Atul K. Bhan

Spontaneous development of inflammatory bowel disease in T cell receptor mutant mice

We describe the spontaneous development of inflammatory bowel disease (IBD) in several immunodeficient mouse strains created via gene targeting in embryonic stem cells. Chronic colitis was observed in T cell receptor (TCR) alpha mutant, TCR beta mutant, TCR beta x delta double mutant, or class II major histocompatibility complex (MHC) mutant mice, but not in recombination-activating gene RAG-1 mutant mice or nude mice kept in the same specific pathogen-free animal facility. This clinical pattern suggests that the disease requires the presence of B lymphocytes and the absence of class II MHC-restricted CD4+ alpha beta T cells. IBD in the mutant mice has some of the features of the human disease ulcerative colitis. Based on these results, we suggest that dysfunction of the mucosal immune system may underly the pathogenesis of some types of IBD in humans.

Chronic intestinal inflammatory condition generates IL-10-producing regulatory B cell subset characterized by CD1d upregulation

B cells possess a variety of immune functions that are involved in normal and abnormal immune responses, including autoimmune disorders. Through murine models of intestinal inflammation, we here demonstrate a B cell subset that is induced in gut-associated lymphoid tissues and is characterized by CD1d upregulation. This B cell subset appears under a chronic inflammatory environment, produces IL-10, and suppresses progression of intestinal inflammation by downregulating inflammatory cascades associated with IL-1 upregulation and STAT3 activation rather than by altering polarized T helper responses. This study indicates that B cells, by producing cytokines such as IL-10, can act as regulatory cells in immunologically mediated inflammatory reactions.

IL-22 ameliorates intestinal inflammation in a mouse model of ulcerative colitis

Expression of IL-22 is induced in several human inflammatory conditions, including inflammatory bowel disease (IBD). Expression of the IL-22 receptor is restricted to innate immune cells; however, the role of IL-22 in colitis has not yet been defined. We developed what we believe to be a novel microinjection-based local gene-delivery system that is capable of targeting the inflamed intestine. Using this approach, we demonstrated a therapeutic potency for IL-22-mediated activation of the innate immune pathway in a mouse model of Th2-mediated colitis that induces disease with characteristics similar to that of IBD ulcerative colitis (UC). IL-22 gene delivery enhanced STAT3 activation specifically within colonic epithelial cells and induced both STAT3-dependent expression of mucus-associated molecules and restitution of mucus-producing goblet cells. Importantly, IL-22 gene delivery led to rapid amelioration of local intestinal inflammation. The amelioration of disease by IL-22 was mediated by enhanced mucus production. In addition, local gene delivery was used to inhibit IL-22 activity through overexpression of IL-22-binding protein. Treatment with IL-22-binding protein suppressed goblet cell restitution during the recovery phase of a dextran sulfate sodium-induced model of acute colitis. These data demonstrate what we believe to be a novel function for IL-22 in the intestine and suggest the potency of a local IL-22 gene-delivery system for treating UC.

The Transcription Factor T-bet Regulates Mucosal T Cell Activation in Experimental Colitis and Crohn's Disease

The balance between pro and antiinflammatory cytokines secreted by T cells regulates both the initiation and perpetuation of inflammatory bowel diseases (IBD). In particular, the balance between interferon (IFN)-γ/interleukin (IL)-4 and transforming growth factor (TGF)-β activity controls chronic intestinal inflammation. However, the molecular pathways that evoke these responses are not well understood. Here, we describe a critical role for the transcription factor T-bet in controlling the mucosal cytokine balance and clinical disease. We studied the expression and function of T-bet in patients with IBD and in mucosal T cells in various T helper (Th)1- and Th2-mediated animal models of chronic intestinal inflammation by taking advantage of mice that lack T-bet and retroviral transduction techniques, respectively. Whereas retroviral transduction of T-bet in CD62L+ CD4+ T cells exacerbated colitis in reconstituted SCID mice, T-bet–deficient T cells failed to induce colitis in adoptive transfer experiments suggesting that overexpression of T-bet is essential and sufficient to promote Th1-mediated colitis in vivo. Furthermore, T-bet–deficient CD62L− CD4+ T cells showed enhanced protective functions in Th1-mediated colitis and exhibited increased TGF-β signaling suggesting that a T-bet driven pathway of T cell activation controls the intestinal balance between IFN-γ/IL-4 and TGF-β responses and the development of chronic intestinal inflammation in T cell–mediated colitis. Furthermore, TGF-β was found to suppress T-bet expression suggesting a reciprocal relationship between TGF-β and T-bet in mucosal T cells. In summary, our data suggest a key regulatory role of T-bet in the pathogenesis of T cell–mediated colitis. Specific targeting of this pathway may be a promising novel approach for the treatment of patients with Crohns disease and other autoimmune diseases mediated by Th1 T lymphocytes.

Epicutaneous sensitization with protein antigen induces localized allergic dermatitis and hyperresponsiveness to methacholine after single exposure to aerosolized antigen in mice.

Our understanding of the pathogenesis of atopic dermatitis (AD) and its relationship to asthma remains incomplete. Herein, we describe a murine model of epicutaneous (EC) sensitization to the protein allergen, chicken egg albumin, ovalbumin (OVA), which results in a rise in total and OVA-specific serum IgE and leads to the development of a dermatitis characterized by infiltration of CD3(+) T cells, eosinophils, and neutrophils and by local expression of mRNA for the cytokines IL-4, IL-5, and interferon-gamma. A single exposure of the EC sensitized mice to aerosolized OVA induced eosinophilia in the bronchoalveolar lavage fluid and airway hyperresponsiveness to intravenous methacholine as assessed by measurement of pulmonary dynamic compliance (Cdyn). These results suggest a possible role for EC exposure to antigen in atopic dermatitis and in the development of allergic asthma.

