MSC spheroids-loaded collagen hydrogels simultaneously promote neuronal differentiation and suppress inflammatory reaction through PI3K-Akt signaling pathway

It is critical for the clinical success to take the anti-inflammatory function into consideration when integrating the neurogenesis into the nerve repair materials. To this aim, we prepared mesenchymal stem cell (MSC) spheroids-loaded collagen (Col) hydrogels with combined superior anti-inflammatory efficacy and neurogenic activity. The size of the MSC spheroids showed a strong modulation effect on both functions, and the MSC spheroids-100 sample exhibited the best neuronal and anti-inflammatory potentials. The observed dual functions were likely based on the elevated intrinsic cell-cell contacts and cell-extracellular matrix interactions from the MSC spheroids. MSC self-assembly as spheroids expedited the secretions of endogenous trophic factors and extracellular matrix (ECM), which was beneficial to drive neural stem cell differentiation into the neuronal lineage. In addition, the formation of the MSC spheroids secreted more amounts and types of cytokines as well as immunomodulatory paracrine factors to suppress LPS-induced inflammatory reaction. LC-MS/MS analysis further demonstrated that MSC spheroids contributed to the activation of neuroactive ligand-receptor interaction, thereby triggering downstream PI3K-Akt signal pathway, which was likely due to the acceleration of ECM-receptor interaction, gap junction and tight junction. Importantly, inhibiting Akt pathway significantly suppressed the neuronal differentiation, indicating that PI3K-Akt signal pathway was critically involved in the Col-MSC spheroid hydrogel mediated neuroprotection and neurogenesis. Such findings not only provided a simple approach for improving MSC-based therapies for neuron-related diseases, but also shed insight on understanding the underlying mechanisms of MSC-mediated neuronal differentiation.