A dominant,coordinated T regulatory cell-IgA response to the intestinal microbiota

A T cell receptor transgenic mouse line reactive to a microbiota flagellin, CBir1, was used to define mechanisms of host microbiota homeostasis. Intestinal IgA, but not serum IgA, was found to block mucosal flagellin uptake and systemic T cell activation in mice. Depletion of CD4⁺CD25⁺ Tregs decreased IgA⁺ B cells, total IgA, and CBir1-specific IgA in gut within days. Repletion of T cell-deficient mice with either CD4⁺CD25⁺ or CD4⁺foxp3⁺ Tregs restored intestinal IgA to a much greater extent than their reciprocal CD4⁺ subsets, indicating that Tregs are the major helper cells for IgA responses to microbiota antigens such as flagellin. We propose that the major role of this coordinated Treg-IgA response is to maintain commensalism with the microbiota.     

Alterations of intestinal immune function and regulatory effects of L-arginine in experimental severe acute pancreatitis rats

ABM: To discuss the changes of intestinal mucosal immune function in rats with experimental severe acute pancreatitis (SAP) and the regulatory effect of L-arginine. METHODS: Male adult Wistar rats were randomly divided into pancreatitis group, sham-operation group, and L-arginine treatment group. Animals were killed at 24, 48, and 72 h after SAP models were developed and specimens were harvested. Endotoxin concentration in portal vein was determined by limulus endotoxin analysis kit. CD3+, CD4+, CD8+ T lymphocytes in intestinal mucosal lamina propria were examined by immunohistochemistry. Secretory immunoglobulin A (SIgA) in cecum feces was examined by radioimmunoassay. RESULTS: Compared to the control group, plasma endotoxin concentration in the portal vein increased, percentage of CD3+ and CD4+ T lymphocyte subsets in the end of intestinal mucosal lamina propria reduced significantly, CD4+/CD8+ ratio decreased, and SIgA concentrations in cecum feces reduced at 24, 48, and 72 h after SAP developed. Compared to SAP group, the L-arginine treatment group had a lower level of plasma endotoxin concentration in the portal vein, a higher CD3+ and CD4+ T lymphocyte percentage in the end of intestinal mucosal lamina propria, an increased ratio of CD4+/CD8+ and a higher SIgA concentration in cecum feces. CONCLUSION: Intestinal immune suppression occurs in the early stage of SAP rats, which may be the main reason for bacterial and endotoxin translocation. L-arginine can improve the intestinal immunity and reduce bacterial and endotoxin translocation in SAP rats.     

The continuum of intestinal CD4+ T cell adaptations in host-microbial mutualism

How a mutualistic relationship between the intestinal microbiota and intestinal T cell compartments is established is important, as a breakdown of intestinal T cell homeostasis may cause inflammatory bowel diseases. A number of studies have shown that different bacterial species modulate the intestinal CD4⁺ T cell compartment in different ways. We performed mechanistic in vivo studies that demonstrated the crucial requirement for regulatory T cells (Treg) and interleukin-10 (IL-10) in the induction of intestinal T cell homeostasis even following colonization with a completely benign microbiota. In the absence of a functional Treg response or IL-10 receptor signaling, the same bacteria that induced a Treg response in wild-type animals now induced T helper type 17 responses, without intestinal inflammation. Therefore, Treg, IL-10 and Th17 are crucial regulatory mechanisms in the intestine not only for controlling inflammation, but also to establish a continuum of CD4⁺ T cell homeostasis upon commensal colonization.     

Gut microbiota amplifies host-intrinsic conversion from the CD8 T cell lineage to CD4 T cells for induction of mucosal immune tolerance

Microbiota has been shown to promote tolerogenic differentiation of T lymphocytes. It remains unclear to what extent microbiota triggers de novo re-programming or amplify pre-existing plasticity intrinsic to T cells. In a study with mouse models to track the clonal fate of CD4 and CD8 T cells, we discovered that CD8 T cells converted to MHC class I-restricted CD4 T cells without regard to selfness of their antigen specificity. In mesenteric lymph nodes (MLN), CD8 T cells converted to CD4⁺Foxp3⁺ regulatory T (T_reg) cells which were enriched in the large intestine lamina propria (LILP) and suppressed chemical- or immune-mediated inflammatory damage. In germ-free conditions, the converted CD4 populations were present in MLN, but absent in LILP. Therefore, an intrinsic plasticity in the host was amplified by the gut microbiota, leading to selfless tolerance induction in the intestinal mucosa. The findings may be relevant to HIV infection, cancer and autoimmune disorders.     

