The role of Q10 engineering mesenchymal stem cell-derived exosomes in inhibiting ferroptosis for diabetic wound healing

Abstract

Background: Ferroptosis plays an essential role in the development of diabetes and its complications, suggesting its potential as a therapeutic target. Stem cell-derived extracellular vesicles (EVs) are increasingly being developed as nano-scale drug carriers. The aim of this study was to determine the role of ferroptosis in the pathogenesis of diabetic wound healing and evaluate the therapeutic effects of coenzyme Q10 (Q10)-stimulated exosmes derived from mesenchymal stem cells (MSCs). Methods: Human keratinocytes (HaCaTs) were exposed to high glucose (HG) conditions in vitro to mimic diabetic conditions, and the ferroptosis markers and expression level of acyl-coenzyme A synthase long-chain family member 4 (ACSL4) were determined. Exosomes were isolated from control and Q10-primed umbilical cord mesenchymal stem cells (huMSCs) and characterized by tramsmission electron microscopy and immunofluorescence staining. The HG-treated HaCaTs were cultured in the presence of exosomes derived from Q10-treated huMSCs (Q10-Exo) and their in vitro migratory capacity was analyzed. Results: Q10-Exo significantly improved keratinocyte viability and inhibited ferroptosis in vitro. miR-548ai and miR-660 were upregulated in the Q10-Exo and taken up by HaCaT cells. Furthermore, miR-548ai and miR-660 mimics downregulated ACSL4-inhibited ferroptosis in the HG-treated HaCaT cells and enhanced their proliferation and migration. However, simultaneous upregulation of ACSL4 reversed their effects. Q10-Exo also accelerated diabetic wound healing in a mouse model by inhibiting ACSL4-induced ferroptosis. Conclusions: Q10-Exo promoted the proliferation and migration of keratinocytes and inhibited ferroptosis under hyperglycemic conditions by delivering miR-548ai and miR-660. Q10-Exo also enhanced cutaneous wound healing in diabetic mice by repressing ACSL4-mediated ferroptosis.

Keywords

Ferroptosis / Exosomes / Coenzyme Q10 / ACSL4 / Diabetic wound healing / Keratinocytes / Mesenchymal stem cells / Keratinocyte

References

[1] Singh N, Armstrong DG, Lipsky BA. Preventing foot ulcers in patients with diabetes. JAMA 2005; 293: 217-28. https://doi.org/10.1001/jama.293.2.217.

[2] Shi Y, Wang S, Zhang W, Zhu Y, Fan Z, Huang Y. et al. Bone marrow mesenchymal stem cells facilitate diabetic wound healing through the restoration of epidermal cell autophagy via the HIF-1α/TGF-β1/SMAD pathway. Stem Cell Res Ther 2022; 13: 314. https://doi.org/10.1186/s13287-022-02996-9.

[3] Guan Y, Niu H, Liu Z, Dang Y, Shen J, Zayed M. et al. Sustained oxygenation accelerates diabetic wound healing by promoting epithelialization and angiogenesis and decreasing inflammation. Sci Adv 2021; 7: eabj0153. https://doi.org/10.1126/sciadv.abj0153.

[4] Liu Y, Shi Y, Zhu J, Chen XD, Yang RH, Shu B. et al. Study on the effect of the five-in-one comprehensive limb salvage Technologies of Treating Severe Diabetic Foot. Advances in wound care 2020; 9: 676-85. https://doi.org/10.1089/wound.2018.0903.

[5] Long M, Rojo de la Vega M, Wen Q, Bharara M, Jiang T, Zhang R. et al. An essential role of NRF2 in diabetic wound healing. Diabetes 2016; 65: 780-93. https://doi.org/10.2337/db15-0564.

[6] Fu Y, Liu Y, Song W, Yang D, Wu W, Lin J. et al. Early monitoring-to-warning internet of things system for emerging infectious diseases via networking of light-triggered point-of-care testing devices. Exploration (Beijing) 2023; 3: 20230028. https://doi.org/10.1002/EXP.20230028.

[7] Xiong Y, Feng Q, Lu L, Qiu X, Knoedler S, Panayi AC. et al. Metal-organic frameworks and their composites for chronic wound healing: from bench to bedside. Advanced materials (Deerfield Beach, Fla) 2024; 36: e2302587. https://doi.org/10.1002/adma.202302587.

