Brain microvascular endothelial cells exhibit lower activation of the alternative complement pathway than glomerular microvascular endothelial cells

Sarah E Sartain,Nancy A Turner,Joel L Moake

doi:10.1074/jbc.ra118.002639

Abstract

Atypical hemolytic uremic syndrome (aHUS) and bone marrow transplantation-associated thrombotic microangiopathy (TA-TMA) are associated with excessive activation of the alternative complement pathway (AP) and with severe renal, but rarely cerebral, microvascular damage. Here, we compared AP activation and regulation in human glomerular and brain microvascular endothelial cells (GMVECs and BMVECs, respectively) unstimulated or stimulated by the proinflammatory cytokine, tumor necrosis factor (TNF). Compared with GMVECs and under both experimental conditions, BMVECs had increased gene expression of the AP-related genes C3, CFB, and C5 and decreased expression of CFD This was associated with increased expression in BMVECs (relative to GMVECs) of the genes for surface and soluble regulatory molecules (CD46, THBD, CD55, CFI, and CFH) suppressing formation of the AP C3 and C5 convertases. Of note, unlike GMVECs, BMVECs generated extremely low levels of C3a and C5a and displayed decreased activation of the AP (as measured by a lower percentage of Ba generation than GMVECs). Moreover, BMVECs exhibited increased function of CD141, mediating activation of the natural anticoagulant protein C, compared with GMVECs. We also found that the C3a receptor (C3aR) is present on both cell types and that TNF greatly increases C3AR1 expression in GMVECs, but only slightly in BMVECs. Higher AP activation and C3a generation in GMVECs than in BMVECs, coupled with an increase in C3aR production in TNF-stimulated GMVECs, provides a possible explanation for the predominance of renal damage, and the absence of cerebral injury, in individuals with episodes of aHUS and TA-TMA.

Highlights

That TNF-stimulated BMVECs released higher concentrations of both C3 and FB than TNF-stimulated GMVECs, a smaller percentage of possible C3 convertases was formed in BMVECs
Our data indicate that BMVECs are more effective in resisting TNF-mediated AP activation than GMVECs, possibly contributing to the explanation for the absence of cerebral microvascular injury and the prominence of renal damage in patients with inflammation/infection-induced episodes of aHUS and TA-TMA
The AP component gene expression levels of C3, CFB, and C5 in BMVECs were much higher than the expression levels in GMVECs

Summary

To whom correspondence should be addressed

Lytic anemia, thrombocytopenia, and microvascular endothelial injury [1,2,3]. Atypical hemolytic uremic syndrome (aHUS) and bone marrow transplantation-associated TMA (TA-TMA) are TMAs associated with abnormalities in the alternative complement pathway (AP) as well as severe renal damage [3,4,5,6]. The AP is one of three pathways of the innate immune complement system responsible for protecting the host from unwanted pathogens or foreign invaders It is initiated when C3b is cleaved from complement component C3 and attaches to an activating surface, releasing a soluble C3a fragment [7, 8]. Receptors for vascular endothelial cell; TNF, tumor necrosis factor; C3aR, complement C3a receptor; FB, factor B; FD, factor D; ULVWF, ultra large von Willebrand factor; HUVEC, human umbilical vein endothelial cell; CP, classical complement pathway; C5aR, complement C5a receptor; FH, factor H; FI, factor I; THBD, thrombomodulin; PC, protein C; BSA, bovine serum albumin; VWF, von Willebrand factor; Ab, antibody; qPCR, quantitative PCR; PE, phycoerythrin. These MVECs serve as ideal models for our studies as they produce and secrete all AP components and regulators, as well as VWF [18, 55]

Results

Discussion

Experimental procedures