Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides
© Copyright 2019, Mary Ann Liebert, Inc., publishers 2019. In diabetes-associated chronic wounds, the normal response to hypoxia is impaired and many cellular processes involved in wound healing are hindered. Central to the hypoxia response is hypoxia-inducible factor-1α (HIF-1α), which activates mu...
| Main Authors: | , , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Mary Ann Liebert Inc
2021
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| Online Access: | https://hdl.handle.net/1721.1/135853 |
| _version_ | 1826200937543237632 |
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| author | Dallas, Anne Trotsyuk, Artem Ilves, Heini Bonham, Clark A Rodrigues, Melanie Engel, Karl Barrera, Janos A Kosaric, Nina Stern-Buchbinder, Zachary A White, Aleksandr Mandell, Kenneth J Hammond, Paula T Mansbridge, Jonathan Jayasena, Sumedha Gurtner, Geoffrey C Johnston, Brian H |
| author_facet | Dallas, Anne Trotsyuk, Artem Ilves, Heini Bonham, Clark A Rodrigues, Melanie Engel, Karl Barrera, Janos A Kosaric, Nina Stern-Buchbinder, Zachary A White, Aleksandr Mandell, Kenneth J Hammond, Paula T Mansbridge, Jonathan Jayasena, Sumedha Gurtner, Geoffrey C Johnston, Brian H |
| author_sort | Dallas, Anne |
| collection | MIT |
| description | © Copyright 2019, Mary Ann Liebert, Inc., publishers 2019. In diabetes-associated chronic wounds, the normal response to hypoxia is impaired and many cellular processes involved in wound healing are hindered. Central to the hypoxia response is hypoxia-inducible factor-1α (HIF-1α), which activates multiple factors that enhance wound healing by promoting cellular motility and proliferation, new vessel formation, and re-epithelialization. Prolyl hydroxylase domain-containing protein 2 (PHD2) regulates HIF-1α activity by targeting it for degradation under normoxia. HIF-1α also upregulates microRNA miR-210, which in turn regulates proteins involved in cell cycle control, DNA repair, and mitochondrial respiration in ways that are antagonistic to wound repair. We have identified a highly potent short synthetic hairpin RNA (sshRNA) that inhibits expression of PHD2 and an antisense oligonucleotide (antimiR) that inhibits miR-210. Both oligonucleotides were chemically modified for improved biostability and to mitigate potential immunostimulatory effects. Using the sshRNA to silence PHD2 transcripts stabilizes HIF-1α and, in combination with the antimiR targeting miR-210, increases proliferation and migration of keratinocytes in vitro. To assess activity and delivery in an impaired wound healing model in diabetic mice, PHD2-targeting sshRNAs and miR-210 antimiRs both alone and in combination were formulated for local delivery to wounds using layer-by-layer (LbL) technology. LbL nanofabrication was applied to incorporate sshRNA into a thin polymer coating on a Tegaderm mesh. This coating gradually degrades under physiological conditions, releasing sshRNA and antimiR for sustained cellular uptake. Formulated treatments were applied directly to splinted full-thickness excisional wounds in db/db mice. Cellular uptake was confirmed using fluorescent sshRNA. Wounds treated with a single application of PHD2 sshRNA or antimiR-210 closed 4 days faster than untreated wounds, and wounds treated with both oligonucleotides closed on average 4.75 days faster. Markers for neovascularization and cell proliferation (CD31 and Ki67, respectively) were increased in the wound area following treatment, and vascular endothelial growth factor (VEGF) was increased in sshRNA-treated wounds. Our results suggest that silencing of PHD2 and miR-210 either together or separately by localized delivery of sshRNAs and antimiRs is a promising approach for the treatment of chronic wounds, with the potential for rapid clinical translation. |
| first_indexed | 2024-09-23T11:43:45Z |
| format | Article |
| id | mit-1721.1/135853 |
| institution | Massachusetts Institute of Technology |
| language | English |
| last_indexed | 2024-09-23T11:43:45Z |
| publishDate | 2021 |
| publisher | Mary Ann Liebert Inc |
| record_format | dspace |
