41例女性慢性输血后丙型肝炎10～15年后的感染状态

目的　了解我国女性输血后丙型肝炎病毒 (HCV)感染的慢性化规律和影响因素。方法　对河北省固安县 1989～ 1993年 4 1例女性慢性输血后丙型肝炎患者的现状进行调查 ,包括临床表现 ,血清生物化学指标 ,病毒学标志检测及B型超声检查。其中 ,HCVRNA的测定采用荧光定量PCR方法 ,抗 HIV ,抗 HCV和HBsAg测定采用酶联免疫吸附试验。结果　 4 1例女性丙型肝炎患者平均年龄 (40± 7)岁 ,随访时间 10～ 15年 ,HCVRNA间隔半年两次检测 ,自然阴转率为 19 5 1% (8 4 1)。 30例(73% )现在有症状 ,以乏力最为常见 (77% )。总的丙氨酸转氨酶 (ALT)和 (或 )天冬氨酸转氨酶 (AST)异常率为 32 % (13 4 1) ,均为轻度异常 ,无中度和重度。B超轻度慢性肝炎占 83% (34 4 1) ,中度占 17%(7 4 1) ,无重度。结论　平均感染 (13± 1)年的女性输血后慢性丙型肝炎患者多数肝脏炎症轻微慢性感染进程中有部分感染者出现HCVRNA自发阴转。     

血清HCV RNA检测在预防输血后丙型肝炎中的意义

目的 探讨血清HCV RNA检测在预防输血后丙型肝炎中的意义。方法 用ELISA法检测2000年1月至2003年12月末抗．HCV阴性的全血标本56400份次，再进行RT-PCR（荧光适时技术）检测HCV RNA，并对输过HCV RNA筛查阴性血液的患者进行追查。结果 HCVRNA阳性率为2．5‰（146／56400）。对输过HCVRNA筛查阴性血液的患者追查结果，无一例发生输血后丙型肝炎。结论 对抗-HCV阴性的血液标本用敏感的试剂进行HCV RNA检测在预防输血后丙型肝炎中不但有效，而且可行。     

The Immune Reactivity Role of HCV-Induced Liver Infiltrating Lymphocytes in Hepatocellular Damage

Liver infiltrating lymphocytes (LIL) were isolated from HCV-positive (+) and HCV-negative (–) end-stage livers. Phenotypic analysis and functional studies using proliferative and lymphocytotoxic assays were performed with the isolated LIL. Two CD3+ lymphocyte populations were found in LIL using FITC anti-CD3 monoclonal antibodies (mAb). One was a bright fluorescence intensity population (as in PBL), and the other dim. We calculated the number of FITC-anti-CD3 mAbs bound per lymphocyte on PBL and LIL and found 80,040 ± 4628 and 39,615 ± 3932, respectively. Therefore, HCV+ and HCV– patient PBL contained approximately twice the number of CD3 molecules per cell than patient CD3+ LIL. LIL also contained approximately a threefold higher concentration of TCR+, CD4–CD8–, and CD56,16 (NK) cells than the patient PBL. Thus, a major subset of LIL is phenotypically similar to mouse NK1.1+ intermediate T cells. LIL freshly isolated from HCV+ livers exhibited weak CTL activity against EBV- or Con A-transformed lymphoblast targets infected with vaccinia–HCV recombinant virus (rHCV) or primary hepatocyte cultured cells. However, after in vitro coculture of LIL with rHCV, these cells developed a strong cytotoxicity for the above targets. In contrast, LIL from HCV– livers were not cytotoxic against the same targets. Histochemical studies (in situ) demonstrated that these hepatocytes express CD95, and stains demonstrated apoptosis. The HCV+ hepatocytes also express class I MHC molecules and ICAM-1. The addition of mAb specific for these adhesion molecules inhibited CML activity. Short-term cultured hepatocytes (targets) from HCV+ and HCV– patients produced low levels of cytokines IL-1, IL-2, IL-6, TNF, and IFN- but a high level of IL-8. It is speculated that LIL expressing reduced numbers of CD3 molecules may even function as immune regulators as proposed for intermediate T cells in mice.     

