Differential tissue-specific damage caused by bacterial epididymo-orchitis in the mouse

Ascending bacterial urinary tract infections can cause epididymo-orchitis. In the cauda epididymidis, this frequently leads to persistent tissue damage. Less coherent data is available concerning the functional consequences of epididymo-orchitis on testis and caput epididymidis. This in vivo study addresses the functional and spatial differences in responsiveness of murine epididymis and testis to infection with uropathogenic Escherichia coli (UPEC). Whole transcriptome analysis (WTA) was performed on testis, caput, corpus and cauda epididymidis of adult C57BL/6 J wildtype mice. Following UPEC-induced epididymo-orchitis in these mice, epididymal and testicular tissue damage was evaluated histologically and semi-quantitatively at 10 days and 31 days post-inoculation. Expression of inflammatory markers and candidate antimicrobial genes were analysed by RT-qPCR. WTA revealed distinct differences in gene signatures between caput and cauda epididymidis, particularly amonst immunity-related genes. Cellular and molecular signs of testicular inflammation and disruption of spermatogenesis were noticed at day 10, but recovery was observed by day 31. In contrast to the cauda, the caput epididymidis did not reveal any signs of gross morphological damage or presence of pro-inflammatory processes despite confirmed infection. In contrast to beta-defensins, known UPEC-associated antimicrobial peptides (AMP), like Lcn2, Camp and Lypd8, were inherently highly expressed or upregulated in the caput following infection, potentially allowing an early luminal protection from UPEC. At the time points investigated, the caput epididymidis was protected from any obvious infection/inflammation-derived tissue damage. Studies addressing earlier time-points will conclude whether in the caput epididymidis a pro-inflammatory response is indeed not essential for effective protection from UPEC.

Next-generation sequencing analysis reveals segmental patterns of microRNA expression in yak epididymis

MicroRNAs (miRNAs) have emerged as potent regulators of gene expression and are widely expressed in biological systems. In reproduction, they have been shown to have a significant role in the acquisition and maintenance of male fertility, whereby deletion of Dicer in mouse germ cells leads to infertility. Evidence indicates that this role of miRNAs extends from the testis into the epididymis, controlling gene expression and contributing to regional variations in gene expression. In this study, RNA sequencing technology was used to investigate miRNA expression patterns in the yak epididymis. Region-specific miRNA expression was found in the yak epididymis. In all, 683 differentially expressed known miRNAs were obtained; 190, 186 and 307 differentially expressed miRNAs were identified for caput versus corpus, corpus versus cauda and caput versus cauda region pairs respectively. Kyoto Encyclopedia of Genes and Genomes results showed endocytosis as the most enriched pathway across region pairs, followed by protein processing in the endoplasmic reticulum, phagosome, spliceosome and biosynthesis of amino acids in region pair-specific hierarchical order. Gene ontology results showed varied enrichment in terms including cell, biogenesis, localisation, binding and locomotion across region pairs. In addition, significantly higher miR-34c expression was seen in the yak caput epididymidis relative to the corpus and cauda epididymidis.

GPX5, the selenium-independent glutathione peroxidase-encoding single copy gene is differentially expressed in mouse epididymis

Using various molecular approaches, including reverse transcription–polymerase chain reaction (RT–PCR), rapid amplification of cDNA ends–PCR, sequencing, northern and western blotting, we found that the mouse GPX5 gene gives rise to at least three different transcripts that are not expressed at the same levels in the mouse epididymis. In addition to the major GPX5 transcript, we show that minor GPX5 transcripts exist, arising either from precocious termination of transcription or an alternative splicing event within intron 4 of the 5 exon-encoding GPX5 single copy gene. Furthermore, we demonstrate that variants of the GPX5 protein that are correlated with the shorter GPX5 transcripts can be detected in caput epididymidis protein extracts and that the various GPX5 isoforms are subject to differential post-transcriptional maturation processes in the mouse epididymis that essentially involve the addition of O-glycosyl extensions. Using a sensitive poly-A+ mRNA tissue blot, as well as RT–PCR and northern assays, we further show that in addition to being expressed in the epididymis, the GPX5 gene is also expressed, albeit at lower levels, in other tissues of the male genital tract, including the testis and prostate. Finally, we present evidence suggesting that the GPX5 gene is expressed in a temporally regulated manner during mouse embryonic development.

11β-Hydroxysteroid dehydrogenase enzymes in the testis and male reproductive tract of the boar (Sus scrofa domestica) indicate local roles for glucocorticoids in male reproductive physiology

11β-Hydroxysteroid dehydrogenase (11βHSD) enzymes modulate the target cell actions of corticosteroids by catalysing metabolism of the physiological glucocorticoid (GC), cortisol, to inert cortisone. Recent studies have implicated GCs in boar sperm apoptosis. Hence, the objective of this study was to characterise 11βHSD enzyme expression and activities in the boar testis and reproductive tract. Although 11βHSD1 and 11βHSD2 mRNA transcripts and proteins were co-expressed in all tissues, cortisol–cortisone interconversion was undetectable in the corpus and cauda epididymides, vas deferens, vesicular and prostate glands, irrespective of nucleotide cofactors. In contrast, homogenates of boar testis, caput epididymidis and bulbourethral gland all displayed pronounced 11βHSD activities in the presence of NADPH/NADP+ and NAD+, and the penile urethra exhibited NAD+-dependent 11β-dehydrogenase activity. In kinetic studies, homogenates of boar testis, caput epididymidis and bulbourethral gland oxidised cortisol with Km values of 237–443 and 154–226 nmol/l in the presence of NADP+ and NAD+ respectively. Maximal rates of NADP+-dependent cortisol oxidation were 7.4- to 28.5-fold greater than the Vmax for NADPH- dependent reduction of cortisone, but were comparable with the rates of NAD+-dependent cortisol metabolism. The relatively low Km estimates for NADP+ -dependent cortisol oxidation suggest that either the affinity of 11βHSD1 has been increased or the cortisol inactivation is catalysed by a novel NADP+-dependent 11βHSD enzyme in these tissues. We conclude that in the boar testis, caput epididymidis and bulbourethral gland, NADP+- and NAD+-dependent 11βHSD enzymes catalyse net inactivation of cortisol, consistent with a physiological role in limiting any local actions of GCs within these reproductive tissues.

TEX101 is shed from the surface of sperm located in the caput epididymidis of the mouse

It is generally believed that cell-to-cell cross-talk and signal transduction are mediated by cell surface molecules that play diverse and important regulatory roles in spermatogenesis and fertilization. Recently, we identified a novel plasma membrane-associated protein, TES101-reactive protein (TES101RP, or TEX101), on mouse testicular germ cells. In this study, we investigate Tex101 mRNA expression in the adult mouse testis using in situ hybridization, and we examine the fate of TEX101 during sperm transport by immunohistochemical and Western blot analyses. Tex101 mRNA was expressed in a stage-specific manner in spermatocytes and in step 1–9 spermatids of the testis, but not in spermatogonia. Although the TEX101 protein remained on the cell surfaces of step 10–16 spermatids and testicular sperm, it was shed from epididymal sperm located in the caput epididymidis. The results of this study provide additional information on germ cell-specific TEX101 expression during spermatogenesis and post-testicular sperm maturation.