Amygdalin (Vitamin B17) pretreatment attenuates experimentally induced acute autoimmune hepatitis through reduction of CD4+ cell infiltration

Abstract

Background

Autoimmune hepatitis (AIH) is an immune-mediated inflammation of the liver characterized by disorganized hepatic parenchyma and inflammatory cell infiltration. Although the increased incidence of AIH, the development of novel therapeutic strategies are impeded by the poor understanding of the accompanied detailed immunopathogenic changes. CD4+ T cells are key mediators of inflammatory cell infiltration in initial phases of liver injuries like AIH. The distribution of CD4+ cells and the histopathological changes accompanying Con A-induced AIH were investigated together with the postulated protective effect of Amygdalin (Amg.).

Materials and methods

30 adult male mice were divided into three groups; control, AIH and AIH-Amg. groups. AIH was induced by a single intravenous injection of Concanavalin A (Con A) (15 mg/kg). The AIH-Amg. group received Amg. 5 mg/kg intraperitoneally once a week for three weeks. Blood samples were examined for ALT and AST. MDA, SOD, and GSH were determined in hepatic homogenates. Liver section stained with hematoxylin and eosin, Masson trichrome and CD4+ immune stain were examined by light and electron microscopy.

Results

AIH group showed a significant increase in levels of ALT, AST and MDA and a significant decline in SOD and GSH compared to the controls. The liver tissue showed distorted hepatic architecture with intercellular hemorrhage, necrosis, and inflammatory cell infiltration. The area percent of CD4+ immune staining was significantly increased. Electron microscopic examination showed massive cellular degenerative changes. Amg. pretreatment in AIH-Amg. group significantly reversed these changes.

Conclusion

AIH induced CD4+ cells infiltration in the liver with subsequent liver tissue damage. Amg. pretreatment inhibited CD4+ cell infiltration and protected the liver tissue. This finding suggests that Amg. could be a therapeutic agent in the management of AIH.

Introduction

Autoimmune hepatitis (AIH) is an immune-mediated inflammation of the liver characterized by disorganized hepatic parenchyma and inflammatory cell infiltration (Christen and Hintermann, 2016). The exact etiology of AIH is still unknown (Aizawa and Hokari, 2017), however, different pathogens as environmental factors, viruses and chemicals have been postulated (Czaja, 2011). AIH reported a progressive increased prevalence and incidence in all ages and ethnic groups (Gatselis et al., 2015). Yet, the management of AIH is still largely restricted to steroids and cytostatic drugs (Czaja, 2011). The poor understanding of the detailed immunopathogenic changes accompanying AIH hampers the development of novel therapeutic strategies (Burak et al., 2013).

A known experimental model of drug-induced AIH in mice is the use of Concanavalin A (Con A) (Mao et al., 2015). Con A-induced acute hepatitis matches the histopathological picture of AIH patients (Ye et al., 2018). Induced autoimmune hepatitis involves the release of inflammatory mediators and inflammatory cell infiltration. These pathologic changes can proceed with time to chronic liver disease (De Biasio et al., 2006). As in other body tissues, the orderly step that follows inflammation is tissue healing by fibrosis which is a beneficial reversible defense mechanism aiming to encapsulate the tissue injury (Ebrahimi et al., 2016). However, the ultimate progression to advanced hepatic fibrosis and/or cirrhosis, which is an irreversible tissue response, disrupt the liver architecture and function (Friedman, 2008).

CD4+ cells are key mediators of inflammatory responses which play a crucial role in initial phases of liver injury, regeneration and graft rejection (Yamamoto et al., 2002). CD4+ cells were supposed to induce cytokine responses that control liver response to injury (Salgame et al., 1991). CD4+ cells were reported to direct the Con A-induced AIH through oxidative stress (Tiegs et al., 1992). Furthermore, the absence of CD4+ cells was found to improve the recovery of the liver from oxidant-mediated damage (Zwacka et al., 1997). In this context, agents that can suppress CD4+ cells infiltration could withhold the subsequent inflammatory cell infiltration, oxidative stress and improve liver tissue histopathological changes.

Amygdalin (Amg., Vitamin B17) is a cyanogenic glycoside compound which belongs to the group widely distributed in plants like apricot and peach (Santos Pimenta et al., 2014; Song and Xu, 2014). Amg. has multiple therapeutic effects as antiatherogenic, antifibrotic, anti-inflammatory and antiulcer agent (Chang et al., 2005; Du et al., 2010; Mirmiranpour et al., 2012). A particular action of Amg. on the liver is the ability to lower levels of AST, ALT, increase hydroxyproline content and inhibit proliferation of connective tissue in CCl4 and D-galactosamine treated rats (Wei et al., 2009).

Exploring the relation between CD4+ cells infiltration and the migration and engraftment of the inflammatory cells with subsequent liver histopathological changes in AIH can help to define new therapeutic agents. The tissue and the cellular distortion accompanying Con A-induced AIH and the postulated protective effect of Amg. were investigated in this study aiming for the implication of nontraditional management procedure.