Comparative Analysis of Caspase Activation and Apoptosis in Renal Tubular Epithelial Cells and Renal Cell Carcinomas

Abstract

Background/Aims: Treatment of renal cell carcinoma (RCC) is limited by its resistance to conventional chemotherapies. This may occur, in part, from resistance to apoptosis. The role of caspase activation in apoptosis resistance in treated RCCs was investigated. Methods: Two human RCC cell lines (ACHN and SN12K1) and renal tubular epithelial cells (HK2) were treated with 5-fluorouracil (0.2–20 µg/ml) or cisplatin (1–100 µM). Activation of caspase-3 and -2 was analysed and compared with levels of apoptosis. Caspase function was analysed using pan-caspase inhibition (z-VAD-fmk) and caspase-2 inhibition (z-VDVAD-fmk). Results: RCC apoptosis was significantly lower (p < 0.05) than in HK2s after treatment, confirming their chemoresistance. Pro-caspase-3 (32 kDa) was detected in all cell lines. Cleaved caspase-3 (19 kDa) was not detected by Western immunoblots in treated RCCs and only minimal activated caspase-3 was detected in treated RCCs using immunohistochemistry. All cells had pro-caspase-2 (48 kDa) and the activated form (33 kDa) appeared in all treated cells. Caspase inhibition caused a reduction in, but not negation of, therapy-induced apoptosis in HK2s and RCCs (p < 0.05 for HK2s and ACHN cells), indicating that a caspase activation pathway must occur in RCC apoptosis but this pathway does not act via caspase-3 cleavage. Inhibition of caspase-2 reduced apoptosis only in HK2s, indicating that the activated caspase-2, identified in treated RCCs, was not responsible for their apoptosis induction. Conclusion: Specific differences in caspase-3 and -2 activation were identified in renal tubular epithelium and RCCs after chemotherapy. Identification of RCC-specific caspase inactivation or redundancy may explain, in part, the resistance of RCCs to cancer therapies and may be useful in targeting apoptotic pathways to overcome RCC resistance to treatment.

Erythropoietin reduces cisplatin-induced apoptosis in renal carcinoma cells via a PKC dependent pathway

“…Lack of correlation for apoptosis in the RCCs with cancer treatments is similar to that reported by others. 15,33 There was no apparent link with NFkB, in contrast with the results of Carvalho et al 13 This may be explained by the different RCCs used in our study, and the individual differences between RCC subtypes and also within an individual subtype, where patient differences have been recorded. 28 The RCC differences were also seen for cyclin-D1.…”

“…However, since cisplatin cytotoxicity is mediated through apoptotic pathways, mechanisms that block these pathways might also contribute to cisplatin resistance. 15,28 Co-treatment with rhEpo may either exacerbate or have a negligible or even a positive effect on the induction of apoptosis by cisplatin.…”

“…We and others have demonstrated that many of the apoptotic pathways, especially those involving the caspases, are either partially disabled, or redundant, in RCCs. 14,15 NFkB has been implicated in RCC resistance to apoptosis. 16 Signal transduction pathways involving protein kinase C (PKC) are also known to act in the development and treatment of RCCs.…”

Role of JAK3 in the Pathogenesis of Oxidative Stress-Induced Kidney Fibrosis

“…The morphological characteristics used to distinguish mitosis were as follows: (i) formation of mitotic spindles occurring during metaphase and remaining visible in anaphase or (ii) cells in the later stages of mitosis, telophase or undergoing cytokinesis. Morphological assessment has been confirmed as a reliable measure of apoptosis and mitosis, in a similar experimental protocol using molecular biomarkers for apoptosis (ApopTag) and mitosis (PCNA) in parallel with morphology (16).…”

Avaliação da interferência do efeito antioxidante do carvedilol, um potencial agente nefroprotetor, na atividade antitumoral da cisplatina

“…Apesar de ser uma linhagem celular imortalizada, as células HK-2 têm sido aceitas em numerosas publicações científicas como modelo de células normais do epitélio tubular renal, sendo especialmente sensíveis ao dano induzido pela cisplatina (DAVIDSON et al, 2005).…”