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Published on: May 10, 2019
Downregulation of cold-inducible RNA-binding protein activates mitogen-activated protein kinases and impairs
Zhi-Ping Xia1, Xin-Min Zheng, Hang Zheng
1Department of Urology, Zhongnan Hospital of Wuhan University, Wuhan 430071, China.
Abstract:
Cold-inducible RNA-binding protein (CIRP) is an RNA-binding protein that is expressed in normal testes and downregulated after heat stress caused by cryptorchidism, varicocele or environmental temperatures. The purpose of this study was to investigate the functions of CIRP in the testes. We employed RNAi technique to knock down the expression of CIRP in the testes, and performed haematoxylin and eosin staining to evaluate morphological changes following knockdown. Germ cell apoptosis was examined by terminal deoxynucleotidal transferase-mediated dUTP nick end labelling (TUNEL) assay, and mitogen-activated protein kinase (MAPK) signalling pathways were investigated by Western blotting to determine the possible mechanism of apoptosis. We found that using siRNA is a feasible and reliable method for knocking down gene expression in the testes. Compared to controls, the mean seminiferous tubule diameter (MSTD) and the thickness of the germ cell layers decreased following siRNA treatment, whereas the percentage of apoptotic seminiferous tubules increased. The p44/p42, p38 and SAPK/JNK MAPK pathways were activated after downregulation of CIRP. In conclusion, we discovered that downregulation of CIRP resulted in increased germ cell apoptosis, possibly via the activation of the p44/p42, p38 and SAPK/JNK MAPK pathways.
Insights
Downregulating cold-inducible RNA-binding protein (CIRP) in testes increases germ cell apoptosis. This occurs possibly through activating p44/p42, p38, and SAPK/JNK mitogen-activated protein kinase (MAPK) pathways.
Area of Science:
- Reproductive Biology
- Molecular Biology
- Cell Biology
Background:
- Cold-inducible RNA-binding protein (CIRP) is crucial in testes, downregulated by heat stress.
- Understanding CIRP's role is vital for male reproductive health.
Purpose of the Study:
- Investigate CIRP's function in testicular physiology.
- Determine the impact of CIRP downregulation on germ cells and associated signaling pathways.
Main Methods:
- Utilized RNA interference (RNAi) via siRNA to reduce CIRP expression in testes.
- Performed histological analysis (H&E staining) and TUNEL assay for apoptosis assessment.
- Analyzed mitogen-activated protein kinase (MAPK) pathway activation using Western blotting.
Main Results:
- siRNA effectively knocked down CIRP expression in testes.
- CIRP downregulation led to decreased seminiferous tubule diameter and germ cell layer thickness.
- Increased germ cell apoptosis and activation of p44/p42, p38, and SAPK/JNK MAPK pathways were observed.
Conclusions:
- CIRP downregulation significantly increases germ cell apoptosis in testes.
- The observed apoptosis is potentially mediated by the activation of p44/p42, p38, and SAPK/JNK MAPK pathways.
- RNAi is a viable method for studying gene function in the testes.
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