重型乙型病毒性肝炎肝组织内T淋巴细胞亚群的观察

本文用抗CD_3,CD_4,CD_8McAb和ABC法检测21例重型乙肝患者肝大片,亚大片坏死区炎性浸润细胞中的T细胞亚群,CD_3~+细胞>70％,其中主要为CD_8~+细胞,CD_4~+细胞减少,CD_4~+/CD_8~+比值显著下降,与急黄肝和慢活肝比较差异显著。相反,非HBV感染性肝病患者非T细胞和CD_4~+细胞增加,CD_4~+/CD_8~+比值>1。提示重肝时T细胞可能参与了肝损伤,CD_8细胞亚群可能是介导肝细胞坏死的重要因素之一。还观察到相当数量的淋巴细胞和肝细胞HLA—DR抗原阳性,淋巴细胞与膜型HBAg(+)肝细胞密切接触。     

利用双扩增聚合酶链反应检测非甲非乙型肝炎患者血清中丙型肝炎病毒RNA

利用丙型肝炎病毒（ＨＣＶ）５’－端序列合成两对引物，建立了灵敏、特异的ＨＣＶＲＮＡ双扩增聚合酶链反应检测方法。用此方法及第二代Ａｂｂｏｔｔ酶联抗－ＨＣＶ检测试剂盒，检测了４４例非甲非乙型肝炎患者血清及１０名抗－ＨＣＶ阴性健康人。在４４例患者中，４１例（９３％）ＨＣＶＲＮＡ阳性，３６例（８２％）抗－ＨＣＶ阳性，３３例（７５％）ＨＣＶＲＮＡ、抗－ＨＣＶ全部阳性。３例ＨＣＶＲＮＡ阴性，但抗－ＨＣＶ阳性，另外，有８例抗－ＨＣＶ阴性，ＨＣＶＲＮＡ阳性。１０名健康人ＨＣＶＲＮＡ均为阴性。结果表明，大部分（９２％）抗－ＨＣＶ阳性患者带有ＨＣＶ，但为了检测所有病毒血症患者，抗－ＨＣＶ检测是不够的，利用双扩增ＰＣＲ方法检测ＨＣＶＲＮＡ对于抗－ＨＣＶ阴性患者的诊断是非常有用的。     

Metabolism of phenylalanine in liver diseases

Summary 1. Amethod for the determination of phenylalaninehydroxylase-activity in needle biopsy material of human liver was developed and tested. The kinetic data of the enzyme were determined. TheK _m for the substrate phenylalanine is 1,32 mM, for the cofactor 0,08 mM.2. The activity of phenylalaninehydroxylase was determined in biopsies from normal liver, liver cirrhosis, alcoholic hepatitis and other liver diseases. In all liver diseases the enzyme activity related to wet weight or DNA is reduced. The capacity for the hydroxylation of phenylalanine (of a cirrhotic liver) amounts to about 20% of normal liver.3. After an oral load with L-phenylalanine (100 mg/kg) the phenylalanine- and tyrosine-concentration in blood plasma was followed for 5 hours. Patients with liver cirrhosis or acute hepatitis show significant higher concentrations of phenylalanine, and significant lower concentrations of tyrosine than normal persons.4. From the decline of phenylalanine-concentration after the maximum the phenylalanine-elimination rate and from the increase of tyrosine concentration the tyrosine-production rate were calculated. Between phenylalanine-elimination rate and tyrosine-production rate a strong correlation exists. In patients with cirrhosis or acute hepatitis significantly reduced phenylalanine-elimination rate and tyrosine production rate were found compared with persons without liver disease. Patients with alcoholi hepatitis show no significant difference compared with normal persons.5. There is no correlation between the activity of phenylalaninehydroxylase in liver tissue and phenylalanine-elimination rate and tyrosine-production rate, respectively, calculated from the concentration of this amino acids in serum after a phenylalanine load.6. We conclude from these findings that the increased serum concentration of phenylalanine in some liver diseases, especially liver cirrhosis, can partly be explained by the diminished metabolism of phenylalanine in the liver. Portocaval shunts probably contribute to the elevated level of phenylalanine in serum.Mit Unterstützung der Deutschen Forschungsgemeinschaft und der Sandoz-Stiftung für medizinische Forschung     

