Isolated glomeruli and cultured mesangial cells as in vitro models to study immunosuppressive agents

Abstract

Immunosuppressive agents, such as cyclosporin A (CsA), by their vasoconstrictive properties, induce in vivo in patients and rodents a dramatic fall in renal hemodynamics. The aim of this study is to review the ability of some physiological and/or pharmacological agents which are supposed to be involved in the renal physiopathology of CsA to prevent the contraction induced by CsA in two in vitro glomerular models. Isolated glomeruli are obtained by a sieving method from male Sprague-Dawley rat superficial cortex. Mesangial cells from these isolated glomeruli are cultured in RPM1 1640 medium with 20% FCS in 5% CO2 atmosphere. The area of isolated glomeruli and cultured mesangial cells is assessed by an image analyzer with a video camera. Each glomerulus and cell is its own control and is photographed before incubation with any drug (T0) and then during incubation at 5, 10, 20, and 30 min. Incubations are performed during 30 min with 10(-6) mol/L CsA either with a 10 min pretreatment with the vasoactive agent or without pretreatment. CsA alone induces a time- and dose-dependent decrease in glomerular structure area (-4.7% at 10 min, -10.3% at 20 min, and -12.0% at 30 min for isolated glomeruli); Cremophore excipient or control solute does not induce any significant decrease in surface area. CsA with 10(-6) mol/L verapamil pretreatment induces only a slight decrease: -1.5% at 10 min, -3.0% at 20 min, and -4.8% at 30 min. Calcium blockers nifedipine and felodipine produce similar results. Likewise, with 10(-8) mol/L prostacyclin analog (iloprost), only a slight area decrease in mesangial cells is noted: -1.3% at 5 min, -1.8% at 10 min, and -3.3% at 20 min; with 10(-6) mol/L TXA2 synthesis inhibitor (CGS 12970) the results are -2.0% at 10 min, -3.6% at 20 min, and -4.3% at 30 min. Finally, a similar protective effect can be noted with 10(-5) mol/L theophylline: -0.4; -1.5 and -1.9% at 10, 20, and 30 min. In conclusion, CsA-induced contraction in two in vitro glomerular models can be partially or even totally prevented by pretreatment with various pharmacological agents.