Endothelial Dysfunction and Vascular Inflammation

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Transfection of monoblastoid U937 cells with human eNOS (endothelial nitric oxide synthase) resulted in a cell line that produces nitric oxide (NO) in response to a calcium ionophore, but little or no NO in the resting state (Blood, 1997). However, after differentiation with phorbol-12-acetate-13-myristate, eNOS expressing cells produced increased amounts of both TNFalpha and reactive oxygen species by mechanisms that were independent of NO. Neither N-omega-methyl-L-arginine, a NOS inhibitor, nor mutation of the L-arginine binding site of eNOS (rendering it incapable of producing nitric oxide) blocked the ability of eNOS to upregulate TNFalpha (J Biol Chem, 2000). Conversely, co-transfection with superoxide dismutase or deletion of the NADPH binding site of eNOS completely prevented eNOS from upregulating TNFalpha production. Ultimately, superoxide produced by eNOS was shown to upregulate TNFalpha via Erk1/2 activation (J Biol Chem, 2001). This series of investigations demonstrated that while high output NO from inducible NOS can enhance inflammation (J Immunol, 1994; J Biol Chem, 1997), dysregulation of eNOS can also contribute to an inflammatory phenotype through reactive oxygen species-based signal transduction pathways. Sp1 was shown to act as a sensor of high-output NO, down regulating eNOS in endothelial cells via a proximal Sp promoter-binding site (J Biol Chem, 2003). This work defined a key negative feedback mechanism that suppresses eNOS and reinforces vascular dysfunction in septic shock. Sickle cell disease was characterized by oxidant and inflammatory stress in the vasculature, even in the absence of an acute crisis (Blood, 2004). Circulatory stress in sickle cell disease was associated with abnormalities of arginine metabolism in platelets (Circulation, 2007), further characterizing this condition as an NO-deficient state. Anti-proliferative effects of NO that prevent the development of a chronic vascular injury phenotype was linked to p38 MAPK activation and p21 mRNA stabilization with subsequent down regulation of polo-like kinase 1 through a CDE/CHR proximal promoter site (BMC Genomics, 2005; J Biol Chem, 2006). This and later investigations (see below) indicate that p38 MAPK may play a protective role in the vasculature under some conditions. Both NO and peroxisome proliferator-activated receptors (PPARs) protect the endothelium and regulate its function. Therefore, we tested for crosstalk between these signaling pathways using human umbilical vein and hybrid EA.hy926 endothelial cells (FASEB J, 2007). PPARgamma was activated by NO through a p38 MAPK dependent signal transduction pathway. This crosstalk mechanism may contribute to the anti-inflammatory and cytoprotective effects of NO in the vasculature. Recent work has shown that p38 MAPK is also necessary for the optimal activation of PPAR gamma by thiazolidinediones (TZDs), drugs used in the treatment of type 2 diabetes mellitus. TZDs, like homeostatic NO production, reduce markers of cardiovascular inflammation. Collectively, these findings suggest that PPAR gamma as well as other nuclear receptors with transrepression activity may be useful targets for reducing endothelial dysfunction and vascular inflammation in sepsis and septic shock.