Nowadays, paracrine regulation is considered as a major tool of mesenchymal stem cell (MSC) involvement in tissue repair and renewal in adults. Aging results in alteration of tissue homeostasis including neovascularization. In this study, we examined the influence of replicative senescence on the angiogenic potential of adipose-derived MSCs (ASCs). Angiogenic activity of conditioned medium (CM) from senescent and “young” ASCs was evaluated in chorioallantoic membrane (CAM) assay in ovo using Japanese quail embryos. Also, the formation of capillary-like tubes by human umbilical vein endothelial cells (HUVECs) in 3D basement membrane matrix “Matrigel” and HUVEC migration capacity were analyzed. Multiplex, dot-blot and gene expression analysis were performed to characterize transcription and production of about 100 angiogenesis-associated proteins. The results point to decreased angiogenic potential of senescent ASC secretome in ovo. A number of angiogenesis-associated proteins demonstrated elevation in CM after long-term cultivation. Meanwhile, VEGF (key positive regulator of angiogenesis) did not change transcription level and concentration in CM. Increasing both pro- (FGF-2, uPA, IL-6, IL-8 etc.) and antiangiogenic (IL-4, IP-10, PF4, Activin A, DPPIV etc.) factors was observed. Some proangiogenic genes were downregulated (IGF1, MMP1, TGFB3, PDGFRB, PGF). Senescence-associated secretory phenotype (SASP) modifications after long-term cultivation lead to attenuation of angiogenic potential of ASC.

Multipotent mesenchymal stem/stromal cells (MSCs) are strongly involved in tissue homeostasis mainly through paracrine regulation. In this study, we examined the influence of simulated microgravity on the angiogenic potential of adipose-derived MSCs (ASCs). The conditioned medium (CM) from random positioning machine (RPM)-exposed ASCs stimulated the formation of vessel network in ovo, endothelial cell (EC) capillary-like network, and nondirected EC migration in vitro. These effects were driven by alteration of both angiogenesis-related gene and protein expression. The elevation of angiogenic regulators Serpin E1, Serpin F1, IGFBP, VEGF, and IL-8 was detected in ASC-CM after 3D-clinorotation. In addition, transcription of genes encoding growth factors with proangiogenic activity were upregulated including VEGF-c and VEGF-a. These data evidenced that besides direct effect on ECs, microgravity could provoke MSC-mediating specific microenvironment for ECs supporting their functions, that is, proliferation and migration via increased production of IL-8 and VEGF as well as other paracrine factors involved in angiogenesis regulation.

Multipotent mesenchymal stromal cells are considered as a perspective tool in cell therapy and regenerative medicine. Unfortunately, autologous cell therapy does not always provide positive outcomes in elder donors, perhaps as a result of the alterations of stem cell compartments. The mechanisms of stem and progenitor cell senescence and the factors engaged are investigated intensively. In present paper, we elucidated the effects of tissue-related O on morphology, functions, and transcriptomic profile of adipose tissue-derived stromal cells (ASCs) in replicative senescence in vitro model. Replicatively senescent ASCs at ambient (20%) O (12-21 passages) demonstrated an increased average cell size, granularity, reactive oxygen species level, including anion superoxide, lysosomal compartment activity, and IL-6 production. Decreased ASC viability and proliferation, as well as the change of more than 10 senescence-associated gene expression were detected (IGF1, CDKN1C, ID1, CCND1, etc). Long-term ASC expansion at low O (5%) revoked in part the replicative senescence-associated alterations.

The expression of 84 focal adhesion genes of multipotent mesenchymal stromal cells (MMSCs) after 96-h microgravity simulation at 3D clinorotation was studied. The upregulation of ITGA6, ITGA7, BCAR1, GRB2, CAV1, and DIAPH1 and the downregulation of ITGA11, ITGAV, ITGB1, PTEN, PTK2 (FAK), ARHGAP5, DOCK1, ROCK2, and AKT3 was found. These changes at the transcriptional level may be a cause of the reduction of the osteogenic potential of MMSCs and their ability to migration and adhesion in microgravity.

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