Effect of anti-C5 antibody on recuperation from ischemia/reperfusion-induced acute kidney injury

Abstract

AbstractAim: The complement system is activated in acute kidney injury (AKI). Anti-C5 antibody targets the common terminal portion of the complement cascade that generate the terminal complex C5b-9 and has a renal-protective effect in paroxysmal nocturnal hemoglobinuria. However, the anti-C5 antibody's role in ischemia/reperfusion (I/R)-induced AKI has not been fully investigated. We therefore evaluated its effect on the pathophysiological cascade of I/R-induced AKI.Methods: Sprague-Dawley rats underwent unilateral right kidney nephrectomies with simultaneous clamping of the contralateral hilum for 60 min (ischemia), followed by reperfusion. In addition to a placebo, two treatment groups received either high or low doses of anti-C5 monoclonal antibody. After 48 h, the rats were euthanized, blood was drawn to evaluate systemic inflammation and to estimate glomerular filtration rate (GFR). The remaining kidney was removed for pathological evaluation and intra-renal complement activation.Results: I/R induced significant intra-renal complement activation and systemic inflammation compared with unilateral nephrectomy group. The anti-C5 antibody ameliorated the intra-renal complement activation (intra-renal C3 and C6), reduced systemic inflammation (C-reactive protein, and systemic C3), decreased intra-renal acute tubular necrosis damage and improved GFR (seen by the sensitive marker, serum cystatin C; 1.63 mg/L (I/R + placebo), 1.36 mg/L (I/R + low dose) and 1.21 mg/L (I/R + high dose), p = .08 and .03 compared with I/R + placebo).Conclusion: In I/R-induced AKI, the monoclonal anti-C5 complement factor ameliorates intra renal complement activation, decreases local and systemic inflammation and may improve GFR.

Keywords: Acute kidney injury; C5-antibody; complement system; ischemia-reperfusion injury; rat model.

Figure 1.

Serum cystatin C levels for sham and ischemia-reperfusion groups. Serum cystatin C levels are in milligram per liter for all groups. Group comparisons were calculated using Student’s independent t-test. A comparison was conducted between both the sham groups: sham + placebo and I/R + placebo and between I/R + placebo and I/R + treatment groups. Data shown are mean ± SD. *p < .05 for I/R + placebo group versus I/R + treatment group. #p < .05 for I/R + placebo group versus Sham + placebo group. I/R: ischemia-reperfusion.

Figure 2.

Serum C-reactive protein levels for sham and ischemia-reperfusion groups. Serum CRP levels are in milligram per liter for all groups. Group comparisons were calculated using Student’s independent t-tests. A comparison was conducted between both the sham groups: sham + placebo and I/R + placebo and between I/R + placebo and I/R + treatment groups. Data shown are mean ± SD. *p < .05 for I/R + placebo group versus I/R + treatment group. #p < .05 for I/R + placebo group versus Sham + placebo group. I/R: ischemia-reperfusion; CRP: C-reactive protein.

Figure 3.

Renal immunofluorescence staining for C3 and C6 in the sham and ischemia/reperfusion groups treated by placebo or anti-C5 antibody. (A) Immunofluorescence of C3 and C6 of sham and I/R groups. Nucleolus is seen in blue. Arrows pointing to immune deposits in tubular cells. (B) Intensity evaluation of C3 and C6 immunofluorescence. *p<.05 for I/R + placebo group versus I/R + treatment group. #p<.05 for I/R + placebo group versus Sham + placebo group. I/R: ischemia-reperfusion.

Figure 4.

Renal pathology in different study groups. Hematoxylin and eosin staining for sham and I/R groups. Arrows point to tubular necrosis and damage. Hematoxylin and eosin staining of renal biopsy for sham and after I/R. All histologic slides show preserved glomeruli, without specific glomerular pathology. Tubular damage is the prominent feature with severe tubular necrosis for I/R + placebo, manifested as tubular cells detached from the tubular structure, and loss of the cell nucleus. Tubular cells in both treatment groups are less prominent. In the higher dose of I/R + 20 mg, tubules demonstrate hyaline casts inside tubular lumen. I/R: ischemia reperfusion.