Abbreviations & AcronymsObjectives: To analyze the crystal components and matrix proteins of urinary stones by proteomic analysis using liquid chromatography-tandem mass spectrometry. Methods: Urinary stones were obtained from patients with gout and hyperuricemia. The outside and inside of the stones were measured non-destructively with a micro area X-ray diffractometer. After stones were powdered , extracted proteins were analyzed by proteomic analysis. Results: Of 17 investigated stones, seven were composed of calcium oxalate monohydrate or calcium oxalate dihydrate, seven were of uric acid , and three were a mixture of calcium oxalate monohydrate and uric acid. In calcium oxalate monohydrate or calcium oxalate dihydrate stones, osteopontin, uromodulin, albumin, protein Z, prothrombin, protein S, hemoglobin and histone H4 were identified. In uric acid stones, uromodulin, albumin, hemoglobin, calgranulins and immunoglobin G fragments were detected. Mixed stones of calcium oxalate monohydrate and uric acid contained both Ca-binding proteins and abundant proteins. Matrix proteins were different when the crystal components of the stone were different, even when from the same patient. Conclusions: Proteins, such as uromodulin and albumin, are often detected in stones, regardless of crystal components. However, osteopontin, prothrombin, protein S and protein Z are identified specifically in calcium oxalate stones. Furthermore, immunoglobin G fragments are detected in uric acid stones. The role of these specific proteins in the different types of stones can be of particular interest.

In this study, we performed proteomic analysis following sequential protein extraction on calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) urinary stones to determine the specific matrix proteins according to the crystal components of the stones. After X-ray and IR analysis of 13 urinary stones, matrix proteins were sequentially extracted with KCl, formic acid, guanidine-HCl, and EDTA, before SDS-electrophoresis followed by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). The electrophoretic patterns of the extracted proteins differed from that of COM and COD stones. LC-MS/MS identified 65 proteins, of which many were cellular plasma proteins, and were frequently detected regardless of the crystal components. However, 6 proteins (protein Z, protein S, prothrombin, osteopontin, fatty acid binding protein 5, and ubiquitin) were detected in the final EDTA fractions of COM stones. These proteins are involved in the coagulation process or osteometabolism, and thus the roles they play are of particular interest.

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