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. 2020 Dec 7;9(12):1238.
doi: 10.3390/antiox9121238.

Paraoxonase-2 Silencing Enhances Sensitivity of A375 Melanoma Cells to Treatment with Cisplatin (VSports在线直播)

Affiliations

Paraoxonase-2 Silencing Enhances Sensitivity of A375 Melanoma Cells to Treatment with Cisplatin

Roberto Campagna (VSports) et al. Antioxidants (Basel). .

Abstract

Melanoma represents the most aggressive skin cancer, being responsible for the majority of deaths related with these neoplasms. Despite chemotherapy represents a frontline approach for management of the advanced stages of the disease, it displayed poor response rates and short-term efficacy due to melanoma cell resistance. Therefore, the discovery of molecules that can be used for effective targeted therapy of melanoma is crucial VSports手机版. In this study, we evaluated the impact of paraoxonase-2 (PON2) silencing on proliferation, viability, and resistance to treatment of the A375 melanoma cell line with chemotherapeutic drugs dacarbazine (DTIC) and cisplatin (CDDP). Due to the enzymes ability to counteract oxidative stress, we also evaluated the effect of enzyme knockdown on reactive oxygen species (ROS) production in cells treated with CDDP. The data reported clearly demonstrated that PON2 knockdown led to a significant reduction of cell proliferation and viability, as well as to an enhancement of A375 sensitivity to CDDP treatment. Moreover, enzyme downregulation was associated with an increase of ROS production in CDDP-treated cells. Although further analyses will be necessary to understand how PON2 could influence melanoma cell metabolism and phenotype, our results seem to suggest that the enzyme may serve as an interesting molecular target for effective melanoma treatment. .

Keywords: cell proliferation; cell viability; chemotherapeutic drugs; melanoma; oxidative stress; paraoxonase-2 V体育安卓版. .

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VSports - Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Evaluation of PON2 silencing. A375 cells were treated with a shRNA-coding plasmid against PON2 (pLKO.1-647), with empty vector (pLKO.1-puro), or with transfection reagent only (mock). PON2 expression was evaluated at mRNA and protein level by Real-Time PCR (panel A) and Western blot (panels B,C). Values are expressed as mean ± standard deviation (* p < 0.05).
Figure 1
Figure 1
Evaluation of PON2 silencing. A375 cells were treated with a shRNA-coding plasmid against PON2 (pLKO.1-647), with empty vector (pLKO.1-puro), or with transfection reagent only (mock). PON2 expression was evaluated at mRNA and protein level by Real-Time PCR (panel A) and Western blot (panels B,C). Values are expressed as mean ± standard deviation (* p < 0.05).
Figure 2
Figure 2
In vitro effect of PON2 silencing on cell proliferation and cell viability. Cell proliferation was analyzed by trypan blue exclusion assay in control samples (mock and pLKO.1-puro) and PON2 downregulating cells (pLKO.1-647) at 0, 24, 48, and 72 h (panel A). Cell viability was evaluated through MTT assay (panel B). Values are expressed as mean ± standard deviation (* p < 0.05).
Figure 3
Figure 3
Effect of chemotherapeutic treatment on A375 cells. MTT assay was used to evaluate the effect of dacarbazine (DTIC) (100 µg/mL) (panel A) and cisplatin (CDDP) (4 and 8 µM) (panels B,C) on cell viability of mock, samples treated with empty vector (pLKO.1-puro), and PON2 downregulating cells (pLKO.1-647). Measurements were performed at different time points (0, 24, 48, and 72 h). All values are expressed as mean ± standard deviation (* p < 0.05).
Figure 4
Figure 4
Intracellular reactive oxygen species (ROS) levels in A375 cells upon CDDP treatment. ROS levels were determined in mock, A375 transfected with empty vector (pLKO.1-puro), and PON2 downregulating cells (pLKO.1-647) upon treatment with CDDP (4 µM) at different time points (0, 24, 48, and 72 h). All values are expressed as the mean ± standard deviation (* p < 0.05).

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