Biomedical subjects

A Willey

Publications and source records attributed to A Willey.

11 recordsLinked to original sources

PubMed2003

Linoleic acid, but not oleic acid, upregulates the production of interleukin-8 by human intestinal smooth muscle cells isolated from patients with Crohn's disease.

BACKGROUND & AIMS: Crohn's disease is a chronic inflammatory bowel disease (IBD) of unknown etiology. In this study, we investigated the hypothesis that dietary fatty acids, linoleic acid (LA) and oleic acid (OA), could be involved in the inflammatory response through stimulation of the neutrophil chemokine, IL-8. METHODS: Human intestinal smooth muscle (HISM) cells were isolated from normal patients and patients with Crohn's disease and cultured for 24h with LA or OA in the presence or absence of oxidative stress. The concentrations of IL-8 were measured in the media and cellular oxidative stress was quantitated by measurement of thiobarbituric acid reactive substances (TBARSs). RESULTS: Spontaneous production of IL-8 was significantly higher in HISM cells isolated from Crohn's bowel compared to control bowel. LA caused a marked, nine-fold, increase in IL-8 secretion by Crohn's cells, an effect that could be simulated in normal HISM cells by co-incubation of LA with an oxidizing solution (Ox) composed of hypoxanthine+xanthine oxidase+FeSO(4) (OxLA). These effects were inhibited by vitamins C and E. Treatment of Crohn's cells with OxLA did not further increase IL-8 over that of LA alone. The effect of LA alone was not associated with an increase in cellular oxidative stress as quantitated by TBARSs. In contrast to the results with LA, treatment with OA or OxOA did not increase IL-8 in either normal or Crohn's cells. In addition, OA protected Crohn's cells from the increase in TBARSs induced by Ox. In contrast to IL-8, spontaneous production of monocyte chemotactic protein (MCP-1) was significantly lower in Crohn's HISM cells as compared to normal cells and exposure to OxLA did not increase its production. CONCLUSIONS: LA, but not OA, increased the production of IL-8 by HISM cells. These results suggest that replacement of LA by OA in the diet of Crohn's patients and increased intake of a diet rich in antioxidants could be beneficial in decreasing inflammatory activity in Crohn's disease.

Analysis of Variance↗

PubMed2001

Redox imbalance in Crohn's disease intestinal smooth muscle cells causes NF-kappaB-mediated spontaneous interleukin-8 secretion.

Interleukin-8 (IL-8), a chemokine secreted by cells at injury sites, has recently been recognized as involved in the pathogenesis of Crohn's disease. However, the pathogenesis of enhanced spontaneous transcription of IL-8 by the bowel in patients with Crohn's disease is undefined. Although IL-8 is secreted primarily by neutrophils, macrophages, and endothelial and epithelial cells, we observed the involvement of mesenchymal cells in the inflammatory process. A smooth muscle cell line isolated from the ileum of a patient with Crohn's disease (CDISM) and maintained in culture exhibited spontaneous transcription and secretion of IL-8 when compared with intestinal smooth muscle cells obtained from a normal subject (NHISM). Furthermore, IL-8 transcription from CDISM cells was associated with remarkable spontaneous activation of the oxidant-sensitive transcription factor NF-kappaB, as assessed by transient transfection assays with an IL-8 promoter reporter construct, Western blot analysis, and electrophoretic mobility shift assays (EMSA). Finally, we report here that CDISM cells exhibit significantly altered redox balance. The antioxidant pyrrolidine dithiocarbamate (PDTC) restored the redox equilibrium by mechanisms that inhibit binding of NF-kappaB to its cognate site on the IL-8 promoter. These findings suggest that restoration of the redox balance could hold promise for therapeutic intervention in Crohn's disease.

Antioxidants↗

PubMed1999

Palmitoyl ascorbate: selective augmentation of procollagen mRNA expression compared with L-ascorbate in human intestinal smooth muscle cells.