Increased numbers of Foxp3-positive regulatory T cells in gastritis, peptic ulcer and gastric adenocarcinoma

AIM: To determine the number of regulatory T cells (Tregs) in gastric mucosa of patients with gastritis, peptic ulcers and gastric cancer.METHODS: This study was a retrospective analysis of gastric antrum biopsy specimens from healthy controls (n = 22) and patients with gastritis (n = 30), peptic ulcer (n = 83), or gastric cancer (n = 32). Expression of CD4, CD25 and Foxp3 was determined by immunohistochemistry in three consecutive sections per sample.RESULTS: Compared with healthy controls, there was an increased number of CD25⁺ and Foxp3⁺ cells in patients with gastritis (P = 0.004 and P = 0.008), peptic ulcer (P < 0.001 and P < 0.001), and gastric cancer (P < 0.001 and P < 0.001). The ratio of CD25⁺/CD4⁺ or Foxp3⁺/CD4⁺ cells was also significantly higher in all disease groups (P < 0.001, respectively). The number of CD4⁺, CD25⁺, and Foxp3⁺ cells, and the ratio of CD25⁺/CD4⁺ and Foxp3⁺/CD4⁺ cells, were associated with the histological grade of the specimens, including acute inflammation, chronic inflammation, lymphoid follicle number, and Helicobacter pylori infection. The number of CD4⁺, CD25⁺ and Foxp3⁺ cells, and the ratio of CD25⁺/CD4⁺ and Foxp3⁺/CD4⁺ cells, were negatively associated with intestinal metaplasia among gastritis (P < 0.001, P < 0.001, P < 0.001, P = 0.002 and P = 0.002) and peptic ulcer groups (P = 0.013, P = 0.004, P < 0.001, P = 0.040 and P = 0.003).CONCLUSION: Tregs are positively associated with endoscopic findings of gastroduodenal diseases and histological grade but negatively associated with intestinal metaplasia in gastritis and peptic ulcer groups.     

Reversal of altered intestinal mucosal immunity in rats fed elemental diet by supplementation of oleic acid

We have previously demonstrated that elemental diet (ED) induces decreased lymphocyte transport in intestinal lymph and significant changes in T cell subsets and the number of IgA-containing cells in gut-associated lymphoid tissues of rats. In order to examine whether the low fat content contributes to the induction of immunological changes in gut-associated lymphoid tissues, the effects of additional fatty acid in the ED were investigated. Rats were divided into four groups: elemental diet alone, elemental diet supplemented with 5% oleic acid (OA), elemental diet with 10% OA and conventional diet as a control. These diets were given at the same daily calorie intake for 4 weeks. The flow rate of intestinal lymph showed no significant difference between the four groups. However, lymphocyte flux as well as the percentage of CD3⁺ and CD4⁺ cells were significantly greater in the control and the 10% OA groups than in the ED and 5% OA groups. Intestinal lymph showed decreased concentrations of IgG and IgA in the ED group, whereas the addition of 10% OA significantly attenuated the decrease in these levels. In mesenteric lymph nodes, the CD4⁺/CD8⁺ ratio was significantly decreased in the ED group, but 10% OA reversed this change. Immunohistochemical analysis of the ileal mucosa showed that in the ED group the population of CD4⁺ cells was decreased, while the number of CD8⁺ cells was increased. Supplementation of OA to ED produced similar stepwise attenuation of the changes in lymphocyte subpopulations in the lamina propria, while the 10% OA group reached levels that were not statistically different from controls. In the elemental diet, group, there was a significant decrease in immunoglobulin-containing cells of the IgA class in the lamina propria of the intestine. Similarly, the addition of OA induced dose-dependent recovery in the number of IgA-containing cells. These results suggest that a low dietary concentration of fat may be closely related to changes in lymphocyte transport in intestinal lymph and mucosal immunity of intestinal mucosa induced by the feeding of a long-term ED.     

