T cell-associated CD18 but not CD62L, ICAM-1, or PSGL-1 is required for the induction of chronic colitis.

The induction and perpetuation of chronic colitis are thought to involve a complex set of adhesive interactions between T cells and endothelial cells located on the vasculature within secondary lymphoid tissue and the intestine. The objective of this study was to assess the roles of T cell-associated CD18, CD62L (L-selectin), ICAM-1, and P-selectin glycoprotein ligand-1 (PSGL-1) in the induction of chronic colitis in mice. CD4(+)CD25(-) T cells derived from either wild-type (WT), CD18-deficient [CD18 knockout (KO)], CD62L KO, ICAM-1 KO, or PSGL-1 KO mice were adoptively transferred into recombinase activating gene-1 (RAG-1)-deficient mice (RAG KO mice) to assess the potential of these T cells to induce chronic colitis. At 8-10 wk following T cell transfer, we observed moderate to severe colitis as assessed by increases in colon weight-to-length ratios and by blinded histopathological analysis. In contrast, we found that transfer of CD18 KO T cells into RAG KO recipients resulted in the significant attenuation of colonic inflammation in these mice. Furthermore, we observed fewer infiltrating CD4(+) T cells in the colonic lamina propria in the CD18 KO-->RAG KO group compared with the WT-->RAG KO group. Finally, message levels of colonic TNF-alpha, IL-1beta, and IFN-gamma were significantly reduced in CD18 KO-->RAG KO mice compared with colitic control animals. We conclude that T cell-associated CD18, but not CD62L, ICAM-1, or PSGL-1, is required for the development of chronic colitis.

Regulation of dextran sodium sulfate induced colitis by leukocyte beta 2 integrins.

Inflammatory bowel diseases (IBD) are chronic inflammatory disorders whose etiology remains unknown. Reports have shown that infiltration of leukocytes into intestinal tissue is a pathognomonic hallmark for this disease. Leukocyte beta(2) integrins are heterodimeric adhesion membrane proteins that are exclusively expressed on leukocytes and participate in immune cell adhesion and activation. In this study, we examined the pathophysiological role of the beta(2) integrins CD18, CD11a, and CD11b in the pathogenesis of dextran sodium sulfte (DSS)-induced experimental colitis. Disease activity was measured by daily assessment of clinical parameters including stool consistency, weight loss, occult blood, and gross rectal bleeding. Histopathological changes including severity of inflammation, surface epithelial/crypt damage, and depth of injury were also determined. The CD18 null and CD11a null mice had significantly lower disease activity and cumulative histopathological scores compared to wild-type mice. Interestingly, CD11b null mice did not show protection against DSS colitis and displayed increased disease activity compared to wild-type mice. Examination of specific leukocyte populations in the distal colon from various mice revealed significant attenuation of neutrophil and macrophage infiltrates in CD18, CD11a, and CD11b null mice. Surprisingly, the CD11b null mice showed a significant increase in plasma cell infiltration in response to DSS suggesting that this molecule may influence plasma cell function during colitis. This study demonstrates that genetic loss of CD18 or CD11a is protective during experimental colitis and that CD11b may serve a regulatory role during development of disease.

Role of T-cell-associated lymphocyte function-associated antigen-1 in the pathogenesis of experimental colitis.

The beta2 integrin lymphocyte function-associated antigen-1 (LFA-1; CD11a/CD18) is important for lymphocyte trafficking and activation as well as recruitment to sites of tissue inflammation. The objective of this study was to assess the role of 'T-cell-associated' LFA-1 in the pathogenesis of chronic colitis in vivo. Transfer of CD4+CD25- T cells isolated from wild-type (wt) mice into immunodeficient recipients [recombinase-activating gene-1-deficient (RAG-1-/-] produced moderate to severe colitis, whereas RAG-1-/- mice injected with CD11a-deficient (CD11a-/-; LFA-1-/-) donor T cells displayed minimal macroscopic and histological evidence of colitis. Surface expression of L-selectin, alpha4, alpha4beta7 and chemokine receptor-7 were similar for wt and CD11a-/- donor T cells. Attenuated disease in the CD11a-/- --> RAG-1-/- animals was associated with decreased numbers of CD4+ T cells in the mesenteric lymph nodes (MLNs), spleen and intestinal lamina propria (LP). In addition, significant reductions in Th1 cytokines were observed following ex vivo stimulation of mononuclear cells obtained from the MLNs and colonic LP. Interestingly, mononuclear cells obtained from the spleens of CD11a-/- --> RAG-1-/- exhibited enhanced pro-inflammatory cytokine production compared with splenocytes obtained from wt --> RAG-1-/- colitic mice. Taken together, our data suggest that T-cell-associated CD11a (LFA-1) expression plays a dual role in the initiation of chronic gut inflammation by facilitating naive T-cell priming/activation and expansion within MLNs and by augmenting pro-inflammatory cytokine production following secondary stimulation by antigen-presenting cells in the colonic interstitium.

