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Published on: December 31, 2015
Molecular Cloning of Genes Related to Apoptosis in Spermatogenic Cells of Mouse
Hong Jiang1, Lu-Yun Li, Guang-Xiu Lu
1Human Reproductive Engineering Research Institute, Hunan Medical University, Changsha 410078, China.
Abstract:
An apoptotic model of spermatogenic cells of mouse has been established by the method of operative cryptorchidism. By using light microscope and electronmicroscope, as well as TUNEL-stain (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling method), it had been confirmed that operative cryptorchidism could successfully induce apoptosis of spermatogenic cells of mouse.On the basis of this research, 24 ESTs (expressed sequence tags) of the genes related to apoptosis in spermatogenic cells have been isolated and cloned by the method of suppression subtractive hybridization(SSH). After being confirmed with PCR and digestion by restriction enzyme PstI, they were sequenced and searched in GenBank for homology analysis. All of those were novel ESTs and were banked into GenBank with accession numbers. With the probes derived from three arbitrary cDNA sequences, Northern hybridization confirmed that there was obvious differential expression between cryptorchid testis and contralateral testis. These results suggest that experimental cryptorchidism is a suitable model for studying gene expression in the process of hypospermatogenesis and spermatogenic cell apoptosis, and ESTs isolated from this work may be involved in this apoptosis.
Insights
Researchers established a mouse model of spermatogenic cell apoptosis using cryptorchidism. This study identified 24 novel expressed sequence tags (ESTs) potentially involved in germ cell apoptosis and hypospermatogenesis.
Area of Science:
- Reproductive biology
- Molecular genetics
- Cellular biology
Background:
- Cryptorchidism, or undescended testes, is associated with impaired spermatogenesis and germ cell apoptosis.
- Understanding the molecular mechanisms underlying germ cell apoptosis is crucial for addressing male infertility.
Purpose of the Study:
- To establish a reliable animal model for studying spermatogenic cell apoptosis.
- To identify novel genes involved in germ cell apoptosis and hypospermatogenesis in mice.
Main Methods:
- Operative cryptorchidism was used to induce apoptosis in mouse spermatogenic cells.
- TUNEL staining, light and electron microscopy confirmed apoptosis.
- Suppression subtractive hybridization (SSH) was employed to isolate expressed sequence tags (ESTs).
- PCR, restriction enzyme digestion, sequencing, GenBank homology analysis, and Northern hybridization were performed.
Main Results:
- Operative cryptorchidism successfully induced apoptosis of spermatogenic cells.
- 24 novel ESTs related to apoptosis in spermatogenic cells were identified and sequenced.
- Northern hybridization revealed differential gene expression between cryptorchid and contralateral testes.
Conclusions:
- Experimental cryptorchidism serves as a suitable model for investigating gene expression during hypospermatogenesis and spermatogenic cell apoptosis.
- The identified ESTs represent potential candidates involved in the apoptotic process of spermatogenic cells.
