In the present study, we employed multiomics analysis to investigate the biological interactions of SF and SS with mesenchymal stem cells (MSCs) to elucidate the triggered cellular responses and pathways. The integrated transcriptomics and proteomics revealed multiple biological processes of MSCs with widespread gene and protein changes initiated by SF and SS. Specifically, both SF and SS strongly potentiated the paracrine activities of MSCs via activating multiple but distinct signaling pathways instead of directly regulating cell behaviors. Additionally, we investigated whether the paracrine functions of MSCs in response to SF and SS could mediate the local signaling processes in vivo and have implications for tissue regeneration. We found that SF and SS triggered specific paracrine signals of MSCs that promote cell proliferation, migration, angiogenesis, immunomodulation, and tissue healing processes. Moreover, the high consistency of transcriptomic and proteomic analyses in vitro and in vivo indicated the strong cellular responses of MSCs to SF and SS and the stable and powerful biological effects of these paracrine signals. This work thus provides a comprehensive and reliable insight into the cellular interactions with SF and SS via multiomics profiling, which enables the further development of silk-based therapeutics for tissue engineering and regenerative medicine