發炎所引起的肝臟疾病，無論是急性或慢性肝炎，一直是全人類的健康問題，而慢性肝炎更是與肝細胞癌的形成有密切相關。因此長久以來科學家即不斷發展實驗動物模式來研究肝臟特異性的免疫調節機制，目前由Concanavalin A ( Con A )所誘發的小鼠肝炎被認為和人類自體免疫性肝炎所造成的反應是類似的。過去已知的研究中，因自體免疫造成肝炎的嚴重程度在兩性中有顯著的差異。我們要利用Con A誘發肝炎的模式，瞭解性荷爾蒙在這個肝炎模式中扮演的角色。Con A是一種源自植物的lectin，會特異性與T細胞表面醣蛋白上的特異糖分子，如T細胞受體或CD3分子結合，經由這些分子刺激T細胞活化，它是一種能使T淋巴細胞活化增生的裂殖原。當不同品系的小鼠以尾靜脈方式給予Con A ( 10 mg/kg )後，在24小時內，藉由測量血清中轉胺酵素可以發現小鼠產生急性肝炎，並會持續至36到48小時左右。然而Con A誘發的肝炎會隨著不同品系與性別的小鼠，其嚴重度有明顯差異。C57BL/6與BALB/c品系小鼠對Con A所誘發之肝炎敏感度較A/J品系小鼠高，而雌鼠則比公鼠產生更嚴重的肝炎。Con A誘發之肝炎會造成大量T淋巴細胞浸潤聚集到肝臟中，其中以CD4+ T淋巴細胞最為顯著，而從流式細胞儀的偵測，發現經誘導後的雌鼠，其浸潤的CD4+與CD8+ T細胞遠較公鼠多。為了更進一步探討性荷爾蒙在此肝炎模式的角色，我們以外科手術去除掉成熟雌鼠之卵巢二週後，再給予Con A刺激，發現卵巢去除後的雌鼠，血清中的轉胺酵素明顯降低，程度近似公鼠，而以雌性荷爾蒙再補充的雌鼠即會回復肝炎的程度，其CD4+與CD8+ T細胞數量較未給予雌激素補充的雌鼠多，並且有顯著的統計上差異。測量一些細胞激素，如IL-4、IL-6、IL-10、IL-12、TNF-α、IFN-γ，我們發現它們的量在正常雌鼠與卵巢切除後再補充雌性激素之雌鼠皆高於公鼠以及只有卵巢去除的雌鼠。另外自然殺手性T細胞的數量，在未受過任何處理的公母鼠中，即具有差異，在給予Con A後，其細胞數目也有變化，顯示自然殺手性T細胞也參與在Con A誘發之肝炎模式中。由以上的結果顯示，性荷爾蒙會參與調節Con A誘發的小鼠肝炎，但雌激素是如何作用及調節細胞激素與免疫細胞則須更進一步的研究。

An inflammation-mediated liver disease, both acute and chronic hepatitis, represents a worldwide health problem in human beings, and chronic hepatitis is associated strongly with the development of hepatocellular carcinoma. Experimental animal models have been used for a long time to study liver-specific immune regulation. Concanavalin A ( Con A ) – induced hepatitis in mice is thought to mimic the autoimmune hepatitis in human. The degree of autoimmune hepatitis is known to be different between genders – female is severe than male. In this study, we used the Con A-induced hepatitis model to investigate the role of sex hormones in autoimmune hepatitis. Con A is a lectin from plant, it binds specifically to certain sugar residues on T cell surface glycoproteins such as TCR or CD3 molecules and then stimulates the T cells to proliferate. It is known as a mitogen for T lymphocytes. When different strains of mice were given a single intravenous injection of Con A (10 mg/kg ). The acute hepatic injury assessed by increased plasma transaminase levels developed within 24 hours and lasted until 36 or 48 hours. The severities of liver injury were different among different strains and gender. C57BL/6 and BALB/c mice were more sensitive to Con A-induced hepatitis than A/J mice. The Con A-induced hepatitis is more severe in female mice than in male mice. The number of T lymphocytes in the liver was increased after Con A injection, especially the CD4+ T cells. There were more infiltrated CD4+ and CD8+ T cells in female than in male mice. We are interested in the hormonal regulation of this hepatitis model. Female mice were ovariectomized for two weeks, then challenged with Con A. The plasma transaminase levels of these levels of these ovariectomized mice were reduced to those of male mice. However after the estrogen supplement in these ovariectomied mice, the severity of Con A-induced hepatitis could be restored to the level of female mice. The CD4+ and CD8+ infiltrating T cells were increased significantly in estradiol supplemented ovariectomied mice compared to ovariectomied non-supplemented mice. The cytokines of IL-4, IL-6, IL-10, IL-12, TNF-α and IFN-γ in liver after Con A stimulation were found higher in female or estradiol supplemented ovariectomized mice than male or ovariectomized mice. Furthermore, the natural killer T (NKT) cells in liver were different in different genders of mice. After Con A stimulation, the number of these NKT cells changed as well. This suggests that NKT may also be affected by estradiol in this Con A-induced hepatitis.

