Osteopontin gene expression in the Holstein bull reproductive tract

The objective of this study was to localize gene expression of osteopontin in the Holstein bull reproductive tract using Northern blot analysis and in situ hybridization. For Northern blot analysis, a digoxigenin-labeled osteopontin complementary deoxyribonucleic acid (cDNA) was used to probe blots containing total ribonucleic acid (RNA) isolated from the testis, epididymis, vas deferens, ampulla, seminal vesicle, prostate, and bulbourethral glands. The digoxigenin-labeled cDNA for the bovine homologue of osteopontin was hybridized to a single band at approximately 1.6 kb to RNA samples from the ampulla and seminal vesicle. For in situ hybridization studies, antisense and sense riboprobes were synthesized and used to hybridize cryosections that had been obtained from bull reproductive tissues. In situ hybridization of the bull testis detected osteopontin messenger RNA in the developing germ cells. Osteopontin gene expression was detected only in seminiferous tubules that contained elongated spermatids, which suggests that expression varies with the stage of the seminiferous epithelium. Within the epididymis, silver grains were distributed over the sperm that were located within the lumen of the caput, corpus, and cauda epididymis. Osteopontin expression was primarily observed in the epithelial cells of the ampulla. Antisense riboprobes also hybridized to sperm that were located within the lumen of the ampulla, confirming the presence of osteopontin transcripts in the haploid male gamete.