去氧核糖核酸酶I對於DSS誘發小鼠腸炎模式之影響

Abstract

發炎性腸道疾病(inflammatory bowel diseases, IBDs) 指的是一種由腸道內菌相之改變以及腸道發炎所引起之慢性疾病。發炎性腸道疾病主要可以分為兩種：潰瘍性結腸炎(Ulcerative colitis) 以及克隆氏症(Crohn’s disease)。當腸道發炎時，哺乳動物體內最多的白血球為多核型白血球(PMNs)，又稱嗜中性白血球，在初級免疫反應當中扮演著第一道防線的角色。嗜中性白血球可藉由吞噬作用(phagocytosis)以及脫顆粒作用(degranulation)來達到對抗病原的目的。而在2004年，Brinkmann等人發現了一個由解縮的染色質絲以及一群蛋白所構成的胞外網狀結構，統稱為嗜中性胞外網狀結構(Neutrophil Extracellular Traps, NETs)，嗜中性胞外網狀結構可以藉由其網狀的結構來捕捉病原體，進而幫助病原體之清除。先前的研究指出，在克隆氏症的病人身上發現嗜中性包外網狀結構之前驅物，過氧化物(Reactive oxygen species, ROS)表現量的上升。在另一篇研究中，作者利用蛋白質體學的方式，在潰瘍性結腸炎的病人身上偵測到大量的嗜中性白血球以及嗜中性胞外網狀結構相關蛋白質的表現，但嗜中性胞外網狀結構對於潰瘍性結腸炎是好是壞並不了解。因此在本篇研究當中，我利用dextran sulfate sodium (DSS) 誘發小鼠腸炎模式，並施打DNase I (一種已知的嗜中性胞外網狀結構之抑制劑)，來探討胞外網狀結構在此模式下扮演的角色。結果顯示在經DSS誘導發炎的小鼠當中，有施打DNase I的小鼠，其體重下降的幅度以及臨床指標分數均較對照組來的低，且其腸子長度縮短的情形也受到抑制。在免疫螢光染色的部分，我們偵測了胞外網狀結構的重要蛋白，瓜胺酸化之組蛋白H3 (citrullinated histone 3)，結果顯示，在經DSS誘導發炎的小鼠當中，再施打DNaseI的小鼠中，其組蛋白有減少的趨勢。最後在組織切片染色的結果顯示，在DSS誘導發炎的小鼠當中，施打DNase I的小鼠，其腸道受到破壞的情形趨於減緩。綜合以上結果，可推論在DSS誘導小鼠發炎的情況之下，抑制胞外網狀結構可減緩腸道發炎之症狀。

Inflammatory bowel disease (IBD) refers to the chronic disease that is due to the disruption of immune homeostasis against gut microbiota. There are two major types of inflammatory bowel diseases: ulcerative colitis (UC) and Crohn’s disease (CD). During the intestinal inflammation, polymorphonuclear leukocytes (PMN), or neutrophils, the most abundant leukocytes in mammals, are considered as the first line of innate immune defense. Neutrophils utilize phagocytosis and degranulation to control microbe infection. In 2004, Brinkmann and colleagues discovered neutrophils also form an extracellular structure composed of decondensed chromatin fibers and a set of proteins, which terms as neutrophil extracellular traps (NETs) when they encounter pathogens. Enhanced ROS production, a precursor of NET formation, is found in the inflamed intestine of patients with Crohn’s disease. A recent proteomic analysis further detected elevated NET-associated proteins in the colons of ulcerative colitis patients compared to those of control groups. However, it is still unknown whether NET formation is beneficial or harmful to ulcerative colitis. In this study, I administered Deoxyribonuclease I (DNase I), an inhibitor of NET into mice with dextran sodium sulfate (DSS)-induced colitis. Body weight and clinical score were examined every day. The results indicated that the group of DSS-induced colitis mice with DNase I treatment manifested a decreased body weight change and lower clinical score when compared to the control group (without DNase I treatment) after DSS administration. Repression of NETs also ameliorate DSS-induced colon shortening. Immunofluorescence staining results suggested that there were lower amount of citrullinated histone 3, a component of NET, in the colons of mice treated with DSS and DNase I when compared to mice with DSS administration. Histological damage result suggested that administration with DNase I protected mice form disruption of intestinal architecture in DSS-treatment mice. Overall, our results indicated DNase I treatment ameliorated the development of DSS-induced colitis in mice.