Localization of hepatitis C and G viruses in liver tissue and its significance

1.原位杂交条件采用体外转录法制备地高辛标记的RNA探针，用于原位杂交。杂交探针的核苷酸序列均选自丙型肝炎病毒和庚型肝炎病毒的NS5区。用地高辛检测试剂盒检测杂交探针。本研究采用石蜡包埋的肝脏标本。HGV在肝组织中的定位对6例慢性HGV感染患者和6例正常对照进行分析。6例HGV感染者中有2例检测到阳性信号，而6例对照组均未检测到阳性信号。2例阳性信号患者肝移植后接受免疫抑制治疗，血清HGV RNA水平较高。HGV RNA阳性链和阴性链在汇管区浸润的单个核细胞中均可检测到，但在肝细胞中未检测到。这些结果表明HGV在肝细胞中不存在复制或仅限于复制，HGV不是肝炎病毒。分析了8例慢性丙型肝炎患者和8例正常对照肝组织中丙型肝炎病毒的定位情况。6例丙型肝炎病毒感染者均检测到原位杂交阳性信号，而6例正常对照均未检测到阳性信号。阳性信号主要见于肝细胞(仅见于胞浆)，少见于胆管上皮。肝细胞内均可检测到HCVRNA的正链和负链。呈阳性信号的肝细胞多见于小叶中央部，远多于周边部。在病毒载量低和高的患者中，阳性信号的强度和位置没有差异。丙型肝炎病毒在肝组织中定位的临床意义正在研究中。

1. Condition of in-situ hybridizationDigoxigenine labeled RNA probe was prepared by in vitro transcription for in-situ hybridization. Nucleotide sequence of hybridization probe was selected from the NS5 region in both HCV and HGV. Hybridized probes were detected by DIG detection kit. Paraffin-embedded liver specimens were used in the present study.2. Localization of HGV in liver tissueSix patients with chronic HGV infection and 6 controls were analyzed. Positive signal of in-situ hybridization was observed in 2 of the 6 patients with HGV infection, while it was observed in none of the 6 controls. The 2 patients with the positive signal were under immuno-suppressive therapy after liver transplantation and had high level of HGV RNA in serum. Both positive and negative strands of HGV RNA were detected in mononuclear cells infiltrated in portal areas, but not in hepatocytes. Those results indicate that HGV replication is absent or limited in hepatocytes, and that HGV is not a hepatitis virus.3. Localization of HCV in liver tissue Eightpatients with chronic hepatitis C and 8 controls were analyzed. Positive signal of in-situ hybridization was observed in all 6 patients with HCV infection, while it was observed in none of the 6 controls. The positive signal was observed mainly in hepatocytes (only in cytoplasm), and rarely in epitherium of bile ducts. Both positive and negative strands of HCV RNA were detected in hepatocytes. Hepatocytes with the positive signal tended to be more common in central part of lobule than in its peripheral part. Strength and localization of the positive signal did not differ between patients with low and high viral load. Clinical significance of HCV localization in liver tissue is now under investigation.