artery disease: a study comparing hypercholesterolaemia and hypertriglyceridaemia in angiographically characterized patients

Fifty-two male patients undergoing coronary angiography were allocated to four groups each consisting of 13 subjects: group I had normal coronary arteries and patients in groups II-IV exhibited coronary artery disease. In group II, plasma cholesterol was below 250 mg dl-1 and triglycerides below 160 mg dl-1; in group III, cholesterol was above 270 mg dl-1 and triglycerides under 160 mg dl-1; and in group IV, cholesterol was under 270 mg dl-1 and triglycerides above 180 mg dl-1. The hypertriglyceridaemic group IV had the highest coronary score. In addition, it had lowest lipoprotein lipase activity, lowest HDL-cholesterol and lowest high-density lipoproteins-2 (HDL-2) levels, suggesting that this type of hypertriglyceridaemia is caused--at least in part--by lipoprotein lipase deficiency with impaired removal of the triglyceride-rich lipoproteins and increased catabolism of HDL-2. Our findings point towards a type of hypertriglyceridaemia strongly associated with coronary artery disease which should therefore be treated accordingly.     

重点中学中学生6307人的抑郁障碍现况调查

目的:了解中学生抑郁障碍的患病情况。方法:在北京、辽宁、安徽3省市各随机取1所重点中学,在同1个月内分别对该校全体非毕业班的在校中学生6307人采用抑郁自评量表进行抑郁障碍的筛查。对筛查结果异常及由班主任提供的筛查结果虽正常但怀疑有情绪问题的学生进行精神科检查,根据ICD-10中F32抑郁发作、抑郁复发和F34.1恶劣心境的诊断标准筛选出抑郁障碍的学生。其中北京市某校参加学生(北京组)3727人;辽宁省某校学生(辽宁组)1637人;安徽省某校高一年级学生943人(安徽组)。结果:发放问卷6307份,收回合格问卷6307份,有效率100%。①在6307人,抑郁自评量表筛查的阳性率为23.50%;诊断为抑郁障碍的学生184例,患病率2.92%(184/6307)。其中北京组89例,辽宁组45例,安徽组50例。男女学生抑郁障碍的发病情况接近,差异无显著性意义(85/3016,99/3291,χ2=0.392,P=0.531)。三所中学高一学生的总检出率以安徽最高(50/943),辽宁次之(32/870),北京最低(25/668),差异无显著性意义(χ2=5.423,P=0.066)。北京组高中学生抑郁障碍患病率与辽宁组接近,差异无显著性意义[3.11%(42/1349),2.75%(45/1647),χ2=0.344,P=0.558]。②北京组中学学生抑郁障碍的总患病率为2.39%。初一学生的总患病率明显低于初二、高一、高二学生,且差异有显著性意义(P=0.001,0.011,0.037)。但各年级中学生抑郁障碍男女性别之间差异均无显著性。③辽宁组中学学生抑郁障碍总检出率为2.75%。高一学生抑郁障碍患病率明显高于高二学生(3.68%,1.84%,χ2=6.016,P=0.0146);各年级男女学生抑郁障碍患病率差异无显著性。④安徽组高一学生抑郁障碍总患病率为5.31%(50/943),男女学生差异无显著性意义(5.20%,5.44%,χ2=0.027,P=0.870)。结论:所调查的重点中学学生中抑郁障碍比较常见,从初二年级开始出现有较明显的增加趋势,男女学生抑郁障碍的患病情况无明显差别。     

The effect of transfusion on cardiac function in patients with chronic anemia

BACKGROUND: The deteriorating cardiac function of patients with chronic anemia may be improved with transfusion. The effect of transfusion on cardiac function was evaluated in patients with chronic anemia. STUDY DESIGN AND METHODS: In a prospective study, ejection fraction (EF) was determined before and after transfusion in 41 patients with chronic anemia. The results were compared and analyzed. RESULTS: The volume of red cells transfused and the levels of pretransfusion hemoglobin, hematocrit, and red cell, white cell, and platelet counts did not affect the posttransfusion EF, whereas the pretransfusion EF of the right or left ventricle inversely affected the posttransfusion change in EF in the respective ventricle (p < 0.001 and r = -0.5022; p = 0.01 and -0.3917, respectively). There was no significant difference in the change in EF in the right and left ventricles. CONCLUSION: Transfusion produced little immediate effect on cardiac function, but did change the EF to an extent that aided cardiac function in chronic anemia patients. The pretransfusion EF itself, but not the degree of anemia or volume of red cells transfused, affected the posttransfusion change in EF.     

