摘要:全球約有1.8億人口為慢性C型肝炎病毒帶原者。根據世界衛生組織估計,每年有超過六百四十萬以上的人遭受到急性C型肝炎病毒感染。急性C型肝炎病毒感染後有50-80%的患者會轉變為慢性C型肝炎病毒感染,慢性C型肝炎病毒感染者中有五分之一的人最終會變成肝硬化,之後每年有1-5%的患者最後會產生肝細胞癌。因此C型肝炎病毒感染是一個影響全球醫療、公共衛生與社會經濟的重要健康課題。C型肝炎病毒感染除了造成肝臟的發炎外,還會影響身體其他的器官與系統,造成C肝患者各種不同的肝外臨床表現(extrahepatic manesfications),其中在近年來最令人注目的是C肝患者與糖尿病及胰島素抗性等臨床表現的相關性。特別在生活型態改變所引起的肥胖症及糖尿病等代謝性疾病的全球大流行,以及代謝性肝臟疾病盛行率大幅增加後,C肝病毒與代謝疾病間的交互作用已成為肝臟醫學研究領域中的顯學。流行病學上除了發現C肝病毒感染會有胰島素抗性或是第二型糖尿病的臨床表現外,也發現慢性C肝患者的糖尿病盛行率較其他肝臟疾病患者的盛行率為高,糖尿病患者的C肝抗體盛行率也較一般族群高。吾人及他人之研究也發現,慢性C肝患者常有較高的胰島素抗性,胰島素抗性為預測C肝患者肝臟組織纖維化進展的獨立因子。同時伴有胰島素抗性或是第二型糖尿病的C肝患者,常會有較嚴重程度的肝臟纖維化,並且對於抗病毒治療的反應較差。基因轉殖動物與細胞實驗的研究則明白指出,C肝病毒感染為引起胰島素抗性的重要獨立因子,C肝病毒的核心蛋白會引起胰島素抗性,並且可能藉由增加肝內TNF-α濃度來引起胰島素抗性。另外最近研究還發現,CD4+輔助T細胞(helper T cell)分泌的介白素21(interleukin-21, IL-21)與受體(IL-21R)與糖尿病相關聯並且可能與C肝病毒轉變為慢性感染的過程有關。由於西方國家基因型第一型之C肝患者接受目前標準治療長效型干擾素合併口服雷巴威林共48週只有約50%的痊癒率,因此許多學者不斷努力尋找與治療有關或是可以用來評估治療反應的相關因子,並研究其可能的相關機轉,希望利用這些機轉來改善當前C肝的治療方式。應用C型肝炎病毒與胰島素抗性及脂肪激素的相關性,藉由改變胰島素抗性及代謝反應或是調控相關脂肪激素來影響C肝病毒感染與複製,進一步來改善現今C肝治療,就成為可行以及重要的研究題目。另一方面,日本Akuta等學者在2005年發現C肝基因型第ㄧ型病毒核心蛋白基因序列多形性與C肝患者抗病毒治療的效果有關。最近的研究更指出,C肝基因型第ㄧ型病毒核心蛋白基因多形性與患者產生肝癌亦有重要的相關性。吾人亦針對台灣C肝基因型第ㄧ型病毒核心蛋白基因序列多形性與臨床因子的相關性進行研究。初步的研究發現,不同的病毒核心蛋白基因序列多形性似乎與宿主的胰島素抗性及脂肪指標有關係。綜上所述,吾人之假說為C肝基因型第ㄧ型病毒核心蛋白基因多形性可能與胰島素抗性及及脂肪激素有交互作用存在。基於此項假說及初步研究發現,吾人欲詳細探討C肝患者介白素21 及基因型第ㄧ型病毒核心蛋白基因多形性對患者治療反應,胰島素抗性、葡萄糖及脂質代謝、脂肪激素等狀態之影響,希望能從另一個角度切入,找出減緩甚至改善C肝患者肝臟組織損傷的機轉,進一步改善C肝患者的治療成效。在此,吾人提出三年之研究計畫。第一年將以病例對照的研究設計(case-control study),比較100例慢性C肝基因型第ㄧ型病毒感染達持續病毒反應者(sustained virologic response, SVR)與100位無持續病毒反應(non-SVR)之對照組,在治療前血中IL-21量、IL-21/IL-21R相關基因多形性及單點核酸變異偵測(single nucleotide polymorphism, SNP)上是否有差異,並進一步分析其差異是否會影響患者治療反應、胰島素抗性或代謝和脂肪激素。第二年將收集300例接受長效型干擾素與雷巴威林合併治療的慢性C肝第ㄧ型病毒感染者,比較不同C肝基因型第ㄧ型病毒核心蛋白基因多形性對C肝患者抗病毒治療反應是否有差異,以及其差異是否會影響患者治療前之臨床因子(包括病毒學、組織學以及生化學相關因子)、胰島素抗性、代謝、脂肪激素和IL-21等。第三年將著重於細胞生物學的研究,利用帶有不同C肝基因型第ㄧ型病毒核心蛋白基因多形性的載體來轉染肝癌細胞株,以這些細胞株為實驗平台進行細胞內胰島素訊息傳遞途徑、糖類代謝途徑、脂肪代謝途徑、脂肪激素及細胞激素訊息傳遞途徑(特別是IL-21與IL-21R)與C肝病毒核心蛋白交互作用之研究,以了解不同C肝基因型第ㄧ型病毒核心蛋白多形性與胰島素抗性、糖類、脂肪代謝、脂肪激素或細胞激素間交互作用的機轉。
Abstract: Chronic hepatitis C virus (HCV) infection affects more than 180 million people worldwide, and is an important etiology of chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). The clinical manifestations of chronic HCV infection vary widely. In addition to hepatic injury, several extrahepatic disorders have been linked to HCV infection. Among them, recent attention is focused on the interactions between chronic HCV infection and metabolic derangements, including glucose, lipid, insulin resistance, adipokines and cytokines. Ample experimental, clinical and epidemiological data all demonstrate a close linkage between chronic HCV infection and metabolic derangements. Chronic HCV infection has been shown to be an independent predictor of metabolic abnormality such as insulin resistance, steatosis and diabetes mellitus. In addition, metabolic profiles can also affect treatment outcomes in chronic hepatitis C patients. On the other hand, interleukin-21 (IL-21) is known to be associated with the pathogenesis of type 1 diabetes, and is rerequired to control chronic viral infection, such as chronic HCV infection.To eradicate HCV infection or to attain a sustained virological