Role of protein kinase C and oxidative stress in interleukin-1beta-induced human proximal tubule cell injury and fibrogenesis.
BACKGROUND
Interleukin (IL)-1beta, a pro-inflammatory macrophage-derived cytokine, is implicated as a key mediator of interstitial fibrosis and tubular loss or injury in progressive renal insufficiency. This study investigates some of the mechanisms of action of IL-1beta on the proximal tubule.


METHODS
Confluent cultures of primary human proximal tubule cells (PTC) were incubated in serum-free media supplemented with either IL-1beta (0-4 ng/mL), phorbol-12-myristate 13-acetate (PMA, protein kinase C activator) (6.25-100 nmol/L), or vehicle (control), together with a non-specific protein kinase C inhibitor (H7), a specific protein kinase C inhibitor (BIM-1), an anti-oxidant (NAC) or a NADPH oxidase inhibitor (AEBSF).


RESULTS
Interleukin-1beta-treated PTC exhibited time-dependent increases in fibronectin secretion (ELISA), cell injury (LDH release) and reactive nitrogen species (RNS) release (Griess assay). Proximal tubule cell DNA synthesis (thymidine incorporation) was also significantly suppressed. The effects of IL-1beta, which were reproduced by incubation of PTC with PMA (6.25-100 nmol/L), were blocked by H7 but not by BIM-1. The anti-oxidant (4 mmol/L) partially blocked IL-1beta-induced fibronectin secretion by PTC, but did not affect IL-1beta-induced LDH release, RNS release or growth inhibition. The NADPH oxidase inhibitor (AEBSF) significantly attenuated all observed deleterious effects of IL-1beta on PTC.


CONCLUSIONS
Interleukin-1beta directly induces proximal tubule injury, extracellular matrix production and impaired growth. The anti-oxidant, NAC, appears to ameliorate part of the fibrogenic effect of IL-1beta on PTC through mechanisms that do not significantly involve protein kinase C activation or nitric oxide release.
Vesey
David A
DA
Department of Renal Medicine, University of Queensland at the Princess Alexandra Hospital, Brisban, Queensland, Australia.
Cheung
Catherine
C
Endre
Zoltan
Z
Gobé
Glenda
G
Johnson
David W
DW
D000328:Adult; D000975:Antioxidants / Q000494:pharmacology; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005109:Extracellular Matrix / Q000378:metabolism; D005260:Female; D005353:Fibronectins / Q000378:metabolism; D005355:Fibrosis; D006801:Humans; D066298:In Vitro Techniques; D007375:Interleukin-1 / Q000494:pharmacology*; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology*; D008297:Male; D008875:Middle Aged; D018384:Oxidative Stress / Q000502:physiology*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D026361:Reactive Nitrogen Species / Q000378:metabolism
D000975:Antioxidants; D004791:Enzyme Inhibitors; D005353:Fibronectins; D007375:Interleukin-1; D026361:Reactive Nitrogen Species; D011493:Protein Kinase C