Donna Rennick

Interleukin-10-deficient mice develop chronic enterocolitis

Interleukin-10 (IL-10) affects the growth and differentiation of many hemopoietic cells in vitro; in particular, it is a potent suppressor of macrophage and T cell functions. In IL-10-deficient mice, generated by gene targeting, lymphocyte development and antibody responses are normal, but most animals are growth retarded and anemic and suffer from chronic enterocolitis. Alterations in intestine include extensive mucosal hyperplasia, inflammatory reactions, and aberrant expression of major histocompatibility complex class II molecules on epithelia. In contrast, mutants kept under specific pathogen-free conditions develop only a local inflammation limited to the proximal colon. These results indicate that the bowel inflammation in the mutants originates from uncontrolled immune responses stimulated by enteric antigens and that IL-10 is an essential immunoregulator in the intestinal tract.

Enterocolitis and colon cancer in interleukin-10-deficient mice are associated with aberrant cytokine production and CD4(+) TH1-like responses.

We have characterized the progressive stages of chronic intestinal inflammation that develops spontaneously in specific pathogen-free (SPF) mice with a targeted disruption in the IL-10 gene (IL-10-/-). Our longitudinal studies showed that inflammatory changes first appear in the cecum, ascending and transverse colon of 3-wk-old mutants. As the disease progressed, lesions appeared in the remainder of the colon and in the rectum. Some aged IL-10-/- mice also developed inflammation in the small intestine. Prolonged disease with transmural lesions and a high incidence of colorectal adenocarcinomas (60%) was observed in 6-mo-old mutants. Mechanistic studies have associated uncontrolled cytokine production by activated macrophages and CD4+ Th1-like T cells with the enterocolitis exhibited by IL-10-/- mice. A major role for a pathogenic Th1 response was further suggested by showing that anti-IFNgamma antibody (Ab) treatment significantly attenuated intestinal inflammation in young IL-10-/- mice. When weanlings were treated with IL-10, they failed to develop any signs of intestinal inflammation. Interestingly, IL-10 treatment of adults was not curative but did ameliorate disease progression. Our studies have also shown that inheritable factors strongly influence the disease susceptibility of IL-10-/- mice. In 3-mo-old mutants, intestinal lesions were most severe in IL-10-/- 129/SvEv and IL-10-/- BALB/c strains, of intermediate severity in the IL-10-/- 129 x C57BL/6J outbreds, and least severe in the IL-10-/- C57BL/6J strain.

IL-25 Induces IL-4, IL-5, and IL-13 and Th2-Associated Pathologies In Vivo

We have characterized a cytokine produced by Th2 cells, designated as IL-25. Infusion of mice with IL-25 induced IL-4, IL-5, and IL-13 gene expression. The induction of these cytokines resulted in Th2-like responses marked by increased serum IgE, IgG(1), and IgA levels, blood eosinophilia, and pathological changes in the lungs and digestive tract that included eosinophilic infiltrates, increased mucus production, and epithelial cell hyperplasia/hypertrophy. In addition, our studies show that IL-25 induces Th2-type cytokine production by accessory cells that are MHC class II(high), CD11c(dull), and lineage(-). These results suggest that IL-25, derived from Th2 T cells, is capable of amplifying allergic type inflammatory responses by its actions on other cell types.

Interleukin-10 is a central regulator of the response to LPS in murine models of endotoxic shock and the Shwartzman reaction but not endotoxin tolerance.

Previous studies in vivo have shown that IL-10 infusion can prevent lethal endotoxic shock. Mice deficient in the production of IL-10 (IL10T) were used to investigate the regulatory role of IL-10 in the responses to LPS in three experimental systems. In a model of acute endotoxic shock, it was found that the lethal dose of LPS for IL10T mice was 20-fold lower than that for wild type (wt) mice suggesting that endogenous IL-10 determines the amount of LPS which can be tolerated without death. The high mortality rate of IL10T mice challenged with modest doses of LPS was correlated to the uncontrolled production of TNF as treatment with anti-TNF antibody (Ab) resulted in 70% survival. Additional studies suggested that IL-10 mediates protection by controlling the early effectors of endotoxic shock (e.g., TNF alpha) and that it is incapable of directly antagonizing the production and/or actions of late appearing effector molecules (e.g., nitric oxide). We also found that IL10T mice were extremely vulnerable to a generalized Shwartzman reaction where prior exposure to a small amount of LPS primes the host for a lethal response to a subsequent sublethal dose. The priming LPS dose for IL10T mice was 100-fold lower than that required to prime wt mice implying that IL-10 is important for suppressing sensitization. In agreement with this assumption, IL-10 infusion was found to block the sensitization step. Interestingly, IL-10 was not the main effector of endotoxin tolerance as IL10T mice could be tolerized to LPS. Furthermore, IL-10 infusion could not substitute for the desensitizing dose of LPS. These results show that IL-10 is a critical component of the hosts natural defense against the development of pathologic responses to LPS although it is not responsible for LPS-induced tolerance.

