Highly-expressed micoRNA-21 in adipose derived stem cell exosomes can enhance the migration and proliferation of the HaCaT cells by increasing the MMP-9 expression through the PI3K/AKT pathway.

Wound healing remains a challenge in burns and trauma fields. Adipose derived stem cells exosomes (AD-exos) had been confirmed to have a positive effect on the wound healing and the migration and proliferation of keratinocyte. However, the mechanism of the AD-exos is still unclear. The objective of this article is to observe the function of the miR-21 expressed in the adipose AD-exos and the effect on migration and proliferation of the HaCaT cells. The full layer dermal wound of BALb/c mouse was used to observe the vitro effect of the AD-exos and detect the expression of miR-21.The co-culture systems were established by transwell plates for observing the migration, proliferation, apoptosis rate, detecting the RNA, and protein expression in different treated groups. MiR-21 plasmid was used to over-express miR-21 by transfection of HaCaT cells. GW4869 was used to inhibit the secreting of exosomes from ADSCs. The results showed that both ADSCs and the AD-exos could improve the wound healing process of BALb/c mouse full layer skin wound at a similar level, especially at the 7th day post surgery when compared to the control group (p < 0.01) and the highly expressed miR-21 was detected (p < 0.01 compared with control group and p < 0.001 compared to other microRNAs) in the treated groups at the same time point. AD-exos could obviously enhance the migration and proliferation of the HaCaT cells (p < 0.01), and fell back to the same level when the exosomes inhibitor--GW4869 was added compared with control group (p > 0.05). Over-expressed miR-21 could also significantly improve the migration and proliferation of HaCaT cells. But both AD-exos and miR-21 had no significantly effect on the apoptosis rate of HaCaT cells (p > 0.05 compared with each other). Over-expression of miR-21 plasmid could decrease the TGF-βI protein level (p < 0.001 vs. control group) in HaCaT cells while TGF-βI protein level increased again when antagomiR-21 was added in (p < 0.01 vs. empty plasmid group, p < 0.001 vs. miR-21 plasmid group). MiR-21 expression of HaCaT cells could be increased by the transfect ion of miR-21 plasmid (p < 0.001 vs. empty plasmid group) and decreased by antagomiR-21 (p < 0.01 vs. empty plasmid group, p < 0.001 vs. miR-21 plasmid group). MiR-21 appeared to have influence on MMP-9 and TIMP-2 (p < 0.001 compared to control group and p < 0.001 compared to TGF-βI group) but not MMP-2 and TIMP-1 (p > 0.05 compared to control group and TGF-βI group). These processes might act through PI3K/AKT pathway. This research provide the experimental evidence that the miR-21 is highly expressed in AD-exos and can significantly accelerate the wound healing process and enhance the migration and proliferation of the HaCaT cells. Over-expressed miR-21 can inhibit the TGF-βI expression and excess TGF-βI can also have negative feedback influence on miR-21. We have found a reliable evidence that these two factors can act on HaCaT cells by influencing MMP-2 and TIMP-1 protein expression through the PI3K/AKT signal pathway. These results may provide a potential perspectives on improving the wound healing.

Yang C ，Luo L ，Bai X ，Shen K ，Liu K ，Wang J ，Hu D ... -  《-》

[The effect and mechanism of exosomes derived from human amniotic epithelial cells on the proliferation and migration of HaCaT in high glucose environment].

Wei P ，Xu ZR ，Chen YM ，Chen XD ，Chen ZH ... -  《-》

Adipose mesenchymal stem cell exosomes promote wound healing through accelerated keratinocyte migration and proliferation by activating the AKT/HIF-1α axis.

Zhang Y ，Han F ，Gu L ，Ji P ，Yang X ，Liu M ，Tao K ，Hu D ... -  《-》

Cell-free therapy based on adipose tissue stem cell-derived exosomes promotes wound healing via the PI3K/Akt signaling pathway.

