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Biomedical subjects

R P MacDermott

Publications and source records attributed to R P MacDermott.

At least 37 records · Page 2Linked to original sources
PubMed1995

Monocyte-chemoattractant protein 1 gene expression in intestinal epithelial cells and inflammatory bowel disease mucosa.

BACKGROUND: Monocyte-chemoattractant protein 1 (MCP-1) activates macrophages and increases the migration of monocytes into tissue during inflammation. It was hypothesized that MCP-1 expression is involved in intestinal inflammation. METHODS: MCP-1 protein was detected by immunohistochemistry and immunoprecipitation. Biological activity of MCP-1 was assessed using a chemotactic assay. MCP-1 messenger RNA (mRNA) levels were measured by quantitative reverse-transcription polymerase chain reaction. RESULTS: In normal mucosa, MCP-1 was predominantly present in surface epithelium. In contrast, inflamed mucosa from patients with ulcerative colitis or Crohn's disease contained multiple cells immunoreactive for MCP-1, including spindle cells, mononuclear cells, and endothelial cells. Furthermore, MCP-1 mRNA expression was markedly increased in inflamed intestinal biopsy specimens from patients with inflammatory bowel disease. MCP-1 was detected in isolated intestinal epithelial cells and in conditioned media from Caco-2 cells. Caco-2 cell-conditioned media stimulated monocyte chemotaxis activity that was inhibited by anti-MCP-1 antibodies. Constituitive MCP-1 mRNA levels in Caco-2 cells were up-regulated by interleukin 1 beta and down-regulated by dexamethasone. CONCLUSIONS: In addition to lamina propria macrophages, endothelial cells, and spindle cells, intestinal epithelial cells are able to produce MCP-1. MCP-1 is expressed constitutively in the intestinal colonic mucosa and is up-regulated during inflammation.

Base Sequence↗
PubMed1994

Butyrate increases colonocyte protein synthesis in ulcerative colitis.

Butyrate promotes epithelial cell healing and improves symptoms when administered rectally in patients with distal ulcerative colitis (UC). It was hypothesized that butyrate may enhance healing in patients with UC by stimulating colonocyte proliferation and/or protein production. Mucosa from the descending colon was obtained from patients with UC (n = 5), Crohn's disease (n = 8), diverticulitis (n = 6), and cancer (normal tissue 10 cm from tumor; n = 10). Epithelial cells were isolated using dispase/collagenase and differential sedimentation and incubated for 4 hr at 37 degrees C with either Na butyrate (10 mM) or NaCl (10 mM). Protein synthesis was assessed by [14C]leucine incorporation and proliferation was determined with [3H]thymidine. Mean viability and purity were >88%. Spontaneous proliferation was significantly increased in UC when compared to diverticulitis and normal controls. Butyrate significantly increased protein synthesis in UC epithelial cells when compared to saline control. The therapeutic effects of butyrate in patients with UC may be due to its use by epithelial cells as a metabolic fuel to increase protein production and promote healing.

Aged↗
PubMed1994

Alterations in the mucosal immune system in ulcerative colitis and Crohn's disease.

Emphasis is now being placed upon obtaining a better understanding of the regulatory cytokines that normally downregulate acute intestinal inflammation. These inhibitory cytokines appear to be missing or not functioning properly in patients with inflammatory bowel disease (IBD), thereby leading to perpetuation of inflammation. As we obtain an increased understanding of immune and inflammatory regulatory processes in the intestine, we will be able to devise better future therapeutic strategies for use in our IBD patients.

Colitis, Ulcerative↗
PubMed1994

Quantitative PCR for detection of femtogram quantities of interleukin-8 mRNA expression.

A quantitative polymerase chain reaction (PCR) assay for mRNA expression of interleukin-8 (IL-8), a neutrophil chemotactant and activator, was developed to examine the expression of this cytokine by colonic mucosa. A synthetic IL-8 RNA deleted in size of native IL-8 mRNA was used as an external control. The synthetic IL-8 RNA was mixed with total RNA from cells and converted to cDNA and amplified by PCR simultaneously. The lower limit of sensitivity for the assay was found to be more than 1 femtogram of IL-8 mRNA. The assay determined IL-8 mRNA expression when the RNA was isolated from either human histiocytic lymphoma cell line U937 cells or human colonic mucosa obtained from colitis patients and healthy controls. The development of the rapid and sensitive assay should provide a means to more fully evaluate the role of this cytokine in diverse disease states with small scale.

