以小鼠模式探討IL-21對原發性膽汁性膽管炎的調控

Abstract

原發性膽汁性膽管炎 (Primary biliary cholangitis, PBC)是一種肝臟自體免疫疾病。先前研究指出在早期診斷的PBC病患血清中發現interleukin (IL)-21的高量表現。IL-21隸屬於IL-2 family，此家族是由一群多效性cytokine組成，可以調控許多不同細胞的功能。然而IL-21如何影響PBC疾病病程尚未被研究清楚，因此我們使用PBC小鼠動物模式以探討IL-21在自體肝臟發炎的情況下之角色。我們每兩週以腹腔注射的方式給予小鼠2-octynoic acid-OVA與complete/incomplete Freund’s adjuvant的混和物使其誘發PBC，在第一週時注射Adeno-associated virus (AAV)-mIL-21使小鼠肝臟表現高量mIL-21，而五週及十一週時犧牲小鼠並進一步分析各組之間的差異。PBC的標誌是血清中存在專一性辨識pyruvate dehydrogenase E2 complex (PDC-E2)的antimitochondrial antibody (AMA)。本實驗中，每個組別均測得AMA表現，因此表示小鼠成功誘發PBC。我們發現給予AAV-mIL-21的組別有較高的liver mononuclear cells (LMNCs) 浸潤，肝臟中促進發炎的IFN-γ亦顯著升高。此外，T、CD8+ T與follicular helper T (Tfh) cells的百分比在mIL-21存在的情況下顯著增加。我們更進一步發現CD8+ T cells高度活化並且高度表現granzyme B。另一方面，我們從Masson’s trichrome stain染色的病理切片觀察到AAV-mIL-21組呈現較嚴重的肝臟纖維化，而collagen III的qPCR結果亦證實了此現象。因此，我們認為IL-21藉由活化CD8+ T cells與促進肝臟纖維化而加劇PBC病程。

Primary biliary cholangitis (PBC) is an autoimmune liver disease. Previous study showed that serum interleukin (IL)-21 is highly expressed in early diagnosed PBC patients. IL-21 is a member of IL-2 family which has been reported to be a pleiotropic cytokine that mediates numerous cell types. However, the mechanism of IL-21 in PBC remains unclear. In this study, we used a mouse model of PBC to evaluate the roles of IL-21 in autoimmune liver inflammation. Mice were immunized with 2-octynoic acid-OVA mixed with complete/incomplete Freund’s adjuvant biweekly through i.p. to form PBC. Adeno-associated virus (AAV)-mIL-21 was given at week 1 by i.v. to express mIL-21 in liver. Mice were euthanized to examine the differences among groups. Antimitochondrial antibody (AMA) against pyruvate dehydrogenase E2 complex (PDC-E2), a specific hallmark of PBC, was detected in all groups. The numbers of liver mononuclear cells (LMNCs), mRNA expression level of proinflammatory cytokine IFN-γ increased in AAV-mIL-21 injected group. In addition, the percentage of T, CD8+ T, follicular helper T (Tfh) cells increased with the existence of mIL-21. Furthermore, CD8+ T cells were highly activated and granzyme B were further upregulated in early-stage PBC. Moreover, the histopathology sections stained by Masson’s trichrome stain showed exacerbated liver fibrosis in AAV-mIL-21 injected group and mRNA expression levels of collagen III revealed the data. The results indicate that IL-21 may exacerbate PBC by upregulating CD8+ T cells and further enhance liver fibrosis.