Molecular Biology of Interleukin 4 and Interleukin 5 Genes and Biology of their Products that Stimulate B Cells, T Cells and Hemopoietic Cells

Lymphokines produced by helper T cells activated by antigen mediate numerous efTector functions of T cells. Unlike immunoglobulin or the T cell receptor, lymphokines do not bear antigen specificity and stimulate proliferation and differentiation of lymphocytes and hemopoietic cells (Arai et al. 1986). Since many lymphokines are composed of single polypeptide chains, their coding sequences can be isolated by functional expression in appropriate host cells. Using a pcD cDNA expression vector (Okayama & Berg 1983), we have developed a screening procedure employing transfection of plasmid DNAs into mammalian cells followed by assay of transfected cell supernatants for lymphokine activities of interest (Yokota et al. 1984. 1985, 1987c). Based on this expression cloning protocol, many T cell lymphokine genes have been isolated and their primary structures determined. These studies have revealed a regulatory network formed between lymphoid cells and hemopoietic cells through the action of multiple lymphokines produced by activated T cells. For example, interleukin 2 (IL-2) stimulates predominantly T cells whereas interleukin 3 (lL-3) and granulocytemacrophage colony stimulating factor (GM-CSF) stimulate hemopoietic cells. A number of molecules stimulate B cells. Among them, interleukin 4 (IL-4), interleukin 5 (IL-5) and B cell stimulatory factor 2 (BSF-2) have been chemically defined by molecular cloning (Lee et al. 1986, Noma et al. 1986, Yokota et al. !986,

A single bone marrow-derived stromal cell type supports the in vitro growth of early lymphoid and myeloid cells

A clonal cell line (ALC) derived from murine bone marrow stroma is capable of supporting the continuous, in vitro growth of early lymphoid and myeloid cell populations. The growth-promoting effects of ALC are in part mediated through M-CSF and a pre-B cell growth factor, both of which accumulate in ALC-culture supernatant. To analyze the lymphoid growth factor produced by ALC cells, we derived a pre-B cell indicator line that is dependent on ALC-growth-conditioned medium. Using a combination of biological and biochemical analyses, we have established that the pre-B cell growth factor produced by ALC cells is distinct from IL-1, IL-2, IL-3, and IL-4 (BSF-1), suggesting that the early stages of B-cell development are regulated by a unique stroma-derived growth factor.

Chronic Colitis in IL-10-/- Mice: Insufficient Counter Regulation of a Th1 Response

IL-10-deficient (IL-10-/-) mice, generated by a gene-targeted mutation, develop abnormal immune responses as a result of uncontrolled interactions between antigen presenting cells and lymphocytes. The studies reviewed herein have focused on the enterocolitis that spontaneously develops in IL-10-/- mice. Not unexpectedly, heightened production of proinflammatory mediators accompanied pathologic changes in the gastrointestinal tract of young mutants. In a series of studies, the proinflammatory mediators responsible for initiating the pathogenic response were distinguished from those that were elicited as a consequence of persistent inflammation. We have also investigated the possibility that different mediators are involved in the inductive versus the maintenance phase of disease. The findings of these mechanistic studies as they relate to our understanding of progressive inflammatory disease and the role of IL-10 in controlling the acute and chronic stages are discussed.

The Role of IL-10 in Inflammatory Bowel Disease: "Of Mice and Men"

Inflammatory bowel disease (IBD) is a generic term typically used to describe a group of idiopathic inflammatory intestinal conditions in humans that are generally divided into Crohns disease and ulcerative colitis. Although the etiology of these diseases remains unknown, a number of rodent models of IBD have recently been identified, all sharing the concept that the development of chronic intestinal inflammation occurs as a consequence of alterations in the immune system that lead to a failure of normal immunoregulation in the intestine. On the basis of these models, it has been hypothesized that the development of IBD in humans may be related to a dysregulated immune response to normal flora in the gut. Immunodeficient scid mice injected with CD4+ CD45RBhigh T cells and mice deficient in interleukin (IL)-10 (IL-10-/-) are among the rodent models of IBD. In both models, there is inflammation and evidence of a Th1-like response in the large intestine, characterized by CD4+ T-cell and macrophage infiltrates, and elevated levels of interferon-γ. Because IL-10 is an immunomodulatory cytokine that is capable of controlling Thl-like responses, the role of IL-10 was investigated in these models. IL-10 was shown to be important in regulating the development of intestinal inflammation in both models. These results provided key data that supported initiation of clinical trials evaluating the efficacy of IL-10 in patients with IBD.

Interleukin 10: An overview

Since the original description of interleukin-10, a wealth of information concerning its biological properties has been gathered. Studies in vitro have rapidly identified both immunostimulatory and immunosuppressive activities for IL-10. Based on these findings, in vivo studies were initiated in a variety of animal disease models to assess the importance of these activities. This review will summarize the pleiotropic properties of IL-10 and will survey current research regarding the potential of IL-10 to regulate acute and chronic inflammatory reactions.

Acquisition of CD24 expression by Lin−CD43+B220lowckithi cells coincides with commitment to the B cell lineage

The present study describes three subsets of bone marrow‐derived Lin−CD43+B220low (B220low) progenitor cells which represent distinct stages in hematopoietic development. These populations differ in their expression of CD24 and ckit and the occurrence of IgH gene rearrangement. B220low CD24−ckithi progenitors have their IgH loci in germ‐line configuration and are multipotent since they can give rise to B cells, T cells and myeloid cells. B220lowckithi cells which have acquired CD24 expression have retained IgH loci in germ‐line configuration and the ability to generate B cells, however, they have lost the ability to generate T cells and myeloid cells. Thus acquisition of CD24 by B220low cells occurs concurrently with the transition from a multipotent to a lineage‐restricted progenitor population. B220lowCD24+ cells expressing low levels of ckit are also lineage restricted, giving rise to only B cells and have begun D‐JH rearrangement, implying that initiation of IgH rearrangement coincides with the down‐regulation of ckit expression.