Astrocytes perform essential functions in the preservation of neural tissue. For this reason, these cells can respond with changes in gene expression, hypertrophy, and proliferation upon a traumatic brain injury event (TBI). Different therapeutic strategies may be focused on preserving astrocyte functions and favor a non-generalized and non-sustained protective response over time post-injury. A recent strategy has been the use of the conditioned medium of human adipose mesenchymal stem cells (CM-hMSCA) as a therapeutic strategy for the treatment of various neuropathologies. However, although there is a lot of information about its effect on neuronal protection, studies on astrocytes are scarce and its specific action in glial cells is not well explored. In the present study, the effects of CM-hMSCA on human astrocytes subjected to scratch assay were assessed. Our findings indicated that CM-hMSCA improved cell viability, reduced nuclear fragmentation, and preserved mitochondrial membrane potential. These effects were accompanied by morphological changes and an increased polarity index thus reflecting the ability of astrocytes to migrate to the wound stimulated by CM-hMSCA. In conclusion, CM-hMSCA may be considered as a promising therapeutic strategy for the protection of astrocyte function in brain pathologies.