IDENTIFICATION AND CHARACTERIZATION OF MIR-126-3P AS A DIAGNOSTIC BIOMARKER, MECHANISTIC DRIVER, AND THERAPEUTIC TARGET FOR KNEE OSTEOARTHRITIS

Document Type

Conference Proceeding

Publication Date

4-1-2024

Publication Title

Osteoarthritis Cartilage

Abstract

Purpose (the aim of the study): To reduce the immense burden of disease associated with knee osteoarthritis (OA), there is an urgent need for molecular biomarkers for earlier diagnosis and therapeutic targets for effective intervention. MicroRNAs (miRNAs) are small non-coding RNAs with the potential to meet both these needs given their specificity and stability in liquid biopsies as well as their ability to regulate key biological processes through direct inhibition of gene targets. Found across tissues, miRNAs are expressed as primary transcripts (pri-miRNAs) that are processed into precursor transcripts (pre-miRNAs) then mature miRNAs. To identify important miRNAs in knee OA, we performed secondary analysis of two miRNA-sequencing datasets and discovered circulating miR-126-3p to be elevated compared to non-OA controls, a finding we validated in our Henry Ford Health OA cohort. Existing literature indicates miR-126-3p is primarily expressed by endothelial cells and regulates angiogenesis through inhibition of its gene target SPRED1. In a rat model of OA, exosomes carrying miR-126-3p were shown to attenuate the severity of disease. Based on these findings, the objectives of this study are to assess miR-126-3p as a diagnostic biomarker, mechanistic driver, and therapeutic target for knee OA. Methods: To investigate the biomarker potential of miR-126-3p, we assessed its predictive ability with area under the receiver operating characteristic curve (AUC) analysis and explored correlations with common confounding variables. To investigate the mechanistic potential of miR-126-3p, we collected subchondral bone, infrapatellar fat pad, synovium, anterior cruciate ligament, meniscus and articular cartilage from N=20 OA patients undergoing knee replacement. We measured pri-mir-126, pre-mir-126, and miR-126-3p in each tissue by real-time PCR and quantified miR-126-3p secretion from tissue explants into culture medium over time. We transfected bone, fat pad and synovium explants and cells with 100nM miR-126-3p mimic, inhibitor, or controls and measured candidate gene expression changes. To investigate the therapeutic potential of miR-126-3p, we performed medial meniscectomy or sham surgery in right knees of 12-week-old mice and delivered 4 weekly injections of miR-126-3p mimic, inhibitor, or controls. At 20 weeks, harvested knee joints were assessed by OARSI grading in a blinded manner. Results: Exploring its potential as a biomarker for radiographic knee OA, we found that circulating miR-126-3p showed greater accuracy for predicting knee OA (AUC = 0.83) compared to age, sex, and BMI alone (AUC = 0.52), and importantly showed no association with age, sex, or BMI. Exploring miR-126-3p in knee OA tissues, we found the highest levels in bone, fat pad and synovium versus cartilage which showed the lowest levels (Figure 1). Fat pad had high levels of both pri- and pre-mir-126, and exhibited the highest rate of miR-126-3p secretion over time, suggesting it may be a source of circulating miR-126-3p. Prioritizing bone, fat pad and synovium, we treated explants with miR-126-3p mimic and found reduced expression of OSX and OCN in bone, ADIPOQ and LEP in fat pad, and IL1b, IL6 and TNF in fat pad and synovium; we also found reduced NGF in all three tissues. For canonical functions of miR-126-3p, we found reduced SPRED1 and increased angiogenesis in endothelial cells isolated from bone, fat pad and synovium. Exploring the therapeutic potential of miR-126-3p in a mouse model, treatment with miR-126-3p mimic reduced the severity of knee OA, while inhibitor increased the severity. Conclusions: Our data-driven strategy identified circulating miR-126-3p in radiographic knee OA and further characterization revealed its potential as a candidate diagnostic biomarker. Systematic profiling of primary human knee OA tissues suggests miR-126-3p impacts specific mechanisms in bone, fat pad and synovium, reducing markers of osteogenesis, adipogenesis, inflammation and pain, with a concomitant increase in angiogenesis potentially indicating anabolism. Ta en together, our findings suggest that miR-126-3p may become elevated during OA as a mechanism to mitigate disease processes and therefore should be further evaluated as a therapeutic target for OA, where delivery at earlier stages may prevent progression to later stages. [Formula presented]

Volume

32

First Page

S52-S53

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