Abstract
Intravascular hemolysis of any cause can induce acute kidney injury (AKI). Hemolysis-derived product heme activates the innate immune complement system and contributes to renal damage. Therefore, we explored the role of the master complement regulator Factor H (FH) in the kidney's resistance to hemolysis-mediated AKI. Acute systemic hemolysis was induced in mice lacking liver expression of FH (hepatoFH−/−, ~20% residual FH) and in WT controls, by phenylhydrazine injection. The impaired complement regulation in hepatoFH−/− mice resulted in a delayed but aggravated phenotype of hemolysis-related kidney injuries. Plasma urea as well as markers for tubular (NGAL, Kim-1) and vascular aggression peaked at day 1 in WT mice and normalized at day 2, while they increased more in hepatoFH−/− compared to the WT and still persisted at day 4. These were accompanied by exacerbated tubular dilatation and the appearance of tubular casts in the kidneys of hemolytic hepatoFH−/− mice. Complement activation in hemolytic mice occurred in the circulation and C3b/iC3b was deposited in glomeruli in both strains. Both genotypes presented with positive staining of FH in the glomeruli, but hepatoFH−/− mice had reduced staining in the tubular compartment. Despite the clear phenotype of tubular injury, no complement activation was detected in the tubulointerstitium of the phenylhydrazin-injected mice irrespective of the genotype. Nevertheless, phenylhydrazin triggered overexpression of C5aR1 in tubules, predominantly in hepatoFH−/− mice. Moreover, C5b-9 was deposited only in the glomeruli of the hemolytic hepatoFH−/− mice. Therefore, we hypothesize that C5a, generated in the glomeruli, could be filtered into the tubulointerstitium to activate C5aR1 expressed by tubular cells injured by hemolysis-derived products and will aggravate the tissue injury. Plasma-derived FH is critical for the tubular protection, since pre-treatment of the hemolytic hepatoFH−/− mice with purified FH attenuated the tubular injury. Worsening of acute tubular necrosis in the hepatoFH−/− mice was trigger-dependent, as it was also observed in LPS-induced septic AKI model but not in chemotherapy-induced AKI upon cisplatin injection. In conclusion, plasma FH plays a key role in protecting the kidneys, especially the tubules, against hemolysis-mediated injury. Thus, FH-based molecules might be explored as promising therapeutic agents in a context of AKI.
Highlights
Intravascular hemolysis is responsible for acute kidney injury (AKI), partly due to cellular damage induced by hemoglobin and its breakdown product heme [1]
The heatmap incorporates for comparison the data from panels (A,B,D,F). (F) Renal expression of VCAM-1 (Vcam1) mRNA after PBS orPHZ injection in wild type C57BL/6 mice (WT) and hepatoFH−/− mice. (G) Validation by VCAM-1 immunohistochemistry staining in embedded paraffin, seen in peritubular capillaries and arteriols (*) in hepatoFH−/− at day 4 post PHZ vs. PBS injection (40x)
P-values are derived from Two way ANOVA test with Sidak correction for multiple comparisons: *p < 0.05; **p < 0.005; ***p < 0.001; ****p < 0.0001. mRNA values are represented in fold change (FC), relatively to the mean expression of PBS treated mice for each strain
Summary
Intravascular hemolysis is responsible for acute kidney injury (AKI), partly due to cellular damage induced by hemoglobin and its breakdown product heme [1]. Renal injury related to hemolysis has been described in genetic hemoglobinopathies (like sickle cell disease, SCD), in malaria or in transfusion of stored red blood cells, where hemolysis-derived products injure renal structures, the proximal tubules [2, 3]. These products and especially heme, have been shown to activate the complement system [4]. Complement activation drives acute tubular injury, which was prevented in C3−/− mice [5, 6, 8]. Heme renders the endothelial cells (EC) susceptible to complement attack [7, 9, 13]
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