Ansar Ahmed S, Penhale WJ, Talal N. Sex hormones, immune responses, and autoimmune diseases. Mechanisms of sex hormone action. Am J Pathol. 1985. 121:531-51.
Beuth J, Ko HL, Steuer M, Pulverer G. Hepatocellular injury inhibits lectin-mediated tumor colonization into BALB/c-mice livers. Experientia. 1993. 49:547-50.
Boelsterli UA. Disease-related determinants of susceptibility to drug-induced idiosyncratic hepatotoxicity. Curr Opin Drug Discov Devel. 2003. 6:81-91.
Burger D, Dayer JM. Cytokines, acute-phase proteins, and hormones: IL-1 and TNF-alpha production in contact-mediated activation of monocytes by T lymphocytes. Ann N Y Acad Sci. 2002. 966:464-73.
Carlsten H, Tarkowski A, Holmdahl R, Nilsson LA. Oestrogen is a potent disease accelerator in SLE-prone MRL lpr/lpr mice. Clin Exp Immunol. 1990. 80:467-73.
Chao TC, Van Alten PJ, Greager JA, Walter RJ. Steroid sex hormones regulate the release of tumor necrosis factor by macrophages. Cell Immunol. 1995. 160:43-9.
Decker K, Keppler D. Galactosamine hepatitis: key role of the nucleotide deficiency period in the pathogenesis of cell injury and cell death. Rev Physiol Biochem Pharmacol. 1974. 71:77-106.
Endo Y, Kikuchi T, Nakamura M. Ornithine and histidine decarboxylase activities in mice sensitized to endotoxin, interleukin-1 or tumour necrosis factor by D-galactosamine. Br J Pharmacol. 1992. 107:888-94.
Finn R, St Hill CA, Govan AJ, Ralfs IG, Gurney FJ, Denye V. Immunological responses in pregnancy and survival of fetal homograft. Br Med J. 1972. 3:150-2.
Fukuda R, Satoh S, Nguyen XT, Uchida Y, Kohge N, Akagi S, Ikeda S, Watanabe M, Fukumoto S. Expression rate of cytokine mRNA in the liver of chronic hepatitis C: comparison with chronic hepatitis B. J Gastroenterol. 1995. 30:41-7.
Gantner F, Leist M, Lohse AW, Germann PG, Tiegs G. Concanavalin A-induced T-cell-mediated hepatic injury in mice: the role of tumor necrosis factor. Hepatology. 1995. 21:190-8.
Goldmann O, Chhatwal GS, Medina E. Immune mechanism underlying host susceptibility to infection with Group A Streptococci. J Infect Dis. 2003. 187:854-61.
Gregory MS, Duffner LA, Faunce DE, Kovacs EJ. Estrogen mediates the sex difference in post-burn immunosuppression. J Endocrinol. 2000. 164:129-38.
Griffith JS, Jensen SM, Lunceford JK, Kahn MW, Zheng Y, Falase EA, Lyttle CR, Teuscher C. Evidence for the genetic control of estradiol-regulated responses. Implications for variation in normal and pathological hormone-dependent phenotypes. Am J Pathol. 1997. 150:2223-30.
Herzog D, Rasquin-Weber AM, Debray D, Alvarez F. Subfulminant hepatic failure in autoimmune hepatitis type 1: an unusual form of presentation. J Hepatol. 1997. 27:578-82.
Hu SK, Mitcho YL, Rath NC. Effect of estradiol on interleukin 1 synthesis by macrophages. Int J Immunopharmacol. 1988. 10:247-52.
Jansson L, Holmdahl R. Estrogen-mediated immunosuppression in autoimmune diseases. Inflamm Res. 1998. 47:290-301.
