Biomimetic Randall’s plaque as an in vitro model system for studying the role of acidic biopolymers in idiopathic stone formation

Abstract

Randall’s plaque (RP) deposits seem to be consistent among the most common type of kidney stone formers, idiopathic calcium oxalate stone formers. This group forms calcium oxalate renal stones without any systemic symptoms, which contributes to the difficulty of understanding and treating this painful and recurring disease. Thus, the development of an in vitro model system to study idiopathic nephrolithiasis, beginning with RP pathogenesis, can help in identifying how plaques, and subsequently stones, form. One main theory of RP formation is that calcium phosphate deposits initially form in the basement membrane of the thin loops of Henle, which then fuse and spread into the interstitial tissue, and ultimately make their way across the urothelium, where upon exposure to the urine, the mineralized tissue serves as a nidus for overgrowth with calcium oxalate into a stone. Our group has found that many of the unusual morphologies found in RP and stones, such as concentrically-laminated spherulites and mineralized collagenous tissue, can be reproduced in vitro using a polymer-induced liquid-precursor (PILP) process, in which acidic polypeptides induce a liquid-phase amorphous precursor to the mineral, yielding non-equilibrium crystal morphologies. Given that there are many acidic proteins and polysaccharides present in the renal tissue and urine, we have put for the hypothesis that the PILP system may be involved in urolithiasis. Therefore, our goal is to develop an in vitro model system of these two stages of composite stone formation in order to study the role that various acidic macromolecules may play. In our initial experiments presented here, the development of “biomimetic” RP was investigated, which will then serve as a nidus for calcium oxalate overgrowth studies. In order to mimic the tissue environment, MatriStem® (ACell, Inc.), a decellularized porcine urinary bladder matrix was used, because it has both an intact epithelial basement membrane surface and a tunica propria layer, thus providing the two types of matrix constituents found associated with mineral in the early stages of RP formation. We found that when using the PILP process to mineralize this tissue matrix, the two sides led to dramatically different mineral textures, and they bore a striking resemblance to native RP, which was not seen in the tissue mineralized via the classical crystal nucleation and growth process. The interstitium side predominantly consisted of collagen associated mineral, while the luminal side had much less mineral, which appeared to be tiny spherules embedded within the basement membrane. Although these studies are only preliminary, they support our hypothesis that kidney stones may involve non-classical crystallization pathways induced by the large variety of macromolecular species in the urinary environment. We believe that mineralization of native tissue scaffolds is useful for developing a model system of stone formation, with the ultimate goal of developing strategies to avoid RP and its detrimental consequences i...

Liquid-Liquid Phase Separation in Nucleation Process of Biomineralization

“…They added poly-L-aspartic acid to the CP solution, and observed multi-laminated CP spherules formed through the PILP pathway by SEM. These spherules were striking resemble kidney stones, which give some clues that PILP could be a necessary process of kidney stone formation (Chidambaram et al, 2015;Lovett et al, 2019;O'Kell et al, 2019).…”

The potential effects of dietary food and beverage intakes on the risk of kidney stone formation

“…Nephrolithiasis is a highly recurring disease and prevention of stone recurrence during treatment process is as important as the treatment itself [25,26]. The study showed that 79.3% of male patients and 80.0% of female patients have kidney stone formation recurrence ( Table 2).…”

Structural Biology of Calcium Phosphate Nanoclusters Sequestered by Phosphoproteins

“…The polymer used in the PILP process, referred to as the process directing agent, was usually polyaspartate but similar anionic polyelectrolytes were also used [242,243]. Model systems were devised for biomimetic mineralization of collagen sponges [241,244,245] and models of bone [42,241,242,246], tendon [247], dentin [248] and soft tissues [249][250][251][252].…”

“…Biomimetic studies of collagen mineralization were also been made with OPN [41,159,[250][251][252][253][254] and interpreted as due to the PILP mechanism. A comparative study showed that collagen mineralization using solutions containing OPN was more homogeneous, rapid and reproducible than with polyaspartate [159].…”