Nephrolithiasis requires formation of crystals followed by their retention and accumulation in the kidney. Crystal retention can be caused by the association of crystals with the epithelial cells lining the renal tubules. The present study investigated the interaction between calcium oxalate monohydrate (COM) crystals and primary cultures of human proximal (PTC) and distal tubular/collecting duct cells (DTC). Both PTC and DTC were susceptible to crystal binding during the first days post-seeding (4.9 +/- 0.8 micro g COM/cm2), but DTC lost this affinity when the cultures developed into confluent monolayers with functional tight junctions (0.05 +/- 0.02 micro g COM/cm2). Confocal microscopy demonstrated the expression of the transmembrane receptor protein CD44 and its ligands osteopontin (OPN) and hyaluronic acid (HA) at the apical membrane of proliferating tubular cells; at confluence, CD44 was expressed at the basolateral membrane and OPN and HA were no longer detectable. In addition, a particle exclusion technique revealed that proliferating cells were surrounded by HA-rich pericellular matrices or "cell coats" extending several microns from the cell surface. Disintegration of these coats with hyaluronidase significantly decreased the cell surface affinity for crystals. Furthermore, CD44, OPN, and HA were also expressed in vivo at the luminal side of tubular cells in damaged kidneys. These results suggest (1) that the intact distal tubular epithelium of the human kidney does not bind crystals, and (2) that crystal retention in the human kidney may depend on the expression of CD44-, OPN-, and-HA rich cell coats by damaged distal tubular epithelium.

Screening for and treating asymptomatic bacteriuria are common in KTRs despite uncertainties around the benefits and harms. In an era of antimicrobial resistance, further studies are needed to address the diagnosis and management of asymptomatic bacteriuria in these patients.

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Erythropoietin (EPO) has been shown to have tissue protective properties by binding to its receptor (EPOR) which is also expressed on non-haematopoietic cells. The mechanisms underlying this protection have not yet been elucidated and the renal cell types mediating these effects remain ill-defined. This study aimed to identify the EPOR expression in human tubular epithelial cells (hTECs) and in rat kidney and to investigate the role of EPOR in EPO-mediated renoprotection. Male Wistar rats were treated with saline or EPO (3000 U/kg, i.p.) 24h prior to sham-operation or 30 min bilateral renal ischemia. Renal morphology and function, tubular regeneration, apoptosis and expression of EPOR, hemeoxygenase-1 (HO-1) and hepatocyte growth factor (HGF) were analyzed. Primary cultures of human proximal (PTC) and distal/collecting duct (DTC) tubular cells were incubated with EPO (5-50-500 ng/mL) either or not in the presence of soluble EPOR. Total RNA was extracted and mRNA expression of HO-1 was investigated by quantitative RT-PCR. EPOR mRNA could be demonstrated in hTECs and in cortical tubules of the rat kidney. Furthermore, EPOR protein was expressed at the membrane and as intracellular vesicles in hTECs. In vivo, EPO treatment attenuated histological and functional renal damage, decreased both cell necrosis and apoptotic cell death, enhanced tubular regeneration and resulted in an upregulation of HO-1 and HGF mRNA. In vitro, EPO administration resulted in an early upregulation of HO-1 mRNA which was restricted to PTC and inhibited by simultaneous addition of supra-equivalent amounts of soluble EPOR. These data strongly suggest that the EPO-mediated renoprotection results from direct interaction of EPO with EPOR on tubular cells.

ABSTRACT. In normal human and rat kidneys, osteopontin (OPN) is present at the apical surface of cells in the distal nephron. After ischemic or toxic renal damage in rats, OPN is upregulated in distal tubular cells (DTC) and expressedde novoin perinuclear vesicles in proximal tubular cells (PTC). In the first phase of this study, OPN localization in ischemic human biopsies was compared with that in ischemic rat kidneys. In the second phase, cultures of PTC and DTC were used to investigate human renal OPN synthesis, secretion, and localization. OPN localization in human biopsies after renal ischemia was comparable to that in ischemic rat kidneys. Microscopic and flow cytometric detection of immunofluorescent OPN staining in tubular cell cultures demonstrated strong plasma membrane localization in DTC, whereas mainly perinuclear intracellular expression was observed in PTC. Northern blotting and reverse transcription-PCR demonstrated production of a single OPN mRNA in PTC and DTC. Detection of OPN by Western blotting and enzyme-linked immunosorbent assay demonstrated that PTC and DTC synthesized and secreted the same three molecular mass OPN forms, in comparable amounts. Finally, confocal microscopy demonstrated different staining patterns for endocytotic/lysosomal vesicles and perinuclear OPN; however, perinuclear OPN exhibited colocalization with the Golgi apparatus. In conclusion, human renal OPN localization in cell cultures demonstrated differences between PTC and DTC comparable to those observed after renal ischemiain vivo. Therefore, these cell cultures represented an excellent model for the study of human OPN synthesis, secretion, and localization in PTCversusDTC. It is reported for the first time that intracellular OPN is located in the Golgi apparatus of both PTC and DTC and that PTC and DTC are able to produce and secrete the same OPN isoforms, in comparable amounts.