Characteristics of intestinal lamina propria dendritic cells in a mouse model of postinfectious irritable bowel syndrome

Background and Aim: Postinfectious irritable bowel syndrome (PI‐IBS), which results from inflammation has been emphasized a lot recently. Dendritic cells (DCs) may contribute to intestinal mucosal immune activation in the pathogenesis of PI‐IBS. This study tested the hypothesis that phenotype and function of intestinal lamina propria DCs (LPDCs) changed in the development of a PI‐IBS mouse model. Methods: Mice infected with Trichinella spiralis underwent abdominal withdrawal reflex (AWR) to evaluate visceral sensitivity. LPDCs were isolated and purified by intestine digestion and magnetic label‐based technique. Surface markers were analyzed by flow cytometry. Endocytic activity, mixed lymphocyte reaction (MLR) and chemotaxis were studied. Cytokine production of the LPDCs cocultured with CD4⁺ T cells was determined. Results: Intestinal inflammation resolved after 8 weeks infection with sustained visceral hyperalgesia. Surface markers CD86 and MHCII were lower in the acute infection group, but increased in the PI‐IBS stage. Enhanced ability of endocytic activity and decreased abilities to attract and stimulate CD4⁺ T cell proliferation were in the acute infection group. However, LPDCs in the PI‐IBS stage showed weakened endocytic ability with enhanced abilities to attract and stimulate CD4⁺ T cell proliferation. Cocultured LPDCs with CD4⁺ T cells showed a predominant Th2 response in the acute infection stage, and more important roles of Th1, Th17 responses in the PI‐IBS stage. Conclusions: The hypothesis was supported that the phenotype and function of LPDCs changed in the development of PI‐IBS, which induced the maintenance of intestinal mucosal immune activation and might provide a clue for the treatment of the disease.     

Recombinant angiopoietin-like protein 4 attenuates intestinal barrier structure and function injury after ischemia/reperfusion

BACKGROUNDIntestinal barrier breakdown, a frequent complication of intestinal ischemia-reperfusion (I/R) including dysfunction and the structure changes of the intestine, is characterized by a loss of tight junction and enhanced permeability of the intestinal barrier and increased mortality. To develop effective and novel therapeutics is important for the improvement of outcome of patients with intestinal barrier deterioration. Recombinant human angiopoietin-like protein 4 (rhANGPTL4) is reported to protect the blood-brain barrier when administered exogenously, and endogenous ANGPTL4 deficiency deteriorates radiation-induced intestinal injury. AIMTo identify whether rhANGPTL4 may protect intestinal barrier breakdown induced by I/R.METHODSIntestinal I/R injury was elicited through clamping the superior mesenteric artery for 60 min followed by 240 min reperfusion. Intestinal epithelial (Caco-2) cells and human umbilical vein endothelial cells were challenged by hypoxia/ reoxygenation to mimic I/R in vitro.RESULTSIndicators including fluorescein isothiocyanate-conjugated dextran (4 kilodaltons; FD-4) clearance, ratio of phosphorylated myosin light chain/total myosin light chain, myosin light chain kinase and loss of zonula occludens-1, claudin-2 and VE-cadherin were significantly increased after intestinal I/R or cell hypoxia/reoxygenation. rhANGPTL4 treatment significantly reversed these indicators, which were associated with inhibiting the inflammatory and oxidative cascade, excessive activation of cellular autophagy and apoptosis and improvement of survival rate. Similar results were observed in vitro when cells were challenged by hypoxia/reoxygenation, whereas rhANGPTL4 reversed the indicators close to normal level in Caco-2 cells and human umbilical vein endothelial cells significantly.CONCLUSIONrhANGPTL4 can function as a protective agent against intestinal injury induced by intestinal I/R and improve survival via maintenance of intestinal barrier structure and functions.     

Interleukin 18 is a potent proliferative factor for intestinal mucosal lymphocytes in Crohn's disease

