Urinary stones are a common medical condition affecting over 10% of adults in the developed world. Whilst many different materials are found in kidney stones, the majority are dominated by deposits of calcium oxalate (CaOx). In addition to the mineral phases, urinary calculi also contain macromolecules. It has been shown that many of these, particularly the proteins, can inhibit CaOx crystallisation and are therefore widely assumed to be relevant to stone pathogenesis. More recently, it has been proposed that certain urinary proteins facilitate the prevention of stone formation through their selective inclusion into the growing crystal and subsequent structural disruption through protease activities. Given that the intracrystalline location of proteins would be structurally challenging, their selective occlusion may support a functional role. This thesis explores these concepts. The crystallisation of CaOx in the urine of stone formers and healthy controls was compared. CaOx crystals grown in human urine and inorganic media were examined using a range of complementary techniques, e.g., electron microscopy, X-ray diffraction and thermogravimatric analysis. Proteins associated with the surface of different mineral phases and/or occluded within the crystal structures were isolated and identified using immunoblotting. The recovered proteins were quantified in terms of their crystal volume occupation, and the proportion of proteins taken up from the urine were investigated. Finally, six common methods used to study CaOx/protein interactions in vitro were compared. Significant differences were noted in the crystallisation parameters which define the urine samples from stone-formers and controls. These could not be attributed to electrolyte balance from which it is inferred that the influence of other urinary constituents such as proteins must be significant. Only a small proportion of the urinary proteins associated with the CaOx crystals and they contributed to less than 5% of the total crystal volume, suggesting that arguments for a preventative role in stone formation cannot be supported. Kinetic experiments demonstrated that two proteins, often proposed as inhibitors of stone formation had little effect on CaOx nucleation and growth rates. Different proteins were found to be surface-associated when compared to those that were occluded within the crystal structures. However, the same proteins all associated with both CaO monohydrate and dehydrate, which suggests unique template-mediated interactions are unlikely. The growth of CaOx in urine using six methods showed that the stabilisation of different hydromorphs and induction of some morphological changes in CaOx were protocol dependant and not, as previously suggested, attributable to protein interactions. The method used did not influence the type of protein associating with CaOx. These results indicate that whilst urinary proteins may be capable of modifying CaOx crystal growth they appear to do so through non-specific interactions. There is little evidence that proteins are present in urine as teleological inhibitors of stone formation.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:606747 |
| Date | January 2003 |
| Creators | Walton, Rachel C. |
| Publisher | University of Manchester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://www.manchester.ac.uk/escholar/uk-ac-man-scw:75546 |
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