Treatment with an Inhibitory Monoclonal Antibody to Mouse Factor B Protects Mice from Induction of Apoptosis and Renal Ischemia/Reperfusion Injury

Abstract

Complement activation in the kidney after ischemia/reperfusion (I/R) seems to occur primarily via the alternative complement pathway. The ability of an inhibitory mAb to mouse factor B, a necessary component of the alternative pathway, to protect mice from ischemic acute renal failure was tested. Treatment with the mAb prevented the deposition of C3b on the tubular epithelium and the generation of systemic C3a after renal I/R. Treated mice had significantly lower increases in serum urea nitrogen and developed significantly less morphologic injury of the kidney after I/R. For gaining insight into potential mechanisms of protection, the activity of caspases within the kidney also was measured, and it was found that caspases-2, -3, and -9 increased in a complement-dependent manner after renal I/R. Apoptotic cells were detected by terminal deoxynucleotidyl transferase catalyzed labeling of DNA fragments, and mice in which the alternative pathway was inhibited demonstrated significantly less apoptosis than control mice. Thus, use of an inhibitory mAb to mouse factor B effectively prevented activation of complement in the kidney after I/R and protected the mice from necrotic and apoptotic injury of the tubules. T he alternative pathway of complement is activated after ischemia/reperfusion (I/R) of the kidneys in rodents and humans (1,2), and complement deficiency protects mice from the full development of ischemic acute renal failure (ARF) (2,3). Complement activation by the classical and lectin pathways has been demonstrated after I/R of the heart (4), intestine (5,6), and skeletal muscle (7). Mannose-binding lectin is deposited in the postischemic kidney (8), perhaps indicating activation of this pathway. Complement activation after I/R of the kidney is critically dependent on an intact alternative pathway (2,3), however, and does not require the protein C4 (3). Selective inhibition of the alternative pathway, therefore, should confer protection against the development of ischemic ARF while leaving the classical and lectin pathways intact.Renal I/R results in both epithelial cell necrosis and apoptosis (9). Although necrosis is often the more prominent morphologic finding, inhibition of apoptosis has been shown to protect mice from ischemic ARF (10). The caspases are a group of cysteine proteases that execute many of the processes that are necessary for cellular apoptosis (11). Studies have demonstrated that complement activation directly activates caspases within renal cells (12) and may also induce caspase activity within other tissues after I/R (13). Furthermore, the use of an anti-C5 antibody (which prevents formation of C5a and the membrane attack complex) has been shown to prevent the development of apoptosis after renal (14) and cardiac (13) I/R. Thus, there is reason to suspect that complement activation in the kidney after I/R mediates the development of apoptosis through the caspases and that this is an important mechanism of complement-mediated injury to the kidney.We recently developed an inhibitory mAb ...

Therapeutic Targeting of Classical and Lectin Pathways of Complement Protects from Ischemia-Reperfusion-Induced Renal Damage

“…4 In agreement with these observations, the use of complement inhibitors was effective in limiting ischemia-reperfusion damage in normal animals. 9,10 These data strongly suggest that the inhibition of complement may represent a potential target for preventing tissue damage. 6,11 Interestingly, data available so far indicate that, in contrast to ischemia-reperfusion in the intestine, heart, and lung, complement activation due to ischemia-reperfusion in kidneys occurs almost exclusively through the alternative pathway.…”

“…8 Moreover, the use of anti-factor B or C5a-receptor antagonists has been shown to reduce renal damage due to ischemia-reperfusion. 9,10 Therefore, prevention of complement activation is currently considered one of the best therapeutic targets to prevent or limit ischemia-reperfusionϪinduced renal damage. 5,6,11 C1-inhibitor (C1INH) is a potent inhibitor of proteases of the classical and lectin complement pathways (C1r, C1s, and MASP2).…”

C3a Is Required for the Production of CXC Chemokines by Tubular Epithelial Cells after Renal Ishemia/Reperfusion

“…To accommodate for variation in the response to serum batches, direct comparisons are only made between cells treated side by side with the same serum batch. We have previously demonstrated that under these conditions the cells do not undergo a significant degree of spontaneous complement-mediated lysis (21), and pilot experiments demonstrated that when exposed to this concentration of serum, the transcription of the target genes increased after 1 h. To explore the pathways by which serum elicits production of the chemokines by these cells, the alternative pathway was blocked in the serum using the highly inhibitory mouse anti-mouse factor B mAb 1379 (0.5 g/l) (27) in medium containing 10% serum. To block specific complement activation fragments, the cells were treated with a small molecule (5 or 50 M of SB290157; Calbiochem).…”

“…For example, hypoxic PTECs synthesize C3 and express decreased complement inhibitory proteins on their basal surface (20). Once activated, the complement system generates the C3a and C5a anaphylatoxins, and systemic levels of C3a are measurably increased in mice after renal I/R (21). Complement activation fragments may induce cells to produce a number of cytokines and chemokines (6,22,23).…”

Mammalian DNA Is an Endogenous Danger Signal That Stimulates Local Synthesis and Release of Complement Factor B

“…It has been previously demonstrated that factor B is an important mediator of tissue damage in the setting of a number of autoimmune disorders, because mice deficient in factor B are protected from inflammation and tissue destruction in several animal models of autoimmunity (14)(15)(16)(17)(18)(19). Previous work has also shown that factor B plays an important role in the pathogenesis of I/R injury (20,21). Factor B-deficient mice developed less C3 deposition, neutrophil infiltration and functional impairment in comparison to wild-type (WT) counterparts in a renal I/R model (20).…”