Base Sequence↗
PubMed1993

Crohn's disease monocytes are primed for accentuated release of toxic oxygen metabolites.

BACKGROUND: Inflammatory bowel disease occurs in regions of the intestine characterized by a bowel content high in bacteria. Intestinal bacteria synthesize cell wall products such as lipopolysaccharide; when normal monocytes or macrophages come in contact with these products, they can be primed to release a number of inflammatory mediators. Mediators such as toxic oxygen metabolites released as part of the respiratory burst may contribute to inflammatory tissue damage. The aim of this study was to determine if monocytes from patients with Crohn's disease are primed by lipopolysaccharide for a greater respiratory burst. METHODS: The generation of superoxide anion was measured by superoxide dismutase inhibitable reduction of ferricytochrome c. RESULTS: Freshly isolated monocytes from active untreated Crohn's disease patients (n = 8) showed enhanced stimulated release of superoxide anion when compared with normal monocytes (n = 15; 3.80 +/- 0.12 vs. 1.02 +/- 0.06 nmol/5 min; P < 0.001). We tested the hypothesis that the monocyte priming factor in Crohn's disease serum may be lipopolysaccharide by showing that Crohn's disease serum lost its ability to prime normal monocytes after lipopolysaccharide was removed (0.25 +/- 0.25 nmol/5 min, P < 0.001). CONCLUSIONS: These studies indicate that bacterial cell wall products may be important proinflammatory molecules involved in the initiation and/or perpetuation of Crohn's disease.

Adolescent↗
PubMed1993

Increased spontaneous secretion of rheumatoid factor by intestinal lamina propria mononuclear cells from Crohn's disease but not ulcerative colitis patients.

Increased levels of rheumatoid factors (RF) have been observed in the serum of Crohn's disease but not ulcerative colitis patients, and have been proposed to relate to an increased state of intestinal lymphocyte activation. We have therefore examined the spontaneous in vitro secretion of RF by intestinal lamina propria mononuclear cells (MNC) isolated from specimens from control and inflammatory bowel disease (Crohn's disease, ulcerative colitis) patients. Normal intestinal lamina propria MNC spontaneously secrete rheumatoid factors of different isotypes during 14 days of in vitro culture (9.7 ng/ml IgA RF, 11.6 ng/ml IgM RF and 64.6 ng/ml IgA anti-Fc (IgG)). In matched studies intestinal MNC isolated from normal large bowel exhibited significantly greater levels of RF synthesis and secretion in vitro than normal small bowel intestinal MNC. A large increase in spontaneous RF secretion was observed from Crohn's disease intestinal MNC (21.4 ng/ml IgA RF, 21.4 ng/ml IgM RF, and 108.15 ng/ml IgA anti-Fc (IgG)), when compared with normal controls. The amount of RF secreted was dependent on the amount of inflammatory activity of the bowel specimens, from which the MNC were isolated (198.3 ng/ml of IgA anti-Fc(IgG) from involved versus 50.0 ng/ml from matched non-involved tissue). Ulcerative colitis MNC released decreased amounts of RF (7.1 ng/ml IgA RF, 6.2 ng/ml IgM RF, and 42.3 ng/ml IgA anti-Fc(IgG)). These observations using isolated intestinal MNC may explain the findings of RF changes in the sera of inflammatory bowel disease patients. Our observations support the hypothesis of a heightened state of activation in normal intestinal lamina propria MNC, which is further increased in active Crohn's disease. The dissimilarities observed between Crohn's disease and ulcerative colitis may indicate fundamental differences in disease pathophysiology and will lead to further studies exploring intestinal immunoregulatory properties of RF.

Antibodies, Anti-Idiotypic↗
PubMed1993

Enhanced secretion of tumour necrosis factor-alpha, IL-6, and IL-1 beta by isolated lamina propria mononuclear cells from patients with ulcerative colitis and Crohn's disease.

