A novel 3D co-culture model augmented with single-cell sequencing to study the response of breast cancer cells to cisplatin

Three-dimensional (3D) cell direct co-culture is an essential method for cancer research, which can overcome the limitations of traditional research based on cell lines. Single-cell RNA sequencing can detect the cell heterogeneity of direct co-culture of 3D cells at the single-cell level,However, there is still lack verification. The purpose of this study is to verify the superiority of single-cell RNA sequencing combined with 3D cell direct co-culture in breast cancer research and prove that it can become a powerful solution for cancer cell culture in vitro.In this study ,we found that the different cell culture modes showed very different responses to cisplatin intervention. In MCF-7 alone culture mode, there was a large difference in the transcriptome of MCF-7 after cisplatin intervention compared with the group not given drug intervention; while in direct co-culture mode, there was also a difference between the cisplatin intervention group and the control group, but this difference was relatively small. To further seek evidence that the direct co-culture mode is closer to the real environment in vivo, we analyzed the differential genes in the direct co-culture mode, and the enriched signaling pathways were similar to the results in the CancerSEA database, and the enriched signaling pathways included apoptosis, DNA damage, hypoxia, and metastasis. Conclusions: Single-cell sequencing can facilitate direct co-culture of 3D cells in tumor research, a model that more closely resembles the real pathophysiological environment in vivo. Overall design: The experimental samples were divided into four groups: cell co-culture control group, cell co-culture cisplatin-induced group, MCF-7 cultured alone group, and MCF-7 cultured alone cisplatin-induced group. For cell co-culture, MCF-7, HMF, and HUVEC were mixed at a ratio of 1:1:1. All the cells were cultured in the previously prepared Matrigel matrix. The cells in the co-culture cisplatin-induced group and the MCF-7 alone cisplatin-induced group were incubated with cisplatin at a concentration of 1.25 mm/ml. The cells were harvested after 2 days of cisplatin treatment and subjected to single-cell RNA sequencing.