The
12-lipoxygenase (12-LO) pathway of
arachidonic acid metabolism is implicated in extracellular matrix (ECM) synthesis, but its role in podocytes has not been studied. This study tested whether 12-LO induction by diabetes or by high
glucose (HG) in cultured podocytes alters glomerular basement membrane by activating signal transduction pathways culminating in ECM synthesis. Sprague-Dawley rats received an injection of diluent (control [C]) or
streptozotocin 65 mg/kg (DM) and were killed at 1 or 4 mo. Glomerular 12-LO
mRNA and
protein levels were higher in DM than in C glomeruli at 1 and 4 mo, and 12-LO localized predominantly in podocytes. Glomerular p38
mRNA and
protein were higher in DM at months 1 and 4, but phospho-p38
mitogen-activated
protein (MAPK) was increased only at month 1. Glomerular
collagen alpha5(IV)/
glutaraldehyde-3-
phosphate dehydrogenase (GAPDH)
mRNA ratio was increased in DM at month 1 but not at month 4, whereas
collagen alpha5(IV)
protein was higher at both 1 and 4 mo. Mouse podocytes were cultured in media with 25 mM
glucose (HG) with or without the 12-LO inhibitor cinnamyl-3,4-dihydroxy-cyanocinnamate (CDC) or with 5.5 mM
glucose + 19.5 mM
mannitol (low
glucose [LG+M]) for 10
d at 37 degrees C. 12-LO
mRNA and
protein levels were higher in HG than in LG+M as was the
p38 MAPK/GAPDH
mRNA ratio. Phospho-p38 MAPK
protein but not total
p38 MAPK was higher in HG compared with LG+M.
Collagen alpha5(IV)/GAPDH
mRNA ratio and
protein were higher in HG than in LG+M. 12-LO inhibition by CDC decreased HG-induced phospho-p38 MAPK and the phospho-p38/total
p38 MAPK ratio,
collagen alpha5(IV)/GAPDH
mRNA ratio, and
collagen alpha5(IV)
protein expression. In summary, diabetes in vivo and exposure of podocytes to HG in vitro stimulated 12-LO,
p38 MAPK, and
collagen alpha5(IV)
mRNA and (activated)
protein. 12-LO inhibition by CDC diminished the expression of podocyte phospho-p38 MAPK and
collagen alpha5(IV)
mRNA and
protein. These findings implicate 12-LO and the
p38 MAPK signaling pathway in the mediation of ECM synthesis by podocytes in diabetes.