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MicroRNA-339-5p inhibits lipopolysaccharide-induced rat mesangial cells by regulating the Syk/Ras/c-Fos pathway
Jia Rong Gao1,2, Miao Miao Shi3, Hui Jiang4
1Department of Pharmacy, The First Affiliated Hospital of Anhui University of Chinese Medicine, Hefei, 230031, Anhui, China. 2020205219007@stu.ahtcm.edu.cn.
Abstract:
Chronic glomerulonephritis (CGN) is a disease occurred in glomeruli. The mechanism of CGN is regarded to be involved in a range of inflammatory responses. MicroRNA-339-5p (miR-339-5p) has been reported to be involved in inflammatory responses in many diseases. However, the role of miR-339-5p in CGN remains unclear. The purpose of this study was to investigate the role of miR-339-5p in lipopolysaccharide (LPS)-induced nephritis injury in vitro. The real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) and western blot (WB) were used to detect the expression of miR-339-5p and Syk/Ras/c-Fos pathway. Double luciferase was performed to identify targeted binding of miR-339-5p to Syk. Cell counting kit-8 (CCK-8) and flow cytometry were used to observe cell viability and cell cycle. Enzyme-linked immunosorbent assay (ELISA) was performed to measure the concentrations of inflammatory cytokines IL-1β, IL-10, IL-6, and TNF-α. Lipopolysaccharide (LPS) could increase HBZY-1 (rat mesangial cells) cell viability, decrease the G2 phase, and promote cell proliferation and accelerate inflammatory cytokine. However, overexpression of miR-339-5p could inhibit LPS-induced HBZY-1 cell viability, decrease the expression of Syk/Ras/c-Fos signaling pathway, downregulate the expression level of inflammatory cytokines, increase the G2 phase, and inhibit cell proliferation. miR-339-5p could inhibit the proliferation and inflammation of the rat mesangial cells through regulating Syk/Ras/c-Fos signaling pathway.
Insights
MicroRNA-339-5p (miR-339-5p) inhibits inflammation and proliferation in rat mesangial cells during nephritis. Overexpression of miR-339-5p suppresses the Syk/Ras/c-Fos pathway, reducing inflammatory cytokine release.
Area of Science:
- Nephrology
- Molecular Biology
- Immunology
Background:
- Chronic glomerulonephritis (CGN) involves complex inflammatory responses.
- MicroRNA-339-5p (miR-339-5p) is implicated in various inflammatory conditions.
- The specific role of miR-339-5p in CGN pathogenesis is not well understood.
Purpose of the Study:
- To investigate the function of miR-339-5p in lipopolysaccharide (LPS)-induced nephritis in rat mesangial cells (HBZY-1).
- To elucidate the molecular mechanisms underlying miR-339-5p's effects on inflammation and cell proliferation.
Main Methods:
- Real-time RT-qPCR and Western blot to assess miR-339-5p and Syk/Ras/c-Fos pathway expression.
- Double luciferase assay to confirm direct targeting of Syk by miR-339-5p.
- Cell Counting Kit-8, flow cytometry, and ELISA to evaluate cell viability, cell cycle, and inflammatory cytokine levels (IL-1β, IL-10, IL-6, TNF-α).
Main Results:
- LPS treatment increased HBZY-1 cell viability, proliferation, and inflammatory cytokine production while altering cell cycle progression.
- Overexpression of miR-339-5p counteracted LPS effects, inhibiting cell viability and proliferation.
- miR-339-5p significantly downregulated the Syk/Ras/c-Fos signaling pathway and reduced inflammatory cytokine levels.
Conclusions:
- miR-339-5p plays a protective role in LPS-induced nephritis.
- The inhibitory effects of miR-339-5p on cell proliferation and inflammation are mediated through the regulation of the Syk/Ras/c-Fos signaling pathway.
