黃怡嘉HUANG, YI-CHIA賴家鈺LAI, CHIA-YUCHIA-YULAI營養學系博士班2026-02-102024-09-0320242024-07-22https://handle.ncl.edu.tw/11296/4n23c9https://scholars.csmu.edu.tw/handle/123456789/5359根據衛生福利部統計處資料，在新冠疫情前，肝硬化為臺灣十大死因中的第十名，就各死因的平均死亡年齡，肝硬化患者約為60歲，是十大死因中平均年齡最低的，在25至44歲及45至69歲這兩個年齡層中，肝硬化為死因的第五名，足見此疾病對家庭社會的影響甚鉅。當肝臟細胞長期暴露於病毒或過量的酒精、脂肪、膽鹽等傷害，持續的慢性發炎造成肝細胞傷害及壞死，受損的肝細胞分泌細胞激素活化肝臟內的免疫巨噬細胞(庫氏細胞，Kupffer cells)，被活化的庫氏細胞進而透過細胞分泌素及發炎介質活化星狀細胞(hepatic stellate cells)，活化的星狀細胞增生並轉分化為肌纖維母細胞(myofibroblasts)，分泌產生膠原纖維及細胞外基質，大量的膠原纖維及細胞外基質堆積形成疤痕組織，導致肝臟纖維化，破壞原有的組織結構，造成門靜脈壓力上升及肝臟生理功能受損，引發門脈高壓的併發症而危害生命。肝硬化的致病過程是緩慢且持續的，慢性發炎反應及受損的肝細胞造成活性氧及活性氮的生成增加，導致體內氧化壓力上升，損耗體內抗氧化物質，使得氧化還原機制失去平衡。除了生活型態的調整及針對特定病因的藥物治療以外，補充抗氧化營養素來改善肝硬化患者體內抗氧化-氧化平衡機制，也是減緩肝硬化疾病進展的治療策略之一。穀胱甘肽為人體細胞內濃度最高的抗氧化物質，可以透過氧化還原的過程，直接清除活性氧及活性氮等氧化產物；同時它也是穀胱甘肽過氧化酶及穀胱甘肽硫轉移酶的重要輔因子，藉由此二相關抗氧化酵素的作用，可更有效率地去除自由基及毒性物質，保護細胞免於氧化壓力的傷害。肝臟為穀胱甘肽生合成最主要的臟器，肝硬化除了直接影響體內的穀胱甘肽生成調節以外，疾病所伴隨的免疫發炎反應及氧化壓力上升，更可能加劇穀胱甘肽的耗用。因此，肝硬化患者的穀胱甘肽與發炎反應、氧化壓力、抗氧化能力和臨床疾病結果的相關性值得被檢視。本研究以橫斷面研究模式進行，比較不同疾病程度的肝硬化患者體內發炎反應、氧化壓力及抗氧化能力，並探討肝硬化患者的發炎反應及氧化壓力指標與血漿穀胱甘肽濃度的關係。於臺中榮民總醫院一般外科、肝膽腸胃科及健康中心分別募集75位肝硬化患者以及110位健康受試者。記錄受試者的基本資料，包括：年齡、性別、抽煙及飲酒習慣)，肝硬化患者有無服用Silymarin治療；測量受試者的身高、體重、收縮壓及舒張壓(休息超過五分鐘後)。於約定時間採集受試者的空腹血液，血液樣本用以分析白血球數目及分類計數、凝血酶原時間、白蛋白、總膽紅素、麩胺酸丙酮酸轉胺酶、肌酐酸、高敏感度C-反應蛋白、脂質過氧化指標(丙二醛)、還原型穀胱甘肽、氧化型穀胱甘肽、總抗氧化能力及穀胱甘肽相關抗氧化酵素活性(穀胱甘肽過氧化酶、穀胱甘肽硫轉移酶、穀胱甘肽還原酶)。結果顯示肝硬化患者體內的發炎反應與抗氧化能力在疾病嚴重度進展時存在顯著的協同作用關係。同時我們也觀察到肝硬化疾病嚴重程度較低的CTP class A組患者，相較於健康對照組及疾病嚴重程度較高的class B-C組患者，其體內有較高濃度的還原型穀胱甘肽、氧化型穀胱甘肽、還原型穀胱甘肽與氧化型穀胱甘肽比例，以及較高活性的穀胱甘肽過氧化酶。此研究結果除了意味著慢性發炎反應會增加體內穀胱甘肽的週轉使用，用以平衡消弭上升的氧化壓力，也反映出肝硬化患者隨著疾病嚴重程度進展，加劇肝臟細胞的傷害，影響穀胱甘肽生合成及調節的功能。穀胱甘肽生合成過程的速率決定步驟受到體內半胱胺酸含量的調節。半胱胺酸的自體生合成則是透過轉硫作用，將同半胱胺酸縮化水解形成半胱胺酸，轉硫途徑的反應皆需維生素B-6作為輔酶。當肝硬化患者體內維生素B-6不足，亦會間接影響穀胱甘肽生合成及其相關的抗氧化酵素活性。因此進一步以雙盲隨機介入研究模式，給予肝硬化患者單一補充或同時補充穀胱甘肽或維生素B-6，以評估及比較介入後對氧化壓力程度、抗氧化能力以及疾病嚴重度的影響。於臺中榮民總醫院一般外科及肝膽腸胃科募集61位肝硬化患者，雙盲隨機分配至安慰組 (服用安慰劑)、維生素B-6組 (服用50 mg/d 磷酸比哆醇，pyridoxine-hydrochloride)、穀胱甘肽組 (服用500 mg/d左旋穀胱甘肽)或維生素B-6合併穀胱甘肽組 (服用50 mg/d磷酸比哆醇及500 mg/d左旋穀胱甘肽)，受試者被要求每天服用補充劑，以雙盲形式介入十二週。記錄受試者的基本資料，包括：年齡、性別、抽煙及飲酒習慣；採集受試者在介入前及介入十二週後的空腹血液，並於每次抽血時測量受試者的身高、體重、收縮壓及舒張壓(休息超過五分鐘後)。空腹採集的血液樣本用以分析白蛋白、總膽紅素、凝血酶原時間、脂質過氧化指標(丙二醛)、半胱胺酸、磷酸比哆醛(維生素B-6指標)、還原型穀胱甘肽、氧化型穀胱甘肽、總抗氧化能力及抗氧化酵素活性(穀胱甘肽過氧化酶、穀胱甘肽還原酶、穀胱甘肽硫轉移酶、過氧化氫酶、超氧化物歧化酶)。 結果顯示每日服用50毫克磷酸比哆醇的維生素B-6組及維生素B-6合併穀胱甘肽組在介入第12週後的血漿磷酸比哆醛濃度有顯著上升。然而，每日服用500毫克左旋穀胱甘肽的穀胱甘肽組及維生素B-6合併穀胱甘肽組的血漿穀胱甘肽濃度及相關的抗氧化酵素活性，在介入12週後並無顯著變化，僅是穩定持平。以多重迴歸分析顯示，無論是補充維生素B-6或穀胱甘肽對肝硬化患者的氧化壓力及抗氧化能力指標皆未有顯著影響。由於補充維生素B-6或穀胱甘肽未能顯著改善肝硬化患者的血漿穀胱甘肽濃度及相關的抗氧化酵素活性，我們進一步追蹤肝硬化患者在補充劑介入前後及介入完成後的各個時間點的疾病嚴重度變化。結果顯示肝硬化患者血漿穀胱甘肽濃度和還原型穀胱甘肽與氧化型穀胱甘肽比例以及穀胱甘肽相關的抗氧化酶活性，在補充劑介入前、中、後，皆與疾病嚴重度指標(CTP分數)存在顯著的負相關。但血漿磷酸比哆醛、半胱胺酸及丙二醛濃度與CTP分數皆未有顯著相關性。總結上述兩個研究，肝硬化患者體內的發炎反應與抗氧化能力在疾病嚴重度進展時存在顯著的協同作用關係。儘管穀胱甘肽及維生素B-6補充劑的介入未能改善肝硬化患者的氧化壓力及提高抗氧化能力，肝硬化患者在補充劑介入前、中、後的血漿穀胱甘肽濃度、還原型穀胱甘肽與氧化型穀胱甘肽比例及穀胱甘肽相關的抗氧化酶活性，皆與疾病嚴重度(CTP分數)存在顯著的負相關。