BACKGROUND & AIMS: Crohn's disease (CD) is characterized by a marked accumulation of activated Th1 type CD4(+) T cells and macrophages in inflamed intestinal mucosa. Interleukin (IL)-18 is a recently described cytokine that mainly exists in activated macrophages and shares biological activities with IL-12 in driving the development of Th1 type CD4(+) T cells by inducing interferon gamma. To clarify the role of IL-18 in intestinal inflammation in CD, we assessed the functional role of IL-18 in regulating intestinal mucosal lymphocytes. METHODS: Serum IL-18 concentration was measured by enzyme-linked immunosorbent assay. Expression of IL-18 and IL-18 receptor in human intestinal mucosa was determined using immunohistochemistry and flow cytometry. The functional activity of IL-18 was assessed by the use of recombinant IL-18 to stimulate both the growth of intestinal mucosal lymphocytes and IL-2 receptor induction activity. RESULTS: The serum IL-18 concentration was significantly higher in patients with CD than normal controls. In the inflamed colonic mucosa of CD, many IL-18(+)CD68(+) macrophages had infiltrated the lamina propria. Intestinal mucosal lymphocytes from CD expressed functional IL-18 receptors. Recombinant IL-18 induced significant proliferative responses in freshly isolated mucosal lymphocytes from CD patients, but not from normal controls. IL-18 up-regulated IL-2 receptor expression in mucosal lymphocytes from patients with CD, but not from normal controls. CONCLUSIONS: These findings suggest that infiltrated macrophages in the inflamed intestinal mucosa in CD produce IL-18, and that macrophage-derived IL-18 may serve as a potent regulatory factor for intestinal mucosal lymphocytes, thereby contributing to chronic intestinal inflammation in CD.     

Identification of activated T cells and the suppressor/inducer subset in patients suffering from severe aplastic anemia

Summary In patients with severe aplastic anemia (SAA), lymphocyte subpopulations were examined for the presence of HLA-DR and 2H4 (suppressor/inducer subset) antigen-expressing cells by flow cytometric analysis. Investigations were performed on peripheral blood lymphocytes before and after therapy with antithymocyte globulin (ATG) and methylprednisolone (MP), as well as on bonemarrow lymphocytes before therapy. Before treatment, only the absolute numbers of CD4⁺ T cells and the CD4⁺HLA^–DR⁺/CD8⁺HLA-DR⁺ activated T cell ratio were significantly decreased (p<0.01 and p<0.001, respectively). Following successful ATG/MP treatment, a decrease in the CD4⁺/CD8⁺T cell ratio was found. Regarding the suppressor/inducer subset, only absolute numbers of CD4⁺/2H4⁺ cells were somewhat higher in treated patients; the percentages were the same in all groups of patients. Studies performed on bonemarrow lymphocytes showed significantly decreased percentages of CD4⁺ and CD8⁺ T lymphocytes, which also express HLA-DR antigen. No significant changes in the distribution of activated T cells following ATG/MP therapy were found, suggesting that these cells play no major role in the pathogenesis of the disease.     

Interleukin-7 activates intestinal lymphocytes

Human intestinal lymphocytes, particularly intraepithelial lymphocytes, proliferate minimally to some agents, like mitogens and stimuli of the CD3 pathway. Thisin vitro finding may be due, in part, to a loss of factors foundin vivo. Three T-cell growth factors, IL-7, IL-9, and IL-12, were tested for their ability to stimulate the proliferation of intestinal lymphocytes. Both intraepithelial lymphocytes and lamina propria lymphocytes proliferated more vigorously to IL-7 than to IL-9 or IL-12, and only IL-7 increased stimulation through the CD3 pathway. The IL-7-induced response was IL-2-dependent: IL-2 receptors appeared on both intestinal lymphocyte types, and antibody to the IL-2 receptor blocked IL-7-induced proliferation. Both CD4⁺ and CD8⁺ T-cell subsets responded to this cytokine as shown by phenotype-depletion experiments and constancy in the CD4/CD8 ratios after culture with IL-7. In addition, the T-cell receptor and subsets responded equally well to IL-7. This newly described selective proliferative response of intestinal lymphocytes to IL-7, but not to IL-9 or IL-12, requires no preactivation and may enhance, growthin vivo.This work was supported by grants from the Crohn's & Colitis Foundation of America and the National Institutes of Health (DK42166).     

