INTRODUCTION. Cerebral malaria (CM) is a life-threatening complication characterized by sequestration of infected erythrocytes (IRBC) in brain microvessels, blood-brain barrier (BBB) damage, increased inflammation and infiltration of macrophages partially laden with hemozoin (HZ, malarial pigment). Although the mechanisms underlying BBB damage in CM are not well understood, in the recent years growing evidence on dysregulation of matrix metalloproteinases (MMP), proteolytic enzymes involved in inflammation, tight junction disruption, and matrix invasion has emerged. Here the effects of HZ on production of MMP and their endogenous inhibitors (TIMP, Tissue inhibitors of Metalloproteinase) were investigated in human endothelial cells (EC) from either large or small caliber vessels. METHODS. Human umbilical vascular EC (HUVEC) and human dermal microvascular endothelial cells (HMEC-1) were treated with HZ for 48 h. Thereafter, the release and activation of MMP-1, -2, -3, -9, along with TIMP-1 and -2, was evaluated in cell supernatants by western blotting or gelatin zymography. RESULTS. In HUVEC, HZ promoted the release of pro-MMP-9 (but not active MMP-9) and TIMP-2, whereas basal levels of all other molecules were not affected. In HMEC-1, HZ induced either pro- or active MMP-9 and enhanced MMP-1, MMP-3 and TIMP-2 protein release, whereas basal levels of proMMP-2 and TIMP-1 were not altered, and active MMP-2 was not found. CONCLUSION. HZ induces proMMP-9 protein expression in human endothelium, independently on vessel caliber. However, HZ appears to switch on the MMP-9 activation machinery in micro-vascular endothelial cells only, from which active MMP-9 is released, as a likely consequence of MMP-1/MMP-3 enhanced levels and TIMP-1 unaltered levels. On the contrary, in both macro- and micro-vascular EC HZ does not affect pro-MMP-2 expression and active MMP-2 is not generated, probably due to increased TIMP-2 levels. We suggest that such a selectivity for microvascular EC displayed by malarial pigment in promoting MMP dysregulation may concur to explain the etiology of BBB damage in CM.
New insights for mechanisms of BBB damage in cerebral malaria: differential MMP regulation by haemozoin in human macro/microvascular endothelial cells.
GIRIBALDI, Giuliana;CARDAROPOLI, Simona;PRATO, Mauro
2012-01-01
Abstract
INTRODUCTION. Cerebral malaria (CM) is a life-threatening complication characterized by sequestration of infected erythrocytes (IRBC) in brain microvessels, blood-brain barrier (BBB) damage, increased inflammation and infiltration of macrophages partially laden with hemozoin (HZ, malarial pigment). Although the mechanisms underlying BBB damage in CM are not well understood, in the recent years growing evidence on dysregulation of matrix metalloproteinases (MMP), proteolytic enzymes involved in inflammation, tight junction disruption, and matrix invasion has emerged. Here the effects of HZ on production of MMP and their endogenous inhibitors (TIMP, Tissue inhibitors of Metalloproteinase) were investigated in human endothelial cells (EC) from either large or small caliber vessels. METHODS. Human umbilical vascular EC (HUVEC) and human dermal microvascular endothelial cells (HMEC-1) were treated with HZ for 48 h. Thereafter, the release and activation of MMP-1, -2, -3, -9, along with TIMP-1 and -2, was evaluated in cell supernatants by western blotting or gelatin zymography. RESULTS. In HUVEC, HZ promoted the release of pro-MMP-9 (but not active MMP-9) and TIMP-2, whereas basal levels of all other molecules were not affected. In HMEC-1, HZ induced either pro- or active MMP-9 and enhanced MMP-1, MMP-3 and TIMP-2 protein release, whereas basal levels of proMMP-2 and TIMP-1 were not altered, and active MMP-2 was not found. CONCLUSION. HZ induces proMMP-9 protein expression in human endothelium, independently on vessel caliber. However, HZ appears to switch on the MMP-9 activation machinery in micro-vascular endothelial cells only, from which active MMP-9 is released, as a likely consequence of MMP-1/MMP-3 enhanced levels and TIMP-1 unaltered levels. On the contrary, in both macro- and micro-vascular EC HZ does not affect pro-MMP-2 expression and active MMP-2 is not generated, probably due to increased TIMP-2 levels. We suggest that such a selectivity for microvascular EC displayed by malarial pigment in promoting MMP dysregulation may concur to explain the etiology of BBB damage in CM.
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