The effect of 6-O-palmitoyl ascorbate on procollagen mRNA levels, collagen synthesis, and collagen secretion was investigated and compared with the effect of L-ascorbate in human intestinal smooth muscle (HISM) cells in vitro. Collagen synthesis, determined by the incorporation of 3H-proline into pepsin-resistant, salt-precipitated collagen, increased in a concentration-dependent manner in response to palmitoyl ascorbate. There was a twofold increase in collagen synthesis at 2.5 and 5 microM. By contrast, L-ascorbate was required at 4-5 times the concentration for the same response. However, at 20 microM, both palmitoyl and L-ascorbate induced similar 2.7-fold increases in collagen synthesis. Palmitoyl ascorbate induced a 1.6- and 3.5-fold increase in steady-state levels of procollagen I and III mRNA levels respectively, whereas L-ascorbate had no effect. Palmitoyl ascorbate and L-ascorbate induced similar increases in the amounts of newly synthesized procollagen secreted into the medium and in the amounts of collagen types I, III and V accumulating in the cell layer. There was no effect of either palmitoyl ascorbate or L-ascorbate on the activity of a procollagen alpha2 (I) promoter construct transiently transfected into HISM cells. Palmitoyl ascorbate augments HISM cell procollagen synthesis and mRNA levels more efficiently than L-ascorbate. This property may be due to the greater resistance of the ascorbate ester to oxidation and suggests that palmitoyl ascorbate could be an important agent for studies of collagen synthesis in vitro.

Ascorbic Acid↗

PubMed1999

Penetration of solar erythemal UV radiation in the shade of two common Australian trees.

The penetration of solar erythemal ultraviolet radiation has been measured in the shade of a gum (Eucalyptus sp.) and a she oak (Casaurina) tree, both on a horizontal plane and with polysulphone dosimeters to human anatomical sites. This has provided new data useful for protection strategies against harmful ultraviolet radiation. For larger solar zenith angles, the relative penetration of solar erythemal ultraviolet in the shade of the trees is higher. On a horizontal plane, at noon, in winter, the shade erythemal ultraviolet ranged from 44 to 55% of that in the sun whereas in spring it ranged from 29 to 37% of the irradiances in the sun. Similarly, at 9:00 EST and 15:00 EST, the shade erythemal ultraviolet in winter ranged from 51 to 81% of the irradiances in the sun whereas in spring and summer they ranged from 35 to 51% of the unshaded irradiances. The shade ratios for specific body sites provided by the shade of the two trees were 0.05 to 0.45 for the solar zenith angles in this research. The shade ratios ranged from 0.14 to 0.45 for the gum tree and from 0.05 to 0.28 for the she oak. The denser foliage of the she oak provided higher ultraviolet protection compared to that of the gum tree.

Australia↗

PubMed1998

Development of pseudointima and stenosis after transjugular intrahepatic portasystemic shunts: characterization of cell phenotype and function.

The clinical utility of transjugular intrahepatic portasystemic shunts (TIPS) is frequently complicated by the ingrowth of tissue into the stent lumen, causing stent stenosis. These studies were undertaken to define the cellular and matrix components of the pseudointima, define the phenotype and function of the mesenchymal cells in the pseudointima and maintain them in culture, and to study the differences between stenotic and nonstenosed stents. A total of 35 stents were evaluated. TIPS pseudointima were examined histologically, by immunohistochemistry and in situ hybridization to determine the cellular and connective tissue constituents. Mesenchymal cells were grown from tissue within the TIPS and around it, and their phenotype was studied and compared with control smooth muscle cells and fibroblasts. Masson's trichrome staining of histological sections demonstrated that TIPS tissue was composed of collagen and palisades of mesenchymal cells and was lined by an endothelium. Immunostaining demonstrated strong and uniform alpha-smooth muscle staining in TIPS mesenchymal cells and peri-TIPS cells. Type I procollagen mRNA expression was demonstrated in mesenchymal cells in and around the stent by in situ hybridization. TIPS mesenchymal cells secreted less radiolabeled fibronectin, and far more type III, relative to type I, collagen compared with peri-TIPS cells. TIPS cells also expressed high levels of type III procollagen mRNA compared with peri-TIPS cells. There was no difference between stenotic stents and nonstenosed stents with respect to clinical features, time from stenting, gross morphology, histology, presence of bile fistulae, and cell phenotype. However, smooth muscle cells (SMC) from stenotic stents demonstrated both greater cell proliferation and collagen I and III secretion compared with those from nonstenosed stents. These data demonstrate that TIPS stenosis results from an accumulation of collagen and proliferation of SMC within the stent lumen.

