Morphological and biochemical effects of immunosuppressive drugs in a capillary tube assay for endothelial dysfunction

Abstract

Immunosuppressive drugs common in clinical transplantation are known to have untoward effects on the vascular system. The effects of some drugs, notably cyclosporin A (CyA), have been studied on the vascular system, while those of others have not. In the vascular system, endothelial cells are the predominant cell type exposed to intravascular concentrations of immunosuppressive drugs. We therefore studied the effects of drugs common in clinical transplantation on endothelial cells in a capillary tube assay. The endothelial cells in the capillary tubes are morphologically more similar to those in the microvasculature than endothelial cells in monolayers. We studied the kinetics and extent of capillary tube formation and prostacyclin (PGI2) and endothelin-1 (ET-1) release from the in vitro capillaries to determine the morphological and biochemical effects of five immunosuppressive agents on endothelial function. We found a significant difference in the morphological and biochemical effects of the two common calcineurin inhibitors, CyA and tacrolimus (FK506) on capillary morphology in vitro. The former had a pronounced injurious effect on the morphology of the in vitro capillaries, while the latter did not. CyA also significantly increased ET-1 release by the capillaries, but FK506 did not. Mycophenolate mofetil (MMF) was the only other agent that had a moderately injurious effect on the morphology of the in vitro capillaries. Sirolimus (rapamycin) and dexamethasone, similar to FK506, had no effect on the capillary morphology. All these agents, except dexamethasone, increased PGI2 release. Our data suggest that CyA adversely affects the morphology of the microvasculature and that this is mediated, at least partly, by an increased ET-1 release by endothelial cells exposed to CyA. These findings describe a novel effect of CyA and MMF on endothelial cells that could be relevant to understanding the mechanisms of immunosuppressive drug-mediated endothelial injury in clinical transplantation.

mTOR Inhibitors and Calcineurin Inhibitors Do Not Affect Adhesion Molecule Expression of Human Macro- and Microvascular Endothelial Cells

“…We could show that long-term incubation of micro- and macrovascular EC with both mTOR inhibitors had no effect on the basal release of IL-8 and MCP-1 [27], as well as on the expression of CAM. Due to different experimental protocols in the literature [5,6,7, 13, 14], using hyperconfluent seeding of EC for 24 h accompanied by drug treatment for an additional 24–48 h, we validated our previous data in a confluent monolayer experiment. The drug effects were independent of the seeding protocol.…”

“…Karlsson and Nassberger [22] confirmed our finding that Tac does not affect EC activation in response to inflammatory stimuli like TNF. Moreover, up to supratherapeutic Tac levels, there seem to be no deleterious effects on capillaries in vitro [7]. …”

“…The impact of immunosuppression on individual endothelial properties has been investigated in several in vitro models [5,6,7]. However, most of these studies used human umbilical veins, immortalized or animal-derived EC often without considering the functional heterogeneity of EC derived from different origins [8, 9].…”

Proliferation Signal Inhibitor–induced Decrease of Vascular Endothelial Cadherin Expression and Increase of Endothelial Permeability In Vitro Are Prevented by an Anti-oxidant

“…To date, only one hypothesis for this phenomenon has been presented: Wilasrusmee et al found that prostacyclin release was stimulated by SRL on human aortic ECs. 30 Pascual et al speculated that prostacyclin-induced vasodilation might be responsible for capillary leakage in SRL-treated patients. 8 Increased prostacyclin release was found after treatment with CsA and FK506.…”

Effects of Immunosuppressive agents on neutrophils inflammatory response in humans: An inflammatory perspective on coronary allograft vasculopathy

“…The difference between CsA and TAC was further evaluated in an in vitro study analyzing the prostaglandin I 2 (PGI 2 ) and ET-1 on microvascular capillaries [87] . In this study, CsA caused some detrimental effects on capillary morphology and resulted in a significant increase in capillary ET-1 release, while TAC did not [66] . Furthermore, CsA yielded a greater impairment in brachial endothelial function using high resolution vascular ultrasound in renal transplant patients, compared with TAC treated patients [67] .…”