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RNA探针进行原位杂交。从HCV和HGV的NS5区选择杂交探针的核苷酸序列。杂交探针采用DIG检测试剂盒检测。本研究采用石蜡包埋肝脏标本。分析6例慢性HGV感染患者和6例对照者的肝组织定位。6例HGV感染患者中2例观察到原位杂交阳性信号，而6例对照均未观察到原位杂交阳性信号。2例阳性信号患者均为肝移植术后免疫抑制治疗，血清HGV RNA水平较高。在门脉区浸润的单核细胞中检测到HGV RNA的阳性和阴性链，而在肝细胞中未检测到。这些结果表明HGV在肝细胞中不存在或有限的复制，并且HGV不是肝炎病毒。分析8例慢性丙型肝炎患者和8例对照者的肝组织定位。6例HCV感染患者均观察到原位杂交阳性信号，而6例对照组均未观察到原位杂交阳性信号。阳性信号主要见于肝细胞（仅见于细胞质），很少见于胆管上皮。在肝细胞中检测到HCV RNA的阳性链和阴性链。呈阳性信号的肝细胞多见于小叶中部而非周围。阳性信号的强度和定位在低病毒载量和高病毒载量的患者之间没有差异。丙型肝炎病毒在肝组织定位的临床意义目前正在研究中。

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.