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Wnt5a, a New Diabetic Corneal Marker Related to Wound Healing

Wnt5a，一种与伤口愈合相关的新糖尿病角膜标记物

基本信息

批准号：
10468981
负责人：
Alexander V Ljubimov
金额：
$ 40.5万
依托单位：
CEDARS-SINAI MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-30 至 2025-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10468981
关键词：
Address Adult Affect Age Antineoplastic Agents Binding Blindness Cathepsins Cell Proliferation Cells Complications of Diabetes Mellitus Cornea Corneal Diseases DNA DNA Methylation Defect Deoxycytidine Development Diabetes Mellitus Disease Epigenetic Process Epithelial Epithelial Cells Eye Eye diseases Family Future Gene Abnormality Gene Expression Genes Goals Human Image Impaired wound healing Impairment Iris Keratopathy Lead MAP3K7 gene MAPK4 gene MCAM gene Mediating Memory Metabolic Methods MicroRNAs Molecular Mechanisms of Action Nanoconjugate Nature Neuropathy Normal Cell Organ Organ Culture Techniques Pain Pathway interactions Phenotype Phosphorylation Population Process Proteins Public Health Applications Research ROR1 gene Receptor Signaling Recurrence Research Project Grants Retina Signal Transduction Signaling Molecule Small Interfering RNA Source Surface System Testing Therapeutic Tissues Transplantation Treatment Efficacy Ulcer Viral Vision research Visual impairment WNT5A gene WNT5A protein Zebularine antagonist base bead chip cell motility corneal epithelial stem cells corneal epithelial wound healing corneal epithelium diabetic diabetic patient effective therapy efficacy testing epigenetic regulation epithelial stem cell epithelial wound gene therapy improved inhibitor innovation lens limbal member nano nanodrug nanopolymer nanotherapy novel protein expression receptor restoration single-cell RNA sequencing stem cell biomarkers stem cell population stem cells stromelysin 2 wound wound healing

项目摘要

Diabetes is the most widespread blinding disease in working-age adults. Up to 70% of diabetic patients suffer from corneal problems including neuropathy and epithelial keratopathy (impaired wound healing, ulcers, recurrent erosions) that impair vision and cause pain and discomfort. Diabetic keratopathy is underdiagnosed, and therapy remains symptomatic. We previously identified markers altered in human diabetic corneas and normalized their levels in human corneal organ cultures by gene and nano therapy, which also restored normal stem cell phenotype and corneal wound healing. Epigenetic changes may contribute to diabetic complications. Therefore, we examined epigenetic DNA methylation in human diabetic corneas and found a set of genes abnormally methylated vs. normal corneas. WNT5A gene, a noncanonical member of Wnt regulators of cell motility, proliferation and differentiation, was hypermethylated in diabetic corneas. Its expression was reduced in diabetic corneas and stem cell-enriched epithelial cultures. Wnt5a addition accelerated wound healing in diabetic (but not in normal cells), with stem cell marker expression increase. Inhibition of diabetes-increased miRNA-203a targeting WNT5A increased Wnt5a and wound healing in diabetic cells. miRNA-203a inhibitor and WNT5A siRNA reduced wound healing in normal cells. We will identify mechanisms of action of new diabetic marker Wnt5a, its effects on diabetic corneal cell populations, and normalize its levels in diabetic corneal cells. We hypothesize that normalization of diabetes-impaired stem cell phenotype and epithelial wound healing may be achieved by restoring normal expression of noncanonical Wnt5a by blocking its inhibiting microRNA with a novel nanoconjugate and using demethylating agents. Specific Aim 1. To identify Wnt5a receptor(s) mediating its effects on diabetic epithelial cells and corneas. Wnt5a receptors in diabetic limbal cells and corneas will be identified by imaging and functionally, by siRNAs, focusing first on ROR2. This will allow modulating their expression and signaling in diabetic cells to boost Wnt5a effects. Specific Aim 2. To examine Wnt5a effects on epithelial wound healing and limbal cell populations in human organ-cultured diabetic corneas. In these corneas, we will confirm normalizing Wnt5a effects using wound healing and stem cell marker expression. Changes in limbal epithelial differentiated and stem cell populations upon Wnt5a (or inhibitors) treatment will be examined by single cell RNA-seq vs. control normal cells. Specific Aim 3. To test the efficacy of therapies aimed at increasing Wnt5a expression using cultured diabetic cells and organ-cultured corneas. Diabetic limbal epithelial cells and corneas will be treated with nanoconjugate with miR-203a specific antagonist and/or with demethylating 5-Aza-2′-deoxycytidine (Decitabine) or Zebularine. Our aims fit well the priorities set in the NEI Vision Research: Needs, Gaps, and Opportunities. These priorities include (1) improving the transplantation of cultured corneal epithelial cells, (2) understanding of the epigenetic regulation of wound healing, and (3) developing methods to enhance the wound healing process.

糖尿病是劳动年龄成年人中最普遍的致盲疾病。高达70%的糖尿病患者患有糖尿病