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Pathological Biomineralization of Calcium Oxalate Kidney Stones

Jeffrey D. Rimer1, Jeffrey A. Wesson2, and Michael D. Ward1. (1) Molecular Design Institute, Department of Chemistry, New York University, 100 Washington Square East, New York, NY 10003, (2) Nephrology Division, Department of Veterans Affairs Medical Center and the Medical College of Wisconsin, 5000 West National Avenue, Milwaukee, WI 53295

The design of new technologies and pharmaceuticals for therapeutic drug delivery requires a fundamental understanding of the pathogenesis of physiological diseases; however, control and prevention of many common diseases, such as kidney stones, which affect 15 % of the U.S. population, are currently hindered by a knowledge gap pertaining to critical steps responsible for stone formation in vivo. Kidney stones derive predominantly from supersaturated calcium oxalate (CaOx), which crystallizes in renal tubules to produce two polymorphs, calcium oxalate monohydrate (COM) and dihydrate (COD). Adverse effects of the disease have been correlated to COM, which forms layered polycrystalline aggregates through a series of events involving nucleation, growth, aggregation, and crystal and/or aggregate attachment to epithelial cells. Our group 1 and others 2 have investigated interfacial aspects of these processes using atomic force microscopy (AFM), probing molecular-level events on the surfaces of kidney stone crystals (e.g. adhesion forces and growth kinetics) in an effort to identify the underlying mechanism(s) of their formation.

Lipids and proteins on cellular membranes can mediate crystal-crystal contact, while a multitude of urinary proteins, polysaccharides, lipids, and cellular debris adsorb to crystal surfaces, likely acting as adhesive glue among aggregates. Previous studies suggested that proteins with substantial anionic moieties serve as COM adhesives 1e; however, a variety of urinary proteins have immerged as potential inhibitors of crystal nucleation, growth, and/or aggregation most notably transferrin, human serum albumin, Tamm-Horsfall protein (THp), and osteopontin (OPN). Research efforts in our group have focused on (i) in vitro investigations of crystal growth to examine face-specific inhibitory effects in the presence of protein and macromolecular additives 1a,c, and (ii) adhesion force measurements using AFM probes modified with biologically relevant functional groups to study surface-adsorbate interactions involved in crystal adhesion and aggregation 3a. We will present recent investigations that examine interactions between the most bioactive surface, COM (100), and functional groups that mimic L-arginine and L-glutamic acid residues. Experiments were performed with additives believed to influence crystal growth and aggregation, focusing on THp and OPN. A systematic study of THp was performed to identify the affect of sialylated and glycoslated side chains on the force of adhesion, while dynamic protein unfolding measurements were conducted on OPN a protein that exhibits uncharacteristically strong adhesion to COM surfaces 3b. Lastly, we have expanded the use of single amino-acid functionalized tips to directly measure protein-crystal and protein-protein interactions on COM surfaces using established protocols in the literature for binding proteins to AFM tips 4. These collective results offer new insights on specific interactions that potentially regulate kidney stone formation, serving as a basis for future development of targeted drug delivery for COM stones.

1. (a) Guo et al., Langmuir 2002, 18, 4284, (b) Sheng et al., J. Am. Chem. Soc. 2003, 125, 2854, (c) Jung et al., Langmuir 2004, 20, 8587, (d) Sheng et al., Proc. Nat. Acad. Sci. 2005, 102, 267, (e) Ward and Wesson, Elements 2007, 3, 415.

2. (a) Qiu et al., Proc. Nat. Acad. Sci. 2004, 101, 1811, (b) Qiu et al., J. Am. Chem. Soc. 2005, 127, 9096, (c) Wang et al., Langmuir 2006, 22, 7279.

3. (a) Sheng et al., J. Am. Soc. Nephrol. 2005, 16, 1904, (b) Wesson et al., J. Am. Soc. Nephrol. 2003, 14, 139.

4. (a) Ebner et al., Current Nanoscience 2007, 3, 49, (b) Lee et al., Proc. Nat. Acad. Sci. 2006, 103, 12999, (c) Lee et al., Micron 2007, 38, 446, (d) Strunz et al., Proc. Nat. Acad. Sci. 1999, 96, 11277.