Brandt JL, Zumoff B, Castelman L, Ruskin HD, Jones ... New York, John Wiley and Sons,. 1985, pp 452- ... Forrest JW, John F, Milk LR, Buxton TB, Moore WL,.
American Journal of Pathology, Vol. 133, No. 3, December 1988 Copyright © American Association of Pathologists
Activation of Glomerular Mesangial Cells by GramNegative Bacterial Cell Wall Components D. H. LOVETT, MD,* S. L. BURSTEN, MD,f D. GEMSA, MD,* W. BESSLER, MD,§ K. RESCH, MD,11 andJ. L. RYAN, MD, PhD¶
From the Medical Service, San Francisco VAMC-University of California, San Francisco, California, * the Medical Service, Seattle VAMC-University of Washington, Seattle, Washingtont the Institutfiir Immunologie, Universidt Marburg, Marburg,* the Institutfir Immunobiologie der Universitdt, Freiburg,§ the
Institutfuir Molekularpharmakologie, Medizinische Hochschule, Hannover, Federal Republic of Germany, I and the Medical Service, West Haven VAMC-Yale University, New Haven, Connecticutl
The cell walls of gram-negative bacteria contain several biologically active components, including lipopolysaccharide (LPS), lipoprotein, and protein 1. The effects of these individual components and a synthetic analog of lipoprotein, TPP, on several activation parameters ofglomerular mesangial cells (MC) were examined. Prostaglandin secretion, synthesis of the autogrowth factor, mesangial interleukin-1 (IL-1), and new synthesis of cellular proteins were assessed as markers of MC activation. All bacterial cell wall components evaluated were active in varying degrees as stimulants of prostaglandin secretion. In general, PGE was the predominant product. TPP and protein 1 also induced substantial secretion of thromboxane. Each cell-wall
component was effective in stimulating mesangial IL1 secretion. The activation of MC was associated with the enhanced synthesis ofmany cellular proteins in addition to IL-1. Stimulation by these bacterial components was dependent on the state of the mesangial cell cycle, because nonproliferating cells did not respond to these factors. Activation of MC by gram-negative bacterial cell wall components, with release of vasoactive prostaglandins and peptide mitogens, may be responsible for some ofthe glomerular hemodynamic alterations and cellular proliferative events associated with sepsis or chronic bacterial infection. (AmJ Pathol 1988, 133:472-484)
IT HAS BEEN LONG recognized that bacterial infection is associated with alterations in renal function. `Severe bacterial infection with gram-negative organisms frequently results in acute renal insufficiency, a condition that is a major complication of the endotoxic shock syndrome. The functional changes in renal glomeruli that accompany the development of renal insufficiency include an increase in renal vascular resistance and a decline in glomerular filtration rate.5-9 These hemodynamic and functional alterations can occur independently of changes in the systemic circulation,8'9 and are in part the result of the activation of several humoral mediator systems.5 These mediator systems include complement, the kinin system, and the coagulation cascade. In addition, platelet or inflammatory cell-derived thromboxane and vasoconstricting leukotrienes have been implicated in the pathogenesis of the systemic and renal he-
modynamic perturbations found in several models of endotoxic shock.4"0"' Structural alterations in glomeruli may also occur during acute gram-negative bacterial infections, including glomerular endothelial cell injury characterized by cellular swelling and formation of intracapillary microthrombi.'2 Patients with Salmonella typhi or Shigella infection can develop the glomerular lesions characteristic of the hemolytic-uremic syn-
472
Accepted for publication July 14, 1988. Supported by Research Funds of the Veterans Administration, grants from the Deutsche Forschungsgemeinschaft (Ge 354/5-2, Re 281/9-1), and by the Alexander von Humboldt Stiftung, Bonn, FRG. Address reprint requests to D. H. Lovett, Medical Service 1 1 1J, San Francisco VAMC-University of California at San Francisco, 4150 Clement Street, San Francisco, CA, 94121.
MC ACIIVATION BY BACrERIAL COMPONENTS
Vol. 133 * No. 3
drome, an event attributed to circulating bacterial endotoxin.'3"4 Although considerably less frequent, chronic gram-negative bacterial infections are associated with the development of glomerulonephritis, primarily of the membranoproliferative form.5" 9 The glomerulus generally has been viewed as a passive target structure during these processes; however, recent evidence derived from a model of endotoxic renal failure indicated that vasoactive prostaglandins and leukotrienes are also derived from an intrinsic, presumably glomerular, source. 1 Of the cellular components of the glomerulus, endothelial cells and mesangial cells (MC) represent the most likely sources of these mediators. Endothelial cells have been shown in both animals models and in vitro to be highly sensitive to the effects of bacterial endotoxin, leading to cellular detachment, lysis, and release of prostaglandins.2024 To date, there is little information available concerning the sensitivity or reactivity of the glomerular mesangial cell to bacterial components. The mesangium is a frequent site for the deposition ofphlogogenic material, due in part to the high glomerular capillary pressures and the absence of a limiting basement membrane. Similar factors could conceivably result in the mesangial localization of bacterial cell wall components during infectious states, an event that has been demonstrated in some models of experimental glomerulonephritis.25 The importance of the intrinsic MC as an effector cell is related to its central localization within the glomerular capillary tuft and the extensive synthetic repertoire of vasoactive prostaglandins, reactive oxygen species and peptide mitogens released by these cells in response to inflammatory stimuli.26 Given the potential pathophysiologic significance of MC activation by bacterial components, it was decided to examine the effects ofpurified gramnegative bacterial cell wall components on several parameters of MC activation. These parameters included induction of cellular protein synthesis and secretion of prostaglandins and mesangial IL- 1. In addition to its action as an autogrowth factor for MC,27 IL- 1 induces the secretion ofprostaglandins and a specific mesangial type IV collagenase, events that could further amplify glomerular inflammatory processes.28'29 Gram-negative bacterial cell wall components, including lipopolysaccharide, are shown in this report to have distinct stimulatory effects upon cultured MC. These findings are of potential relevance to the renal pathophysiologic alterations characteristic ofbacterial infection.
Materials and Methods Animals Male Sprague-Dawley rats, 125-150 g were used as a source of thymocytes and mesangial cells.
473
Media and Reagents
Actively growing rat MC were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 20% heat-inactivated fetal calf serum (FCS, Gibco, Paisley, UK), 100 U/ml penicillin, 100 ,ug/ml streptomycin, and 300 ,tg/ml glutamine (growth medium). Medium (rest medium) to induce viable, noncycling, Go/GI mesangial cells consisted of Medium 199 (Gibco), 0.5% bovine serum albumin (BSA), 1 X 10-6 M bovine insulin, 50 U/ml penicillin, 50 ,ug/ml streptomycin, and 300 ,ug/ml glutamine.27'30 All media components were screened for the presence of exogenous endotoxin using a Limulus amoebocyte lysate assay sensitive to