T cells of the human intestinal lamina propria are high producers of interleukin-10

Background and aim—Some of therecently observed functional features characteristic of immunocompetentcells residing in the human intestinal lamina propria could be mediatedby interleukin- 10 (IL-10). To investigate the role of IL-10 in thehuman intestinal mucosa, the regulation of IL-10 production by laminapropria T lymphocytes (LPL-T) was determined and compared with that ofperipheral blood T lymphocytes (PBL-T).
Methods—Following activation by usingdifferent stimuli, IL-10 release by LPL-T and PBL-T into thesupernatant was measured by enzyme linked immunosorbent assay (ELISA).In parallel, cell growth was determined by [³H]-thymidine incorporation.
Results—Neither LPL-T nor PBL-Trelease IL-10 constitutively. Triggering through CD2 or the T cellreceptor (TCR)/CD3 complex in the presence of autologous monocytesinduces significantly greater IL-10 secretion by LPL-T than by PBL-T.Engagement of the CD45 receptor enhances IL-10 release andproliferation of CD2 triggered CD45RO+ PBL-T. In contrast, it reducesCD2 induced IL-10 production by LPL-T without altering cellgrowth significantly.
Conclusions—Activated LPL-T release relativelyhigh amounts of IL-10. Enhanced IL-10 production by activatedLPL-T, in comparison with activated PBL-T, is not only related to thepresence of a higher proportion of CD45RO+ T cells in theintestinal lamina propria, but is also caused by increased sensitivityof LPL-T to CD2 co-stimulation. The differential responsiveness ofLPL-T, compared with PBL-T, to CD45 engagement demonstrates that CD45 could be involved in the altered CD2 reactivity of LPL-T.

Keywords:CD2; CD45; interleukin 10; lamina propria; T cellsubsets; T lymphocytes

     

Population alterations of l-arginase- and inducible nitric oxide synthase-expressed CD11b+/CD14−/CD15+/CD33+ myeloid-derived suppressor cells and CD8+ T lymphocytes in patients with advanced-stage non-small cell lung cancer

Background

Immune aberrations have been demonstrated in tumorogenesis, and myeloid-derived suppressor cells (MDSC) have shown to play a pivotal role in mediating immune suppression in animal models of human tumors. In the present study, we explored the clinical relevance of CD11b⁺/CD14^?/CD15⁺/CD33⁺ MDSCs and the association of MDSCs with CD8⁺ cytotoxic T lymphocytes in patients with non-small-cell lung cancer (NSCLC).

Patients and methods

The population of CD11b⁺/CD14^? cells in peripheral blood mononuclear cells (PBMNC) was determined in 173 patients with NSCLC and 42 control subjects. The expression of CD15, CD33, IL-4R, INF-γR, iNOS and l-arginase were analyzed. Cocultures with CD8⁺ T lymphocytes and Jurkat cells were developed to determine the impact of MDSCs on the expression of CD3ζ of CD8⁺ T lymphocytes.

Results

Patients with treatment-naïve, advanced-stage NSCLC (n = 87) had an increased subpopulation of CD11b⁺/CD14^?/CD15⁺/CD33⁺ cells in the PBMNCs with characteristics of MDSCs (P < 0.0001). The CD11b⁺/CD14^? cells in PBMNC also express IL-4R and INF-γR and can suppress CD3ζ expression in CD8⁺ T lymphocytes. The subpopulation of CD11b⁺/CD14^? cells in PBMNC was decreased in the advanced-stage NSCLC patients who had responsiveness to chemotherapy (n = 41, P < 0.0001) and in the early-stage NSCLC patients after removal of tumor (n = 8, P = 0.0391). Notably, a negative association existed between the population of CD11b⁺/CD14^? cells in PBMNC and the frequency of CD8⁺ T lymphocytes (n = 48, r = ?0.3141, P = 0.0297).

Conclusions

Our study provided evidence of an increased pool of CD11b⁺/CD14^?/CD15⁺/CD33⁺ MDSCs in the peripheral blood of NSCLC patients. For the suppressive effect of the cells on CD8⁺ T lymphocytes, these findings suggest the important role of the CD11b⁺/CD14^?/CD15⁺/CD33⁺ MDSCs in mediating immunosuppression in NSCLC.     

