Klaus Vosbeck

Interleukin 1 and tumor necrosis factor synergistically stimulate prostaglandin synthesis and phospholipase A2 release from rat renal mesangial cells

Treatment of rat glomerular mesangial cells with recombinant human interleukin 1 alpha (rIL-1 alpha), recombinant human interleukin 1 beta (rIL-1 beta) or recombinant human tumor necrosis factor (rTNF) induces prostaglandin E2 (PGE2) synthesis and the release of a phospholipase A2 (PLA2) activity. rIL-1 beta is significantly more potent than rIL-1 alpha or rTNF in stimulating PGE2 as well as PLA2 release from mesangial cells. When given together, rTNF interacts in a synergistic fashion with rIL-1 alpha and rIL-1 beta to enhance both, PGE2 synthesis and PLA2 release. The released PLA2 has a neutral pH optimum and is calcium-dependent. Pretreatment of cells with actinomycin D or cycloheximide inhibits basal and cytokine-stimulated PGE2 and PLA2 release.

Transforming growth factor β2 inhibits interleukin 1β- and tumour necrosis factor α-induction of nitric oxide synthase in rat renal mesangial cells

Treatment of mesangial cells with interleukin 1 beta (IL-1 beta) or tumour necrosis factor alpha (TNF alpha) has been shown to induce nitric oxide (NO) synthase with subsequent autocrine stimulation of soluble guanylate cyclase (Pfeilschifter and Schwarzenbach, 1990, FEBS Lett. 273, 185-187). Here we report that transforming growth factor beta 2 (TGF beta 2) dose-dependently inhibits IL-1 beta- and TNF alpha-stimulated cGMP formation in mesangial cells. Half-maximal inhibition is observed at concentrations of 0.4 and 0.06 ng/ml of TGF beta 2, respectively. Maximum inhibition of cGMP formation over a 24 h period requires the presence of TGF beta 2 during the first 4 h of induction. In addition, the inhibitory effect of TGF beta 2 on cytokine-induced cGMP formation is not affected by the potent cyclo-oxygenase inhibitor indomethacin, thus excluding prostaglandins as mediators.

Hydrogen peroxide as a potent bacteriostatic antibiotic: Implications for host defense

Host defense against bacterial pathogens in higher organisms is mediated in part by the generation of reactive oxygen species (ROS) by PMN. In this study, we determined the following effects of exposure of constant concentrations of H2O2 on E. coli in a culture continuously monitored for H2O2 concentration, numbers, and viabilities of cells: (1) E. coli growth rates monitored for 1 h were profoundly affected by concentrations of H2O2, between 25-50 microM. (2) Complete bacteriostasis was observed at 100 microM. (3) Significant cell killing was not observed until the concentration of H2O2 was greater than 500 microM. (4) Bacteriostatic (25-50 microM) concentrations of H2O2 appeared not to be toxic to human skin fibroblasts for a 2-h exposure. (4) Bacteriostasis by H2O2 could not be explained by metabolic inhibition, because intracellular ATP levels were not compromised at bacteriostatic doses of H2O2. (5) Measurements of H2O2 concentrations in subcutaneous abscess fluid infected with both E. coli and S. aureus indicated prevailing concentrations of the oxidant consistent with a proposed role of H2O2 in host defense.

PDGF suppresses the activation of group II phospholipase A2 gene expression by interleukin I and forskolin in mesangial cells

Treatment of rat mesangial cells with interleukin 1β (IL‐1β) and forskolin greatly enhanced the expression of group II phospholipase A2 (PLA2) mRNA, with subsequent increased synthesis and secretion of PLA2 as detected by PLA2 activity measurements and immunoprecipitation of culture media of [35S]methionine‐labelled mesangial cells. PDGF—BB dose‐dependently suppressed the IL‐1β‐ and forskolin‐induced elevation of PLA2 mRNA, as well as PLA2 synthesis and secretion. In contrast, PDGF—AA had no inhibitory effect. The tyrosine kinase inhibitor genistein dose‐dependently antagonized the inhibitory effect of PDGF‐BB on IL‐1β‐stimulated PLA2 secretion, thus suggesting that tyrosine phosphorylation may be required for PDGF‐BB inhibition of PLA2 gene expression in mesangial cells.

