The preliminary study of exosomes derived from thymosin beta 4-treated adipose-derived stem cells in fat grafting

The retention rate in autologous fat grafting is an increasing concern for surgeons and patients. Our previous research verified that thymosin beta 4 (Tβ4) positively affected fat survival, while the mechanism was unknown. The endothelial cells (ECs) and exosomes derived from adipose-derived stem cells (ADSCs) were regarded to play a critical role in fat transplantation.

This study aimed to evaluate the effect of exosomes derived from Tβ4-treated ADSCs on EC proliferation and to identify the exosomal microRNA (miRNA) profile compared with the Tβ4-untreated group. Additionally, this research intended to recognize the related molecules and signaling pathways in the Tβ4-treated group with potential roles in fat transplants.

ADSCs were collected from patients who underwent liposuction surgery. Depending on whether the medium was supplemented with exogenous Tβ4 or not, exosomes derived from cultured ADSCs were divided into the Tβ4-Exos group and Con-Exos group. Exosome uptake and cell counting kit-8 (CCK-8) assays assessed the influence of Tβ4-Exos on EC proliferation. The exosomal miRNAs of the two groups were analyzed by next-generation sequencing. With the criteria at the |log₂ (fold change)| ≥ 1 and p-value < 0.05, up-regulated and down-regulated differentially expressed miRNAs (DEMs) were obtained. Prediction databases were used to predict the downstream mRNAs for DEMs. And then, overlapping genes for the up-regulated DEMs and the down-regulated were screened out, followed by enrichment analysis, protein-protein interaction network construction, and the gene cluster and hub gene identification.

ADSCs were obtained from four female patients. The exosome uptake and CCK-8 assays showed that the Tβ4-Exos could increase cell growth rate compared with the control group (DMEM-H + PBS). In Tβ4-Exos and Con-Exos groups, 2651 exosomal miRNAs were recognized, with 80 up-regulated and 99 down-regulated DEMs according to the criteria. After the prediction, 621 overlapping genes for the up-regulated and 572 for the down-regulated DEMs were screened. The subsequent bioinformatics analysis found specific molecules and pathways related to the positive effect on fat survival.

The exosomes derived from Tβ4-treated ADSCs probably positively affect EC proliferation. Compared with the Con-Exos group, several exosomal DEMs, genes, and pathways were distinguished. These findings of this exploratory study provide the potential direction for future in-depth research on fat grafting.