Dendritic cells frequency and phenotype in Egyptian type 1 diabetic patients

This study was performed to investigate changes in the dendritic cells (DCs) frequency and phenotype in the peripheral blood in Egyptian Type 1 diabetic children. Also to study the level of B, T lymphocytes, activated T lymphocytes, and costimulatory molecules expression on B lymphocytes. Twenty five children with T1DM and 25 healthy controls were enrolled. Flow cytometric detection of DCs, B-lymphocytes and T-lymphocytes, CD19⁺CD80⁺, CD19⁺CD86⁺, CD19⁺HLA-DR⁺ and CD3⁺ HLA-DR⁺ was preformed. The frequencies of monocytoid dendritic cells (mDCs) and plasmacytoid dendritic cells (pDCs) were significantly decreased in diabetic patients than the controls and the mDCs/pDCs ratio was significantly higher in diabetic patients. The expression of costimulatory molecules CD80 and CD86 on the entire DCs was significantly higher in diabetic children. The frequency of pDCs was negatively correlated with the age in diabetic patients and positively correlated with the level of insulin C-peptide. The percentage of CD80 expressing B lymphocytes and of activated T lymphocytes was significantly higher in the patients. Dendritic cells are reduced in number and display more mature phenotype in T1DM children. The higher expression of CD80 on B lymphocytes and activation of T lymphocytes may reflect the ongoing autoimmune process in this disease. Modulation of the DCs could have beneficial effect in T1DM.     

Gegen Qinlian decoction enhances immunity and protects intestinal barrier function in colorectal cancer patients via gut microbiota

BACKGROUNDWe previously showed, using the Traditional Chinese Medicine System Pharmacology Database, that Gegen Qinlian decoction (GQD) had a direct antitumor effect, and was combined with programmed cell death protein (PD)-1 inhibitors to treat microsatellite stable (MSS) tumor-bearing mice. However, the effect of GQD on patients with colorectal cancer (CRC) is not clear.AIMTo determine the therapeutic mechanism of GQD in improving immune function, reducing inflammation and protecting intestinal barrier function.METHODSSeventy patients with CRC were included in this study: 37 in the control group and 33 in the treatment group. The proportions of CD4⁺ T, CD8⁺ T, natural killer (NK), NKT and T regulatory cells were measured by flow cytometry. Levels of the cytokines tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-2, IL-6, IL-10 and serotonin (5-hydroxytryptamine; 5-HT) in serum were assessed by enzyme-linked immunosorbent assay (ELISA). The expression of zonula occludens (ZO)-1, occludin, nuclear factor (NF)-κB and TNF-α in tumor and normal tissues was measured by immunohistochemistry. The composition of gut microbiota from patients in the treatment group was assessed using 16S rDNA analysis. RESULTSThere were no adverse events in the treatment group. The proportion of CD4⁺ T cells and NKT cells in the post-treatment group was significantly higher than that in the pre-treatment and control groups (P < 0.05). The level of TNF-α in the post-treatment group was significantly lower than that in the pre-treatment and control groups (P < 0.05). The concentration of 5-HT in the post-treatment group was significantly lower than that in the pre-treatment group (P < 0.05). The expression of ZO-1 and occludin in tumor tissues in the treatment group was significantly higher than that in the control group (P < 0.05). The expression of ZO-1 in normal tissues of the treatment group was significantly higher than that in the control group (P = 0.010). Compared with the control group, expression of NF-κB and TNF-α in tumor tissues of the treatment group was significantly decreased (P < 0.05). Compared with the pre-treatment group, GQD decreased the relative abundance of Megamonas and Veillonella. In addition, GQD increased the relative abundance of Bacteroides, Akkermansia and Prevotella. CONCLUSIONGQD enhances immunity and protects intestinal barrier function in patients with CRC by regulating the composition of gut microbiota.     

Intestinal microbes direct CX3CR1+ cells to balance intestinal immunity

ABSTRACT

Intestinal damage driven by unrestricted immune responses against the intestinal microbiota can lead to the development of inflammatory diseases including inflammatory bowel disease. How such breakdown in tolerance occurs alongside the mechanisms to reinforce homeostasis with the microbiota are a focus of many studies. Our recent work demonstrates coordinated interactions between intact microbiota and CX₃CR1 expressing intestinal antigen presenting cells (APCs) that limits T helper 1 cell responses and promotes differentiation of regulatory T cells (Treg) against intestinal antigens including pathogens, soluble proteins and the microbiota itself. We find a microbial attachment to intestinal epithelial cells is necessary to support these anti-inflammatory immune functions. In this addendum, we discuss how our findings enhance understanding of microbiota-directed homeostatic functions of the intestinal immune system and implications of modulating this interaction in ameliorating inflammatory disease.     

