Indirubin induces apoptosis in ovarian cancer cells via the mitochondrial pathway

To investigate the pro-apoptotic effects of Indirubin, a traditional Chinese medicine, on ovarian cancer SKOV3 cell line and to explore its underlying mechanisms.

Ovarian cancer SKOV3 cells were divided into a control group (cells cultured normally) and an experimental group (cells cultured in medium containing Indirubin). SKOV3 cells at the logarithmic phase were treated with Indirubin at various concentrations. Cell proliferation was assessed using the Cell Counting Kit-8 (CCK-8) assay and 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay, while apoptosis was detected by flow cytometry, TdT-mediated dUTP Nick-End Labeling (TUNEL) staining, and immunofluorescence. Transcriptome sequencing was conducted to screen for apoptosis-related factors. qPCR and western blot were used to detect the mRNA and protein expression on added of molecules related to mitochondrial permeability transition pores.

MTT assay showed that Indirubin inhibited the growth of SKOV3 cells in both plate and sphere cultures, with IC₅₀ values of 3.003 μM (plate culture) and 4.253 μM (sphere culture), respectively. Indirubin showed a lower inhibitory concentration than cisplatin (IC₅₀ 3.687 μM in plate culture and 7.023 μM in sphere culture), and its effect was comparable to adriamycin. Flow cytometry revealed an increase in apoptosis rates in SKOV3 cells treated with Indirubin. Transcriptome sequencing indicated significant changes in the transcription of various apoptosis-related genes, particularly those involved in the mitochondrial apoptosis pathway, such as Bcl-2-associated X protein (Bax) and Bcl2 associated agonist of cell death (Bad). After Indirubin treatment, the mRNA and protein expression levels of mitochondrial channel-related genes Cyclophilin D (CyPD), adenine nucleotide translocator 1 (ANT1), and voltage-dependent anion channel (VADC) were significantly elevated (all P < 0.05). By regulating the mitochondrial membrane permeability through the Bcl-2 family members, Indirubin promoted apoptosis in SKOV3 cells.

Indirubin inhibits the proliferation and promotes the apoptosis of ovarian cancer cells, exerting an anti-tumor effect. Its pro-apoptotic action is closely related to the mitochondrial apoptosis pathway.