Hypoxia Alters the Proteome Profile and Enhances the Angiogenic Potential of Dental Pulp Stem Cell-Derived Exosomes

Abstract

Dental pulp stem cells (DPSCs) and their exosomes (Exos) are effective treatments for regenerative medicine. Hypoxia was confirmed to improve the angiogenic potential of stem cells. However, the angiogenic effect and mechanism of hypoxia-preconditioned DPSC-Exos are poorly understood. We isolated exosomes from DPSCs under normoxia (Nor-Exos) and hypoxia (Hypo-Exos) and added them to human umbilical vein endothelial cells (HUVECs). HUVEC proliferation, migration and angiogenic capacity were assessed by CCK-8, transwell, tube formation assays, qRT-PCR and Western blot. iTRAQ-based proteomics and bioinformatic analysis were performed to investigate proteome profile differences between Nor-Exos and Hypo-Exos. Western blot, immunofluorescence and immunohistochemistry were used to detect the expression of lysyl oxidase-like 2 (LOXL2) in vitro and in vivo. Finally, we silenced LOXL2 in HUVECs and rescued tube formation with Hypo-Exos. Hypo-Exos enhanced HUVEC proliferation, migration and tube formation in vitro superior to Nor-Exos. The proteomics analysis identified 79 proteins with significantly different expression in Hypo-Exos, among which LOXL2 was verified as being upregulated in hypoxia-preconditioned DPSCs, Hypo-Exos, and inflamed dental pulp. Hypo-Exos partially rescued the inhibitory influence of LOXL2 silence on HUVEC tube formation. In conclusion, hypoxia enhanced the angiogenic potential of DPSCs-Exos and partially altered their proteome profile. LOXL2 is likely involved in Hypo-Exos mediated angiogenesis.

Small extracellular vesicles derived from hypoxic preconditioned dental pulp stem cells ameliorate inflammatory osteolysis by modulating macrophage polarization and osteoclastogenesis

“…However, the effects of hypoxia on the therapeutic potential of DPSC-sEV remain largely unknown. Our recent study revealed that hypoxia-induced DPSC-sEV have enhanced angiogenic potential through the enrichment of LOXL2 compared with control DPSC-sEV [ 26 ]. Here, we showed that hypoxia promotes the release of sEV from DPSCs, which is in line with recent studies about the effects of hypoxia on the release of exosomes in tumor cells and umbilical cord MSCs [ 42 , 47 ].…”

“…Our previous study indicated that the proteomics of hypoxia-preconditioned DPSC-derived sEV were significantly different from those of control DPSC-sEV. More importantly, hypoxia-preconditioned DPSC-derived sEV showed increased angiogenic potential [ 26 ]. However, whether DPSC-sEV produced under hypoxic conditions exert better therapeutic effects on inflammatory osteolysis by inhibiting the macrophage inflammatory response and osteoclastogenesis remains largely unknown.…”

MicroRNA-195-3p as a potential regulator of hypoxic injury in HUVECs

“…The ECs are activated during collateral artery formation (arteriogenesis) (18), resulting in their proliferation, migration, and differentiation into tubular structures. The surrounding tissues wrap around the ECs to form vascular walls, and vascular networks coincide, initiating the collateral circulation (19)(20)(21). Researchers have a contrary opinion regarding the role of hypoxia in arteriogenesis.…”

Research progress of odontogenic extracellular vesicles in regeneration of dental pulp

“…Furthermore, hypoxia partially altered the proteomic profile of odontogenic EVs and increased their vasculogenesis capacity, and researchers hypothesized that LOXL2 could be involved in Hypo-EVs mediated angiogenesis (Li, Xian, et al, 2022). In terms of cell apoptosis, hDPSC-apoVs activated ECs autophagy signaling through the transfer of mitochondrial Tu translation elongation factor (TUFM),…”

“…Pulp tissue is exposed to ischemic–hypoxic environment due to the rigid dentin wall and the narrow apical foramina (Li, Wu, et al, 2022; Li, Xian, et al, 2022). At present, the biological effects of DSC‐EVs under hypoxia, as well as the specific molecular mechanisms regulating angiogenesis, have become research priorities.…”