人TH基因重组腺病毒的构建及生物学活性研究

将人酪氨酸羟化酶cDNA表达盒克隆于质粒型腺病毒载体p△Elsp1A，得到重组质粒pAd－TH。随后用脂质体法将重组质粒pAd－TH和拯救型腺病毒质粒pBHG11一起共转染293细胞，通过体内同源重组生成重组腺病毒AdCMVth，THcDNA重组进入腺病毒E1区并受CMV启动子控制。采用形态学、病毒核酸酶切和PCR／RT－PCR等方法进行鉴定正确。重组腺病毒滴度达到1010pfu／ml。初步结果表明，该重组腺病毒感染MN9D细胞后可使细胞内多巴胺水平增加1倍，显示出明显的TH生物学活性。提示TH重组腺病毒AdCMVth可作为高效的基因转移载体用于帕金森氏病基因治疗。     

肾移植取肾方法的改进

报道了自1985年2月至今共摘取供肾132例,取得尸肾263只(1例独立肾),其中251只供肾用于临床。分侧切取肾38例,整块切肾41例,改进后整块切肾53例。文中着重介绍改进后的整块切取尸肾的手术方法,讨论了术式的优缺点。     

SAGE遗传分析系统的功能及应用

SAGE是集多功能于一体的医学遗传学群体与家系资料计算机分析系统。本文概述SAGE系统的主要功能及应用环境。重点介绍了FCOR2和TDTEX两个功能模块的数学原理和使用方法。应用TDTEX模块 ,我们发现微卫星标记 85ca与小儿失神症存在连锁不平衡 ,提示在该位点附近存在小儿失神症的易感基因     

Porcine small intestine submucosa (SIS) is not an acellular collagenous matrix and contains porcine DNA: possible implications in human implantation

Porcine small intestinal submucosa (SIS) has been recommended as a cell-free, biocompatible biomaterial for the repair of rotator cuff tendon tear. However, we have observed noninfectious edema and severe pain in patients who have undergone SIS implantation for tendon repair. The aim of this study was to conduct an independent assessment of the safety and efficacy of Restore SIS membrane. The Restore orthobiologic implant was examined by histology and the nested PCR technique using porcine immunoreceptor DAP12 gene to examine if SIS membrane contained porcine cells or DNA, respectively. The material was also implanted into mice and rabbits for the evaluation of biological reaction and inflammatory response. Restore SIS was found to contain multiple layers of porcine cells. Chloroacetate esterase staining showed that some of these cells were mast cells. Nested PCR of the DAP12 gene demonstrated that Restore SIS contained porcine DNA material. Subcutaneous implantation of Restore SIS membrane in mice, and in rabbits for rotator cuff tendon repair, showed that the membrane caused an inflammatory reaction characterized by massive lymphocyte infiltration. In conclusion, Restore SIS is not an acellular collagenous matrix, and contains porcine DNA. Our results contradict the current view that Restore SIS is a cell-free biomaterial, and that no inflammatory response is elicited by its implantation. We suggest that further studies should be conducted to evaluate the clinical safety and efficacy of SIS implant biomaterials.     

Cloning and expression of MP13 gene from rat hippocampus, a new factor related to guanosine triphosphate regulation

C-Fos and the Fos-related antigens (FRA) are induced by various stimuli. A novel 35-37 kDa FRA was induced much longer after the treatment using kainic acid (KA) and may be very important for neuronal survival after brain damage. To identify this long-term FRA, we have constructed a cDNA library derived from hippocampus after KA treatment and screened it with an antibody highly conserved M-peptide region of FRAs. One gene, MP13, was cloned with a 1662 bp open reading frame and coded for a 554-amino acid protein. MP13 has a leucine zipper region, a glutamine repeat region, and has high similarity to the activator of the small guanosine triphosphate (GTP)ase Rab5. Gel retardation analysis revealed that MP13 functions as a GTP regulation related factor.     

N-氨基-D-门冬氨酸诱导大鼠下丘脑视上核、室旁核和弓状核内c-fos基因表达

实验用foS蛋白免疫组织化学方法,研究了中枢神经系统兴奋性介质N-氨基-D-门冬氨酸诱导大鼠下丘脑内c-fos的表达.N-氨基-D-门冬氨酸注射大鼠皮下后,观察了fos阳性细胞在下丘脑内开始出现与消失的时程相关以及在下丘脑内的分布.结果表明:给N-氨基-D-门冬氨酸后30分开始出现fos阳性细胞,1～2小时达高峰,4～8小时消失.fos阳性细胞主要分布于视上核、室旁核和弓状核,视上核和室旁核中fos阳性细胞分别占细胞总数的57.8%和63.6%,在弓状核中占细胞总数的60.6%.     

