Lidocaine could promote the cuproptosis through up-regulating the long noncoding RNA DNMBP-AS1 in Hep-2 cells

Abstract

Background: Lidocaine is a traditional local anesthetic, which has been reported to trigger apoptosis through the mitochondrial pathway, independent of death receptor signaling. Cuproptosis is a copper triggered mitochondrial cell death mode. In this study, we explored the biological effects of lidocaine on cuproptosis in Hep-2 cells and studied the relevant mechanisms.

Methods: quantitative RT-PCR was used to measure the expression level of long noncoding RNA (IncRNA) DNMBP-AS1. DNMBP-AS1 siRNA (si-DNMBP-AS1) were transfected into Hep-2 cells to verify the roles of DNMBP-AS1 in cuproptosis. 24 h treatment with 20 nM elesclomol and 2 µM CuCl₂ was performed to promote the occurrence of Cuproptosis. Cell proliferation, migration and apoptosis assays ware utilized to analyze biological effect of lidocaine and DNMBP-AS1 on Hep-2 cells. Active caspase-3 were also determined after treatment.

Results: DNMBP-AS1 was significantly upregulated during cuproptosis in Hep-2 cells. The si-DNMBP-AS1 significantly increased the cell viability with nonactivated caspase-3, promoted the cell migration and suppress the cuproptosis. Lidocaine was cytotoxic to the Hep-2 cells in a dose- and time-dependent manner. Exposure to 10 µM of lidocaine for 24 h did not reduce the viability or activated the caspase-3, but significantly increased the expression of DNMBP-AS1, and promote the cuproptosis. Anymore, si-DNMBP-AS1 reversed the pro-cuproptosis function of lidocaine.

Conclusions: lidocaine was cytotoxic to Hep-2 cells in a time- and dose-dependent manner, promoted the cuproptosis through up-regulating DNMBP-AS1. The results of this study offered initial optimism that lidocaine could be used in an adjuvant or neoadjuvant fashion in cancer treatment.

Keywords: Cuproptosis; Hep-2 cells; IncRNA DNMBP-AS1; Lidocaine.

Fig. 1

DNMBP-AS1 was significantly upregulated by exogenous introduction of copper ions. (A) Changes of the lncRNA DNMBP-AS1 expression in the constructed cuproptosis cell model was assessed by qRT-PCR. (B) Efficiency of DNMBP-AS1 knockdown (si-DNMBP-AS1-1 and si-DNMBP-AS1-2) was assessed by qRT-PCR. (C) Effect of si-DNMBP-AS1 on the proliferation of Hep-2 cells was detected by CCK-8 assays. (D) Migration of Hep-2 cells was detected via wound healing assay. (E) Concentration of active caspase-3 was analyzed by ELISA. (F) Cell death of Hep-2/si-ctrl and Hep-2/si-DNMBP-AS1 cells induced by exogenous introduction of copper ions were analyzed via flow cytometry assay. Data are shown as the mean ± SD of three independent experiments. *P < 0.01, ^#P > 0.05

Fig. 2

Lidocaine could promote the cuproptosis through up-regulating the DNMBP-AS1 (A) Viability of Hep-2 cells treated with lidocaine was assessed by CCK-8. (B) Concentration of active caspase-3 was analyzed by ELISA. (C) Effect of lidocaine on the DNMBP-AS1 expression was assessed by qRT-PCR. (D) effect of lidocaine on the cuproptosis induced by exogenous introduction of copper ions were analyzed via flow cytometry assay. Data are shown as the mean ± SD of three independent experiments. *P < 0.01, ^#P > 0.05