骨髓间充质干细胞向胰岛素分泌细胞诱导分化的动态观察

目的:探讨体外诱导骨髓间充质干细胞向胰岛素分泌细胞分化的可能性,并观察其动态变化。方法:实验于2005-09/2007-03在山东大学齐鲁医院完成。①标本来源:骨髓标本15例来自山东大学齐鲁医院成人骨髓检查结果正常者,均签署捐献同意书。②实验方法:无菌条件下取骨髓2.0～5.0mL,采用percoll分离液和贴壁法获得纯化的成人骨髓间充质干细胞。③实验评估:流式细胞仪行细胞表面抗原检测,在适当的条件下诱导其分化为成骨细胞和脂肪细胞。采用两步法向胰岛素分泌细胞诱导,观察其在碱性成纤维细胞生长因子、活化素A、胰岛素样生长因子、尼克酰胺等因子刺激下向胰岛素分泌细胞分化的动态变化。双硫踪染色鉴定胰岛样细胞团,酶联免疫吸附试验检测细胞分泌胰岛素的情况,RT-PCR检测胰岛细胞特异基因的表达。结果:①骨髓间充质干细胞的生长特性及免疫表型:分离培养获得的贴壁细胞,呈形态均一的梭形,流式细胞仪检测CD34、CD45表达阴性,CD29、CD44表达阳性。②向成骨细胞和脂肪细胞的诱导分化:此类细胞经茜素红染色、油红O染色均呈阳性,可诱导分化为成骨细胞和脂肪细胞。③向胰岛素分泌细胞的诱导分化:第1步诱导后出现细胞簇,双硫腙染色不着色,胰岛素分泌量少,仅检测到PDX-1基因的表达,证实其为胰岛前体细胞。第2步诱导后细胞簇数目逐渐上升,至诱导14d大部分细胞簇经双硫腙染色都呈红色。④诱导后培养上清中胰岛素含量:诱导第3,7,14,21天的胰岛素分泌量分别为(15.3±4.9),(34.1±5.6),(40.4±5.3),(39.8±5.1)mU/L。⑤胰岛细胞特异基因的表达:诱导7d仅检测到PDX-1基因的表达,insulin1、insulin2和Glut2基因均不表达。诱导14,21d检测到insulin2、PDX-1基因表达,insulin1基因弱表达,Glut2基因不表达。结论:体外分离、纯化得到的骨髓间充质干细胞诱导7d可分化出胰岛前体细胞,不具功能性;诱导14d后可成功地分化出成熟的具有功能性的胰岛素分泌细胞。     

国人全膝关节假体置换胫骨近端截骨面至腘血管距离的测量及意义

目的:测量国人全膝关节假体置换术胫骨近端截骨面后缘至腘窝血管之间的距离,以期为临床全膝关节置换术中避免损伤腘窝血管提供参考数据。方法:选择2006-06/12于解放军第二军医大学长征医院体检的50名正常成人(53膝),男29名(31膝),女21名(22膝)。所有观察对象均知情同意,且得到医院伦理道德委员会批准。对所有膝关节进行MRI扫描,在胫骨外侧平台以下10mm水平横断面上辨认腘动静脉,并测量胫骨近端截骨面后缘至腘窝动静脉的距离。结果:53膝全部进入结果分析,无脱落。①男性胫骨近端截骨面后缘至腘动脉、腘静脉平均距离为(6.7±2.5,7.3±2.3)mm,95%可信区间分别为5.8～7.6mm,6.5～8.1mm。②女性胫骨近端截骨面后缘至腘动脉、腘静脉平均距离为(6.6±1.9,7.1±2.7)mm,95%可信区间分别为:5.8～7.4mm,5.9～8.3mm。③不同性别观察对象胫骨近端截骨面后缘至腘血管的距离差异无显著性意义(P>0.05)。结论:腘窝血管紧邻全膝关节假体置换术胫骨近端截骨面后缘,不同性别间无明显差异。全膝关节假体置换术中进行胫骨近端截骨,特别是后方操作时需特别谨慎,以避免损伤腘窝血管。     

