Alpha-1-microglobulin: inhibitory effect on calcium oxalate crystallization in vitro and decreased urinary concentration in calcium oxalate stone formers

In the past few years, alpha-1-microglobulin (alpha 1m) has been copurified from human urine with bikunin, a potent inhibitor of calcium oxalate (CaOx ) crystallization in vitro. In this study, we have purified alpha 1m withou t bikunin contamination and investigated its possible role in CaOx crystall ization by in vitro and in vivo studies. Alpha-1m was purified with an anti -aim antibodies CNBr-activated sepharose column. Two molecular species of a im of respectively 30 and 60 kDa were purified. For each protein, two blots of 30 and 60 kDa cross-reacted with anti-alpha 1m antibodies, suggesting t hat these two forms were derived one from the other, Both protein species i nhibited CaOx crystallization in a dose-dependent manner in two in vitro te sts. In the first test, the presence of alpha 1m of 30 kDa (8 mu g/ml) in a medium containing 0.76 mM CaCl2 (with Ca-45) and 0.76 mM OX(NH4)(2) inhibi ted CaOx crystallization by 38% as estimated by supernatant radioactivity a fter 1 h of agitation. In the second test, CaOx kinetics were examined for 3 to 10 min in a turbidimetric model at 620 nm. The presence of slim of 30 kDa in a medium containing 4 mM CaCl2 and 0.5 mM Na2Ox inhibited CaOx cryst allization by 41.5%, as estimated by the slope modification of turbidimetri c curve. Alpha-lm can be considered as another inhibitor of urinary CaOx cr ystal formation, as shown by the present in vitro studies. Using an ELISA a ssay, we found that urinary alpha 1m concentration was significantly lower in 31 CaOx stone formers than in 18 healthy subjects (2.95 +/- 0.29 vs 5.34 +/- 1.08 mg/l respectively, P = 0.01). The decreased concentration of alph a 1m in CaOx stone formers could be responsible in these patients, at least in part, for an increased risk of CaOx crystalluria.