miRNA-382-5p Carried by Extracellular Vesicles in Osteoarthritis Reduces Cell Viability and Proliferation, and
Hanyu Lu1, Yixin Yang1, Shuanji Ou1
1Department of Orthopedics, Guangdong Second Provincial General Hospital, Guangzhou, People's Republic of China.
Abstract:
The objective of the study was to identify extracellular vesicle (EV) microRNAs (miRNAs) that play important roles in knee osteoarthritis (OA). Models of knee OA were surgically induced in nine male Sprague-Dawley rats. Tissue samples were collected at 0 weeks (Control), 6 weeks (6 weeks), and 12 weeks (12 weeks). The EVs were isolated and analyzed for size. Various biomarkers, including recombinant tetraspanin 30 cluster of differentiation (CD)63 and CD9 were detected. An Agilent array was used to screen for differentially expressed (DE) miRNAs. The levels of DE miRNAs and their target mRNAs were evaluated by quantitative reverse transcription-polymerase chain reaction and western blotting. The viability, proliferation, and apoptosis of lipopolysaccharide (LPS)-induced human synovial cells (HSCs) were examined by using Cell Counting Kit-8, EdU (5-ethynyl-2'-deoxyuridine), and TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling) assays, respectively. The OA model rats had significantly increased levels of inflammatory activity, damaged cells, and rough articular cartilage when compared with rats in the control group. The EVs from the model rats appeared as round vesicle-like structures with a mean diameter of ∼145 nm. Five miRNAs that showed gradual increases in the model rats were selected for further analysis; those miRNAs included miR-127-3p, miR-132-3p, miR-141-3p, miR-345-5p, and miR-382-5p. miR-382-5p was found to reduce the viability and proliferation and promote the apoptosis of LPS-induced HSCs. Moreover, phosphatase and tensin homolog deleted on chromosome 10 (PTEN) was negatively regulated by miR-382-5p. Our findings revealed that EVs produced by the OA rats contained miR-382-5p, which might reduce cell viability and proliferation, and promote cell apoptosis by targeting PTEN.
Insights
Extracellular vesicles (EVs) from knee osteoarthritis (OA) rat models contain miR-382-5p. This microRNA (miRNA) may reduce cell viability and promote apoptosis by targeting PTEN, offering insights into OA pathogenesis.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Knee osteoarthritis (OA) is a degenerative joint disease characterized by cartilage breakdown and inflammation.
- Extracellular vesicles (EVs) are emerging as key mediators of intercellular communication in various pathological conditions, including OA.
- Identifying specific microRNAs (miRNAs) within EVs involved in OA pathogenesis is crucial for understanding disease mechanisms.
Purpose of the Study:
- To identify specific extracellular vesicle (EV)-derived microRNAs (miRNAs) implicated in the development of knee osteoarthritis (OA).
- To investigate the functional role of identified miRNAs in regulating cellular processes relevant to OA.
- To explore the molecular targets of these miRNAs in the context of OA.
Main Methods:
- Surgically induced knee OA model in Sprague-Dawley rats.
- Isolation and characterization of EVs from tissue samples.
- Screening for differentially expressed miRNAs using Agilent arrays.
- Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and western blotting to evaluate miRNA and mRNA levels.
- In vitro assays (Cell Counting Kit-8, EdU, TUNEL) to assess human synovial cell viability, proliferation, and apoptosis.
Main Results:
- OA model rats exhibited increased inflammatory activity, cellular damage, and articular cartilage degradation compared to controls.
- Isolated EVs displayed characteristics of vesicles with a mean diameter of approximately 145 nm.
- Five miRNAs (miR-127-3p, miR-132-3p, miR-141-3p, miR-345-5p, and miR-382-5p) showed increased levels in OA model rats.
- miR-382-5p significantly reduced viability and proliferation while promoting apoptosis in lipopolysaccharide (LPS)-induced human synovial cells (HSCs).
- miR-382-5p was found to negatively regulate phosphatase and tensin homolog deleted on chromosome 10 (PTEN).
Conclusions:
- Extracellular vesicles derived from OA rat models contain elevated levels of specific miRNAs, including miR-382-5p.
- miR-382-5p plays a pro-apoptotic and anti-proliferative role in human synovial cells, potentially contributing to OA pathogenesis.
- The targeting of PTEN by miR-382-5p represents a key molecular mechanism underlying its effects in OA.
- These findings highlight the potential of EV-derived miRNAs as biomarkers and therapeutic targets for knee OA.