[8] Qiang L, Yang S, Cui YH, He YY Keratinocyte autophagy enables the activation of keratinocytes and fibroblastsand facilitates wound healing. Autophagy 2021; 17: 2128-43. https://doi.org/10.1080/15548627.2020.1816342.

[9] Hu SC, Lan CE. High-glucose environment disturbs the physiologic functions of keratinocytes: focusing on diabetic wound healing. J Dermatol Sci 2016; 84: 121-7. https://doi.org/10.1016/j.jdermsci.2016.07.008.

[10] Liang D, Lin WJ, Ren M, Qiu J, Yang C, Wang X. et al. M(6)a reader YTHDC1 modulates autophagy by targeting SQSTM1 in diabetic skin. Autophagy 2022; 18: 1318-37. https://doi.org/10.1080/15548627.2021.1974175.

[11] Hu H, Jiang H, Ren H, Hu X, Wang X, Han C AGEs and chronic subclinical inflammation in diabetes: disorders of immune system. Diabetes Metab Res Rev 2015; 31: 127-37. https://doi.org/10.1002/dmrr.2560.

[12] Xiong Y, Chen L, Liu P, Yu T, Lin C, Yan C. et al. All-in-one: multifunctional hydrogel accelerates oxidative diabetic wound healing through timed-release of exosome and fibroblast growth factor. Small (Weinheim an der Bergstrasse, Germany) 2022; 18: e2104229. https://doi.org/10.1002/smll.202104229.

[13] Li X, Wang W, Gao Q, Lai S, Liu Y, Zhou S. et al. Intelligent bacteria-targeting ZIF-8 composite for fluorescence imaging-guided photodynamic therapy of drug-resistant superbug infections and burn wound healing. Exploration 2024; 20230113. https://doi.org/10.1002/EXP.20230113.

[14] Sha W, Hu F, Xi Y, Chu Y, Bu S Mechanism of Ferroptosis and its role in type 2 diabetes mellitus. Journal of diabetes research 2021; 2021: 9999612.

[15] Jiang QW, Kaili D, Freeman J, Lei CY, Geng BC, Tan T. et al. Diabetes inhibits corneal epithelial cell migration and tight junction formation in mice and human via increasing ROS and impairing Akt signaling. Acta Pharmacol Sin 2019; 40: 1205-11. https://doi.org/10.1038/s41401-019-0223-y.

[16] Chen CY, Rao SS, Ren L, Hu XK, Tan YJ, Hu Y. et al. Exosomal DMBT1 from human urine-derived stem cells facilitates diabetic wound repair by promoting angiogenesis. Theranostics 2018; 8: 1607-23. https://doi.org/10.7150/thno.22958.

[17] Xiong Y, Chen L, Yan C, Zhou W, Endo Y, Liu J. et al. Circulating Exosomal miR-20b-5p inhibition restores Wnt9b Signaling and reverses diabetes-associated impaired wound healing. Small 2020; 16: 1904044. https://doi.org/10.1002/smll.201904044.

[18] Liu X, Li X, Wu G, Qi P, Zhang Y, Liu Z. et al. Umbilical cord mesenchymal stem cell-derived small extracellular vesicles deliver miR-21 to promote corneal epithelial wound healing through PTEN/PI3K/Akt pathway. Stem Cells Int 2022; 2022: 1252557.

[19] Zhu J, Liu Z, Wang L, Jin Q, Zhao Y, du A. et al. Exosome Mimetics-loaded hydrogel accelerates wound repair by transferring functional mitochondrial proteins. Front Bioeng Biotechnol 2022; 10: 866505. https://doi.org/10.3389/fbioe.2022.866505.

[20] Zhang Y, Pan Y, Liu Y, Li X, Tang L, Duan M. et al. Exosomes derived from human umbilical cord blood mesenchymal stem cells stimulate regenerative wound healing via transforming growth factor-β receptor inhibition. Stem Cell Res Ther 2021; 12: 434. https://doi.org/10.1186/s13287-021-02517-0.

[21] Fang S, Xu C, Zhang Y, Xue C, Yang C, Bi H. et al. Umbilical cord-derived mesenchymal stem cell-derived Exosomal MicroRNAs suppress Myofibroblast differentiation by inhibiting the transforming growth factor-β/SMAD2 pathway during wound healing. Stem Cells Transl Med 2016; 5: 1425-39. https://doi.org/10.5966/sctm.2015-0367.