In Taiwan, according to the Ministry of Health and Welfare, chronic liver disease and cirrhosis remained the 10th most common cause of death before the pandemic of COVID-19. The median age of death from cirrhosis was around sixty, the youngest on the list of the top ten causes of death, while cirrhosis was escalated to the 5th cause of death for the age groups of 25 to 44 and 45 to 69, those who were supposed to be the mainstay of the family. A variety of chronic liver diseases contribute to the development of cirrhosis, including chronic infection of hepatitis B or C virus, excess alcohol consumption, metabolic dysfunction-associated fatty liver disease, autoimmune liver disease, chronic disease of biliary tracts, and genetic predisposition disorders. On exposure to these insults, persistent inflammation resulted in hepatocyte damage and apoptosis. Injured hepatocytes produce cytokines and chemokines to activate liver-resident macrophages, Kupffer cells, which in turn activate the hepatic stellate cells via cytokines and mediators. On activation, the hepatic stellate cells proliferate and transdifferentiate to myofibroblast, producing collagen and extracellular matrix. Accumulation of excessive amounts of collagen and extracellular matrix precipitates the formation of scar tissue, which leads to the disruption of the normal liver architecture, the development of portal hypertension, liver function impairment, and life-threatening complications. The fibrogenesis of cirrhosis is dynamic and insidious and during the process, oxidative stress increases due to the overproduction of reactive oxygen species (ROS) and reactive nitrogen species (RNS) and disrupts the redox balance. Besides withdrawing or eliminating causative agents of conditions as well as dietary and lifestyle modification, antioxidant supplementation is considered a therapeutic perspective to restore the imbalanced redox status and mitigate the disease.Glutathione (GSH) represents the most abundant intracellular antioxidant to protect organisms from deleterious oxidative stress, which not only quenches the ROS and free radicals directly but also acts as the essential cofactor of two important relevant antioxidant enzymes, glutathione peroxidase (GPx) and glutathione S-transferase (GST), to detoxify the exogenous and endogenous toxins more effectively. The liver produces a remarkable amount of GSH and releases it into plasma and bile, which is crucial for interorgan GSH homeostasis. Liver cirrhosis affects the endogenous generation and distribution of GSH, and the accompanying persisting inflammation and oxidative stress can further aggravate the depletion of GSH. Therefore, the inflammatory