Karpuzoglu-Sahin E, Hissong BD, Ansar Ahmed S. Interferon-gamma levels are upregulated by 17-beta-estradiol and diethylstilbestrol. J Reprod Immunol. 2001. 52:113-27.
Kaneko Y, Harada M, Kawano T, Yamashita M, Shibata Y, Gejyo F, Nakayama T, Taniguchi M. Augmentation of Valpha14 NKT cell-mediated cytotoxicity by interleukin 4 in an autocrine mechanism resulting in the development of concanavalin A-induced hepatitis. J Exp Med. 2000. 191:105-14.
Knolle PA, Gerken G, Loser E, Dienes HP, Gantner F, Tiegs G, Meyer zum Buschenfelde KH, Lohse AW. Role of sinusoidal endothelial cells of the liver in concanavalin A-induced hepatic injury in mice. Hepatology. 1996. 24:824-9.
Kusters S, Gantner F, Kunstle G, Tiegs G. Interferon gamma playas critical role in T cell-dependent liver injury in mice initiated by concanavalin A. Gastroenterology; 1996. 111: 462-71.
Li ZG, Danis VA, Brooks PM. Effect of gonadal steroids on the production of IL-1 and IL-6 by blood mononuclear cells in vitro. Clin Exp Rheumatol. 1993. 11:157-62.
Lohse AW, Manns M, Dienes HP, Meyer zum Buschenfelde KH, Cohen IR. Experimental autoimmune hapetitis: disease induction, time course and T-cell reactivity. Hepatology. 1990. 11:24-30.
Louis H, Le Moine O, Peny MO, Gulbis B, Nisol F, Goldman M, Deviere J. Hepatoprotective role of interleukin 10 in galactosamine/lipopolysaccharide mouse liver injury. Gastroenterology. 1997. 112:935-42.
Louis H, Le Moine A, Flamand V, Nagy N, Quertinmont E, Paulart F, Abramowicz D, Le Moine O, Goldman M, Deviere J. Critical role of interleukin 5 and eosinophils in concanavalin A-induced hepatitis in mice. Gastroenterology. 2002. 122:2001-10.
Llorent L, Richaud-Patin Y, Alcocer-Castillejos N, Ruiz-Soto R, Mercado MA, Orozco H, Gamboa-Dominguez A, Alcocer-Varela J. Cytokine gene expression in cirrhotic and non-cirrhotic human liver. J Hepatol. 1996. 24:555-63.
McFarlane IG. Pathogenesis of autoimmune hepatitis. Biomed Pharmacother. 1999. 53:255-63.
Medina KL, Smithson G, Kincade PW. Suppression of B lymphopoiesis during normal pregnancy. J Exp Med. 1993. 178:1507-15.
Medina J, Garcia-Buey L, Moreno-Otero R. Review article: immunopathogenetic and therapeutic aspects of autoimmune hepatitis. Aliment Pharmacol Ther. 2003. 17:1-16.
Mendiratta SK, Martin WD, Hong S, Boesteanu A, Joyce S, Van Kaer L. CD1d1 mutant mice are deficient in natural T cells that promptly produce IL-4. Immunity. 1997. 6:469-77.
Mizuhara H, O'Neill E, Seki N, Ogawa T, Kusunoki C, Otsuka K, Satoh S, Niwa M, Senoh H, Fujiwara H. T Cell Activation-associated Hepatic Injury : Mediation by Tumor Necrosis Factors and Protection by Interleukin 6. J Exp Med. 1994. 179:1529-37.
Mizuhara H, Uno M, Seki N, Yamashita M, Yamaoka M, Ogawa T, Kaneda K, Fujii T, Senoh H, Fujiwara H. Critical involvement of interferon gamma in the pathogenesis of T-cell activation-associated hepatitis and regulatory mechanisms of interleukin-6 for the manifestations of hepatitis. Hepatology. 1996. 23:1608-15.
Nicoletti F, Di Marco R, Zaccone P, Salvaggio A, Magro G, Bendtzen K, Meroni P. Murine concanavalin A-induced hepatitis is prevented by interleukin 12 (IL-12) antibody and exacerbated by exogenous IL-12 through an interferon-gamma-dependent mechanism. Hepatology. 2000. 32:728-33.
Nilsson N, Carlsten H. Estrogen induces suppression of natural killer cell cytotoxicity and augmentation of polyclonal B cell activation. Cell Immunol. 1994. 158:131-9.