不同相对分子质量玻璃酸钠对膝关节镜术后关节功能恢复的作用比较

目的:观察关节腔内留置不同相对分子质量玻璃酸钠对膝关节镜术后早期疼痛及功能恢复的影响。方法:于2005-11/2006-05选择北京大学人民医院骨关节科收治的行膝关节镜手术患者60例。关节镜手术中根据不同诊断分别行半月板成形术、游离体取出术以及软骨成形术。60例患者按随机数字表法分为3个实验组,分别为Mr1.5×106～2.5×106玻璃酸钠组,Mr3×106玻璃酸钠组,Mr6×106玻璃酸钠组。术后关节腔内注入不同相对分子质量玻璃酸钠2.0～2.5mL,并被动屈伸膝关节20次,使玻璃酸钠均匀分布于关节内。术后第1天开始股四头肌力量锻炼,坐在床边屈膝活动,并可下床。术后1周拆除缝线,术后6周门诊复查。分别于术前、术后1,2,3d,1,6周采用同一评分量表进行自觉疼痛程度、日常生活活动能力、膝关节屈曲角度测评,评分越高,功能恢复越好。结果:纳入患者60例,均进入结果分析。①自觉疼痛程度测定:术后6周Mr1.5×106～2.5×106,3×106,6×106玻璃酸钠组自觉疼痛程度评分均高于术前[分别为(8.5±1.3),(7.3±2.2)分;(8.5±1.3),(7.3±2.2)分;(8.5±1.3),(7.3±2.2)分]。②日常生活活动能力测定:术后6周Mr1.5×106～2.5×106,3×106,6×106玻璃酸钠组日常生活活动能力评分均高于术前[分别为(60.5±8.4),(59.3±7.0)分;(63.4±8.2),(59.4±8.3)分;(66.9±3.8),(53.8±19.0)分]。③膝关节屈曲角度评分:术后6周Mr1.5×106～2.5×106,3×106,6×106玻璃酸钠组膝关节屈曲角度评分均高于术前[分别为(9.1±1.4),(5.8±2.7)分;(8.1±3.1),(7.2±3.5)分;(6.3±3.8),(5.5±3.1)分]。④综合评分:术后6周Mr1.5×106～2.5×106,3×106,6×106玻璃酸钠组综合评分均高于术前[分别为(88.1±7.7),(79.8±11.1)分;(91.4±6.8),(84.9±13.7)分;(91.2±10.7),(73.5±23.7)分]。关节腔内留置3种不同相对分子质量玻璃酸钠在膝关节镜术后近期各项评分差异均无显著性意义(P>0.05)。结论:关节腔内留置不同相对分子质量玻璃酸钠在膝关节镜术后近期康复中具有相似的效果。     

应用三维有限元法评估聚甲基丙烯酸甲酯和磷酸钙骨水泥铸件的力学特征

目的:聚甲基丙烯酸甲酯和磷酸钙骨水泥作为经皮椎体后凸成形术填充物,应用三维有限元分析法比较两种材料治疗骨质疏松性椎体压缩骨折的生物学效应。方法:实验于2003-03/09在南通医学院第一附属医院骨科和上海大学上海生物力学工程研究所合作完成。①利用WD-5型万能材料机测量聚甲基丙烯酸甲酯和磷酸钙骨水泥铸件力学性质。②选取骨质疏松患者1例(患者知情同意),排除其他疾患,应用CT扫描技术、图形数字化方法获取胸腰段的三维坐标,利用ANSYS5.0有限元分析软件等工具建立骨质疏松患者胸腰段的三维有限元模型。③模拟L1经皮椎体后凸成形术,对其和相邻椎体及椎间盘的负荷传递、应力、位移等进行分析。结果:①获取了精确的T11￣L3的多排螺旋CT扫描断层影像数据,经ANSYS5.0软件处理后得到4718个节点,1642个薄极单元,4495个八节点等参单元的骨质疏松患者胸腰段三维有限元力学模型。②采用聚甲基丙烯酸甲酯作为经皮椎体后凸成形术的填充物,比骨质疏松椎体应力增加了近15%。后部结构的应力平均增加13.2%,其中椎弓根增加5.9%,峡部增加6.25%,关节点增加27.6%。聚甲基丙烯酸甲酯在稳定椎体、恢复强度和刚度的同时,可能使其后部结构及相邻腰椎出现应力集中现象。③采用磷酸钙骨水泥作为椎体成形术填充物,比骨质疏松椎体应力平均增加了7%,应力集中现象明显小于聚甲基丙烯酸甲酯。结论:三维有限元力学分析表明两种材料均提高了椎体的抗变形能力,有利于椎体功能的重建。与聚甲基丙烯酸甲酯相比,磷酸钙骨水泥能减少成形椎体和相邻椎体之间的应力梯度,从长远看,能减少椎体退变和相邻椎体骨折的机会。     

Correlates of Anal Intercourse Vary by Partner Type Among Substance-Using Women: Baseline Data from the UNITY Study

Data are limited about anal intercourse among women at risk of HIV infection. HIV-negative non-injection drug using women at sexual risk (N = 404) were recruited. At baseline, 41.7% reported anal intercourse in the prior 3 months; of these, 88.2% reported unprotected anal intercourse (UAI). Factors associated with UAI varied by partner type: UAI with a steady partner was associated with younger age, depressive symptoms, and experience of battering; UAI with casual partners was associated with younger age, cocaine use and negative outcome expectancies for condom use; UAI with exchange partners was associated with cocaine use, negative outcome expectancies for condom use and depressive symptoms. Younger women were more likely to report unprotected anal intercourse if they did not use birth control. Specific counseling messages are needed to identify and address this risk and associated factors, including partner relationships, substance use, birth control, mental health issues and domestic violence.     

