NOD.c3c4 congenic mice develop autoimmune biliary disease that serologically and pathogenetically models human primary biliary cirrhosis

Primary biliary cirrhosis (PBC) is an autoimmune disease with a strong genetic component characterized by biliary ductular inflammation with eventual liver cirrhosis. The serologic hallmark of PBC is antimitochondrial antibodies that react with the pyruvate dehydrogenase complex, targeting the inner lipoyl domain of the E2 subunit (anti–PDC-E2). Herein we demonstrate that NOD.c3c4 mice congenically derived from the nonobese diabetic strain develop an autoimmune biliary disease (ABD) that models human PBC. NOD.c3c4 (at 9–10 wk, before significant biliary pathology) develop antibodies to PDC-E2 that are specific for the inner lipoyl domain. Affected areas of biliary epithelium are infiltrated with CD3+, CD4+, and CD8+ T cells, and treatment of NOD.c3c4 mice with monoclonal antibody to CD3 protects from ABD. Furthermore, NOD.c3c4-scid mice develop disease after adoptive transfer of splenocytes or CD4+ T cells, demonstrating a central role for T cells in pathogenesis. Histological analysis reveals destructive cholangitis, granuloma formation, and eosinophilic infiltration as seen in PBC, although, unlike PBC, the extrahepatic biliary ducts are also affected. Using a congenic mapping approach, we define the first ABD (Abd) locus, Abd1. These results identify the NOD.c3c4 mouse as the first spontaneous mouse model of PBC

The interplay of type I and type II interferons in murine autoimmune cholangitis as a basis for sex-biased autoimmunity

We have reported on a murine model of autoimmune cholangitis, generated by altering the AU-rich element (ARE) by deletion of the interferon gamma (IFN-γ) 3\u27 untranslated region (coined ARE-Del−/−), that has striking similarities to human primary biliary cholangitis (PBC) with female predominance. Previously, we suggested that the sex bias of autoimmune cholangitis was secondary to intense and sustained type I and II IFN signaling. Based on this thesis, and to define the mechanisms that lead to portal inflammation, we specifically addressed the hypothesis that type I IFNs are the driver of this disease. To accomplish these goals, we crossed ARE-Del−/− mice with IFN type I receptor alpha chain (Ifnar1) knockout mice. We report herein that loss of type I IFN receptor signaling in the double construct of ARE-Del−/− Ifnar1−/− mice dramatically reduces liver pathology and abrogated sex bias. More importantly, female ARE-Del−/− mice have an increased number of germinal center (GC) B cells as well as abnormal follicular formation, sites which have been implicated in loss of tolerance. Deletion of type I IFN signaling in ARE-Del−/− Ifnar1−/− mice corrects these GC abnormalities, including abnormal follicular structure. Conclusion: Our data implicate type I IFN signaling as a necessary component of the sex bias of this murine model of autoimmune cholangitis. Importantly these data suggest that drugs that target the type I IFN signaling pathway would have potential benefit in the earlier stages of PBC. (Hepatology 2018;67:1408-1419)

Induction of Shrimp Tropomyosin-Specific Hypersensitivity in Mice

Background: Shellfish hypersensitivity is amongst the most common food allergies. The major shellfish allergen was identified as tropomyosin. Here, we investigated the immediate hypersensitivity responses, IgE and cell-mediated immune response in mice sensitized with recombinant shrimp tropomyosin. Methods: Shrimp tropomyosin was cloned and expressed as a His -tagged fusion recombinant protein in Escherichia coli. Three- to 4-week- old BALB/c mice were sensitized by intragastric administration of recombinant tropomyosin ( 0.1 mg) plus cholera toxin ( 10 mu g) on days 0, 12, 19 and 26 and challenged on day 33. Mice fed with phosphate-buffered saline plus cholera toxin were included as controls. Animals were monitored for immediate hypersensitive responses and tropomyosin-specific IgE over time. In addition, shrimp tropomyosin-specific CD4+ T cells, interleukin-4 and interferon-gamma levels were determined from in vitro splenocyte cultures. A passive cutaneous anaphylaxis assay was also conducted. Results: Mice fed with shrimp tropomyosin developed swelling of the snout, increased scratching behavior and shrimp tropomyosin- specific IgE. Sera from tropomyosin-sensitized mice elicited vascular leakage in nave mice in the passive cutaneous anaphylaxis assay. Shrimp tropomyosin- specific CD4+ T cell proliferations and elevated interleukin-4 over interferon- gamma levels were evident in splenocyte cultures of tropomyosin-fed mice upon tropomyosin stimulation. In contrast, shrimp tropomyosin-specific IgE, CD4+ T cells and hypersensitive responses were absent in the control mice. Conclusion: We have generated a BALB/c model of shrimp allergy. This model provides a useful tool for evaluating the immuno-pathogenic mechanisms involved in shellfish hypersensitivity