Okuyama R, Abo T, Seki S, Ohteki T, Sugiura K, Kusumi A, Kumagai K. Estrogen administration activates extrathymic T cell differentiation in the liver. J Exp Med. 1992. 175:661-9.
Roberts EA. Autoimmune hepatitis. Indian J Pediatr. 1995. 62:525-31.
Rubanyi GM, Freay AD, Kauser K, Sukovich D, Burton G, Lubahn DB, Couse JF, Curtis SW, Korach KS. Vascular estrogen receptors and endothelium-derived nitric oxide production in the mouse aorta. Gender difference and effect of estrogen receptor gene disruption. J Clin Invest. 1997. 99:2429-37.
Sakakibara S, Shibayama Y, Ueda I, Kagamiyama H. The protective effect of polyriboinosinic acid-polyribocytidylic acid against the occurrence of galactosamine-induced liver cell injury in rat. Experientia. 1983. 39:174-6.
Samy TS, Schwacha MG, Cioffi WG, Bland KI, Chaudry IH. Androgen and estrogen receptors in splenic T lymphocytes: effects of flutamide and trauma-hemorrhage. Shock. 2000. 14:465-70.
Sass G, Koerber K, Bang R, Guehring H, Tiegs G. Inducible nitric oxide synthase is critical for immune-mediated liver injury in mice. J Clin Invest. 2001. 107:439-47.
Sass G, Heinlein S, Agli A, Bang R, Schumann J, Tiegs G. Cytokine expression in three mouse models of experimental hepatitis. Cytokine. 2002. 19:115-20.
Satoh M, Kobayashi K, Ishii M, Igarashi T, Toyota T. Midzonal necrosis of the liver after concanavalin A-injection. Tohoku J Exp Med. 1996. 180:139-52.
Sun Y, Tokushige K, Isono E, Yamauchi K, Obata H. Elevated serum interleukin-6 levels in patients with acute hepatitis. J Clin Immunol. 1992. 12:197-200.
Tholen H., Zimmerli W, and Rajacic Z. Effect of coenzyme-A, NAD, alpha lipoic-acid and cocarboxylase on survival of rats with galactosamine-induced severe hepatitis. Experientia. 1985. 41:1042-1045.
Thong YH, Steele RW, Vincent MM, Hensen SA, Bellanti JA. Impaired in vitro cell-mediated immunity to rubella virus during pregnancy. N Engl J Med. 1973. 289:604-6.
Tiegs G, Hentschel J, Wendel A. A T cell-dependent Experimental Liver Injury in Mice Inducible by Concanavalin A. J Clin Invest. 1992. 90:196-203.
Tiegs G. and Gantner F. Immunotoxicology of T cell-dependent experimental liver injury. Exp.& Toxicol. Pathol. 1996. 48:471-476.
Tornwall J, Carey AB, Fox RI, Fox HS. Estrogen in autoimmunity: expression of estrogen receptors in thymic and autoimmune T cells. J Gend Specif Med. 1999. 2:33-40
Toyabe S, Seki S, Iiai T, Takeda K, Shirai K, Watanabe H, Hiraide H, Uchiyama M, Abo T. Requirement of IL-4 and liver NK1+ T cells for concanavalin A-induced hepatic injury in mice. J Immunol. 1997. 159:1537-42.
Trop S, Nagler A, Ilan Y., Role of NK1.1+ and AsGm-1+ cells in oral immunoregulation of experimental colitis. Inflamm Bowel Dis. 2003. 9:75-86.
Tsuji T, Shinohara T. Pathological study of chronic D-galactosamine induced hepatitis in mice by administration of adjuvants - an animal model of the chronic active hepatitis. Gastroenterol Jpn. 1981. 16:9-20.
Tung BY, Carithers RL Jr. Cholestasis and alcoholic liver disease. Clin Liver Dis. 1999. 3:585-601.
Vernon-Robert B. The effects of steroid hormones on macrophage function. Int Rev Cytol.1969. 25:131-58.
Unno R, Matsuzaki Y, Itoh S, Doy M, Shoda J, Tanaka N. Novel murine autoimmune-mediated liver disease model induced by graft-versus-host reaction and concanavalin A. J Gastroenterol Hepatol. 2001. 16:1149-57.
Wu Y, Brouet I, Calmels S, Bartsch H, Ohshima H. Increased endogenous N-nitrosamine and nitrate formation by induction of nitric oxide synthase in rats with acute hepatic injury caused by Propionibacterium acnes and lipopolysaccharide administration. Carcinogenesis. 1993. 14:7-10.