Schepers Marieke S J, van der Boom Burt G, Romijn Johannes C, Schröder Fritz H, Verkoelen Carl F
Department of Urology, Erasmus University Rotterdam, Rotterdam, The Netherlands.
J Urol. 2002 Apr;167(4):1844-7.
Renal stone formation requires the persistent retention of crystals in the kidney. Calcium oxalate monohydrate (COM) crystal binding to Madin Darby canine kidney strain I (MDCK-I), a cell line that resembles the epithelium in the renal distal tubule/collecting duct, is developmentally regulated, while LLC-PK1 cells (American Type Tissue Collection), which are widely used as a model of the renal proximal tubule, bind crystals irrespective of their stage of epithelial development. Whereas to our knowledge the binding molecules for COM at the surface of LLC-PK1 cells are still unknown, crystals adhere to the hyaluronan (HA) rich pericellular matrix transiently expressed by mobile MDCK-I cells. In the current study we investigated whether crystal binding to either cell type is influenced by urinary substances, including glycoprotein inhibitors of crystallization
We studied crystal binding to MDCK-I cells during wound repair, to confluent LLC-PK1 cells and to HA immobilized on a solid surface using [14C] COM pretreated or not pretreated with urine from healthy male volunteers. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis were performed to assess whether the crystals became coated with urine derived proteins
Western blot analysis demonstrated that pretreated COM crystals were covered with protein inhibitors of crystallization. However, this protein coat had no significant effect on the level of crystal binding to either cell type. In contrast, the adherence of urine treated crystals to immobilized HA was significantly reduced
The adherence of crystals to pericellular matrixes may encompass more than their simple fixation to the polysaccharide HA. Calcium oxalate crystal retention is not prevented by coating crystals with urinary constituents such as glycoproteins and, therefore, may predominantly depend on the surface properties of the renal tubular epithelium.
肾结石的形成需要晶体在肾脏中持续留存。一水草酸钙（COM）晶体与马-达二氏犬肾I型（MDCK-I）细胞结合，该细胞系类似于肾远端小管/集合管中的上皮细胞，其结合过程受发育调控，而广泛用作肾近端小管模型的LLC-PK1细胞（美国模式培养物集存库），无论其上皮发育阶段如何，均可结合晶体。据我们所知，LLC-PK1细胞表面COM的结合分子尚不清楚，晶体可黏附于移动的MDCK-I细胞瞬时表达的富含透明质酸（HA）的细胞周基质。在本研究中，我们调查了晶体与这两种细胞类型的结合是否受尿液物质影响，包括结晶的糖蛋白抑制剂。
我们研究了在伤口修复过程中晶体与MDCK-I细胞的结合、与汇合的LLC-PK1细胞的结合以及与固定在固体表面的HA的结合，使用经健康男性志愿者尿液预处理或未预处理的[14C] COM。进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和蛋白质印迹分析，以评估晶体是否被尿液衍生的蛋白质包被。
蛋白质印迹分析表明，预处理的COM晶体被结晶的蛋白质抑制剂覆盖。然而，这种蛋白质包被对晶体与两种细胞类型的结合水平均无显著影响。相反，经尿液处理的晶体与固定化HA的黏附显著降低。
晶体与细胞周基质的黏附可能不仅仅是其简单地固定于多糖HA。用糖蛋白等尿液成分包被晶体并不能阻止草酸钙晶体的留存，因此，晶体留存可能主要取决于肾小管上皮的表面特性。
