Bactericidal activity of renal tubular cells: the putative role of human beta-defensins.

Renal tubular epithelial cells (RTC) form a barrier between the host and ascending microbes in upper urinary tract infection. Previous studies have shown the ability of the kidney to produce defensins--antimicrobial peptides that play a pivotal role in unspecific host defense. To further clarify the role of renal epithelium for direct antibacterial activity we investigated the expression, regulation and production of antimicrobial peptides by cultured human RTC. Cell culture supernatants of RTC exert strong bactericidal activity against Escherichia coli and Klebsiella pneumoniae, two of the most important pathogens in urinary tract infections. The antimicrobial effect depends on salt concentration, a typical feature of human defensins. RT-PCR of RNA from cultured proximal and distal RTC showed constitutive expression of human beta-defensin 1 (hbd-1) and human beta-defensin 2 (hbd-2) whereas only hbd-1 expression could be detected in RNA preparation from renal biopsy material. Hbd-2 expression of RTC was induced by inflammatory processes as shown by semiquantitative competitive RT-PCR. Coincubation of the cultured cells with IL-1alpha or E. coli promote the strongest hbd-2 induction whereas TNF-alpha and LPS lead to a weaker or no (IL-6) hbd-2 induction. This is the first evidence that human RTC are able to produce antibacterial substances in a biologically relevant amount and that beta-defensins are candidate proteins responsible for this effect.

Tamm-Horsfall protein inhibits binding of S- and P-fimbriated Escherichia coli to human renal tubular epithelial cells.

Escherichia coli is the predominant pathogen in urinary tract infections. Fimbriae are one of the major virulence factors of these bacteria, since these protein appendices contribute towards bacterial adhesion to epithelial cells. In clinical E. coli isolates from urinary tract infections, P fimbriae are more frequently present than S fimbriae. However, these two types of fimbriae mediate adhesion to cultured tubular epithelial cells equally well. Tamm-Horsfall protein, which is the most abundant protein in normal human urine, inhibits hemagglutination by E. coli expressing S fimbriae, but does not interfere with hemagglutination by P-fimbriated E. coli. Therefore, it has been speculated that Tamm-Horsfall protein may serve as a clearance factor for S-fimbriated E. coli in human urine. In our experiments, adherence of purified S fimbriae and of S-fimbriated E. coli to tubular epithelial cells was inhibited by Tamm-Horsfall protein, but the protein also decreased binding of P-fimbriated E. coli to approximately the same degree. We found less adherence of both types of fimbriae to a Madin-Darby canine kidney cell line expressing soluble and membrane-bound Tamm-Horsfall protein as compared with the control cell line. In conclusion, our in vitro data suggest that urinary Tamm-Horsfall protein may serve as a clearance factor for E. coli expressing both S and P fimbriae. In the light of these findings, the low clinical relevance of S-fimbriated E. coli for urinary tract infections may be readily explained; however, the predominance of P fimbriae remains unresolved.

Promotion of Salmonella typhimurium adherence and membrane ruffling in MDCK epithelia by staurosporine.

Infection of Madin-Darby canine kidney epithelial cell monolayers with Salmonella typhimurium SL1344 for 60 min results in widespread bacterial invasion which is associated with remodelling of the apical cell membrane to form "membrane ruffles'. Treatment of Madin-Darby canine kidney cell monolayers with the protein kinase inhibitor staurosporine resulted in a 12-fold increase in the number of adhered bacteria without significantly affecting bacterial invasion. Staurosporine treatment also significantly increased both the number and size of membrane ruffles. As S. typhimurium adhere preferentially to these areas of membrane lacking microvilli, the increased extent of membrane ruffling may explain the increased bacterial adherence. These data provide evidence that the propagation of membrane ruffles during S. typhimurium infection is modulated by changes in the phosphorylation state of host proteins.