【作者】 阎蓉；

【导师】 曾水林；

【作者基本信息】 东南大学 ， 人体解剖学与组织胚胎学， 2006， 硕士

【摘要】 第一部分不同年龄大鼠脑组织中MHC、CD4及CD8分子的表达及其意义目的:观察不同年龄大鼠脑组织中MHCI、MHCII、CD4及CD8分子的表达,并探讨其意义。方法:将实验大鼠按胎龄和日龄分为7组:E15d、E19d、P0d、P7d、P14d、P28d及老年组(24月龄),每组6只,共42只大鼠。经灌流取材冰冻切片,选取顶叶大脑皮质、海马、黑质、纹状体、小脑及脉络丛等部位,采用免疫组化染色方法,检测MHCI、MHCII、CD4和CD8分子在上述部位的表达情况。结果:(1)E15d大鼠脑组织未见MHCI类分子的表达;其他年龄组在结缔组织和脉络丛有MHCI类分子的表达。其中从P0～P14d组海马和顶叶大脑皮质神经元上MHCI类分子的表达随着日龄的增加而逐渐升高,P14d组的表达最高(P<0.05);P28d组的表达明显下降(P<0.05);老年组大鼠海马神经元上的表达再次升高(P<0.05)。(2)E15d组大鼠脑组织中未见MHCII、CD4和CD8分子的表达;其他年龄组MHCII、CD4和CD8除在结缔组织和脉络丛表达外,在神经元上未见表达。(3)各年龄组在小脑皮质神经元、黑质、纹状体等区域未见MHCI、MHCII、CD4和CD8分子的表达。结论:(1)胚胎后期脑内开始有MHC、CD4和CD8分子的表达,表明此时脑内已有可能出现免疫反应;(2)在新生鼠及出生早期,部分皮质和海马神经元上有MHCI类分子的表达,可能与调节神经元活动和突触可塑性连接有关;(3)老年组大鼠MHCI类分子在海马神经元上的表达升高,可能与神经元的功能衰退和记忆识别能力下降有关。第二部分帕金森病大鼠及NSCs联合GDNF移植后脑组织中MHC、CD4及CD8分子的表达变化目的:观察帕金森病(PD)模型大鼠黑质区MHCI、CD4和CD8免疫分子的表达变化和NSCs联合GDNF移植后移植区及免疫反应情况并探讨其意义。方法:(1)将成功PD模型大鼠按时程分为3组:7d、14d、30d,每组4只,共12只;正常对照组4只。经灌流取材冰冻切片,选取黑质区,采用免疫组化染色方法,检测MHCI、CD4和CD8分子表达情况。(2)取E14.5d大鼠腹侧中脑(VM)组织进行原代培养,经鉴定为具有增殖分化能力的神经干细胞(NSCs)后,制备成细胞悬液,加入GDNF,借助脑立体定位仪移植到PD模型大鼠的纹状体内。按照移植的时间点分为3组:2w、4w、6w,经行为学检测后分批处死,每组4只,共12只;假手术对照组4只。经灌流取材冰冻切片,选取移植针道区及周围区域,采用免疫组化染色方法,检测MHCII、CD4和CD8分子表达情况。结果:(1)PD模型大鼠组,MHCI、CD4和CD8阳性细胞在黑质区可见表达,与7d组比较,14d和30d组这三种阳性细胞的表达量升高(P<0.05),其中30d组最高(P<0.05);纹状体区未见表达;对照组在各时间点均未见上述三种阳性细胞。(2)NSCs联合GDNF组,移植区及周围区域MHCII、CD4和CD8阳性细胞在2w组大量出现,4w组下降(P<0.05),6w组基本消失(P<0.05);假手术对照组在各时间点均未见阳性细胞。结论: PD模型大鼠损毁侧黑质区存在MHCI、CD4和CD8阳性细胞,可能是由于该侧多巴胺(DA)能神经元受到损害,引起周围的小胶质细胞等激活表达MHCI分子,同时出现T淋巴细胞渗透进入黑质区,因为这三种阳性细胞的表达变化与同时期DA能神经元丧失程度呈正相关。NSCs更多还原

【Abstract】 Part 1:Expression of MHC、CD4 and CD8 molecules in rat brain tissues of differential ages and its significanceObjective: To investigate the expression of MHC、CD4 and CD8 molecules in rat brain tissues of differential ages and its significance.Methods: The expression of MHC、CD4 and CD8 molecules in rat brain tissues of E15、E19、P0、P7、P14、P28 and aged rat was detected with immunohistochemistry.Results: 1、MHCI molecules was detected in connective tissues、choroid plexus from E19 to aged rat. MHCI molecules also expressed in the neurons of hippocampus and neocortex during the first weeks after born with a trend of stable increase from P0 to P14,P14 at the summit (P<0.05), then that of P28 decreased (P<0.05) and that of the aged hippocampus of rat brain increased again (P<0.05). 2、MHCII、CD4 and CD8 molecules were only detected in connective tissues、choroid plexus without obvious change during all the period (p>0.05) except E15d. 3、MHCI、MHCII、CD4 and CD8 molecules was not observed in cerebellum cortex .Conclution: MHCI molecules expressed in the neurons of hippocampus and neocortex partly during the first weeks after born with a trend of stable increase from P0 to P14, P14 at the summit, it is possibly related with neuronal activity and plasticity of synapses. High MHCI expression of aged rat hippocampus possibly implies it is related with neurological disorders , declination of memory and recognition.Part2:Expression of MHC、CD4 and CD8 molecules in PD rat brain and co-graft of neural stem cells and GDNF for Parkinson disease.Objective :to investigate the occurrence of immunological rejection in PD brain tissue and in co-graft of NSCs and GDNF for PD.Methods:(1)The expression of MHCI、CD4and CD8 molecules in PD rat brain tissues of 7d、14d and 30d was detected with immunohistochemistry .(2)NSCs and GDNF were grafted into the striata of Parkinson’s disease model rats. Surviving animals were sacrificed at 2,4and 6w after transplantation,and the brain tissues were stained with immunohistochemistry to inspect the expression of MHCII、CD4 and CD8 molecules.Results: (1)The numbers of MHCI、CD4 and CD8 positive cells in PD rat brain tissue were low at 7d,but increased at 14d and 30d(P<0.05),and 30d at the summit(P<0.05).(2)High MHCII、CD4 and CD8 expression were observed within and around the grafts at 2w after grafting compared with control.These expressions then decreased at 4w (P<0.05) and 6w only few immunopositive cells remained.Conclusions: (1)MHCI、CD4 and CD8 positive cells were found in PD rat brain tissue,it is maybe because that many dopamine neurons were damaged,activated glial cells around expressed MHCI increased,and T cells entered the damaged postions .The更多还原