Charge-dependent binding of polymeric IgA1 to human mesangial cells in IgA nephropathy

Abstract

Background: IgA nephropathy (IgAN) is characterized by raised serum IgA1 and predominant mesangial IgA1 deposits of polymeric nature. The mechanism of polymeric IgA1 (pIgA1) deposition in the kidney mesangium is poorly understood in IgAN. It has been suggested that increased sialic acid content and anionic charge of the pIgA1 molecules may be operational in the IgA1 deposition in human mesangial cells (HMCs). The present study examined the binding of pIgA1 with different surface charges to HMCs. The binding characteristics of IgA1 to HMCs in the presence of polycation (poly-L-lysine) or polyanion (heparin) were also investigated.

Methods: IgA1 was purified in sera from patients with IgAN and from healthy controls by jacalin affinity chromatography. IgA1 was further separated into pIgA1 and monomeric IgA1 (mIgA1) by fast protein liquid chromatography (FPLC). pIgA1 or mIgA1 with different net charges on their surface were resolved by ion exchange chromatography (IEC) with a Mono Q column. The binding characteristics of pIgA1 and mIgA1 to HMCs in the presence or absence of polycation or polyanion were examined by flow cytometry.

Results: In patients with IgAN, the absolute amount of mIgA1 and pIgA1 is significantly higher than that of healthy controls (P < 0. 001). There was significant increase in binding of pIgA1 from patients with IgAN to HMC and cell lysate. pIgA1 that interacted strongly with the ion exchanger also bound more to HMCs when compared with IgA1 interacted weakly with the ion exchanger (P < 0. 001). The anionic charged pIgA1 from patients was significantly higher than that of healthy controls (P < 0.001). Preincubation with poly-L-lysine increased the binding of pIgA1 to HMCs. The binding of pIgA1 to HMCs was decreased by preincubation with heparin.

Conclusions: The binding of IgA to HMCs is charge dependent. Polymeric IgA with the highest net negative charge binds more to HMCs. Preincubation with polyanion decreased the binding of polymeric IgA to HMCs. These results suggest an important role for anionic charge in IgA1 deposition onto the kidney mesangial cells.