1,鲁凤民 北京大学感染病中心 北京大学医学部病原生物学系,HAV,HBV,HCV,HDV,HEV,黄热病病毒（YFV）,EB病毒（EBV）,巨细胞病毒（CMV）,单纯疱疹病毒（HSV）,风疹病毒（RV）,GBV-C（HGV）,TTV,病毒性肝炎 viral hepatitis：由病毒感染引起的肝脏炎症病变,是一组主要感染肝细胞并在肝细胞中复制、增殖，并引起急、慢性肝脏炎症病变的病毒,HAV,HBV,HCV,HDV,HEV,肝炎病毒的定义：,0.05,34.1,抗-HBc,2011年度全国法定传染病报告发病情况 ( /10万),卫生部发布2011年度全国法定传染病疫情概况 http:/www.moh.gov.cn/publicfiles/,病毒性肝炎发病情况,资料来源：卫生部法定传染病报告系统,急性丙肝,HCV感染的自然史,慢性丙肝（85%）,肝硬化（230%，20 yrs）,失代偿（6%）,肝癌（4%）,死亡,死亡,36%,www.youtube.com, Hepatitis C From A to Z,中国HCV感染的诊断和治疗现况,估计HCV感染者: 500万1000万,2011年CDC报告发病数: 173,872,仅占:1.6%3.2%,http:/www.thewellingtonliverunit.com/treatments-hepatitis-c.asp; Schlosser et al. EASL 2009; Vermehren et al. DGVS 2010; Gane. New Zealand Doctor. 2007; http:/www.hepatitiscmsg.org/hep-c-facts-stats.html.,挑战： HCV感染绝大部分并未被诊断出来,1975,Description of non-A, non-B hepatitis,1989,Identification of hepatitis C virus,Courtesy of Charlie Rice,HCV 病毒学研究的重要时间节点,丙型肝炎病毒的发现,1970s: Hepatitis A and B diagnostics Most transfusion associated hepatitis not caused by HAV and HBV or other known viral agents (NEJM 292: 767; Gastro. 69:1278; JID 139:511; Vox Sang. 44:231) non-A, non-B hepatitis (NANBH) Might be a small, enveloped virus transmissible to chimpanzees (Gastro 88:773; JID 156:636) Standard immunological methods failed to identify viral antibodies or antigens Chiron group Thought that there was not enough viral antigen in NANBH So they decided to try and concentrate the virus,High titer NANBH chimpanzee plasma,Ultracentrifugation at 104,000g for 5 hours,Extract nuclei acid, random primed cDNA into gt11 phage,Immunoscreened with NANBH patient serum,Clone 5-1-1,Larger overlapping Clone 81,丙型肝炎病毒的发现, Phage with portions of HCV genome,Infect E.coli,Phage kills E.coli and leaves plaques,Screen with HCV Patient serum,Library Screening,丙型肝炎病毒的发现,Southern Blots with Clone 81,Placenta,Infected Chimp Liver,Acute Chronic,Placenta,Clone 81 probe,IFN- probe,Clones 5-1-1 and 81 not from host genome and DNA replication intermediates related to these clones not detected,Northern Blots with Clone 81,Clones Hybridize to RNA from infected animals, RNA is single stranded, size approx 5-10,000 nucleotides,Infected Chimp Liver,Uninfected Chimp Liver,Uninfected Chimp Liver,2 4 12 g,DNaseI,RNase A,Strand 1 Clone 81 probe,Strand 2 Clone 81 probe,Viral RNA ds 81,Oligo dT,unbound,Ctrl,Western blot,Clone 5-1-1 expressed as SOD fusion protein called PS5,PS5,SOD,NANBH patient serum,Experimentally infected chimp serum,Protein from clone 5-1-1 closely associated with NANBH infections,HCV was the first virus to be isolated and characterized solely by molecular methods, ie without culture methods,黑猩猩易感,HCV病原学,Hepatitis C Genome Regions Coding for Structural & Non-Structural Proteins,5 Non-coding Region,3 Non-coding Region,Genes Coding for Structural Proteins,Genes Coding for Non-Structural (NS) Proteins,HCV Genome,Hepatitis C Genome Structural and Nonstructural Genes,C,A,NS2,NS3,E1,p7,E2,B,NS5,A,B,NS4,5 Non-coding Region,3 Non-coding Region,Genes Coding for Structural Proteins,Genes Coding for Non-Structural (NS) Proteins,HCV Genome,IRES,Internal Ribosomal Entry Site (IRES) - Consists of Domains II, III, IV of the 5 NCR and first 24-40 nucleotides of core region - IRES involved in processing of polypeptide precursor,Hepatitis C Virus Translation,HCV Genome,C,A,NS2,NS3,E1,p7,E2,B,5 NCR,3 NCR,NS5,A,B,NS4,Polyprotein Precursor,Translation,Hepatitis C Virus Protein Processing,HCV Genome,C,A,NS2,NS3,E1,p7,E2,B,NS5,A,B,Signal Peptidase,NS2/3 Protease,NS3/4A Protease,C,A,NS2,NS3,E1,p7,E2,B,NS5,A,B,NS4,Polyprotein Precursor,Protein Processing,Translation,NS4,Hepatitis C Virus Protein Processing,HCV Genome,C,A,NS2,NS3,E1,p7,E2,B,NS5,A,B,NS4,Polyprotein Precursor,Protein Processing,Translation,C,NS4B,NS5A,NS2,NS3,E1,NS4A,p7,E2,NS5B,Proteins,Protein Processing,Hepatitis C Genome Translation,C,A,NS2,NS3,E1,p7,E2,B,NS5,A,B,NS4,Polyprotein Precursor,Signal Peptidase,NS2/3 Protease,NS3/4A Serine Protease,Protein Processing Role of NS3/4A Serine Protease,C,NS4B,NS5A,NS2,NS3,E1,NS4A,p7,E2,NS5B,Proteins,Hepatitis C Virus NS3/4A Serine Protease Inhibitors,C,A,NS2,NS3,E1,p7,E2,B,NS5,A,B,NS4,Polyprotein Precursor,Signal Peptidase,NS2/3 Protease,NS3/4A Serine Protease,NS3/4A Serine Protease Inhibitors,C,NS2,NS3,E1,p7,E2,Proteins,A,B,NS5,A,B,NS4,Genomic RNA is infectious,HCV Replication,HCV Life Cycle,Typical virus life cycle: Binding, internalization, uncoating, protein expression/replication, assembly, release,HCV Replication- 病毒的入侵需要细胞膜表面的受体,HCV receptors/entry factors,Intracellular,Linear polysaccharides on proteins of all human cell surfaces,Tetraspanin superfamily member Expressed in all nucleated cells Part of B-cell receptor complex,Scavenger receptor class B member I HDL receptor (multiple other ligands) Expressed in hepatocytes, adrenal cortex, gonads (less elsewhere),