The perpetuation of inflammation in ulcerative colitis and Crohn's disease may be regulated in part by an increased secretion of proinflammatory cytokines due to either an appropriate response to initial stimulating agents, and/or due to an impaired down-regulation of cytokine secretion. The aim of this study was to determine the secretion patterns of the proinflammatory cytokines tumour necrosis factor-alpha (TNF-alpha), IL-6 and IL-1 beta, from isolated lamina propria mononuclear cells (LPMNC) isolated from colonic biopsies from patients with untreated ulcerative colitis or Crohn's disease. LPMNC isolated from involved inflammatory bowel disease (IBD) mucosa spontaneously produced increased amounts of TNF-alpha, and IL-6, and IL-1 beta. The TNF-alpha secretion from IBD LPMNC could be further enhanced by pokeweed mitogen stimulation. The secretion patterns of TNF-alpha and IL-1 beta by LPMNC from patients with either ulcerative colitis or Crohn's disease demonstrated a close correlation with the degree of tissue involvement and mucosal inflammation. LPMNC from non-involved ulcerative colitis mucosa secreted markedly increased levels of IL-6 compared with non-involved Crohn's disease mucosa or control mucosa. The heightened IL-6 secretion from LPMNC from non-involved ulcerative colitis mucosa without visible or microscopic signs of inflammation indicates that the pathophysiologic mechanisms involved in the initiation of inflammation may differ between ulcerative colitis and Crohn's disease. The determination of proinflammatory cytokine secretion by isolated LPMNC from colonoscopic biopsies may be a sensitive method for monitoring the severity of mucosal inflammation in IBD patients.

Adult↗
PubMed1993

Enhancement of macrophage candidacidal activity by interferon-gamma. Increased phagocytosis, killing, and calcium signal mediated by a decreased number of mannose receptors.

In contrast to its macrophage-activating capacity, IFN-gamma downregulates expression of the macrophage mannose receptor (MMR), which mediates uptake of Candida and other microorganisms. We found that IFN-gamma induced a concentration-dependent increase in the capacity of human monocyte-derived macrophages to ingest and kill both opsonized and unopsonized Candida albicans and to release superoxide anion upon stimulation with Candida. Mannan or mannosylated albumin inhibited this activated uptake of unopsonized Candida, but glucan did not. Addition of mAb to complement receptor (CR) 3 did not inhibit ingestion; macrophages that lacked CR3 (leukocyte adhesion defect) showed normal upregulation of ingestion by IFN-gamma. The increased candidacidal activity of IFN-gamma-activated macrophages was associated with reduced expression of MMR by a mean of 79% and decreased pinocytic uptake of 125I-mannosylated BSA by 73%; K(uptake) of pinocytosis was not changed. Exposure of resident macrophages to unopsonized Candida did not elicit a transient increase in intracellular free Ca2+ ([Ca2+]i); macrophages activated by IFN-gamma expressed a brisk increase in [Ca2+]i on exposure to Candida. These data suggest that macrophage activation by IFN-gamma can enhance resistance to C. albicans infection in spite of downregulation of the MMR, perhaps through enhanced coupling of the MMR to microbicidal functions.

Calcium↗
PubMed1993

Giant gastric ulcers: an unusual manifestation of Crohn's disease.

We report a patient with Crohn's disease manifesting as recurrent giant gastric ulcers, with subsequent perforation and gastric outlet obstruction. The ulcers contained granulomas, and the patient was achlorhydric. To our knowledge, this is the first report of giant ulcers in a patient with gastric Crohn's disease. This case demonstrates another gastric manifestation of Crohn's disease, it documents nonmalignant gastric ulcers in the setting of achlorhydria, and it raises the possibility of Crohn's disease in the differential diagnosis of giant gastric ulcers refractory to medical management.

Adult↗
PubMed1992

Identification of the uncultured bacillus of Whipple's disease.

BACKGROUND: Whipple's disease is a systemic disorder known for 85 years to be associated with an uncultured, and therefore unidentified, bacillus. METHODS: We used a molecular genetic approach to identify this organism. The bacterial 16S ribosomal RNA (rRNA) sequence was amplified directly from tissues of five unrelated patients with Whipple's disease by means of the polymerase chain reaction, first with broad-range primers and then with specific primers. We determined and analyzed the nucleotide sequence of the amplification products. RESULTS: A unique 1321-base bacterial 16S rRNA sequence was amplified from duodenal tissue of one patient. This sequence indicated the presence of a previously uncharacterized organism. We then detected this sequence in tissues from all 5 patients with Whipple's disease, but in none of those from 10 patients without the disorder. According to phylogenetic analysis, this bacterium is a gram-positive actinomycete that is not closely related to any known genus. CONCLUSIONS: We have identified the uncultured bacillus associated with Whipple's disease. The phylogenetic relations of this bacterium, its distinct morphologic characteristics, and the unusual features of the disease are sufficient grounds for naming this bacillus Tropheryma whippelii gen. nov. sp. nov. Our findings also provide a basis for a specific diagnostic test for this organism.

Actinobacteria↗
PubMed1992

Increased in vitro release of soluble interleukin 2 receptor by colonic lamina propria mononuclear cells in inflammatory bowel disease.