response (SVR) in CHC patients, peginterferon (PEG-IFN) plus ribavirin (RBV) combination therapy is the current standard of care. However, the regimen has many undesirable side effects, is expensive and not effective in a substantial proportion of patients with HCV genotype 1 (GT1) infection, especially in Western countries. Therefore, identifying patients who are difficult to achieve SVR before or during the therapy is clinically important. In our clinical practice, several baseline and on-treatment factors have been used to predict the disease progression and therapeutic response in CHC patients. They are viral factors, host factors, metabolic factors, histological factors, type of regimens, and duration of infection. In 2005, Akuta et al. identified the substitution of amino acids (aa) 70 and/or 91 in the HCV core region (HCV-CR) as a new independent and significant negative predictor associated with virological responses, and also a risk factor of hepatocellular carcinoma. In Taiwan, we evaluated the distribution of amino acid substitution patterns in a pilot study of 53 HCV genotype 1 (GT1) CHC patients treated with combination therapy and found that patients with different amino acid substitutions on the HCV-CR had different HOMA-IR levels as well as lipid profiles. Considering the reported associations among metabolic derangements, therapeutic response to combination therapy and substitution of amino acids (aa) 70 and/or 91 in HCV-CR, we therefore hypothesize that the substitution or polymorphism of amino acids in HCV-CR may affect the response to combination therapy through their actions on metabolic profiles of HCV patients.Herein, we propose a 3-year study to examine the association and underlying mechanisms by which substitution or polymorphism of amino acids (aa) 70 and/or 91 in HCV-CR interacts with host metabolic factors, cytokines and adipokines by using clinical and molecular methods. In the first year study, by using the case-control study design, we will compare the differences of serum IL-21 level and single nucleotide polymorphisms (SNPs) of IL-21/IL-21R between 100 GT1 CHC patients with SVR and 100 patients without SVR, and examine the impact of these polymorphisms on the therapeutic response, insulin resistance, adipokine and metabolic profiles of CHC patients. In the second year study, we will examine the impacts of HCV core gene polymorphisms on the antiviral response in 300 CHC GT1 patients by comparing these polymorphisms between responders, relapsers and non-responders. Further investigation will be done to clarify the interactions among insulin resistance, adipocytokine, IL-21 and metabolic profiles. In the third year study, we will explore the mechanisms involved in HCV core gene polymorphism and metabolic derangements by comparing the expression of genes (RNA/protein) associated with lipid, glucose metabolism, insulin, adipokine and cytokine signaling pathway in cell lines transfected with plasmids harboring different amino acid substitutions in HCV core protein. The effects of various interventions (e.g.: chemicals or siRNA) on the expression of these genes will also be performed.