新生与成年土拨鼠肝组织中部分差异表达的基因比较分析

目的新生土拨鼠感染土拨鼠肝炎病毒后，大部分发展为慢性肝炎，而成年土拨鼠感染后则多发生急性自限性肝炎。本实验目的就是寻找其肝组织中可能导致这种预后差异的关键基因。方法采用全基因组表达谱芯片技术，对比新生与成年小鼠肝组织基因表达差异，选取目的基因，再通过多个物种序列比对，设计简并引物，在土拨鼠肝组织eDNA中扩增对应基因，测序，再次设计引物，进行实时荧光定量PCR。结果与新生土拨鼠相比，成年土拨鼠肝细胞中与钙离子重吸收相关基因DNMl（Dynamin1）、DNM3（Dynamin3）及Prkcc（proteinkinaseC，gamma）表达率明显升高，分别上升2．65±0．25倍、1．90±0．34倍、2．94±0．54倍。结论在钙离子重吸收通路中，在两组小鼠肝脏中表达差异最明显的上述三个基因，在新生组与成年组土拨鼠之间也有明显差异。此类基因造成肝细胞内钙离子浓度的差别，间接影响其中肝炎病毒的复制。这种表达差异很可能是导致两个年龄段动物感染土拨鼠肝炎病毒后转归不同的原因之一。

Comparison of some differentially expressed genes in the liver of neonatal and adult woodchucks

Objective To find some key genes which may account for the different sequelae （chronic woodchuck hepatitis and acute self-limiting woodchuck hepatitis ） of woodchuck hepatitis virus infection in the neonatal and adult woodchucks. Methods Applying the genome-wide expression microarray, some related genes were selected. With the degenerate primers designed on the iCODEHOP from clustal of some different species, some products were amplified from the woodchuck cDNAs. After the products sequencing was performed, some accurately designed primers generated from the Primer Premier 5. 0, and real time quantitative PCR was finally performed. Results Compared with the neonatal woodchucks, the expression of calcium reabsorption-related genes DNM1 （ Dynamin 1 ）, DNM3 （ Dynamin 3） and Prkcc （ protein kinase C, gamma） in theadult woodchuck liver cells were significantly increased by 2. 65 ± 0. 25 times, 1.90 ± 0. 34 times, 2. 94 ± 0. 54 times, respectively. Conclusions In the endocrine and other factor-regulated calcium reabsorption pathway, these three genes show the most significant changes in mouse livers, as well as in the neonatal and adult woodchucks. Those genes contribute to the differential calcium concentration, and promote woodchuck hepatitis virus replication indirectly. In a word, these genes could have an effect on the sequelae of woodchuck hepatitis virus infection in different ages.