Cells, Cultured↗

PubMed1997

Corticosteroids repress the interleukin 1 beta-induced secretion of collagenase in human intestinal smooth muscle cells.

BACKGROUND & AIMS: The cytokine interleukin (IL)-1 beta induces collagenase expression and inhibits collagen expression in human intestinal smooth muscle (HISM) cells. Corticosteroids cause transrepression of certain genes, including the collagenase gene. The aim of this study was to determine if corticosteroids repress the induction of collagenase expression and the inhibition of collagen secretion by IL-1 beta in HISM cells. METHODS: HISM cells were exposed to IL-1 beta (1-100 pmol/L) for 24 hours in the presence or absence of dexamethasone (10(-6) mol/L). Collagenase messenger RNA (mRNA) levels were determined by ribonuclease protection assay. Collagenase and tissue inhibitor of metalloproteinase protein secretion were determined by enzyme-linked immunosorbent assay of culture medium. Procollagen and collagen secretion were determined by polyacrylamide slab gel analysis of radiolabeled proteins in culture medium. RESULTS: A 30-fold induction of collagenase mRNA and collagenase protein secretion by IL-1 beta was completely abrogated by dexamethasone. Dexamethasone at 10(-6) mol/L also reduced basal levels of collagenase mRNA by 50% and blocked the IL-1 beta-induced inhibition of collagen secretion. CONCLUSIONS: Corticosteroids repress the collagenolytic action of IL-1 beta on HISM cells in vitro and therefore should promote healing in the inflamed intestine.

Collagen↗

PubMed1996

Interleukin 1 beta down-regulates collagen and augments collagenase expression in human intestinal smooth muscle cells.

BACKGROUND & AIMS: Smooth muscle cells resident in the intestinal wall play a significant role in the healing of the injured intestine and in the fibrosis that complicates Crohn's disease. The cytokine interleukin 1 beta (IL-1 beta) is involved in inflammatory bowel disease. The aim of this study was to determine the action of IL-1 beta on proliferation and collagen metabolism in human intestinal smooth muscle cells. RESULTS: IL-beta caused a three-fold increase in [3H]thymidine uptake at 100 pmol/L. This mitogenic effect was equipotent with that of platelet-derived growth factor when cells were exposed to IL-beta for 48 vs. 24 hours. IL-beta inhibited the secretion of procollagen into culture medium by 70% and the accumulation of newly synthesized procollagen in cells by 55%. In addition, IL-beta caused a concentration-dependent inhibition of steady-state levels of procollagen I and III messenger RNA (85% inhibition at 100 pmol/L) and a 3-5-fold augmentation of collagenase messenger RNA levels. CONCLUSIONS: IL-beta is mitogenic for human intestinal smooth muscle cells, but this action is associated with a concomitant down-regulation of collagen synthesis and secretion and an augmention of collagenase expression.

Blotting, Northern↗

PubMed1995

Role of ascorbic acid in procollagen expression and secretion by human intestinal smooth muscle cells.

