Abstract
PurposeAutophagy has attracted attentions as a novel mechanism for tumor development. In this study Human ovarian carcinoma cell line SKOV3 and multidrug-resistant phenotype SKVCR cells were used and the roles of autophagy in radiation-induced cell death were analyzed.Methods and materialsCell viability was examined by colony formation and cell counting kit-8 (CCK-8) assay, 3MA and ZVAD were used to block autophagy and apoptosis, respectively. Quantitative real-time PCR was used to detect mRNA level and Western blot was used to detect protein expression, monodansylcadaverine (MDC) staining and flow cytometery were used for autophagy, apoptosis and cell cycle dynamics, respectively.Results(1) The radiosensitivity exhibited differently in SKOV3 and SKVCR cells (SKOV3: D0=3.37, SKVCR: D0= 4.18); compared with SKOV3 the constitutive expression of MAPLC3 in SKVCR was higher, but no change of Caspase-3 and cleaved Caspase-3. (2) The ionizing radiation (IR)- induced apoptosis and autophagy were significant in both cells (P<0.05); inhibition of apoptosis with ZVAD showed no impact on survival of SKOV3 and SKVCR cells after radiation, while inhibition of autophagy significantly decreased viability in SKVCR cells, for SKVO3 cells only low level of radiation (2 Gy and 4 Gy) could decrease the viability(P<0.05). (3) ZVAD inhibited apoptosis and autophagy in both cells, 3MA inhibit apoptosis in SKOV3, and promote apoptosis in SKVCR, together with inhibition of autophagy. (4) G2/M arrest was induced by radiation in both cells; the accumulation of G2/M was more significant in SKOV3, 3MA attenuated the radiation-induced S phase delay in SKVCR.ConclusionIR-induced autophagy provides a self-protective mechanism against radiotherapy in SKVCR cells, the use of autophagy inhibitor, 3MA, increases the killing effects of radiation by inhibiting autophagy and radiation- induced S phase delay, also by the increase of apoptosis, which suggests a better therapeutic strategy in drug- resistant SKVCR ovarian cancer cells.
Highlights
Epithelial ovarian cancer(EOC)is the most lethal Gynecologic malignancy, with 21,990 estimated new cases and 15, 460 deaths in USA in 2011[1]
The radiosensitivity in human ovarian carcinoma SKOV3 and multidrug-resistant phenotype SKVCR cells SKOV3 and SKVCR cells were exposed to various doses of radiation (0, 2, 4, 6 or 8 Gy), cell viability was measured by Colony formation and CCK8 assay
SKOV3 and SKVCR cells After we found a variation in the radiosensitivity between the two cell lines, we sought to find the contributions of apoptosis and autophagy to the total cell death
Summary
Epithelial ovarian cancer(EOC)is the most lethal Gynecologic malignancy, with 21,990 estimated new cases and 15, 460 deaths in USA in 2011[1]. Tumor cells have the capacity to respond to chemotherapy and radiation through multiple growth arrest and cell death pathways [3,4,5]. The type I programmed cell death, has been widely investigated under different circumstance including radiotherapy and the one of the most important strategies for cancer treatment over the past decades. Autophagy, the type II programmed cell death, has been recently reported to play roles in the development of cancer. Dual roles of autophagy have been reported, promotion of cell survival or leading to cell death [18,19]. Elucidation of autophagy roles in treatment responsiveness at different stages of cancer progression is a complex and challenging task. Better understanding of autophagy regulation and its impact on treatment outcomes will potentially provide novel therapeutic targets in cancer
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