| Project/Area Number |
08457430
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Urology
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| Research Institution | Wakayama Medical College |
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Principal Investigator |
OHKAWA Tadashi Wakayama Medical College, Urology, Professor, 医学部, 教授 (90073733)
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| Co-Investigator(Kenkyū-buntansha) |
MORITA Teruo Wakayama Medical College, Urology, Associate, 医学部, 助手 (50291612)
OGURA Hideaki Wakayama Medical College, Urology, Associate, 医学部, 助手 (60264496)
KOHJIMOTO Yasuo Wakayama Medical College, Urology, Associate, 医学部, 助手 (50295820)
MORIMOTO Shigeyoshi Wakayama Medical College, Urology, Assistant Professor, 医学部, 講師 (20094683)
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| Project Period (FY) |
1996 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥8,000,000 (Direct Cost: ¥8,000,000)
Fiscal Year 1998: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1997: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1996: ¥6,100,000 (Direct Cost: ¥6,100,000)
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| Keywords | renal calculus / calcium oxalate crystal / MDCK cell / crystal-cell interaction / crystal adhesion / endocytosis / human urine / urinary macromolecule / ラット結石モデル / ゲンタマイシン / 尿中蓚酸 / 尿中クエン酸 / 走査型電子顕微鏡 |
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| Research Abstract |
We established the quantitative assay of calcium oxalate monohydrate (COM) crystal adhesion to Madin-Darby canine kidney (MDCK) cells. Cultures of MDCK cells were exposed to COM crystal suspension for 5 minutes. After washing them with Hnk's balanced salt solution, the adherent COM crystals on MDCK cells were disolved by 5N hydrochloric acid. Calcium concentration of the solution was measured by atomic absorption analysis to quantify the volume of adherent COM crystals. In morphological examination using light and scanning electron microscopy, COM crystals adhered to the intact surface of MDCK cells, and some crystals were endocytosed into the cells.
However, the cell uptake of COM crystals were significantly reduced by pre-treatment with glcosaminoglycans (sodium pentsan polysuiphate, heparin, and chondroitin sulphate) or surfactant (Triton X 100). The adhesion to MDCK cells was, furthermore, specific to COM crystals as compared with other types of crystals.
Histological study of the ki
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dney in rat model of nephrolithiasis, induced by intraperitoneal injection of sodium oxalate and high calcium diet, also showed that most COM crystals adhered to the tubular cells and some of them were engalfed, via endocytosis.
Theses findings suggest that the crystal-cell interactions might be one of the earliest processes in the formation of kidney stones, and some substances in urine might inhibit this interaction physiologically.
The effects of human urine on the adhesion of COM crystals to cultures of MDCK cells were studied further. It was shown that human urine had a strong inhibitory effect on the adhesion of crystal, which was considered to be mediated by acting on the crystal surface. From human urine, we isolated a macromolecular fraction (MW 60,000) that had strong capacity to inhibit cellular adhesion of COM crystals hrough ion exchange and gel filtration chromatography, although we have not yet identified this macromolecular substance. Comparison of the inhibitory activities of the urine from stone formers and healthy controls revealed that there was no significant difference between them, wheras some urine samples from stone formers demonstrated extreme low inhibitory activities. Less
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