慢性乙肝患者NK,ADCC活性及其对IFN—α,IFN—γ的反应

本文用Hela细胞乳酸脱氢酶释放法测定了42例慢性乙肝病人外周血NK、ADCC活性及其对IFN-α100U/ml、500U/ml及IFN-γ500U/ml预处理4小时后的反应。结果显示,病人PBL NK,ADCC活性与正常对照有显著差别,经IFNs处理后,其NK或/和ADCC活性均有不同程度的升高反应,从而提示了IFNs在慢性肝病治疗中的重要作用。     

乙/丙型肝炎病毒双重感染患者前C区终止变异低频率

目的了解乙型肝炎病毒（ＨＢＶ）与丙型肝炎病毒（ＨＣＶ）双重感染患者前Ｃ区基因变异，及其可能的临床意义。方法用聚合酶链反应（ＰＣＲ）与限制片段长度多态性（ＲＦＬＰ）来分析２５例ＨＢＶＤＮＡ和ＨＣＶＲＮＡ均阳性（Ａ组）和３１例ＨＢｓＡｇ和ＨＢＶＤＮＡ阳性但抗－ＨＣＶ和ＨＣＶＲＮＡ均阴性（Ｂ组）的慢性肝病患者前Ｃ区密码２８终止变异（终２８）。结果ＨＢＶ和ＨＣＶ双重感染患者（Ａ组）血清ＨＢＶＤＮＡ第１次ＰＣＲ阳性率（１６％）明显低于单独ＨＢＶ感染组（６５％）（Ｐ＜０．００１）；前Ｃ终２８检出率（２８％）亦明显低于单独ＨＢＶ感染（６８％）（Ｐ＜０．００１）。结论提示双重感染患者ＨＢＶ前Ｃ终止变异低频率可能与ＨＢＶ低水平复制有关     

C基因截短的HBV复制与包装

目的 探讨C基因截短型HBV变异体的复制与包装。方法 采用分子克隆、人工定点突变等技术构建C基因截短型HBV变异体质粒，用脂质体法转染HepG2细胞，提取细胞内及培养上清液中DNA分别进行Southem杂交，PCR及实时定量荧光PCR分析。结果 经DNA测序及酶切鉴定证实C基因截短型HBV质粒载体构建成功；C基因截短型HBV为复制缺损型，与辅助质粒共转染HepG2细胞，可在细胞内及培养上清液中检测到HBV各种DNA构型；DNA定量分析提示C基因截短型HBV的包装效率较野生型HBV提高3～40倍。结论 C基因截短型HBV变异体为复制缺损型，单独转染后不能在肝细胞内包装与复制，但在缺失包装信号ε的相应辅助病毒辅助下可有效复制并包装成子代病毒颗粒分泌到胞外，且包装效率大大提高。     

Hepatitis B virus genotype C prevails among chronic carriers of the virus in Korea

Hepatitis B virus (HBV) is one of the major causative agents of chronic liver diseases in Korea. HBV has been classified into 8 genotypes by a divergence of >8% in the entire genomic sequence, and have distinct geographic distributions. There are limited data on the relevance between HBV genotypes and clinical outcomes in Korea. To investigate the clinical feature relating to HBV genotype in Korea, a total 120 serum samples with HBsAg (65 from Seoul and 55 from the other city in Korea) were obtained from each 30 chronic HBV carriers with asymptomatic carrier (ASC), chronic hepatitis (CH), liver cirrhosis (LC) and hepatocellular carcinoma (HCC). HBV genotype was determined by either enzyme-linked immunosorbent assay (ELISA) using monoclonal antibodies against genotype-specific epitopes in the preS2-region or the direct sequencing of small S gene. HBV genotypes were determined in 105 (87.5%) of 120 samples. HBV genotype C was identified in all HBV carriers with ASC, CH, LC, and HCC. Genotypes A, B, D, E, F and G were not detected in any of them. Genotype C HBV prevails predominantly among chronic carriers of the virus in Korea, irrespective of their clinical stages of liver disease and geographic origin.