Superfund Research Program

Molecular Biomarkers for Assessing Testicular Toxicity

Project Leader: Kim Boekelheide
Grant Number: P42ES013660
Funding Period: 2009-2021

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Final Progress Reports

Year:   2020  2014 

The Molecular Biomarkers for Assessing Testicular Toxicity Project continued its elucidation of sperm biomarkers of male reproductive tract injury using a rat model of methotrexate (MTX) exposure. This ongoing study established a low-dose subchronic MTX exposure model in the rat with the ultimate goal of identifying distinct sperm RNA expression patterns associated with testicular injury and creating novel biomarker panels for testicular toxicity assessment. MTX was administrated at a dose of 2, 5, and 10mg/kg to adult male Fisher rats (n=10) weekly for 13 weeks. To characterize the testicular injury, testes/epididymal weights, sperm count, sperm motility, and testes/epididymal histology were assessed. Moreover, total RNA was isolated from epididymal sperm and small RNA was profiled using next generation RNA-sequencing. Sperm count and motility were similar among the control and treated groups. MTX treatment at 10mg/kg significantly decreased testes weight. There was a clear dose-related effect of MTX exposure on testicular histopathology, including a loss of spermatocytes in late stages of the seminiferous epithelial cycle and a loss of round spermatids in early stages of the cycle, resulting in seminiferous epithelial thinning and decreased seminiferous tubule diameter. Significant dose-dependent changes in piRNA length and piRNA length-distributions were identified from sequencing analysis. These data showed that sub-chronical low-dose MTX cause testicular histopathological abnormalities, and that sperm piRNAs can be developed as useful biomarkers of MTX-induced testicular injury.