Increased activation of isolated intestinal lamina propria mononuclear cells in inflammatory bowel disease

Normal human lamina propria lymphocytes are in a heightened state of activation compared with peripheral blood with regard to cell-surface activation antigen expression (transferrin receptor, interleukin-2 receptor, 4F2) and the increased spontaneous secretion of immunoglobulins in vitro. This study evaluates the cell-surface expression of activation-associated antigens in different subpopulations of isolated colonic lamina propria mononuclear cells in inflammatory bowel disease. In pilot studies using three-color flow cytometry, autofluorescence was observed that was emitted by unstained lamina propria mononuclear cells, which interfered with both the sensitivity and the specificity of the analyses. Because a major portion of the intestinal lymphocyte populations of interest were autofluorescent, a method to remove autofluorescence signals was developed by designing a computer program for the subtraction of autofluorescence from the emissions of each individual cell. This technique increases both the sensitivity and specificity of flow-cytometric analyses of intestinal lamina propria mononuclear cells. Using fluorescence-activated cell-sorter analyses with subtraction of autofluorescence on a single-cell basis, increased expression of lymphocyte activation antigens (interleukin-2 receptor, transferrin receptor, 4F2) was found on the cell surface of isolated intestinal B cells, T cells, CD4+ T cells, and CD8+ T cells in both Crohn's disease and ulcerative colitis. Therefore, markedly increased intestinal lymphocyte activation is a major immunological alteration in inflammatory bowel disease and includes all lymphocyte subpopulations investigated in this study. In addition, 5-aminosalicylic acid, which is used for the treatment of intestinal inflammation in inflammatory bowel disease, inhibits the expression of cell-surface activation antigens on mitogen-activated peripheral blood lymphocytes in a dose-dependent manner. These observations suggest that lymphocyte activation may play an important role in underlying immune processes that lead to chronicity and perpetuation of inflammatory bowel disease and may implicate an additional mechanism for the therapeutic action of 5-aminosalicylic acid.     

Human CD8+ T Cells Clear Cryptosporidium parvum from Infected Intestinal Epithelial Cells

Intracellular protozoans of the genus Cryptosporidium are a major cause of diarrheal illness worldwide, especially in immunocompromised individuals. CD4⁺ T cells and interferon-gamma are key factors in the control of cryptosporidiosis in human and murine models. Previous studies led us to hypothesize that CD8⁺ T cells contribute to clearance of intestinal epithelial Cryptosporidium infection in humans. We report here that antigen expanded sensitized CD8⁺ T cells reduce the parasite load in infected intestinal epithelial cell cultures and lyse infected intestinal epithelial cells. These effects are most likely mediated by the release of cytotoxic granules. Elimination of parasites seems to require antigen presentation through both human leukocyte antigen (HLA)-A and HLA-B. These data suggest that cytotoxic CD8⁺ T cells play a role in clearing Cryptosporidium from the intestine, a previously unrecognized feature of the human immune response against this parasite.     

Immunohistological characterization of intraepithelial and lamina propria lymphocytes in control ileum and colon and in inflammatory bowel disease

Using monoclonal antibodies to T and B lymphocytes, to natural killer cells, and to HLA-DR antigen, we characterized the lymphocyte population within the epithelial and lamina propria regions in control intestine and colon, and in grossly involved and in grossly uninvolved intestine and colon of patients with active inflammatory bowel disease. There were significantly more intraepithelial T cells in control ileum than in control colon. In comparison to control, there was a heterogeneity of alterations in intraepithelial and lamina propria T lymphocyte subsets (T11⁺, T8⁺, T4⁺) in inflammatory bowel disease. B lymphocytes were not detected within the lamina propria, except when found in and adjacent to lymphoid aggregates. Leu 7⁺ cells were uncommon in the lamina propria of control ileum and colon and in diseased tissues. The majority of intraepithelial lymphocytes did not express HLA-DR. Epithelial cells of control colon did not express HLA-DR while epithelial cells of control ileal tissues and of diseased colonic and ileal specimens expressed HLA-DR antigen. Only small numbers of lamina propria T cells expressed HLA-DR in both control and disease tissues. There was intense expression of HLA-DR by monocytes and modest expression of HLA-DR by capillary and lymphatic endothelial cells. The induction of HLA-DR expression by diseased colonic epithelium and the observation that lymphatic endothelium expresses HLA-DR are new observations, and we established that Leu 7⁺ cells are present in very small numbers in both normal and diseased intestine and colon.Supported by funding from the National Foundation for Ileitis and Colitis and by Merit Review funds from the Veterans Administration.