The role of ascorbate in the production and secretion of procollagen by human intestinal smooth muscle cells and the conditions in culture for optimal ascorbate bioefficacy were studied. Procollagen synthesis and secretion were determined by the incubation of cells with L-[5-3H]proline, and the quantitation of radiolabelled procollagen bands in the cell layer and the culture medium by polyacrylamide slab gel electrophoresis and densitometry. When cells were cultured without ascorbate in the culture medium, procollagen secretion into the medium was 75% less than in cells receiving fresh ascorbate daily. In the cell layer, in contrast, procollagen accumulation was fourfold greater in the scorbutic cells than in the ascorbate-replete cells. These findings contrasted with those in a control line of scorbutic human dermal fibroblasts in which a 95% decrease in procollagen secretion was not associated with any procollagen accumulation in the cells. In the intestinal smooth muscle cells, the absence of ascorbate resulted in a 25 and 50% decrease in steady-state levels of procollagen I and III mRNA, respectively, compared to a 40 and 75% decrease in fibroblasts. Heat inactivation of the serum in the culture medium augmented the promotion of procollagen secretion by ascorbate two- to fourfold. L-ascorbate phosphate did not increase the activity of L-ascorbate when replaced in medium either daily or every 4 days, and its efficacy was not augmented by serum heat inactivation. The changing of culture medium induced collagen secretion in the absence of ascorbate, but this process was markedly enhanced by ascorbate and induced a transient decrease in the steady-state levels of both procollagen and nonprocollagen mRNAs. The predominant action of L-ascorbate on HISM cells in vitro is to promote procollagen secretion and not procollagen synthesis. L-ascorbate-phosphate is not an adequate substitute for L-ascorbate in this cell line.

Ascorbic Acid↗

PubMed1995

Corticosteroids increase procollagen gene expression, synthesis, and secretion by human intestinal smooth muscle cells.

BACKGROUND & AIMS: Collagen synthesis by smooth muscle cells plays an important role in intestinal fibrosis. Corticosteroids inhibit collagen synthesis in fibroblasts. The aim of this study was to examine the effect of corticosteroids on the expression of collagen by human intestinal smooth muscle (HISM) cells in vitro. METHODS: Collagen synthesis was determined by the sensitivity of radiolabeled protein to collagenase. Secretion was determined by polyacrylamide gel electrophoresis of radiolabeled procollagen in the medium. Procollagen messenger RNA was determined by Northern blot analysis. RESULTS: Collagen synthesis by confluent HISM cells was not affected by corticosteroids at 10(-10) to 10(-5) mol/L but, in subconfluent cultures, was nonspecifically increased 50% at 10(-5) mol/L. Procollagen secretion was nonspecifically increased 60% at 10(-6) mol/L dexamethasone without any effect on the type III/I ratio. Procollagen I and III messenger RNA levels responded in a biphasic manner: a 45%-65% increase at 10(-10) mol/L and a 15% and 30% decrease at 10(-8) and 10(-6) mol/L. In fibroblasts, collagen synthesis was inhibited 85% by dexamethasone, procollagen secretion was decreased 70%, the type III/I ratio decreased from 70:1 to 18:1, and procollagen messenger RNA was inhibited 25% and 60% (types I and III). CONCLUSIONS: Collagen expression by HISM cells is refractory to corticosteroids and, at certain concentrations, is augmented.

Blotting, Northern↗

PubMed1988

Effects of antibodies against odorant binding proteins on electrophysiological responses to odorants.

Monoclonal antibodies against two olfactory mucosal proteins, one with affinity for anisole-like and the other for benzaldehyde-like compounds, were applied to mouse olfactory epithelium. Responses to three odorants (anisole, benzaldehyde and amyl acetate) were measured. Of 26 antibodies, three (12%) inhibited responses only to the odorant with affinity for the antigen, nine (35%) inhibited responses to all three odorants, and 14 (54%) were without effect. None reduced responses by as much as 50%. The data support the hypothesis that there is a class of related proteins in olfactory neuronal cell membranes that function as receptor molecules and that other mechanisms also mediate odorant stimulation.

Animals↗

PubMed1984

NALL-1, an acute lymphoblastic leukemia cell line with peroxidase activity.

All cells examined from the non-B, non-T acute lymphoblastic leukemia cell line, NALL-1, stained positive both for terminal deoxynucleotidyl transferase and for common ALL antigen. In addition, peroxidase activity was detected by light microscopy in 55 to 75% of cells and peroxidase-positive granules were detected ultrastructurally in greater than 80% of cells. Peroxidase activity in NALL-1 may result from derepression of peroxidase genes or clonal proliferation of a biphenotypic precursor cell.

Adolescent↗