Adipose tissue-derived stem cells (ADSCs) have been shown to enhance wound healing via their paracrine function. Exosomes, as one of the most important paracrine factors, play an essential role in this process. However, the concrete mechanisms that underlie this effect are poorly understood. In this study, we aim to explore the potential roles and molecular mechanisms of exosomes derived from ADSCs in cutaneous wound healing. Normal human skin fibroblasts and ADSCs were isolated from patient skin and adipose tissues. ADSCs were characterized by using flow cytometric analysis and adipogenic and osteogenic differentiation assays. Exosomes were purified from human ADSCs by differential ultracentrifugation and identified by electron microscopy, nanoparticle tracking, fluorescence confocal microscopy and western blotting. Fibroblasts were treated with different concentrations of exosomes, and the synthesis of collagen was analyzed by western blotting; the levels of growth factors were analyzed by real-time quantitative PCR (RT-PCR) and ELISA; and the proliferation and migration abilities of fibroblasts were analyzed by real-time cell analysis, CCK-8 assays and scratch assays. A mouse model with a full-thickness incision wound was used to evaluate the effect of ADSC-derived exosomes on wound healing. The level of p-Akt/Akt was analyzed by western blotting. Ly294002, a phosphatidylinositol 3-kinases (PI3K) inhibitor, was used to identify the underlying mechanisms by which ADSC-derived exosomes promote wound healing. ADSC-derived exosomes were taken up by the fibroblasts, which showed significant, dose-dependent increases in cell proliferation and migration compared to the behavior of cells without exosome treatment. More importantly, both the mRNA and protein levels of type I collagen (Col 1), type III collagen (Col 3), MMP1, bFGF, and TGF-β1 were increased in fibroblasts after stimulation with exosomes. Furthermore, exosomes significantly accelerated wound healing in vivo and increased the level of p-Akt/Akt in vitro. However, Ly294002 alleviated these exosome-induced changes, suggesting that exosomes from ADSCs could promote and optimize collagen deposition in vitro and in vivo and further promote wound healing via the PI3K/Akt signaling pathway. This study demonstrates that ADSC-derived exosomes can promote fibroblast proliferation and migration and optimize collagen deposition via the PI3K/Akt signaling pathway to further accelerate wound healing. Our results suggest that ADSCs likely facilitate wound healing via the release of exosomes, and the PI3K/Akt pathway may play a role in this process. Our data also suggest that the clinical application of ADSC-derived exosomes may shed new light on the use of cell-free therapy to accelerate full-thickness skin wound healing and attenuate scar formation.

Zhang W ，Bai X ，Zhao B ，Li Y ，Zhang Y ，Li Z ，Wang X ，Luo L ，Han F ，Zhang J ，Han S ，Cai W ，Su L ，Tao K ，Shi J ，Hu D ... -  《-》

Adipose mesenchymal stem cell-derived exosomes accelerate skin wound healing via the lncRNA H19/miR-19b/SOX9 axis.

It has been reported that adipose mesenchymal stem cells (ADSCs) accelerate wound healing. Moreover, exosomes, which serve as paracrine factors, play a vital role in wound healing. However, the mechanism remains unclear. This research aimed to determine the roles of exosomes derived from ADSCs (ADSC-Exos) in wound skin tissue repair. Flow cytometry and electron microscopy were carried out to identify ADSCs and ADSC-Exos, respectively; RT-qPCR was performed to assess the lncRNA H19 (H19), microRNA19b (miR-19b) and SRY-related high-mobility-group box 9 (SOX9) levels; Western blotting was carried out to evaluate collagen and β-catenin expression; CCK-8, scratch and transwell assays were conducted to evaluate human skin fibroblast (HSF) cell proliferation, migration and invasion, respectively; the potential binding sites between H19 and miR-19b, miR-19b and SOX9 were detected by dual-luciferase reporter gene assay and RIP assay; and H&E staining was conducted to observe skin wound tissues. ADSC-Exos accelerated the proliferation, migration and invasion of HSF cells via H19. H19 acts as a molecular sponge towards miR-19b, which targets SOX9. ADSC-Exos inhibited miR-19b expression via H19, resulting in accelerated HSF proliferation, migration and invasion. ADSC-Exos upregulated SOX9 to activate the Wnt/β-catenin pathway, resulting in accelerated HSF cell proliferation, migration and invasion, and ADSC-Exos promoted skin wound healing via H19 in mice.The high expression of H19 in ADSC-Exos may upregulate SOX9 expression via miR-19b to accelerate wound healing of skin tissues. Our study may provide novel perspectives for therapy to accelerate skin wound healing.

Qian L ，Pi L ，Fang BR ，Meng XX ... -  《-》