Transforming growth factors type-β and dexamethasone attenuate group II phospholipase A2 gene expression by interleukin-1 and forskolin in rat mesangial cells

Treatment of rat mesangial cells with interleukin‐1β(IL‐1β) and forskolin induced, in a synergistic fashion,the expression of group II phospholipase A2 (PLA2) mRNA, with subsequent increased synthesis and secretion of PLA2. In contrast, interleukin‐6 did not increase PLA2 mRNA levels of PLA2 activity. Transforming growth factor (TGF) β1, TGFβ2 and TGFβ3 equipotently attenuated the IL‐1β‐ and forskolin‐induced elevation of PLA2 mRNA, as well as PLA2 synthesis and secretion. The glucocorticoid dexamethasone only partially suppressed the IL‐1β‐ and forskolin‐induced elevation of PLA2 mRNA, but totally inhibited PLA2 synthesis and secretion.

Magnetic resonance imaging and morphometric quantitation of cartilage histology after chronic infusion of interleukin 1 in rabbit knees.

Abstract Cartilage pathology in rabbit knees was monitored by noninvasive magnetic resonance imaging (MRI) and evaluated using morphometric histologic measurements. Infusion of rabbit knees with the cytokine interleukin 1 induces cartilage degradation and inflammation. A miniosmotic pump was implanted subcutaneously to deliver interleukin 1 through a polyethylene catheter inserted into the rabbit knee. Rabbit knees were imaged using MRI and prepared for histologic examination at 5 and 12 days after chronic infusion of interleukin 1. MRI obtained 0.7-mm sections for three-dimensional reconstruction of cartilage image. Cartilage deterioration near the site of infusion was visible on MRI. MRI measurements indicated a reduction in cartilage thickness. Histology revealed a loss of staining of cartilage matrix proteoglycan, synovial hypertrophy, and perichondral bone resorption. Morphometric analysis of cartilage histology indicated a reduction in both cellularity (chondrocytes/mμ2 area) and cell to matrix area ratio. These observations suggest that a loss of proteoglycan, an early event in cartilage degeneration, can be detected by MRI. [P.S.E.B.M. 1993, Vol 203]

Anti-inflammatory effects of CGP 47969A, a novel inhibitor of proinflammatory cytokine synthesis, in rabbit immune colitis☆

BACKGROUND & AIMS Proinflammatory cytokines such as interleukin (IL) 1, IL-8, and tumor necrosis factor (TNF) have been implicated as primary mediators of intestinal inflammation. The aim of the present study was to determine the effects of a novel cytokine antagonist (CGP 47969A) in a rabbit model of acute colitis. METHODS Colitis was induced using the formalin-immune complex technique. Animals were pretreated intrarectally with CGP 47969A (30, 10, or 3 mg/kg), hydrocortisone (0.8 mg/kg), or vehicle (4 mL saline) 2 hours before the induction of colitis and twice daily thereafter until death 48 hours after the induction of colitis. The severity of inflammation of colonic tissue was assessed using histological analysis and myeloperoxidase activity assay, and IL-1 alpha, IL-8, TNF-alpha, and IL-1 receptor antagonist levels were determined. RESULTS Compared with vehicle, CGP 47969A (10 mg/kg) significantly reduced the acute inflammatory index by 58%, edema by 67%, necrosis by 99%, and myeloperoxidase activity by 49% (all P < 0.02) with efficacy similar to that of steroids. These effects were associated with a significant inhibition of colonic IL-1 alpha and IL-8 by 56% and 90%, respectively (p < 0.01). CONCLUSIONS Administration of CGP 47969A reduces inflammation and tissue damage in rabbit immune complex colitis through mechanisms involving the inhibition of mucosal proinflammatory cytokines.

Quantitation of the rate of ingestion of Escherichia coli by human polymorphonuclear neutrophils using [3H]uracil

Abstract A method using [ 3 H]uracil to simply and accurately measure the rate of ingestion of E. coli by human polymorphonuclear neutrophils (PMN) is described. The method is based on the observation that [ 3 H]uracil penetrates bacterial membranes and is incorporated into RNA, whereas it is not taken up to a significant extent by PMN. Therefore, extracellular bacteria can be selectively labeled during pulses with [ 3 H]uracil whereas phagocytized bacteria are not labeled. Using this [ 3 H]uracil incorporation method along with viable count determinations, it was shown that although about 5% of ingested E. coli B-1385 remained viable for at least 3 h after being ingested by PMN, the surviving bacteria may have become damaged as shown by a decreased ability to incorporate [ 3 H]uracil into RNA when released from PMN. Similarly, it was shown that PMN treated with cytochalasin B continued to ingest E. coli B-1385, but the bacteria were not killed and, in fact, continued to grow. This method, together with other techniques of assessing bacterial phagocytosis, may therefore be useful for a more detailed analysis of the process of bacterial phagocytosis than was hitherto possible.