[22] Casado-Díaz A, Quesada-Gómez JM, Dorado G. Extracellular vesicles derived from mesenchymal stem cells (MSC) in regenerative medicine: applications in skin wound healing. Front Bioeng Biotechnol 2020; 8: 146. https://doi.org/10.3389/fbioe.2020.00146.

[23] Wei Q, Wang Y, Ma K, Li Q, Li B, Hu W. et al. Extracellular vesicles from human umbilical cord mesenchymal stem cells facilitate diabetic wound healing through MiR-17-5p-mediated enhancement of angiogenesis. Stem Cell Rev Rep 2021; 18: 1025-40. https://doi.org/10.1007/s12015-021-10176-0.

[24] Tao SC, Guo SC, Zhang CQ. Modularized extracellular vesicles: the Dawn of prospective personalized and precision medicine. Advanced science (Weinheim, Baden-Wurttemberg, Germany) 2018; 5: 1700449. https://doi.org/10.1002/advs.201700449.

[25] Liu W, Yu M, Xie D, Wang L, Ye C, Zhu Q. et al. Melatonin-stimulated MSC-derived exosomes improve diabetic wound healing through regulating macrophage M1 and M2 polarization by targeting the PTEN/AKT pathway. Stem Cell Res Ther 2020; 11: 259. https://doi.org/10.1186/s13287-020-01756-x.

[26] Hu Y, Tao R, Chen L, Xiong Y, Xue H, Hu L. et al. Exosomes derived from pioglitazone-pretreated MSCs accelerate diabetic wound healing through enhancing angiogenesis. Journal of nanobiotechnology 2021; 19: 150. https://doi.org/10.1186/s12951-021-00894-5.

[27] Kalluri R, LeBleu VS. The biology, function, and biomedical applications of exosomes. Science (New York, NY) 2020; 367: eaau6977. https://doi.org/10.1126/science.aau6977.

[28] Bo Y, Yang L, Liu B, Tian G, Li C, Zhang L. et al. Exosomes from human induced pluripotent stem cells-derived keratinocytes accelerate burn wound healing through miR-762 mediated promotion of keratinocytes and endothelial cells migration. Journal of nanobiotechnology 2022; 20: 291. https://doi.org/10.1186/s12951-022-01504-8.

[29] Toma MA, Liu Z, Wang Q, Zhang L, Li D, Sommar P. et al. Circular Rna signatures of human healing and non-healing wounds. The Journal of investigative dermatology 2022; 142: 2793-2804.e26. https://doi.org/10.1016/j.jid.2022.03.024.

[30] Yang C, Luo L, Bai X, Shen K, Liu K, Wang J. et al. 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. Arch Biochem Biophys 2020; 681: 108259. https://doi.org/10.1016/j.abb.2020.108259.

[31] Ding J, Wang X, Chen B, Zhang J, Xu J Exosomes derived from human bone marrow mesenchymal stem cells stimulated by Deferoxamine accelerate cutaneous wound healing by promoting angiogenesis. Biomed Res Int 2019; 2019: 9742765. https://doi.org/10.1155/2019/9742765.

[32] Huang P, Wang L, Li Q, Tian X, Xu J, Xu J. et al. Atorvastatin enhances the therapeutic efficacy of mesenchymal stem cells-derived exosomes in acute myocardial infarction via up-regulating long non-coding RNA H19. Cardiovasc Res 2020; 116: 353-67. https://doi.org/10.1093/cvr/cvz139.

[33] Ogawa M, Udono M, Teruya K, Uehara N, Katakura Y Exosomes derived from Fisetin-treated keratinocytes mediate hair growth promotion. Nutrients 2021; 13: 2087. https://doi.org/10.3390/nu13062087.

[34] Liao Z, Li S, Lu S, Liu H, Li G, Ma L. et al. Metformin facilitates mesenchymal stem cell-derived extracellular nanovesicles release and optimizes therapeutic efficacy in intervertebral disc degeneration. Biomaterials 2021; 274: 120850. https://doi.org/10.1016/j.biomaterials.2021.120850.