| spelling | mit-1721.1/1358532022-03-30T14:26:24Z Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides Dallas, Anne Trotsyuk, Artem Ilves, Heini Bonham, Clark A Rodrigues, Melanie Engel, Karl Barrera, Janos A Kosaric, Nina Stern-Buchbinder, Zachary A White, Aleksandr Mandell, Kenneth J Hammond, Paula T Mansbridge, Jonathan Jayasena, Sumedha Gurtner, Geoffrey C Johnston, Brian H © Copyright 2019, Mary Ann Liebert, Inc., publishers 2019. In diabetes-associated chronic wounds, the normal response to hypoxia is impaired and many cellular processes involved in wound healing are hindered. Central to the hypoxia response is hypoxia-inducible factor-1α (HIF-1α), which activates multiple factors that enhance wound healing by promoting cellular motility and proliferation, new vessel formation, and re-epithelialization. Prolyl hydroxylase domain-containing protein 2 (PHD2) regulates HIF-1α activity by targeting it for degradation under normoxia. HIF-1α also upregulates microRNA miR-210, which in turn regulates proteins involved in cell cycle control, DNA repair, and mitochondrial respiration in ways that are antagonistic to wound repair. We have identified a highly potent short synthetic hairpin RNA (sshRNA) that inhibits expression of PHD2 and an antisense oligonucleotide (antimiR) that inhibits miR-210. Both oligonucleotides were chemically modified for improved biostability and to mitigate potential immunostimulatory effects. Using the sshRNA to silence PHD2 transcripts stabilizes HIF-1α and, in combination with the antimiR targeting miR-210, increases proliferation and migration of keratinocytes in vitro. To assess activity and delivery in an impaired wound healing model in diabetic mice, PHD2-targeting sshRNAs and miR-210 antimiRs both alone and in combination were formulated for local delivery to wounds using layer-by-layer (LbL) technology. LbL nanofabrication was applied to incorporate sshRNA into a thin polymer coating on a Tegaderm mesh. This coating gradually degrades under physiological conditions, releasing sshRNA and antimiR for sustained cellular uptake. Formulated treatments were applied directly to splinted full-thickness excisional wounds in db/db mice. Cellular uptake was confirmed using fluorescent sshRNA. Wounds treated with a single application of PHD2 sshRNA or antimiR-210 closed 4 days faster than untreated wounds, and wounds treated with both oligonucleotides closed on average 4.75 days faster. Markers for neovascularization and cell proliferation (CD31 and Ki67, respectively) were increased in the wound area following treatment, and vascular endothelial growth factor (VEGF) was increased in sshRNA-treated wounds. Our results suggest that silencing of PHD2 and miR-210 either together or separately by localized delivery of sshRNAs and antimiRs is a promising approach for the treatment of chronic wounds, with the potential for rapid clinical translation. 2021-10-27T20:29:38Z 2021-10-27T20:29:38Z 2019 2019-08-20T18:11:15Z Article http://purl.org/eprint/type/JournalArticle https://hdl.handle.net/1721.1/135853 en 10.1089/TEN.TEA.2017.0484 Tissue Engineering Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf Mary Ann Liebert Inc Mary Ann Liebert |
| spellingShingle | Dallas, Anne Trotsyuk, Artem Ilves, Heini Bonham, Clark A Rodrigues, Melanie Engel, Karl Barrera, Janos A Kosaric, Nina Stern-Buchbinder, Zachary A White, Aleksandr Mandell, Kenneth J Hammond, Paula T Mansbridge, Jonathan Jayasena, Sumedha Gurtner, Geoffrey C Johnston, Brian H Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides |
| title | Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides |
| title_full | Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides |
| title_fullStr | Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides |
| title_full_unstemmed | Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides |
| title_short | Acceleration of Diabetic Wound Healing with PHD2- and miR-210-targeting Oligonucleotides |
| title_sort | acceleration of diabetic wound healing with phd2 and mir 210 targeting oligonucleotides |
| url | https://hdl.handle.net/1721.1/135853 |
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