Hepatic Overexpression of Hemopexin Inhibits Inflammation and Vascular Stasis in Murine Models of Sickle Cell Disease

Abstract

Sickle cell disease (SCD) patients have low serum hemopexin (Hpx) levels due to chronic hemolysis. We hypothesized that in SCD mice, hepatic overexpression of hemopexin would scavenge the proximal mediator of vascular activation, heme, and inhibit inflammation and microvascular stasis. To examine the protective role of Hpx in SCD, we transplanted bone marrow from NY1DD SCD mice into Hpx -/-or Hpx +/+ C57BL/6 mice. Dorsal skin fold chambers were implanted 13 wks post-transplant, and microvascular stasis (% nonflowing venules) was evaluated in response to heme infusion. Hpx -/-sickle mice had significantly greater microvascular stasis in response to heme infusion than Hpx +/+ sickle mice (p < 0.05), demonstrating the protective effect of Hpx in SCD. We utilized Sleeping Beauty (SB) transposon-mediated gene transfer to overexpress wild-type rat Hpx (wt-Hpx) in NY1DD and Townes-SS SCD mice. Control SCD mice were treated with lactated Ringer's solution (LRS) or a luciferase (Luc) plasmid. Plasma and hepatic Hpx were significantly increased compared with LRS and Luc controls. Microvascular stasis in response to heme infusion in NY1DD and Townes-SS mice overexpressing wt-Hpx had significantly less stasis than controls (p < 0.05). Wt-Hpx overexpression markedly increased hepatic nuclear Nrf2 expression, HO-1 activity and protein, and the heme-Hpx binding protein and scavenger receptor CD91/LRP1, and decreased NF-κB activation. Two missense (ms)-Hpx SB constructs that bound neither heme nor the Hpx receptor CD91/LRP1 did not prevent heme-induced stasis. In conclusion, increasing Hpx levels in transgenic sickle mice via gene transfer activates the Nrf2/HO-1 antioxidant axis and ameliorates inflammation and vasoocclusion.

Intravascular hemolysis and the pathophysiology of sickle cell disease

“…This was recently confirmed in a study of SCD patients correlating high plasma hemoglobin levels appreciated as an additional important mediator of inflammation and vascular injury (102,103). In sickle cell mice, free heme drives inflammation, vaso-occlusion, and coagulation that are blocked by the heme scavenger hemopexin (104)(105)(106)(107)(108)(109). In cultured cells, heme promotes secretion of high levels of placenta growth factor (110), which in turn induces release of the potent vasoconstrictor endothelin 1 (111), a common mediator of PH.…”

Intravascular hemolysis activates complement via cell-free heme and heme-loaded microvesicles

“…Nevertheless, Hx, but not HSA, was effective at preventing tissue injury and inflammation in mouse models of hemolysis (5,(36)(37)(38)(39). Previous reports have also shown that hepatic overexpression of Hx inhibits inflammation and vascular stasis in murine SCD (40), while Hx -/mice develop a severe renal damage after experimental hemolysis. This points to Hx as a guardian of kidney function during intravascular hemolysis (41).…”

Heme as a Target for Therapeutic Interventions

“…The major function of Hx appears to be neutralization and scavenging of excess free heme from the circulation. Up-take of heme-Hx complexes in the liver (Potter et al, 1993) is mediated via the scavenger receptor low-density lipoprotein receptor-related protein-1 (LRP1, synonymous with CD91) (Hvidberg et al, 2005; Vercellotti et al, 2016). Hx belongs to the acute-phase reactants, which include a number of plasma proteins such as C-reactive protein, α2-macroglobulin and α1-antitrypsin that are up-regulated in the liver as part of a systemic inflammatory response (Heinrich et al, 1990; Baumann and Gauldie, 1994).…”