T cell-induced inflammation of the small and large intestine in immunodeficient mice.

It is well known that transfer of CD4+CD45RBhigh (naïve) T cells into syngeneic lymphocyte-deficient mice induces chronic colitis. However, no studies have reported the presence of small bowel inflammation in this T cell-dependent model. Therefore, the objective of this study was to evaluate and compare small and large bowel inflammation induced by transfer of naïve T cells into two different immunodeficient recipient mice. T and B cell-deficient recombinase activating gene 1-deficient [RAG knockout (KO)] and T cell-deficient T cell receptor-beta x T cell receptor-delta double-deficient (TCR KO) mice were reconstituted with wild-type naïve T cells and observed for signs of disease. We found that reconstituted RAG KO mice developed moderate to severe colitis and inflammation of the entire small intestine at 6-8 wk after T cell transfer. Adoptive transfer of naïve T cells into TCR KO mice induced a milder form of chronic colitis and small bowel inflammation that was confined primarily to the duodenum at 10-12 wk after T cell transfer. T helper cell 1 and macrophage-derived proinflammatory cytokine mRNA levels correlated well with the localization and severity of the chronic large and small bowel inflammation. In addition, we observed comparable homing and expansion of donor lymphocytes in the gut and secondary lymphoid tissues of both recipients. Taken together, our data demonstrate that transfer of naïve T cells into immunodeficient recipient mice induces both chronic small and large bowel inflammation and that the presence of B cells in the TCR KO recipients may play a role in regulating chronic intestinal inflammation.

Estrogen receptor-alpha, sexual dimorphism and reduced-size liver ischemia and reperfusion injury in mice.

Estrogen (E(2)) exerts its effect on target organs principally by interacting with specific estrogen receptors (ER) such as ER-alpha or ER-beta. The role that these E(2) receptors play in mediating the protective effects observed in RSL+I/R induced injury remains to be defined. To study the role of ER-alpha, we anesthetized female and male wild type (wt; C57Bl/6) and ER-alpha-deficient (alphaERKO) mice and subjected them to 70% liver ischemia for 45 min followed by resection of the remaining 30% nonischemic lobes and reperfusion of the ischemic tissue. For some experiments, wt and alphaERKO male mice were injected with E(2). Survival was monitored on a daily basis while liver injury was assessed by quantifying serum alanine aminotransferase (ALT) levels and histopathology. Hepatic eNOS mRNA levels were evaluated using semi-quantitative RT-PCR. Our data showed that untreated females or males treated with E(2) survived RSL+I/R surgery indefinitely whereas all male mice given vehicle died within 3-5 days following surgery. This protective effect was diminished in alphaERKO female mice such that only 40% of alphaERKO females survived 7 d following RSL+I/R. Furthermore, liver injury was significantly higher in alphaERKO females compared with their wt counterparts and similar to those seen in wild type males and alphaERKO males. The protective effect observed in wild type females or E(2) treated males correlated well with increases in hepatic eNOS message whereas both male and female alphaERKO mice exhibited significantly lower levels of eNOS mRNA. We conclude that this protection may in part be due to the E(2)/ER-alpha-mediated activation of eNOS.

Regulation of chronic colitis in athymic nu/nu (nude) mice.

The objective of this study was to assess the roles of NK cells, B cells and/or intraepithelial lymphocytes (IEL) in suppressing the development of colitis in nude mice reconstituted with CD4(+)CD45RB(high) T cells. BALB/c nude mice were lethally irradiated and reconstituted with bone marrow from different immunodeficient mice to generate athymic chimeras devoid of one or more lymphocyte populations. Transfer of CD4(+)C45RB(high) T cells into chimeric recipients devoid of B cells, T cells and IEL produced severe colitis within 6-8 weeks, whereas transfer of these same T cells into B cell- and T cell-deficient or T cell-deficient chimeras produced little to no gut inflammation. In addition, we found that nude mice depleted of NK cells or RAG-1(-/-) mice reconstituted with IEL failed to develop colitis following transfer of CD45RB(high) T cells. Severe colitis could, however, be induced in nude mice by transfer of activated/T(h)1 CD4(+)CD45RB(low) T cells. Taken together, our data suggest that IEL, but not B cells or NK cells, play an important role in suppressing the development of chronic colitis in this model. In addition, our data demonstrate that suppression of disease may be due to polarization of naive CD4(+) cells toward a non-pathogenic and/or regulatory phenotype.

Sexual dimorphism in reduced-size liver ischemia and reperfusion injury in mice: role of endothelial cell nitric oxide synthase.