Effects of rheumatoid arthritis on employment and social participation during the first years of disease in The Netherlands

OBJECTIVE: To study the effect of rheumatoid arthritis (RA) on working capabilities and social participation, including non-paying jobs, during the first 6 yr of disease. DESIGN: Cross-sectional study. METHODS: In April 1996, a self-reporting questionnaire was sent to 424 participants of a population-based clinical trial of therapeutic strategies for early RA initiated in 1990. RESULTS: A total of 363 completed questionnaires were returned (response = 86%). Disease duration varied from < 1 to 6 yr (mean 2.8 yr). The employment rate was low in the RA population compared to the Dutch population. In the male 45- to 64-yr-old group, 63% of RA patients were not employed compared to 32% of the Dutch population (P < 0.01). In the female 45- to 64-yr- old group, 76% of the RA population vs 67% of the Dutch were not employed (P < 0.05). Of the employed patients, 59% reported that RA affected their working capabilities, e.g. they worked an average of 21 h per week less due to RA. Of the patients without a paying job, 41% believed that this was (partly) due to RA. In addition, fewer RA patients had non-paying jobs and they performed fewer household activities compared to the general Dutch population. CONCLUSION: RA already has a negative influence on the working capabilities, social participation and household activities of these patients during the first 6 yr of disease.      

Expression and biochemical characterization of human glucose-6- phosphate dehydrogenase in Escherichia coli: a system to analyze normal and mutant enzymes

We have developed a system to characterize normal and mutated glucose-6- phosphate dehydrogenase (G6PD) enzymes in vitro. Normal or mutant G6PD cDNA was subcloned into a pGEX-3X vector, which allowed production of a functional fusion protein in Escherichia coli. When we compared the recombinant normal enzyme with authentic human G6PD, indistinguishable Km values for glucose-6-phosphate (G6P) and NADP were obtained, and the utilization rates for two substrate analogues (2-deoxy G6P and deamino NADP) also showed no difference between the enzymes. This system was used to assay a biochemically uncharacterized variant, G6PD Taipei (493 A-->G; 165 Asn-->Asp), plus two other known mutations (487 G-->A; 163 Gly-->Ser and 592 C-->T; 198 Arg-->Cys) that are located close to or within the putative G6P binding domain. Our results show that the G6PD activities of these three mutants were greatly reduced. No significant alteration in G6PD kinetics was observed for both 487 and 493 mutations. However, a drastic reduction in the Km for G6P (4-fold decrease) and tremendous increases in utilization rates of 2-deoxy G6P (32-fold increase) and deamino NADP (6-fold increase) were associated with the 592 mutation. This results suggests that arginine 198 in human G6PD, possibly located within the putative G6P binding domain, may play an important role in binding the substrate G6P. In addition, we and others have recently identified that at least nine different types of mutations are responsible for G6PD deficiency in Chinese. In this report, we also present the occurrence rate of each mutation present in the population of Taiwan.     

Identifying and characterizing casein kinase II in human platelets

We have recently shown that inhibition of protein phosphatases in platelets causes increases in protein phosphorylations with a concomitant inhibition of platelet responses. The burst in protein phosphorylation appears to be catalyzed by messenger-independent protein kinases. The aim of the present study was to characterize the presence of broad families of protein kinases found in platelets. Lysates of control and thrombin-stimulated platelets were prepared, and proteins were separated on MONO Q fast protein liquid chromatography. In addition to the presence of histone protein kinase and tyrosine kinase activities, human platelets contain casein kinase II (CKII) activity as assessed by phosphorylation of a specific substrate peptide. Western blot analysis and immunogold electron microscopy studies further showed the presence of alpha-, alpha'-, and beta- subunits of CKII. The enzyme appears to be distributed throughout the cytosol and not secreted after thrombin treatment. Immunoprecipitation studies suggest that at least some of the holoenzymes exist as an alpha alpha' beta 2 complex. Although no activation of the enzyme was detected after thrombin treatment, our results show that CKII is a major messenger-independent protein kinase in platelets.