Increased concentrations of the soluble form of the interleukin 2 receptor have been observed in the sera of Crohn's disease and ulcerative colitis patients. In this study we have observed the spontaneous release of soluble interleukin 2 receptor by unstimulated, isolated normal and inflammatory bowel disease colonic lamina propria mononuclear cells. Lamina propria mononuclear cells from Crohn's disease patients (median = 204 U/ml (interquartile range 126-396, n 17) secreted significantly (p less than 0.01) more soluble interleukin 2 receptor than normal controls (median = 124.5 U/ml (108-131), n 12). No statistically significant differences were seen between ulcerative colitis (median = 135 U/ml (92-196), n 20) and normal controls. Moreover, significantly (p less than 0.01) increased amounts of soluble interleukin 2 receptor were secreted by colonic diverticulitis lamina propria mononuclear cells (median = 259 U/ml (149-282), n 15) which were used as disease specificity controls. Time course experiments showed that the majority of soluble interleukin 2 receptor was released by isolated lamina propria mononuclear cells in the first six days of culture. Upon stimulation with pokeweed mitogen, Crohn's disease (median = 2258 U/ml (1435-3584), n 14), normal control (median = 2622 U/ml (2030-3180), n 14) and diverticulitis lamina propria mononuclear cells (median = 2745 U/ml (1733-3192), n 10) reached similar maximal soluble interleukin 2 receptor secretion levels, while ulcerative colitis lamina propria mononuclear cells secreted significantly (p less than 0.005) less soluble interleukin 2 receptor (median = 912 U/ml (494-1259), n 17). These results suggest that enhanced shedding/secretion of soluble interleukin 2 receptor by intestinal lymphocytes may account in part for increased serum soluble interleukin 2 receptor concentrations during chronic intestinal inflammatory reactions.

Cells, Cultured↗
PubMed1992

The role of the mucosal immune system in inflammatory bowel disease.

Continued delineation of the major factors that lead to intestinal inflammation will provide critical insights into many of the pathophysiologic events leading to tissue destruction in IBD. The exploration of exciting and important new areas, such as the role of adhesion molecules, proinflammatory cytokines, and the activation of lymphocytes and phagocytes, will contribute significantly to a better understanding of the mechanisms that sustain the intestinal inflammatory process. Determining the mechanisms of amplification and perpetuation of intestinal inflammation as well as learning more about the natural suppression of intestinal inflammation by the normal cellular and cytokine networks of the mucosal immune system will open exciting new therapeutic approaches. It is encouraging to see realistic and testable working models emerge from the combined efforts of many committed investigators who have been engaged in studying the role of the mucosal immune system in the pathophysiology of IBD. A great deal more remains to be learned in this rapidly advancing area, and we can look forward with confidence to continued advances in the study of IBD.

Antibody Formation↗
PubMed1991

Aggregated bovine IgG inhibits mannose receptor expression of murine bone marrow-derived macrophages via activation.

We previously described the presence of an inhibitory protein contained in the 20 to 40% (NH4)2SO4 precipitable fraction of FCS that down-regulates expression of mannose receptors on bone marrow-derived macrophages. We now identify aggregated bovine IgG as the main inhibitory component. Heat-aggregated bovine IgG was capable of down-regulating expression of the macrophage mannose receptor in a dose-dependent manner without inducing changes in ligand affinity whereas neither F(ab')2 fragments nor nonaggregated IgG displayed any inhibitory effect. Depleting of IgG from heat inactivated FCS by protein G affinity chromatography completely removes the inhibitory activity. Moreover, readdition of the Ig eluate from the protein G chromatography column restored inhibition in a dose-dependent manner. Macrophages were able to clear exogenously added aggregated bovine IgG, thus leading to loss of inhibitory activity in macrophage-conditioned media as compared to sham-conditioned media containing aggregated IgG. These results indicate that aggregated IgG down-regulates mannose receptor expression by macrophage activation via interaction with Fc-gamma R.

Animals↗
PubMed1991

Monomeric IgG2a promotes maturation of bone-marrow macrophages and expression of the mannose receptor.

