TCMK-1

CircVmn2r1 Promotes TCMK-1 Cell Senescence In Vitro

Kidney aging exacerbates kidney diseases and affects other organs. CircRNAs are crucial in aging, but their roles in kidney aging are unknown. Here, Gao's team identified circVmn2r1 as a highly expressed circRNA in mouse kidneys, significantly upregulated in 24-month-old mice compared to 3-month-old mice. They demonstrated that circVmn2r1 overexpression promotes kidney aging in senescence-accelerated mice.

To explore the function of circVmn2r1 in TCMK-1 cells, two siRNAs targeting the junction site of circVmn2r1 and an overexpression vector were designed. qRT-PCR confirmed that circVmn2r1 expression was significantly downregulated or upregulated in TCMK-1 cells transfected with the respective siRNA or vector. Among the siRNAs, si-circVmn2r1-2 had the highest silencing efficiency and was selected for further study. Importantly, neither overexpression nor knockdown of circVmn2r1 affected the linear Vmn2r1 transcript levels (Fig. 1B), making these reagents suitable for functional studies. To demonstrate that circVmn2r1 regulates the senescence of TCMK-1 cells, Gao's team conducted western blot experiments to determine the protein levels of P53 and P21 after circVmn2r1 was overexpressed or knocked down. They found that overexpression of circVmn2r1 increased the expression of P53 and P21, while knockdown of circVmn2r1 decreased the expression of P53 and P21 (Fig. 1C). SA-β-gal staining showed that SA-β-gal activity was increased by overexpression of circVmn2r1 and reduced by knockdown of circVmn2r1 (Fig. 1D and E). Flow cytometry was used to determine DNA content by staining with PI. Gao's team found that upregulation of circVmn2r1 increased the G1 phase and reduced the G2 + S phase, while downregulation of circVmn2r1 had the opposite effects (Fig. 1F and G). In summary, these results suggest that circVmn2r1 promotes ageing and inhibits proliferation of TCMK-1 cells in vitro.