Circ-PRKCI Alleviates Lipopolysaccharide-induced Human Kidney 2 Cell Injury by Regulating miR-106b-5p/GAB1 Axis
Yueli Xiong1, Yang Wang2, Hui Tian1
1Department of Infectious Diseases, Huaihe Hospital of Henan University, Kaifeng, Henan, China; and.
Abstract:
Circular RNAs act as vital regulators in diverse diseases. However, the investigation of circular RNAs in sepsis-engendered acute kidney injury remains dismal. We aimed to explore the effects of circular RNA protein kinase C iota (circ-PRKCI) in lipopolysaccharide (LPS)-mediated HK2 cell injury. Sepsis in vitro model was established by LPS treatment. Quantitative real-time polymerase chain reaction assay was conducted for determining the levels of circ-PRKCI, microRNA-106b-5p (miR-106b-5p), and growth factor receptor binding 2-associated binding protein 1 (GAB1). Cell viability and apoptosis were evaluated using Cell Counting Kit-8 assay and flow cytometry analysis, respectively. The concentrations of interleukin-6, interleukin-1β, and tumor necrosis factor-α were measured with enzyme-linked immunosorbent assay kits. The levels of oxidative stress markers were determined using relevant commercial kits. Western blot assay was conducted for B-cell lymphoma-2 (Bcl-2), BCL2-Associated X (Bax), and GAB1 protein levels. Dual-luciferase reporter assay and RNA immunoprecipitation assay were used to verify the association between miR-106b-5p and circ-PRKCI or GAB1. We found the Circ-PRKCI level was decreased in sepsis patients and LPS-induced human kidney 2 (HK-2) cells. LPS exposure inhibited cell viability and facilitated apoptosis, inflammation, and oxidative stress in HK-2 cells. Circ-PRKCI overexpression abrogated the effects of LPS on cell apoptosis, inflammation, and oxidative stress in HK-2 cells. Furthermore, circ-PRKCI was identified as the sponge for miR-106b-5p to positively regulate GAB1 expression. Overexpression of circ-PRKCI relieved LPS-mediated HK-2 cell damage by sponging miR-106b-5p. MiR-106b-5p inhibition ameliorated the injury of HK-2 cells mediated by LPS, whereas GAB1 knockdown reversed the effect. Collectively, Circ-PRKCI overexpression attenuated LPS-induced HK-2 cell injury by regulating miR-106b-5p/GAB1 axis.
Insights
Circular RNA protein kinase C iota (circ-PRKCI) protects against sepsis-induced acute kidney injury. Overexpressing circ-PRKCI in kidney cells reduced damage by regulating the microRNA-106b-5p/GAB1 pathway.
Area of Science:
- Molecular Biology
- Cell Biology
- Pathology
Background:
- Circular RNAs (circRNAs) are crucial in disease pathogenesis.
- The role of circRNAs in sepsis-induced acute kidney injury (AKI) is understudied.
- This study investigates circ-PRKCI in lipopolysaccharide (LPS)-induced kidney cell injury.
Purpose of the Study:
- To explore the function of circ-PRKCI in LPS-mediated human kidney 2 (HK-2) cell injury.
- To elucidate the molecular mechanism involving circ-PRKCI, microRNA-106b-5p (miR-106b-5p), and growth factor receptor binding 2-associated binding protein 1 (GAB1).
Main Methods:
- Established an in vitro sepsis model using LPS treatment on HK-2 cells.
- Quantified circ-PRKCI, miR-106b-5p, and GAB1 levels using quantitative real-time polymerase chain reaction.
- Assessed cell viability, apoptosis, inflammation, oxidative stress, and protein expression via various assays including Western blot and dual-luciferase reporter assays.
Main Results:
- Circ-PRKCI levels were reduced in sepsis patients and LPS-treated HK-2 cells.
- LPS exposure impaired HK-2 cell viability while increasing apoptosis, inflammation, and oxidative stress.
- Circ-PRKCI overexpression counteracted LPS-induced injury by regulating the miR-106b-5p/GAB1 axis, suggesting circ-PRKCI acts as a sponge for miR-106b-5p to upregulate GAB1.
Conclusions:
- Circ-PRKCI plays a protective role in LPS-induced HK-2 cell injury.
- The circ-PRKCI/miR-106b-5p/GAB1 pathway is a potential therapeutic target for sepsis-related AKI.
- Circ-PRKCI overexpression attenuates kidney cell damage by modulating miR-106b-5p and GAB1 expression.
