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Please use this identifier to cite or link to this item: http://ntour.ntou.edu.tw:8080/ir/handle/987654321/30939

Title: Clozapine透過NF-кB誘導肝癌細胞株HepG2產生發炎反應機制之探討
Clozapine-Induced HepG2 Inflammation via NF-кB
Authors: Hao-Ting, Chien
簡豪廷
Contributors: NTOU:Institute of Bioscience and Biotechnology
國立臺灣海洋大學:生物科技研究所
Keywords: 抗精神病藥物;發炎反應
clozapine;NF-kB;HepG2;inflammation
Date: 2012
Issue Date: 2012-04-16T03:13:18Z
Abstract: 第二代抗精神病藥物有很好的臨床治療效果,但易導致體重的增加。clozapine為臨床最後一線抗精神病藥物,然而引發體重的增加也最為嚴重。肥胖常誘導組織及細胞發炎,而增加糖尿病、癌症等疾病的罹患率。許多文獻指出抗精神病藥物與組織發炎反應具相關性,而NF-kB (nuclear factor-kB)為發炎反應的主要調控者。故本論文以人類肝癌細胞株HepG2探討clozapine是否透過NF-kB造成發炎反應。 初步結果顯示, clozapine處理肝癌細胞株HepG2,會造成發炎因子NF-кВ (nuclear factor-κB p65 Ser536)磷酸化增加,IкВa(inhibitor of nuclear factor-кBa)蛋白量降低,利用real-time PCR證明clozapine會造成發炎細胞素IL-1B、IL-6 mRNA表現量上升、抗發炎細胞素IL-10 mRNA表現量下降,以及造成接受器TNF-R1 mRNA表現量上升、TNF-R2 mRNA表現量下降。此外,在HepG2大量表現dominant negative的IKKa或IKKb,只有IKKb可減少clozapine引起的NF-кВ的磷酸化,也證實clozapine活化NF-кB是透過IKKb。在luciferase報導基因實驗(luciferase activity assay)中,clozapine在HepG2細胞株中增加NF-кВ-Luc (含NF-кB-response element的 luciferase reporter construct)的活性,顯示clozapine可增加NF-кВ 的DNA結合能力。實驗中抑制NF-кВ活性,未有細胞凋亡蛋白PARP cleavage的產生。 綜合上述推測clozapine可能透過影響細胞素及接受器TNFR1,經由IKKb導致磷酸化的NF-кВ增加產生發炎反應,然而利用IKKb dominant negative mutant抑制clozapine導致的NF-kB活化,並未發現cleaved PARP的產生,而cleaved PARP是caspase-dependent途徑,故clozapine誘發的NF-kB活化是否有抗細胞凋亡作用,則有賴進一步偵測caspase-independent途徑的變化。
The second generation antipsychotic is effective in clinical treatment, but it can easily induce weight gain in schizophrenic patients. Clozapine is the last line of antipsychotics in treating clinical refractory schizophrenia, but the side effect of weight gain is worst. Many studies indicated that obesity could induce inflammation in tissues and cells and increase the incidence of type 2 diabetes and cancer. Increasing evidence suggests that antipsychotics are associated with tissue inflammation. NF-кB (nuclear factor-кB) plays a master regulator of inflammation. The aim of this study was to examine whether clozapine can directly induce human hepatocyte inflammation by NF-кB activation. Preliminary data showed that clozapine increased phosphorylation of inflammatory factor NF-кВ p65 at Ser536 and decreased the protein expression of IкВa. At the same time, the mRNA expressions of inflammatory cytokine, IL-1B and IL-6 were increased, but anti-inflammatory cytokine IL-10 was decreased.  The mRNA expression of TNF-R1 was increased but TNF-R2 was decreased in response to clozapine treatment by real-time PCR. Using kinase inactive mutants IKKa and IKKb also demonstrated clozapine activate NF-кB via IKKb. In addition, inactivate clozapine-induced NF-кB activation did not cause cleavage of the apoptosis-related protein PARP (Poly (ADP-ribose) polymerase (PARP)). Clozapine increasing NF-кВ-Luc transcriptional activity by luciferse activity assay suggested that clozapine could augment NF-кВ DNA binding activity. Taken together, clozapine maybe affect cytokines and TNFR1 receptor, and increase phosphorylation of NF-кВ through IKKb. However, overexpression of the dominant negative IKKb inhibiting clozapine-induced NF-кВ activation did not cause cleavage of the apoptosis-related protein PARP. Cleaved PARP is caspase-dependent pathway, whether clozapine-induced NF-кВ activation has anti-apoptosis property via caspase-independent pathway. It needs further study.
URI: http://ethesys.lib.ntou.edu.tw/cdrfb3/record/#G0M98360019
http://ntour.ntou.edu.tw/handle/987654321/30939
Appears in Collections:[生命科學暨生物科技學系] 博碩士論文

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