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Novel immunotherapeutic effects of topically administered ripasudil (K-115) on corneal allograft survival.

ABSTRACT: Corneal allograft survival is mediated by the variety of immunological reactions and wound healing process. Our aim was to explore the effects of topical administration of ripasudil, a selective Rho-associated coiled-coil protein kinase inhibitor, on corneal allograft survival. Ripasudil was administered to mice thrice a day after allogeneic corneal transplantation. Corneal graft survival, opacity, neovascularization, re-epithelization, immune cell infiltration, and mRNA levels of angiogenic and pro-inflammatory factors in the grafted cornea and draining lymph nodes (dLNs) were evaluated with slit-lamp microscopy, immunohistochemistry, flow cytometry, and polymerase chain reaction. Graft survival was significantly prolonged with lower graft opacity and neovascularization scores in 0.4% and 2.0% ripasudil-treated groups, and mRNA levels of angiogenic and pro-inflammatory factors in ripasudil-treated grafted corneas were reduced. Moreover, 0.4% and 2.0% ripasudil reduced CD45+-infiltrated leukocyte frequency, Cd11b and Cd11c mRNA levels, and the frequencies of mature dendritic cells, IFN?-, and IL-17- producing CD4+T cells in the dLNs of recipients. Re-epithelization rate of the grafted cornea was significantly higher in the 0.4% and 2.0% ripasudil groups than in the control. Topically applied ripasudil prolonged graft survival by downregulating neovascularization and inflammation factors, while promoting corneal re-epithelization, suggesting that ripasudil may be useful for suppressing immunological rejection in corneal transplantation.

SUBMITTER: Inomata T

PROVIDER: S-EPMC7666179 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Novel immunotherapeutic effects of topically administered ripasudil (K-115) on corneal allograft survival.

Inomata Takenori T Fujimoto Keiichi K Okumura Yuichi Y Zhu Jun J Fujio Kenta K Shokirova Hurramhon H Miura Maria M Okano Mikiko M Funaki Toshinari T Sung Jaemyoung J Negishi Naoko N Murakami Akira A

Scientific reports 20201113 1

Corneal allograft survival is mediated by the variety of immunological reactions and wound healing process. Our aim was to explore the effects of topical administration of ripasudil, a selective Rho-associated coiled-coil protein kinase inhibitor, on corneal allograft survival. Ripasudil was administered to mice thrice a day after allogeneic corneal transplantation. Corneal graft survival, opacity, neovascularization, re-epithelization, immune cell infiltration, and mRNA levels of angiogenic and ...[more]

PMID: 33188243

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Project description:Rationale: Corneal transplantation is an effective treatment to corneal blindness. However, the immune rejection imperils corneal allograft survival. An interventional modality is urgently needed to inhibit immune rejection and promote allograft survival. In our previous study, subconjunctival injections of bone marrow-derived mesenchymal stem cells (BM-MSCs) into a rat model of corneal allograft rejection extended allograft survival for 2 d. In this study, we sought to generate IL-10-overexpressing BM-MSCs, aiming to boost the survival-promoting effects of BM-MSCs on corneal allografts and explore the molecular and cellular mechanisms underlying augmented protection. Methods: A population of IL-10-overexpressing BM-MSCs (designated as IL-10-BM-MSCs) were generated by lentivirus transduction and FACS purification. The self-renewal, multi-differentiation, and immunoinhibitory capabilities of IL-10-BM-MSCs were examined by conventional assays. The IL-10-BM-MSCs were subconjunctivally injected into the model of corneal allograft rejection, and the allografts were monitored on a daily basis. The expression profiling of long noncoding RNA (lncRNA) in the allografts was revealed by RNA sequencing and verified by quantitative real-time PCR. The infiltrating immune cell type predominantly upregulating the lncRNA expression was identified by RNAscope in situ hybridization. The function of the upregulated lncRNA was proved by loss- and gain-of-function experiments both in vivo and in vitro. Results: The IL-10-BM-MSCs possessed an enhanced immunoinhibitory capability and unabated self-renewal and multi-differentiation potentials as compared to plain BM-MSCs. The subconjunctivally injected IL-10-BM-MSCs reduced immune cell infiltration and doubled allograft survival time (20 d) as compared to IL-10 protein or plain BM-MSCs in the corneal allograft rejection model. Further, IL-10-BM-MSCs significantly upregulated lncRNA 003946 expression in CD68+ macrophages infiltrating corneal allografts. Silencing and overexpressing lncRNA 003946 in macrophage cultures abolished and mimicked the IL-10-BM-MSCs' suppressing effects on the macrophages' antigen presentation, respectively. In parallel, knocking down and overexpressing the lncRNA in vivo abrogated and simulated the survival-promoting effects of IL-10-BM-MSCs on corneal allografts, respectively. Conclusion: The remarkable protective effects of IL-10-BM-MSCs support further developing them into an effective interventional modality against corneal allograft rejection. IL-10-BM-MSCs promote corneal allograft survival mainly through upregulating a novel lncRNA expression in graft-infiltrating CD68+ macrophages. LncRNA, for the first time, is integrated into an IL-10-BM-MSC-driven immunomodulatory axis against the immune rejection to corneal allograft.

Project description:PurposeTo assess whether topical application of VEGFR1R2 Trap after corneal transplantation can impair corneal (lymph)angiogenesis and promote murine corneal allograft survival in eyes at high risk of rejection.MethodsWe used the murine model of suture-induced neovascularization and subsequent keratoplasty in eyes at high risk of rejection, which is an established model for local drug application. After transplantation, the mice were treated with either VEGFR1R2 Trap (aflibercept) or human IgG Fc as eye drops for 2 weeks (three times/d). Deposition of VEGFR1R2 Trap in corneal tissue was detected by immunohistochemistry. Two and 8 weeks after transplantation, corneal (lymph)angiogenesis was assessed morphometrically. Dendritic cells (DCs) and regulatory T cells (Tregs) in the draining lymph nodes (dLNs) were examined by flow cytometry. Allograft survival was determined by corneal graft opacity scores.ResultsTopically applied VEGFR1R2 Trap penetrated into corneal host and graft stroma after keratoplasty in eyes at high risk of rejection. Additional postsurgical corneal hemangiogenesis (P < 0.0001) and lymphangiogenesis (P < 0.01) as well as infiltrating CD45+ leukocytes (P < 0.001) and macrophages (P < 0.01) were significantly reduced in the VEGFR1R2 Trap group compared to controls. VEGFR1R2 Trap eye drops significantly decreased the frequency of total CD11c+ DCs (P < 0.01), as well as activated CD11c+MHC II+ DCs (P < 0.01) and CD11c+CD40+ DCs (P < 0.05). In contrast, the frequency of CD200R+ regulatory DCs (P < 0.05) and Tregs in dLNs (P < 0.01) was enhanced. Moreover, long-term allograft survival was also improved (P < 0.05).ConclusionsTemporary, topical application of VEGFR1R2 Trap after corneal transplantation can achieve sufficient anti-VEGF activity, inhibit additional (lymph)angiogenesis, and significantly improve corneal allograft survival in eyes at high risk of rejection.Translational relevanceVEGFR1R2 Trap eye drops after transplantation present a new therapeutic option for patients undergoing corneal transplantation and are at high risk of graft rejection.

Dataset Information

Novel immunotherapeutic effects of topically administered ripasudil (K-115) on corneal allograft survival.

Publications

Novel immunotherapeutic effects of topically administered ripasudil (K-115) on corneal allograft survival.

Similar Datasets

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