The macrophage mannose receptor, a 172-kDa lineage-specific glycoprotein, partakes in nonopsonin-mediated phagocytosis by recognition of terminal mannose residues on targeted particles. Because appearance of the receptor progresses with monocyte/macrophage differentiation, its expression is indicative of the maturational state of the cell. Monomeric IgG2a and IgG2b up-regulate mannose-receptor surface expression and biosynthesis by murine bone-marrow macrophage precursors as much as 7- to 12-fold in a dose-dependent manner. IgG2a accelerates macrophage mannose-receptor expression by several days during in vitro bone-marrow differentiation; however, treated and control cells ultimately express equivalent levels of receptor. Moreover, the effect is independent of cell cycle or ambient levels of colony-stimulating factor 1. The coinduction of another maturation-dependent lineage-specific antigen, F4/80, and the fact that macrophage precursors respond to IgG2a only within the first day of culture, indicate that the targeted cell is an early myelomonocytic precursor, responsive only during a short, early developmental window. The effect is specific for immunoglobulin molecules of the IgG2a and IgG2b subclasses and probably involves an Fc gamma-receptor signal-transduction pathway but not macrophage priming or activation. Most importantly, a paracrine mechanism of immunoglobulin-mediated bone-marrow macrophage differentiation is suggested by experiments in which basal levels of mannose-receptor expression are reduced by continual removal of B-cell-generated IgG from marrow cultures. Thus, IgG2a and IgG2b prompt mannose-receptor synthesis and bone-marrow macrophage differentiation and may, therefore, play a role in the regulation of macrophage differentiation in host defense.

Animals↗
PubMed1991

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.

Aminosalicylic Acids↗
PubMed1991

Human rib bone marrow mononuclear cells spontaneously synthesize and secrete IgE in vitro.

We have examined spontaneous secretion of IgE by human rib bone marrow mononuclear cells (MNC). Bone marrow MNC from nine out of 12 rib specimens synthesized and secreted substantial amounts of IgE during 14 days of in vitro culture. The 14-day supernatants from these bone marrow MNC contained a mean of 2589 pg/ml of IgE (n = 12) with a maximum production of 15,408 pg/ml of IgE compared with small amounts of IgE (80-200 pg/ml) produced by similarly cultured normal and inflammatory bowel disease intestinal lamina propria MNC. Using two rib specimens, time-course studies revealed spontaneous secretion of IgE to be minimal during the first 2 days of culture (152 pg/ml), followed by a steady increase between days 4 (517 pg/ml) and 14 (3588 pg/ml). The addition of pokeweed mitogen resulted in 72% suppression of spontaneous IgE production by bone marrow MNC. The bone marrow MNC isolated from the ribs consisted of 22% Leu12+ (B) cells of which 3.2% were surface IgE positive. Staining for cytoplasmic immunoglobulin revealed 1% of the bone marrow MNC to be cytoplasmic IgE+. The presence of IgE-bearing and IgE-secreting MNC in human bone marrow is consistent with the observation that allergen-specific IgE-mediated hypersensitivity is adoptively transferred by human bone marrow transplantation and demonstrates the usefulness of human bone marrow MNC for examination of IgE secretory and regulatory events.

Adult↗
PubMed1990

Oxidant defense mechanisms in the human colon.

Reactive oxygen metabolites have been implicated as important mediators of inflammation-induced intestinal injury associated with ischemia (and reperfusion), radiation, and inflammatory bowel disease. Because the colonic mucosa may be subjected to significant oxidant stress during times of acute and chronic inflammation, knowledge of the oxidant defense mechanisms in the colon is of biologic and potential clinical importance. Therefore, the objective of this study was to quantify the specific activities of superoxide dismutase (SOD), catalase, and GSH peroxidase in the normal human colon. We found low, but significant, amounts of all three enzymes in the mucosa, submucosa, and muscularis/serosa of the human colon. However, the mucosal, levels of SOD (3.6 +/- 0.3 units/mg protein), catalase (11 +/- 3 units/mg), and GSH peroxidase (15.2 +/- 0.8 mU/mg) represented only 8%, 4%, and 40%, respectively, of those values determined for human liver. Colonic epithelial cells derived from mucosal biopsies exhibited significantly higher specific activities for SOD (12 +/- 0.5 units/mg) and catalase (26 +/- 6 units/mg) when compared to whole mucosa, suggesting most of the mucosal activity was associated with the epithelial cells and not the lamina propria. In a comparative study, we found that a human colonic carcinoma cell line (CaCo-2) contained significantly lower SOD (6 +/- 0.5 units/mg) and catalase (6 +/- 0.6 units/mg) activities when compared to colonic epithelial cells. Taken together, our data suggest that: (1) the colonic mucosa is relatively deficient in antioxidant enzymes when compared to liver, and (2) most of the protective enzyme activity is localized within the epithelium and not the mucosal interstitium.

Adenocarcinoma↗