Hybridization between different genomes is a source of genomic instability, sometimes associated with transposable element (TE) mobilization. Previous work showed that hybridization between the species Drosophila buzzatii and Drosophila koepferae induced mobilization of different (TEs), the Osvaldo retrotransposon being the most unstable. However, we ignore the mechanisms involved in this transposition release in interspecific hybrids. In order to disentangle the mechanisms involved in this process, we performed Osvaldo expression studies in somatic and germinal tissues from hybrids and parental species. There was a trend towards increased Osvaldo expression in the somatic tissues of hybrid females and males, which was always significant in males compared to the parental species D. buzzatii but, not in females compared to maternal species D. koepferae. There were massive changes of Osvaldo expression in the testes, which varied depending on the hybrid generation and family. Moreover, Osvaldo hybridization signals, restricted to the apical and primary spermatocyte regions in parents, occupied broader region in the hybrids. In ovaries, there were no significant differences in Osvaldo expression rates between hybrids and the maternal species D. koepferae. The transcript location was restricted to ovarian nurse cells in both parents and hybrids, undetectable in some hybrids. This research highlights first, the existence of putative complex deregulation mechanisms different between sexes and cell types and second, disruption of Osvaldo activity particularly evident in testes from sterile hybrid males. Deeper studies of the total transcriptome in hybrids and parental species are necessary to gain a better knowledge of the TE deregulation pathways in hybrids.