Cisplatin-induced Apoptosis in Auditory, Renal, and Neuronal Cells is Associated with Nitration and Downregulation of LMO4

Cytotoxic effects of cisplatin occur primarily through apoptosis. Though several pro- and anti-apoptotic signaling molecules have been identified to play an important role in mediating the ototoxic, nephrotoxic, and neurotoxic side-effects of cisplatin, the underlying mechanism is yet to be fully characterized. We reported that nitration of LIM domain only 4 (LMO4), a transcriptional regulator, facilitates cochlear apoptosis in cisplatin-induced ototoxicity. However, its role in cisplatin-mediated nephrotoxicity and neurotoxicity is poorly understood. Therefore, HK2, and SH-SY5Y cells were employed along with UBOC1 cells, to investigate the perturbations of LMO4 in cisplatin-induced cytotoxicity, in renal, neuronal, and auditory cells, respectively. Cisplatin induced an increase in the expression of active caspase-3, indicating cellular apoptosis, and increased the nitration of proteins, 24 h post-treatment. Immunostaining with anti-nitrotyrosine and anti-LMO4 indicated that nitrotyrosine co-localized with LMO4 protein in cisplatin treated cells. Immunoblotting with anti-LMO4 indicated that cisplatin induced a decrease in LMO4 protein levels. However, a corresponding decrease in LMO4 gene levels was not observed. Inhibition of protein nitration with SRI110, a peroxynitrite decomposition catalyst, attenuated cisplatin-induced downregulation of LMO4. More importantly, overexpression of LMO4 mitigated the cytotoxic effects of cisplatin in UBOC1 cells while a dose-dependent decrease in LMO4 protein strongly correlated with cell viability in UBOC1, HK2, and SH-SY5Y cells. Collectively, these findings suggested a potential role of LMO4 in facilitating the cytotoxic effects of cisplatin in auditory, renal, and neuronal cells.