response and GSH status on the severity of liver cirrhosis and the relationship between these two factors are worth investigating.We conducted a cross-sectional study to measure and compare inflammatory markers and GSH status of patients in different stages of liver cirrhosis and explore the relationship among inflammation, oxidative stress, and antioxidant capacity in patients with cirrhosis. Results showed a significant interaction between levels of serum high sensitivity C-reactive protein (hs-CRP) and plasma GSH were found, suggesting possible synergistic effects of GSH and hs-CRP in the progression of liver cirrhosis. In addition, patients in CTP class A had significantly higher levels of GSH, GSSG, GSH/GSSG ratio, and activity of GPx when compared to patients in CTP class B-C. Our finding not only implies that the increased oxidative stress from chronic inflammation may enhance the utilization and turnover of GSH but also reflects the biosynthesis and homeostasis of GSH are affected alongside the progression of cirrhosis, considering more hepatocytes were injured.The rate-controlling step of GSH biosynthesis is modulated by the amount of cysteine. De novo synthesis of cysteine comes from the transsulfuration of methionine catabolism via condensation and hydrolyzation reactions catalyzed by the vitamin B-6 dependent enzymes. Inadequate vitamin B-6 status in patients with liver cirrhosis would limit GSH synthesis and its relevant antioxidant capacities. The double-blind, randomized controlled trial and follow-up study were conducted to investigate the effectiveness of vitamin B-6 (pyridoxine-hydrochloride, 50 mg/d), GSH (500 mg/d), or combined supplementation on oxidative stress and antioxidant capacity of cirrhotic patients. Results indicated that neither vitamin B-6 nor GSH supplementation had significant effects on the indicators of oxidative stress and antioxidant capacities in cirrhotic patients. Nonetheless, there were significant negative associations between the GSH level, GSH/GSSG ratio, related antioxidant enzyme activity, and CTP score before, during, and after the supplementation in our cirrhotic patients.To sum up our studies, in patients of liver cirrhosis, possible synergistic effects of GSH and the inflammatory marker, hs-CRP, were noted in the progression of the disease. Although GSH or a large dose of vitamin B-6 supplementations had non-significant beneficial effects on oxidative stress and antioxidant capacities, the negative associations of GSH level, GSH/GSSG ratio and antioxidant enzyme activity, and disease severity score of cirrhosis were revealed.肝硬化穀胱甘肽維生素B-6發炎反應氧化壓力抗氧化能力CirrhosisGlutathioneVitamin B-6InflammationOxidative stressAntioxidant capacitiesthesis