Oxalate inhibits renal proximal tubule cell proliferation via oxidative stress, p38 MAPK/JNK, and cPLA₂ signaling pathways

Abstract

Exposure of renal proximal tubule cells to oxalate may play an important role in cell proliferation, but the signaling pathways involved in this effect have not been elucidated. Thus the present study was performed to examine the effect of oxalate on (3)H-labeled thymidine incorporation and its related signal pathway in primary cultured rabbit renal proximal tubule cells (PTCs). The effects of oxalate on [(3)H]thymidine incorporation, lactate dehydrogenase (LDH) release, Trypan blue exclusion, H(2)O(2) release, activation of mitogen-activated protein kinases (MAPKs), and (3)H-labeled arachidonic acid (AA) release were examined in primary cultured renal PTCs. Oxalate inhibited [(3)H]thymidine incorporation in a time- and dose-dependent manner. However, its analogs did not affect [(3)H]thymidine incorporation. Oxalate (1 mM) significantly increased H(2)O(2) release, which was blocked by N-acetyl-l-cysteine (NAC) and catalase (antioxidants). Oxalate significantly increased p38 MAPK and stress-activated protein kinase (SAPK)/c-Jun NH(2)-terminal kinase (JNK) activity, not p44/42 MAPK. Oxalate stimulated [(3)H]AA release and translocation of cytosolic phospholipase A(2) (cPLA(2)) from the cytosolic fraction to the membrane fraction. Indeed, oxalate significantly increased prostaglandin E(2) (PGE(2)) production compared with control. Oxalate-induced inhibition of [(3)H]thymidine incorporation and increase of [(3)H]AA release were prevented by antioxidants (NAC), a p38 MAPK inhibitor (SB-203580), a SAPK/JNK inhibitor (SP-600125), or PLA(2) inhibitors [mepacrine and arachidonyl trifluoromethyl ketone (AACOCF(3))], but not by a p44/42 MAPK inhibitor (PD-98059). These findings suggest that oxalate inhibits renal PTC proliferation via oxidative stress, p38 MAPK/JNK, and cPLA(2) signaling pathways.

Role of ATP in DNA synthesis of renal proximal tubule cells: involvement of calcium, MAPKs, and CDKs

“…Our recent report that oxalate and high glucose inhibited cell proliferation in primary cultured PTCs is consistent with the results obtained from intact renal tissue (21,42). Therefore, PTCs in hormonally defined, serum-free culture conditions would be a powerful tool for studying the effect of ATP on cell proliferation.…”

Extracellular adenosine triphosphate protects oxidative stress‐induced increase of p21WAF1/Cip1 and p27Kip1 expression in primary cultured renal proximal tubule cells: Role of PI3K and Akt signaling

“…The PTC culture system used in this study retains the differentiated phenotype in vitro that is typical of renal proximal tubules, including a polarized morphology and distinctive proximal tubule transport and hormone response (Chung et al, 1982;Han et al, 2000;Park et al, 2001). Recently, we reported that oxalate inhibited cell proliferation in PTCs, which is consistent with the results obtained using intact renal tissue (Han et al, 2004). The results of studies using this culture system in the PTCs are directly comparable with results obtained using the original renal tissue Han, 2005, 2006).…”

Mitochondrial H2O2 generated from electron transport chain complex I stimulates muscle differentiation

“…The levels of mitochondrial H 2 O 2 were determined using a modified method that monitors the absorbance of resorufin, a reaction product of Amplex Red [43,44]. A total of 100 µl of reaction buffer (120 mM KCl, 3 mM HEPES free acid, 1 mM EGTA, and 0.3% BSA, pH 7.2, at 37 °C), including 50 µM Amplex Red (Molecular Probe, Invitrogen), 6 U/ml HRP, and 30 U/ml SOD, was added to each microplate well and then pre-warmed at 37 °C for 10 min.…”