Toward solving the etiological mystery of primary biliary cholangitis

Primary biliary cholangitis (PBC) is considered a model autoimmune disease due to its signature anti-mitochondrial antibody (AMA) autoantibody, female predominance, and relatively specific portal infiltration and cholestasis. The identification and cloning of the major mitochondrial autoantigens recognized by AMA have served as an immunologic platform to identify the earliest events involved in loss of tolerance. Despite the relatively high concordance rate in identical twins, genome-wide association studies have not proven clinically useful and have led to suggestions of epigenetic events. To understand the natural history and etiology of PBC, several murine models have been developed, including spontaneous models, models induced by chemical xenobiotic immunization, and by “designer” mice with altered interferon metabolism. Herein, we describe five such models, including 1) NOD.c3c4 mice, 2) dominant negative form of transforming growth factor receptor type II mice, 3) interleukin-2R α−/− mice, 4) adenylate-uridylate-rich element Del−/− mice, and 5) 2-octynoic acid-conjugated bovine serum albumin immunized mice. Individually there is no perfect murine model, but collectively the models point to loss of tolerance to PDC-E2, the major mitochondrial autoantigen, as the earliest event that occurs before clinical disease is manifest. Although there is no direct association of AMA titer and PBC disease progression, it is noteworthy that the triad of PBC monocytes, biliary apotopes, and AMA leads to an intense proinflammatory cytokine burst. Further, the recurrence of PBC after liver transplantation indicates that, due to major histocompatibility complex restriction, disease activity must include not only adaptive immunity but also innate immune mechanisms. We postulate that successful treatment of PBC may require a personalized approach with therapies designed for different stages of disease. (Hepatology Communications 2017;1:275–287)

Therapeutic and immunological interventions in primary biliary cholangitis: From mouse models to humans

Primary biliary cholangitis (PBC) is a chronic cholestatic liver disease that predominantly affects women in their fifth and sixth decades. The diagnostic hallmarks of PBC are detection of anti-mitochondrial antibodies (AMAs) and chronic non-suppurative destructive cholangitis (CNSDC) of small- and medium-sized intrahepatic bile ducts in liver histological examination [1, 2]. A significant amount of data suggests that immunological activity against small biliary epithelial cells (BECs), found histologically as portal inflammation, leads to clinical disease. In PBC, as with other autoimmune diseases, both genetic and environmental factors contribute to the development of pathology [3–8]. The first-line therapy of PBC is ursodeoxycholic acid (UDCA), although obeticholic acid (OCA) has been approved recently for patients with an incomplete response to UDCA [9–11]. Unfortunately, unlike other autoimmune diseases, no successful clinical trials of biologics have been conducted, and the mechanisms of action of UDCA and OCA are not fully understood [12–16]. The clinical phenotype and the natural history of PBC vary between patients and can have other overlapping autoimmune diseases [12, 17–19]. For example, some patients may have mild elevation of liver enzyme levels and remain asymptomatic for life. By contrast, other patients can develop signs of liver failure and rapidly decompensate despite therapy, requiring liver transplantation. A presumption is that these differences are due to genetic and environmental factors, both contributing to the development of PBC to various degrees in each patient [4, 7, 20]. Although multiple genome- wide association studies (GWASs) have been reported differences in several genes [21–24], their clinical implications and relevance remain elusive [25]. In fact, in PBC, in autoimmune diseases, including PBC, the results of GWASs have been disappointing, and efforts have been made recently for both deep sequencing and study of epigenetic events [26–30]. To understand the importance of developing a useful mouse model, other aspects of PBC such as spectra of disease and gender dominance should be considered [18, 31]

Modulating Shrimp Tropomyosin-Mediated Allergy: Hypoallergen DNA Vaccines Induce Regulatory T Cells to Reduce Hypersensitivity in Mouse Model

Shellfish allergy is one of the most common food allergies, with tropomyosin as the major cross-reactive allergen. However, no allergen-specific immunotherapy is clinically available. Recently, we designed two shrimp hypoallergens MEM49 and MED171. This study aimed to examine and compare the efficacy of the MEM49- and MED171-based DNA vaccines (pMEM49 and pMED171) in modulating shrimp allergy in a murine model of shrimp tropomyosin sensitivity. Intradermal immunization of BALB/c mice with pMEM49 or pMED171 effectively down-modulated allergic symptoms, tropomyosin-specific IgE levels, intestinal Th2 cytokines expression, and inflammatory cell infiltration. Both pMEM49 and pMED171 increased the frequency of regulatory T cells, but to a greater extent by pMED171 with upregulation of gut-homing molecules integrin-α4β7. The functionality of the pMED171-induced Treg cells was further illustrated by anti-CD25-mediated depletion of Treg cells and the adoptive transfer of CD4+CD25+Foxp3+Treg cells. Collectively, the data demonstrate that intradermal administration of pMED171 leads to the priming, activation, and migration of dermal dendritic cells which subsequently induce Treg cells, both locally and systemically, to downregulate the allergic responses to tropomyosin. This study is the first to demonstrate the potency of hypoallergen-encoding DNA vaccines as a therapeutic strategy for human shellfish allergy via the vigorous induction of functional Treg cells

Overcoming Shellfish Allergy: How Far Have We Come?

Shellfish allergy caused by undesirable immunological responses upon ingestion of crustaceans and mollusks is a common cause of food allergy, especially in the Asia-Pacific region. While the prevalence of shellfish allergy is increasing, the mainstay of clinical diagnosis for these patients includes extract-based skin prick test and specific IgE measurement while clinical management consists of food avoidance and as-needed use of adrenaline autoinjector should they develop severe allergic reactions. Such a standard of care is unsatisfactory to both patients and healthcare practitioners. There is a pressing need to introduce more specific diagnostic methods, as well as effective and safe therapies for patients with shellfish allergy. Knowledge gained on the identifications and defining the immuno-molecular features of different shellfish allergens over the past two decades have gradually translated into the design of new diagnostic and treatment options for shellfish allergy. In this review, we will discuss the epidemiology, the molecular identification of shellfish allergens, recent progress in various diagnostic methods, as well as current development in immunotherapeutic approaches including the use of unmodified allergens, hypoallergens, immunoregulatory peptides and DNA vaccines for the prevention and treatment of shellfish allergy. The prospect of a “cure “for shellfish allergy is within reach