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Published on: December 8, 2021
Identification of differentially expressed genes in human cryptorchid testes using suppression subtractive
Kentaro Mizuno1, Yoshiyuki Kojima, Satoshi Kurokawa
1Department of Nephro-Urology, Nagoya City University Graduate School of Medical Sciences, Nagoya, Aichi, Japan.
Purpose:
To restore fertility the current consensus suggests early orchiopexy for cryptorchidism. However, despite early orchiopexy it is reported that transformation of gonocytes into adult dark spermatogonia is already impaired at the time of surgery and consequently affects future fertility. To elucidate the biological processes occurring during germ cell maturation in the cryptorchid testis, we identified the genes affected by testicular maldescent using polymerase chain reaction based suppression subtractive hybridization, and investigated differentially expressed genes to determine whether they are related to cell differentiation and spermatogenesis.
Materials And Methods:
Testicular tissues were excised from 24 boys 12 to 59 months old who underwent orchiopexy or hydrocelectomy at our hospital. Two-way subtraction was performed to compare their tissue samples and those from age matched boys with ipsilateral cryptorchidism and a descended testis. Differential expression was validated by real-time reverse transcriptase polymerase chain reaction. To clarify the distribution of candidate genes, immunohistochemical and western blot analysis was performed.
Results:
We obtained 84 clones corresponding to transcripts representing differential expression. Basic local alignment search tool searches revealed 32 different known genes with 98% to 100% similarity. Among these genes we further investigated 3 genes, TPT1, EEF1A1 and NuMA1, which were significantly more highly expressed in cryptorchidism than in descended testes and were detected in the spermatogonia from immature to adult testes.
Conclusions:
TPT1, EEF1A1 and NuMA1 have cell growth related functions, suggesting that they have certain roles in germ cell differentiation and maintenance of stem cell potential. Changes in the expression levels of these genes in the testes might enable novel evaluation of spermatogenic failure caused by cryptorchidism.
Insights
Undescended testes (cryptorchidism) impair fertility even after early surgery. This study identified key genes (TPT1, EEF1A1, NuMA1) involved in germ cell development, offering new ways to evaluate fertility issues.
Area of Science:
- Reproductive Biology
- Developmental Biology
- Genetics
Background:
- Cryptorchidism, or undescended testes, is a common congenital condition.
- Early orchiopexy is recommended to restore fertility, but germ cell maturation may already be compromised.
- Understanding molecular changes in cryptorchid testes is crucial for improving fertility outcomes.
Purpose of the Study:
- To identify genes affected by testicular maldescent in cryptorchidism.
- To investigate the role of differentially expressed genes in germ cell differentiation and spermatogenesis.
- To explore potential molecular markers for evaluating spermatogenic failure.
Main Methods:
- Gene expression analysis using polymerase chain reaction (PCR) based suppression subtractive hybridization.
- Validation of differential gene expression via real-time reverse transcriptase PCR.
- Immunohistochemical and Western blot analyses to determine gene distribution.
Main Results:
- Identified 84 differentially expressed transcripts, with 32 known genes.
- Found significantly higher expression of TPT1, EEF1A1, and NuMA1 in cryptorchid testes compared to descended testes.
- Detected these genes in spermatogonia across developmental stages.
Conclusions:
- TPT1, EEF1A1, and NuMA1 are implicated in cell growth, germ cell differentiation, and stem cell maintenance.
- Altered expression of these genes may serve as novel indicators for evaluating spermatogenic failure in cryptorchidism.
