| Project/Area Number |
23K07689
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Multi-year Fund |
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| Section | 一般 |
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| Review Section |
Basic Section 53040:Nephrology-related
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| Research Institution | Shonan Kamakura General Hospital, Medical Corporation Tokushukai (Center for Clinical and |
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Principal Investigator |
ヤン ジュンジー 医療法人徳洲会湘南鎌倉総合病院(臨床研究センター), 湘南先端医学研究所 再生医療開発研究部, 主任研究員 (20962137)
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| Co-Investigator(Kenkyū-buntansha) |
浅原 孝之 医療法人徳洲会湘南鎌倉総合病院(臨床研究センター), 湘南先端医学研究所, 副所長 (20246200)
小林 修三 医療法人徳洲会湘南鎌倉総合病院(臨床研究センター), 湘南先端医学研究所, 所長 (60195782)
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| Project Period (FY) |
2023-04-01 – 2026-03-31
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| Project Status |
Granted (Fiscal Year 2023)
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| Budget Amount *help |
¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000)
Fiscal Year 2025: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2024: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2023: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
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| Keywords | acute kidney injury / modified RNA / proximal tubular cells / Pkm2 / miRNA / cell type specificity |
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| Outline of Research at the Start |
Pkm2をPTEC特異的に送達・発現させるPkm2 SMRTsを構築し、modRNAの動態およびPkm2によるPTEC増殖促進作用を確認した後、AKIマウスにてPkm2 SMRTsによるPTEC増殖促進および腎回復促進作用を評価する。腎機能の変化は血中尿素窒素（BUN）と血清クレアチニン比により評価する。
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| Outline of Annual Research Achievements |
We initially identified cell-specific microRNAs in proximal tubular cells and endothelial cells. Our investigation revealed that miR-192, miR-194, and miR-30e are expressed at comparable levels in both proximal tubular cells and endothelial cells, although significantly lower than U6. Subsequently, we synthesized PKM2 and GFP modRNA and confirmed their integrity using a bioanalyzer. We then determined the optimal dose and timing for modRNA transfection, finding that transfection with 3 μg of modRNA and a 48-hour incubation period resulted in the highest protein translation efficiency.
Following this, we established an acute kidney injury (AKI) model and administered PKM2 modRNA to the model. The AKI model involved ischemia/reperfusion of the left kidney, followed by nephrectomy of the right kidney and modRNA injection one week later. Kidney samples were collected three weeks post-surgery, with a survival ratio of 90%. At 48 hours post-modRNA injection, we observed a trend towards recovery in blood urea nitrogen (BUN) and creatinine (Cre) levels with PKM2 mRNA treatment. Additionally, at 3 weeks post-modRNA injection, we observed a trend towards improvement in interstitial fibrosis with PKM2 mRNA treatment.
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| Current Status of Research Progress |
Current Status of Research Progress
2: Research has progressed on the whole more than it was originally planned.
Reason
Our focus is on creating cell type-specific modified RNA and leveraging lipid nanoparticles (LNPs) for mRNA delivery. This approach aims to precisely target specific cell populations and enhance the efficacy of therapeutic interventions. By combining tailored RNA modifications with advanced delivery systems, we aim to revolutionize nucleic acid-based therapeutics and address the challenges in acute kidney injury.
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| Strategy for Future Research Activity |
We will isolate tubular epithelial cells and endothelial cells from kidney samples before and after acute kidney injury by flow cytometry activated-cell sorting, and isolate miRNA from these cells for cell-type specific miRNA identification. Following this, we will synthesize cell-type specific modified RNA for Pkm2 and apply it in vivo in acute kidney injury model.
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