Relationship between 12/15-lipoxygenase and COX-2 in mesangial cells: potential role in diabetic nephropathy.
The 12/15-lipoxygenase (12/15-LO) and cyclooxygenase-2 (COX-2) pathways of arachidonate metabolism have been implicated in the pathogenesis of diabetic nephropathy (DN). In this study, we evaluated whether there is an interplay between 12/15-LO and COX-2 pathways in mesangial cells (MC). We utilized MC, microdissected glomeruli and renal cortical tissues. Transfections with cDNAs or short hairpin RNAs (shRNAs) were performed to overexpress or knockdown 12/15-LO and COX-2, respectively. Reverse transcription-polymerase chain reactions and Western blotting were used for evaluating mRNA and protein expression, respectively. We observed that the expression of both 12/15-LO and COX-2 were increased in high glucose stimulated rat MC relative to normal glucose, and also in cortical tissues from diabetic db/db and streptozotocin-injected mice relative to corresponding control mice. Treatment of rat MC with the 12/15-LO product, 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE), significantly increased COX-2 expression as well as levels of the COX-2 product, prostaglandin E(2) (PGE(2)). Interestingly, treatment of rat MC with PGE(2) led to a reciprocal increase in 12/15-LO expression as well as levels of 12(S)-HETE. The 12/15-LO shRNA could significantly attenuate COX-2 protein expression and vice versa. Furthermore, COX-2 expression levels were lower in MC and glomeruli from 12/15-LO knockout mice relative to control. Conversely, mouse MC stably overexpressing 12/15-LO had greater levels of COX-2 expression relative to mock-transfected cells. These new results indicate for the first time that 12/15-LO and COX-2 pathways can cross-talk and activate each other in MC. These novel interactions may amplify their effects on the progression of DN.
Xu
Z-G
ZG
Gonda Diabetes Research Center, Beckman Research Institute of the City of Hope, Duarte, California, USA.
Li
S-L
SL
Lanting
L
L
Kim
Y-S
YS
Shanmugam
N
N
Reddy
M A
MA
Natarajan
R
R
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid / Q000494:pharmacology; D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D001093:Arachidonate 15-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D051546:Cyclooxygenase 2 / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology / Q000473:pathology / Q000503:physiopathology; D003928:Diabetic Nephropathies / Q000201:enzymology / Q000209:etiology / Q000503:physiopathology*; D015232:Dinoprostone / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D007672:Kidney Cortex / Q000201:enzymology / Q000473:pathology / Q000502:physiology; D008297:Male; D050527:Mesangial Cells / Q000201:enzymology* / Q000473:pathology / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D020929:Mitogen-Activated Protein Kinase Kinases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection
D012333:RNA, Messenger; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D001092:Arachidonate 12-Lipoxygenase; D001093:Arachidonate 15-Lipoxygenase; D051546:Cyclooxygenase 2; C489691:Ptgs2 protein, rat; D020929:Mitogen-Activated Protein Kinase Kinases; D005947:Glucose; D015232:Dinoprostone