Unique hypoxia-tolerant subpopulations of adipose-derived stem cells: ITGB3⁺ cells.

Abstract

We found by accident that stem cells could still be isolated from adipose tissue stored for 14 days in sealed tubes, which was distinct from previous protocols. The morphology of these hypoxia-tolerant stem cells also differs from that of conventional adipose-derived stem cells (ADSCs). In this study, we aim to define the newly found subsets. Stem cells were isolated from adipose tissue that was aspirated immediately or stored for 14 days. The stem cells were then harvested for flowcytometric analysis and differentiation potentials. The expression of hypoxia-inducible factor 1 alpha (HIF-1α) was assayed to confirm the hypoxia-tolerant ability. RNA sequencing (RNA-seq) was performed to find the common signatures of the hypoxia-tolerant cells. The result of bioinformatics was tested by quantitative real-time reverse transcription-polymerase chain reaction (qPCR) and western blotting. Certain subsets of ADSCs can be isolated from adipose tissue stored for 14 days. These survived cells were positive for CD90, CD105, and CD73 and showed multilineage differentiation potentials. The hypoxic condition was evidenced by up-regulation of HIF-1α for 2.0-fold changes (p < 0.05). The hypoxia-tolerant stem cells were distinct from multilineage-differentiating stress-enduring (Muse) cells, previously found stress-enduring stromal cells. RNA-seq suggested that integrin beta 3 (ITGB3) was highly expressed in hypoxia-tolerant subpopulations. The result was further confirmed at transcription and translation levels by qPCR and western blotting (mRNA: 2.9 ± 0.4, p < 0.05; protein: 1.5 ± 0.2, p < 0.05; respectively). The conventional ADSCs are positive for ITGB3, which implies that ITGB3+ cells are subpopulations of heterogeneous ADSCs. Our study reveals the ITGB3⁺ subsets with potent hypoxia tolerance, which has significant implications for improving fat retention rates and curing obesity-related diseases.