DQCAR 113 and DQCAR 115 in combination with HLA-DRB1 alleles are significant markers of susceptibility to rheumatoid arthritis in the Korean population

We have investigated HLA region microsatellite polymorphisms in rheumatoid arthritis (RA) which are known to be associated with HLA class II alleles in the Korean population. Ninety patients with RA and 106 controls were employed for this study, in which TAP1CA, DQCAR, D6S273, HLA-DRB1, -DQA1 and -DQB1 allele typing were performed. DQCAR 113 (RR = 3.2, P<0.0002), DQCAR 115 (RR = 3.6, P<0.0001) and heterozygous DQCAR 113/115 (RR = 11.2, P<0.0001) frequencies were significantly increased in the RA group compared with the control group. The HLA-DRB1 genotypes of patients who had DQCAR 113/115 alleles were defined as DRB1*04 and/or DRB1*09. There was no significant difference between RA and controls in D6S273 and TAP1CA allele frequencies. We demonstrated that HLA-DRB1*0405 (RR = 6.6, P<10(-6)), DQA1*03 (RR = 5.2, P<10(-6)), DQB1*04 (RR = 3.5, P<0.002) alleles were useful markers of susceptibility to RA in Koreans. The frequency of HLA-DRB1*0405 was higher in DQCAR 113 allele-positive RA (68.1%) than in DQCAR 113 allele-negative (16.3%) and total RA (43.3%) groups, and the susceptibility risk of DQCAR 113 allele to RA was more increased in the DRB1*0405-positive group (RR = 5.5, P<0.04). On the other hand, DQCAR 115 allele was more significantly associated with susceptibility to RA in HLA-DRB1*0405-negative patients (RR = 5.1, P<0.0005), and the association between RA and HLA-DRB1*0405 was also significantly associated with DQCAR 115 allele-negative patients (RR = 13.2, P<0.00001) as compared with DQCAR 115 allele-negative control groups. HLA-DRB1*0405-DQA1*03-DQCAR113-DQB1*03 haplotype showed high relative risk value (RR= 17.7, P<0.0002). In conclusion, the DQCAR allele in combination with HLA class II, especially DR, is probably a useful risk marker for RA susceptibility in the Korean population.     

活体亲属供肾肾移植的临床分析

目的　总结分析活体亲属供肾肾移植的手术和治疗经验 ,探讨其临床效果 .方法　回顾性分析 33例活体亲属供肾肾移植的临床资料 ,包括手术方法和创新、免疫抑制药物的用药方案及临床效果 .结果　本组全部切取左肾 ,经腹手术 ,手术顺利 ,移植肾在开放血液循环后 1～ 10分钟内分泌尿液 .供体肾功能在 1周内恢复正常 ,未出现严重并发症 .受者仅 2例出现急性排斥反应 .全部受者至今存活 ,肾功能良好 .结论　活体亲属供肾 ,移植效果明显优于尸体供肾肾移植 .排斥反应发生率低 ,恢复顺利     

Cord blood leucocyte expression of functionally significant molecules involved in the regulation of cellular immunity

The cellular immune system of the newborn infant is immature and hypo-responsive when compared with adults. The extent to which immaturity of the leucocyte function underlies hyporesponsiveness in the newborn is incompletely understood. In this study flow cytometric techniques were applied to investigate the concurrent expression of a range of surface and intracellular leucocyte functional molecules and cytokines in resting and stimulated cord and adult blood. Production of interleukin (IL)-2 and expression of the components of its receptor, IL-2R alpha/beta/gamma, were investigated. No differences in the proportion of leucocytes producing IL-2R alpha and IL-2R gamma were observed for newborns and adults. A lower proportion of T cells and natural killer (NK) cells from newborns expressed IL-2R beta and upregulation of expression was slower. We hypothesize that reduced IL-2R beta may curtail early autocrine IL-2 activation of immune responses in the newborn. This hypothesis was supported by the observation that an increased proportion of stimulated T cells from newborns produced IL-2 at 4 h poststimulation, but at 24 h the proportion was lower than for adult T cells. The very low levels of interferon (IFN)-gamma produced by neonatal T cells and NK cells may also be partly explained by a curtailment of early autocrine activation of T cells. Expression and kinetics of upregulation for other functional molecules were studied. CD71, HLA-DR, tissue factor and CD152 levels were not significantly different for adults and newborns, suggesting that cord blood leucocytes, in some respects, may demonstrate functional maturity. IL-6 secretion by stimulated monocytes was also comparable in cord and adult blood. However, IL-1 alpha and IL-1 beta were produced by a lower proportion of monocytes from newborns than adults. Similarly, tumour necrosis factor (TNF)-alpha production for monocytes and T cells was lower in cord blood. The mean fluorescence intensity for IL-1 alpha, IL-1 beta and TNF-alpha was also lower for leucocytes from cord blood. These findings are significant in relation to the inability of newborn infants to mount a febrile response to infection. The findings of lower expression of IL-2R beta and lower production of inflammatory cytokines IL-1 alpha, IL-1 beta and TNF-alpha is a basis for improved understanding of the immunological immaturity of leucocytes in the newborn.     

中药髓复康对大鼠脊髓GFAP表达的影响

为了探讨中药制剂髓复康对急性脊髓半横断损伤诱发的星形胶质细胞反应性增生是否具有抑制作用,本实验选用8周龄的清洁级雄性SD大鼠54只,其中48只用于制备T12胸髓右侧半横断损伤模型,并将其随机分为髓复康组(S)和空白对照组(B)。另6只设为正常对照组(N组)。不同时间点取材,采用免疫组织化学法比较各组脊髓损伤区GFAP表达的变化。结果显示,B组在术后3、7、15和30d四个时间点GFAP免疫反应阳性细胞数和GFAP免疫阳性产物的OD值均明显高于N组,术后15d达高峰,各时间点两组之间比较均有显著性差异(P<0.05);在脊髓损伤后3d,S组的GFAP免疫反应细胞数明显少于B组(P<0.05),而GFAP免疫阳性产物的OD值与B组接近。在以后的各个时间点,S组的GFAP免疫反应细胞计数和GFAP免疫阳性产物的OD值都明显低于B组(P<0.05),而高于N组(P<0.05)。研究结果提示,髓复康能减轻大鼠脊髓半横断损伤所诱发的星形胶质细胞反应性增生。