Ultracentrifugation of serum samples allows detection of hepatitis C virus RNA in patients with occult hepatitis C
Ultracentrifugation of serum samples allows detection of hepatitis C virus RNA in patients with occult hepatitis C
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DOI：
10.1128/jvi.02750-06
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发表时间：
2007-07-01
影响因子：
5.4
通讯作者：
Carreno, Vicente
中科院分区：
文献类型：
--
作者：
Bartolome, Javier;Manuel Lopez-Alcorocho, Juan;Carreno, Vicente
已经描述了不明原因的肝功能检查异常、抗 HCV 和血清 HCV RNA 阴性但肝脏中有 HCV RNA 的患者隐匿性丙型肝炎病毒 (HCV) 感染。由于 HCV 在这些患者的肝细胞中复制，循环病毒颗粒的数量可能低于最敏感技术的检测极限。为了证明这一假设，对 106 名隐匿性 HCV 感染患者的血清样本进行了分析。将 2 毫升血清在 10% 蔗糖垫上以 100,000 x g(av) 超速离心 17 小时，其中 av 表示平均值，并通过链特异性实时 PCR 进行 HCV RNA 检测。在 106 名患者中，62 名 (58.5%) 超速离心后可检测到血清 HCV RNA 水平，中位负荷为 70.5 拷贝/ml（范围为 18 至 192）。碘克沙醇密度梯度研究显示，HCV RNA 在密度为 1.03 至 1.04 和 1.08 至 1.19 g/ml 时呈阳性，这与经典慢性 HCV 感染患者血清中发现的结果非常相似。超速离心后血清 HCV RNA 水平可检测到的 62 名患者中有 56 名 (90.3%) 的肝脏中发现了反基因组 HCV RNA，而 44 名阴性患者中只有 27 名 (61.4%) (P < 0.001)。未发现血清 HCV RNA 的患者之间的反基因组 HCV RNA 中位量没有差异（4.5 x 10(4) [范围，7.9 x 102 至 1.0 x 10(6)] 与 2.3 x 10(4) [范围，4.0 x 10(2) 至 2.2 x 10(5)]）。两组之间的丙氨酸转氨酶和γ-谷氨酰转肽酶水平、肝脏坏死性炎症活动和纤维化没有差异。总之，通过超速离心浓缩循环病毒颗粒后，可以在隐匿性HCV感染患者的血清中检测到HCV RNA。
Occult hepatitis C virus (HCV) infection of patients with abnormal liver function tests of unknown origin who are anti-HCV and serum HCV RNA negative but who have HCV RNA in the liver has been described. As HCV replicates in the liver cells of these patients, it could be that the amount of circulating viral particles is under the detection limit of the most sensitive techniques. To prove this hypothesis, serum samples from 106 patients with occult HCV infection were analyzed. Two milliliters of serum was ultracentrifuged over a 10% sucrose cushion for 17 h at 100,000 x g(av), where av means average, and HCV RNA detection was performed by strand-specific real-time PCR. Out of the 106 patients, 62 (58.5%) had detectable serum HCV RNA levels after ultracentrifugation, with a median load of 70.5 copies/ml (range, 18 to 192). Iodixanol density gradient studies revealed that HCV RNA was positive at densities of 1.03 to 1.04 and from 1.08 to 1.19 g/ml, which were very similar to those found in the sera of patients with classical chronic HCV infection. Antigenomic HCV RNA was found in the livers of 56 of 62 (90.3%) patients with detectable serum HCV RNA levels after ultracentrifugation, compared to 27 of 44 (61.4%) negative patients (P < 0.001). No differences in the median loads of antigenomic HCV RNA between patients with an those without serum HCV RNA (4.5 x 10(4) [range, 7.9 x 102 to 1.0 x 10(6)] versus 2.3 x 10(4) [range, 4.0 x 10(2) to 2.2 x 10(5)]) were found. Alanine aminotransferase and gamma-glutamyl transpeptidase levels, liver necroinflammatory activity, and fibrosis did not differ between both groups. In conclusion, HCV RNA can be detected in the sera of patients with occult HCV infection after circulating viral particles are concentrated by ultracentrifugation.