【摘要】 目的　观察在移植人造血干细胞的山羊肝脏组织中是否有人特异蛋白的表达。方法　从脐带血中获取人造血干 /祖细胞 ,移植入胎山羊体内 ,产生嵌合体山羊 ;取 4头出生后 10个月的嵌合体山羊的肝脏 ,应用免疫组化技术检测其是否有人增殖细胞核抗原 (PCNA)和肝细胞特异性抗原(HSA)的表达 ,并用FACS检测山羊肝细胞标本中是否存在有人HSA阳性细胞 ;应用RT PCR技术分析嵌合体山羊的肝组织是否有人特异的肝细胞核因子 (hHNF 3β)和人血清白蛋白 (hALB)mRNA表达 ;应用荧光原位杂交 (FISH)技术鉴定嵌合体山羊肝组织的间期细胞核是否存在特异于人 17号染色体基因的杂交信号。结果　在受体山羊血循环中发现有人血细胞嵌合 ,证明移植成功。受体山羊肝脏切片中发现有人PCNA和HSA表达 ,但对照组未见 ;FACS分析结果显示在嵌合体山羊的肝脏组织中有表达人HSA的阳性细胞 ;RT PCR结果表明嵌合体山羊肝细胞表达hHNF 3β和hALBmRNA ;FISH结果显示嵌合体山羊肝组织的个别间期细胞核具有两个人特异的杂交信号。结论　人造血干细胞移植于胎山羊后可以产生嵌合型肝脏 ,其在受体羊肝组织中生长和扩增 ,分化成人的肝细胞 ,并具备了人肝细胞特异性的转录活性。更多还原

【Abstract】 Objective To investigate the expression of human specific proteins in liver tissue obtained from goats engrafted with human hematopoietic stem cells (hHSC). Methods hHSCs derived from cord blood were transplanted into fetal goats for production of chimerism. Liver specimens were obtained from four 10 month old chimeric goats and examined for their reactivity with monoclonal antibodies against human antigens: proliferating cell nuclear antigen (PCNA) and hepatocyte specific antigen (HSA). The presence of human HSA positive cells in the goat liver tissue was determined by fluorescence assisted cell sorting (FACS). The expression of human hepatocyte nuclear factor (hHNF 3β) and human serum albumin (hALB) mRNAs was determined by RT PCR. Meanwhile, FISH experiment was also performed to detect the human cells in the chimeric livers with the probe of human p17H8. Results Successful engraftment was confirmed by the detection of human blood cell chimerism in the circulation of the transplanted goats. Human PCNA and HSA were found in liver specimens of transplanted goats but not of normals. FACS analysis showed the presence of human HSA positive cells in the liver of chimeric goats. RT PCR results demonstrated that hHNF 3β and hALB mRNAs were specifically expressed in liver tissues of chimeric goats. FISH experiment showed two positive hybridized signals in some liver cells of the chimeric goats, indicating that human liver like cells were present there. Conclusion Goats engrafted with hHSC are capable to produce chimeric livers. Growth and propagation of human cells in the liver tissue of such transplanted goats were possible. Particularly, the human liver like cells in chimeric goat livers had transcriptional activity specific to human hepatocytes.更多还原