We have recently reported that female mice are protected to a much greater extent from the injurious effects of reduced-size liver ischemia and reperfusion (RSL+I/R) than are males by an estrogen-dependent mechanism. The objective of this study was to examine the possibility that the protective effect observed in female mice depends on the up-regulation and/or activation of endothelial cell NO synthase (eNOS). Anesthetized female and male wild-type or eNOS-deficient C57BL/6 mice were subjected to 70% liver ischemia for 45 min followed by resection of the remaining 30% nonischemic lobes and reperfusion of ischemic tissue. Survival was monitored daily, whereas liver injury was quantified by using serum alanine aminotransferase determinations and histopathology. Hepatic eNOS mRNA, protein, and enzymatic activity were determined in male and female mice subjected to RSL+I/R. We found that liver injury was reduced and survival increased in female mice compared with males. This protective effect correlated with significant increases in hepatic eNOS message levels and enzyme activity but not protein expression compared with males subjected to the surgery. Furthermore, N(omega)-nitro-L-arginine methyl ester-treated or eNOS-deficient female mice responded to RSL+I/R with dramatic increases in liver injury and 100% mortality within 2 days of surgery. Finally, we found that pravastatin pretreatment significantly attenuated hepatocellular injury and increased survival of male mice, which was associated with enhanced expression of eNOS message. We conclude that the protective effect afforded female mice is due to the activation of hepatic eNOS activity and enhanced NO production.

Regulation of postischemic liver injury following different durations of ischemia.

The objective of this study was to define the relationship among Kupffer cells, O(2)(-) production, and TNF-alpha expression in the pathophysiology of postischemic liver injury following short and long periods of ischemia. Using different forms of superoxide dismutase with varying circulating half-lives, a monoclonal antibody directed against mouse TNF-alpha, and NADPH oxidase-deficient mice, we found that 45 or 90 min of partial (70%) liver ischemia and 6 h of reperfusion (I/R) produced time-dependent increases in liver injury and TNF-alpha expression in the absence of neutrophil infiltration. Furthermore, we observed that hepatocellular injury induced by short periods of ischemia were not dependent on formation of TNF-alpha but were dependent on Kupffer cells and NADPH oxidase-independent production of O(2)(-). However, liver injury induced by extended periods of ischemia appeared to require the presence of Kupffer cells, NADPH oxidase-derived O(2)(-), and TNF-alpha expression. We conclude that the sources for O(2)(-) formation and the relative importance of TNF-alpha in the pathophysiology of I/R-induced hepatocellular injury differ depending on the duration of ischemia.

Role of nitric oxide in liver ischemia and reperfusion injury.

The present study was designed to assess the role of endothelial cell and inducible nitric oxide synthase (eNOS, iNOS)-derived NO in ischemia/reperfusion (I/R)-induced pro-inflammatory cytokine expression and tissue injury in a murine model of hepatic I/R. Forty-five min of partial hepatic ischemia and 3 h of reperfusion resulted in a significant increase in liver injury as assessed by serum alanine aminotransferase and histopathology which occurred in the absence of neutrophil infiltration. Both iNOS and eNOS deficient mice exhibited enhanced liver injury when compared to their wild type (wt) controls again in the absence of neutrophil infiltration. Interestingly, message expression for both tumor necrosis factor-alpha (TNF-alpha) and interleukin 12 (IL-12) were enhanced in eNOS, but not iNOS-deficient mice at 1 h post-ischemia when compared to their wt controls. In addition, eNOS message expression appeared to be up-regulated between 1 and 3 h ofreperfusion in wt mice while iNOS deficient mice exhibited substantial increases at I but not 3 h. Taken together, these data demonstrate the ability of eNOS and iNOS to protect the post-ischemic liver, however their mechanisms of action may be very different.

Nitric oxide and chronic gut inflammation: controversies in inflammatory bowel disease.

One of the most consistent and dramatic findings in both experimental and human inflammatory bowel disease (e.g., Crohn's disease, ulcerative colitis) is the enhanced expression of the inducible isoform of nitric oxide synthase (iNOS) and the sustained overproduction of the free radical nitric oxide (NO). The role that iNOS-derived NO plays in the pathophysiology of inflammatory bowel disease remains the subject of intense investigation and active debate. Although several different studies using a variety of animal models of acute and chronic gut inflammation suggest that NO may promote intestinal inflammation, an equally impressive number of investigations suggest that iNOS may play no role or may act to attenuate or to limit the extent of inflammatory tissue injury. This review discusses some of the basic concepts related to the immunoregulation of chronic gut inflammation and summarizes the current state of knowledge of the role that NO may play in modulating inflammatory tissue injury.

Role of reactive metabolites of oxygen and nitrogen in inflammatory bowel disease.

The inflammatory bowel diseases (IBD; Crohn's disease, ulcerative colitis) are a collection of chronic idiopathic inflammatory disorders of the intestine and/or colon. Although the pathophysiology of IBD is not known with certainty, a growing body of experimental and clinical data suggests that chronic gut inflammation may result from a dysregulated immune response to normal bacterial antigens. This uncontrolled immune system activation results in the sustained overproduction of reactive metabolites of oxygen and nitrogen. It is thought that some of the intestinal and/or colonic injury and dysfunction observed in IBD is due to elaboration of these reactive species. This review summarizes the current state-of-knowledge of the role of reactive oxygen species and nitric oxide in the pathophysiology of IBD.