[35] Zhang D, Yan B, Yu S, Zhang C, Wang B, Wang Y. et al. Coenzyme Q10 inhibits the aging of mesenchymal stem cells induced by D-galactose through Akt/mTOR signaling. Oxidative Med Cell Longev 2015; 2015: 867293. https://doi.org/10.1155/2015/867293.

[36] Duan M, Zhang Y, Zhang H, Meng Y, Qian M, Zhang G Epidermal stem cell-derived exosomes promote skin regeneration by downregulating transforming growth factor-β1 in wound healing. Stem Cell Res Ther 2020; 11: 452. https://doi.org/10.1186/s13287-020-01971-6.

[37] Cui S, Liu X, Liu Y, Hu W, Ma K, Huang Q. et al. Autophagosomes defeat Ferroptosis by decreasing generation and increasing discharge of free Fe(2+) in skin repair cells to accelerate diabetic wound healing. Advanced science (Weinheim, Baden-Wurttemberg, Germany) 2023; 10: e2300414. https://doi.org/10.1002/advs.202300414.

[38] Feng H, Liu Q, Deng Z, Li H, Zhang H, Song J. et al. Human umbilical cord mesenchymal stem cells ameliorate erectile dysfunction in rats with diabetes mellitus through the attenuation of ferroptosis. Stem Cell Res Ther 2022; 13: 450. https://doi.org/10.1186/s13287-022-03147-w.

[39] Doll S, Proneth B, Tyurina YY, Panzilius E, Kobayashi S, Ingold I. et al. ACSL4 dictates ferroptosis sensitivity by shaping cellular lipid composition. Nat Chem Biol 2017; 13: 91-8. https://doi.org/10.1038/nchembio.2239.

[40] Sawaya AP, Stone RC, Brooks SR, Pastar I, Jozic I, Hasneen K. et al. Deregulated immune cell recruitment orchestrated by FOXM1 impairs human diabetic wound healing. Nat Commun 2020; 11: 4678. https://doi.org/10.1038/s41467-020-18276-0.

[41] Sawaya AP, Stone RC, Mehdizadeh S, Pastar I, Worrell S, Balukoff NC. et al. FOXM1 network in association with TREM1 suppression regulates NET formation in diabetic foot ulcers. EMBO Rep 2022; 23: e54558. https://doi.org/10.15252/embr.202154558.

[42] Iglesias-Bartolome R, Uchiyama A, Molinolo AA, Abusleme L, Brooks SR, Callejas-Valera JL. et al. Transcriptional signature primes human oral mucosa for rapid wound healing. Sci Transl Med 2018; 10: eaap8798. https://doi.org/10.1126/scitranslmed.aap8798.

[43] Yang R, Huang H, Cui S, Zhou Y, Zhang T, Zhou Y IFN-γ promoted exosomes from mesenchymal stem cells to attenuate colitis via miR-125a and miR-125b. Cell Death Dis 2020; 11: 603. https://doi.org/10.1038/s41419-020-02788-0.

[44] Feng J, Wang J, Wang Y, Huang X, Shao T, Deng X. et al. Oxidative stress and lipid peroxidation: prospective associations between Ferroptosis and delayed wound healing in diabetic ulcers. Front Cell Dev Biol 2022; 10: 898657. https://doi.org/10.3389/fcell.2022.898657.

[45] Xiong Y, Lin Z, Bu P, Yu T, Endo Y, Zhou W. et al. A whole-course-repair system based on neurogenesis-angiogenesis crosstalk and macrophage reprogramming promotes diabetic wound healing. Advanced materials (Deerfield Beach, Fla) 2023; 35: e2212300. https://doi.org/10.1002/adma.202212300.

[46] Allen RM, Vickers KC. Coenzyme Q10 increases cholesterol efflux and inhibits atherosclerosis through microRNAs. Arterioscler Thromb Vasc Biol 2014; 34: 1795-7. https://doi.org/10.1161/ATVBAHA.114.303741.

[47] Song Y, Wang B, Zhu X, Hu J, Sun J, Xuan J. et al. Human umbilical cord blood-derived MSCs exosome attenuate myocardial injury by inhibiting ferroptosis in acute myocardial infarction mice. Cell Biol Toxicol 2021; 37: 51-64. https://doi.org/10.1007/s10565-020-09530-8.