Tissue distribution of a coelomic-epithelium-related antigen recognized by the monoclonal antibody OC125

OC125, a murine monoclonal antibody, recognizes an antigenic determinant (CA125) that is associated with >80% of epithelial ovarian neoplasms of serous, endometrioid, clear cell, and undifferentiated types. In the present report, a sensitive biotin-avidin immunoperoxidase technique was used to determine reactivity of OC125 with normal adult and fetal tissues, as well as with neoplasms of nonovarian origin. In fetal tissues, the antibody reacted with amnion and with derivatives of the coelomic epithelium, i.e., the mül-lerian epithelium and the lining cells of the peritoneum, pleura, and pericardium. Among adult tissues, OC125 reacted with the epithelium of fallopian tubes, endometrium, and endocervix. The CA125 determinant was also detected on mesothelial cells in the adult pleura, pericardium, and peritoneum, particularly in areas of inflammation and adhesion. Curiously, the surface ep-ithelium of normal fetal and adult ovaries, thought to be derived from coelomic epithelium, did not express the determinant, except in inclusion cysts, areas of metaplasia, and papillary excrescences. Of neoplastic tissues of nonovarian origin, OC125 reacted consistently only with adenocarcinomas of the endocervix, endometrium, and fallopian tube, and with mesotheliomas. Only seven of 64 nongynecological tumors tested reacted with OC125. Thus, OC125 detects a differentiation antigen shared by fetal coelomic epithelium and its derivatives in the fetus and the adult. Apparently, this antigen disappears early in the course of formation of the ovarian epithelium and is reexpressed in certain reactive and neoplastic lesions, a process that could be termed “molecular metaplasia.”

A Case for Regulatory B Cells

B cells are typically characterized by their ability to produce Abs, including autoantibodies. However, B cells possess additional immune functions, including the production of cytokines and the ability to function as a secondary APC. As with T cells, the B cell population contains functionally distinct subsets capable of performing both pathogenic and regulatory functions. Recent studies indicate that regulatory B cells develop in several murine models of chronic inflammation, including inflammatory bowel disease, rheumatoid arthritis, and experimental autoimmune encephalomyelitis. The regulatory function may be directly accomplished by the production of regulatory cytokines IL-10 and TGF-β and/or by the ability of B cells to interact with pathogenic T cells to dampen harmful immune responses. In this review, we make a case for the existence of regulatory B cells and discuss the possible developmental pathways and functional mechanisms of these B cells.

Wiskott-Aldrich Syndrome Protein-Deficient Mice Reveal a Role for WASP in T but Not B Cell Activation

The Wiskott-Aldrich syndrome (WAS) is a human X-linked immunodeficiency resulting from mutations in a gene (WASP) encoding a cytoplasmic protein implicated in regulating the actin cytoskeleton. To elucidate WASP function, we disrupted the WASP gene in mice by gene-targeted mutation. WASP-deficient mice showed apparently normal lymphocyte development, normal serum immunoglobulin levels, and the capacity to respond to both T-dependent and T-independent type II antigens. However, these mice did have decreased peripheral blood lymphocyte and platelet numbers and developed chronic colitis. Moreover, purified WASP-deficient T cells showed markedly impaired proliferation and antigen receptor cap formation in response to anti-CD3epsilon stimulation. Yet, purified WASP-deficient B cells showed normal responses to anti-Ig stimulation. We discuss the implications of our findings regarding WASP function in receptor signaling and cytoskeletal reorganization in T and B cells and compare the effects of WASP deficiency in mice and humans.

Roles of TH1 and TH2 cytokines in a murine model of allergic dermatitis

Skin lesions in atopic dermatitis (AD) are characterized by hypertrophy of the dermis and epidermis, infiltration by T cells and eosinophils, and expression of the cytokines IL-4, IL-5, and IFN-gamma. The role of these cytokines in the pathogenesis of AD is not known. We took advantage of a recently described murine model of AD elicited by epicutaneous sensitization with ovalbumin (OVA) (1) and of the availability of mice with targeted deletions of the IL-4, IL-5, and IFN-gamma cytokine genes to assess the role of these cytokines in this model.OVA-sensitized skin from IL-5(-/-) mice had no detectable eosinophils and exhibited decreased epidermal and dermal thickening. Sensitized skin from IL-4(-/-) mice displayed normal thickening of the skin layers but had a drastic reduction in eosinophils and a significant increase in infiltrating T cells. These findings were associated with a reduction in eotaxin mRNA and an increase in mRNA for the T-cell chemokines macrophage inflammatory protein-2 (MIP-2), MIP-1beta, and RANTES. Sensitized skin from IFN-gamma-/- mice was characterized by reduced dermal thickening. These results suggest that both the TH2 cytokines IL-4 and IL-5 and the TH1 cytokine IFN-gamma play important roles in the inflammation and hypertrophy of the skin in AD.

Development of chronic colitis is dependent on the cytokine MIF

The cytokine macrophage-migration inhibitory factor (MIF) is secreted by a number of cell types upon induction by lipopolysaccharide (LPS). Because colitis is dependent on interplay between the mucosal immune system and intestinal bacteria, we investigated the role of MIF in experimental colitis. MIF-deficient mice failed to develop disease, but reconstitution of MIF-deficient mice with wild-type innate immune cells restored colitis. In addition, established colitis could be treated with anti-MIF immunoglobulins. Thus, murine colitis is dependent on continuous MIF production by the innate immune system. Because we found increased plasma MIF concentrations in patients with Crohns disease, these data suggested that MIF is a new target for intervention in Crohns disease.