The protein osteopontin (OPN) plays an important role in preventing the formation of calcium oxalate monohydrate (COM) kidney stones. To gain insight into these mechanisms, crystallization was induced by addition of human kidney OPN to artificial urine (ionic strength comparable to urine; without citrate), and the OPN-COM interaction studied using a combination of scanning electron (SEM) and confocal microscopy. By SEM, we found that increasing OPN concentrations formed large monoclinic penetration twins (no protein added) and, at higher concentrations (1-, 2μg/ml OPN), super and hyper twins with crystal habits not found in previous studies. For instance, the hyper twins indicate well-facetted gearwheel-like habits with "teeth" developed in all crystallographic <h0l> directions. At OPN concentrations ≥2μg/ml, a switching to small dumbbell-shaped COM habits with fine-textured surfaces occurred. Confocal microscopy of these dumbbells indicates protein incorporation in almost the entire crystal structure (in contrast to facetted COM), proposing a threshold concentration of ∼2μg/ml OPN for the facetted to the non-facetted habit transformation. Both the gearwheel-like and the dumbbell-shaped habit are again found side-by-side (presumably triggered by OPN concentration gradients within the sample) in in-vitro formed conglomerates, which resemble cross-sections of papillary kidney stones. The abrupt transformation from facetted to non-facetted habits and the unique compliance of the two in-vitro formed habits with the two main morphologies found in papillary kidney stones propose that OPN is a main effector in direct stone-forming processes. Moreover, stone structures which exhibit these two morphologies side-by-side might serve as a novel indicator for OPN concentrations surrounding those structures.
Keywords: Artificial urine; Calcium oxalate; Crystal morphology switching; Osteopontin; Papillary kidney stones; Physicochemical conditions; Protein incorporation.
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