Care of post-traumatic spinal cord injury patients in India: An analysis

Background:

The spinal cord injured patients if congregated early in spinal units where better facilities and dedicated expert care exist the outcome of treatment and rehabilitation, can be improved. The objective of this study is to find out the various factors responsible for a delay in the presentation of spinal injury patients to the specialized spinal trauma units and to suggest steps to improve the quality of care of the spinal trauma patients in the Indian setup.

Materials and Methods:

Sixty patients of traumatic spinal cord injury admitted for rehabilitation between August 2005 and May 2006 were enrolled into the study and their data was analyzed.

Results:

Eighty-five per cent of the spinal cord injured patients were males and the mean age was 34 years (range 13-56 years). Twenty-nine (48.33%) of the spinal injuries occurred due to fall from height. There was an average of 45 days (range 0-188 days) of delay in presentation to a specialized spinal unit and most of the time the cause for the delay was unawareness on the part of patients and/or doctors regarding specialized spinal units. In 38 (62.5%) cases the mode of transportation of the spinal cord injured patient to the first visited hospital was by their own conveyance and the attendants of the patients did not have any idea about precautions essential to prevent neurological deterioration. Seventeen (28.33%) patients were given injection solumedrol with conservative treatment, 35 (60%) patients were given only conservative treatment and seven patients were operated (11.66%) upon at initially visited hospital. Of the seven patients operated five were fixed with posterior Harrington instrumentation (71.42%) and two (28.57%) were operated by short segment posterior pedicle screw fixation. None of the patients were subjected to physiotherapy-assisted transfers or wheel chair skills or even basic postural training, proper bladder/ bowel training program and sitting balance.

Conclusion:

Awareness on the part of the general population, attendants of the patients, clinical and paraclinical team regarding spinal cord injury needs to be addressed. Safe mode of transportation of spinal cord injured patient and early presentation at tertiary spinal care center with comprehensive spinal trauma care team should be stressed upon.     

Impaired fear response in mice lacking GIT1

G protein-coupled receptor kinase interacting protein 1 (GIT1) belongs to the family of Arf GAP proteins and has been implicated in the regulation of G protein-coupled receptor (GPCR) sequestration, cell migration, synapse formation and dendritic spine morphogenesis in neurons. To extend these cellular studies on GIT1 to an in vivo system, we generated mice with globally inactivated Git1 gene by breeding mice carrying a conditional Git1^flox allele with mice expressing the CMV-Cre transgene. Although many GIT1 knockout (GIT1-KO) animals died shortly after birth, homozygous mutants that survived the early post-partum period developed normally into adulthood and were fertile. Behavioral analyses of adult GIT1-KO mice revealed normal exploratory, anxiety- and depressive-like behaviors. However, GIT1-KO mice show impaired responses to fear conditioning and fear-potentiated startle. Overall, these findings suggest that GIT1 is involved in the regulation of amygdala-mediated experience-based emotional behaviors.     

Decreased prefrontal, anterior cingulate, insula, and ventral striatal metabolism in medication-free depressed outpatients with bipolar disorder

This study explored whether cerebral metabolic changes seen in treatment resistant and rapid cycling bipolar depression inpatients are also found in an outpatient sample not specifically selected for treatment resistance or rapid cycling. We assessed 15 depressed outpatients with bipolar disorder (six type I and nine type II) who were medication-free for at least 2 weeks and were not predominantly rapid cycling. The average 28-item Hamilton Depression Scale (HAM-D) total score was 33.9. The healthy control group comprised 19 age-matched subjects. All participants received a resting quantitative ¹⁸F-fluoro-deoxyglucose positron emission tomography scan. Data analyses were performed with Statistical Parametric Mapping (SPM5). Analyses revealed that depressed patients exhibited similar global metabolism, but decreased absolute regional metabolism in the left much more than right dorsolateral prefrontal cortex, bilateral (left greater than right) insula, bilateral subgenual prefrontal cortex, anterior cingulate, medial prefrontal cortex, ventral striatum, and right precuneus. No region exhibited absolute hypermetabolism. Moreover, HAM-D scores inversely correlated with absolute global metabolism and regional metabolism in the bilateral medial prefrontal gyrus, postcentral gyrus, and middle temporal gyrus. Analysis of relative cerebral metabolism yielded a similar, but less robust pattern of findings. Our findings confirm prefrontal and anterior paralimbic metabolic decreases in cerebral metabolism outside of inpatients specifically selected for treatment resistant and rapid cycling bipolar disorder. Prefrontal metabolic rates were inversely related to severity of depression. There was no evidence of regional hypermetabolism, perhaps because this phenomenon is less robust or more variable than prefrontal hypometabolism.