Introduction: Adipose tissue-derived stem cells (ATSCs) have been used as an alternative to bone marrow mesenchymal stem cells (BMSCs) for bone tissue engineering. However, several studies have reported that the efficacy of ATSCs in bone regeneration in comparison with BMSCs remains inferior 1,2. The aim of this study was to investigate the mechanism underlying differences in ATSCs versus BMSCs osteogenicity in tissue-engineered constructs by focusing on the innate immune response mediated by the ATSCs.
Methodology: Bone formation induced by transplanted human BMSCs and ATSCs combined with calcium carbonate ceramic granules (named tissue constructs) was evaluated in vivo in an ectopic mouse model. Explants were analyzed by µCT and histology. Kinetic analyses of both the expressed human and murine genes pertaining to osteogenesis, angiogenesis, and inflammatory response in tissue constructs explanted at 0, 7, 14, and 28 days post-implantation were performed. The gene expression and secretome profiles of pro-inflammatory cytokines/chemokines in both ATSC and BMSC were analyzed.
Results: All constructs containing BMSCs induced ectopic bone formation and histological observation of explants revealed the presence of new bone with the presence of osteoclastic (TRAP +) multinucleated cells on contact with the ceramic particles. On the contrary, the constructs containing ATSCs did not generate (or minimally) bone tissue; they were infiltrated with fibrous tissue, and numerous TRAP- multinucleated giant cells (MNGC) were observed. Gene expression analysis of explants revealed that implanted human BMSCs differentiated into the osteogenic lineage in vivo concomitantly with the osteogenic differentiation of host murine progenitors. In contrast, the osteogenic differentiation in construct-contained ATSCs started later than in BMSCs, when only less than 1% of implanted ATSC were present; no osteogenic differentiation in host murine cells occurred. Expressions of genes pertaining to vascularization were not significantly different between both groups. Regarding the inflammatory response, compared with BMSCs, the expressions of human IL1b and IL6 genes were highly upregulated in implanted ATSCs during the first-week post-implantation and then decreased; In parallel, murine IL1b was also upregulated in ATSC-containing constructs as were the M1/MNGC-associated iNOS and CD86 murine genes. An extensive analysis of gene expression of human cytokines and chemokines comparing the ATSC and BMSC contained in constructs at day 0 (before implantation) was conducted. This revealed up-regulation of 23 inflammatory mediators out of 84 tested in ATSC compared to BMSC (the highest (> 30-fold) upregulated genes were CSF3, CXCL10, CXCL5, CXCL11). Only CXCL12 (SDF1), RANKL, and BMP4 were slightly (3-7 fold) upregulated in BMSCs. Such a pro-inflammatory profile of ATSCs was confirmed at the protein level after quantification in the construct-contained cell supernatant.
Conclusion: In contrast to BMSCs, ATSCs display no/weaker osteogenic potential in vivo. ATSCs are a transient source of proinflammatory cytokines and chemokines that promote an inflammatory environment within the cell-containing constructs. This event correlates with impaired osteogenic differentiation of both implanted ATSCs and host osteoprogenitors.
1. Brennan, M. A. et al. Stem Cells Transl. Med. 6, 2160–2172 (2017).
2. Mohamed-Ahmed, S. et al. Stem Cell Res. Ther. 9, 168 (2018)