Basic and clinical andrology

Effects of lipopolysaccharide-induced inflammation on hypoxia and inflammatory gene expression pathways of the rat testis.

PMID 30473791


Bacterial infection and inflammation of the testis impairs fertility, yet an understanding of inflammatory responses of the testis is incomplete. We are interested in identifying gene pathways involved in the detection and clearance of infectious microbes in the male reproductive tract. In previous studies in our lab focused on hypoxia-responsive genes of the testis, preliminary experiments suggested that genes classically categorized as hypoxia genes are also activated during antimicrobial responses. The purpose of this study was to identify hypoxia and inflammatory gene pathways that contribute to antimicrobial protection of the testis and to consider possible cross-talk and interactions between these pathways. Inflammation was induced in Sprague-Dawley rats using P. aeruginosa or E. coli lipopolysaccharide (LPS). Levels of hypoxia-inducible factor-1 (HIF-1α) protein and nuclear factor kappa B (NF-κB) were measured, and hypoxia and inflammatory gene expression patterns in testis were analyzed by gene expression profiling using real-time quantitative PCR arrays. In LPS-treated rats, HIF-1α protein increased with no change in Hif-1α mRNA. Western Blot analysis also demonstrated no change in NF-κB and inhibitory NFKB alpha (IκBα) protein levels following LPS treatment. Five hypoxia pathway genes (Angptl4, Egr1, Ier3, Pai1, and Glut1), and 11 inflammatory pathway genes (Ccl12, Cc13, Cd14, Cxcl10, Icam1, Il10, Il1b, Il6, Nfkbia, Tlr2, Tnf) up-regulated after 3 h of inflammation. Angptl4, Ccl12, Cc13, Cd14, Egr1, Nfkbia, Tlr2, and Tnf remained elevated at 6 h. Six genes were up-regulated at 6 h only (Bhlhe40, C3, Jak2, Nlrp3, Slc11a1, Tlr1). One gene (Tlr5) was down-regulated after 3 h and no genes at 6 h. Electrophoretic mobility shift assay results suggest a decrease in NF-κB binding activity following LPS treatment. Testicular HIF-1α is up-regulated following LPS-induced inflammation. In contrast to other tissues, in which HIF-1α is up-regulated through transcriptional activation via NF-κB, we conclude that HIF-1α in the testis is not up-regulated through an increase in Hif-1α mRNA or through NF-κB-dependent mechanisms. Hypoxia pathway genes and genes involved in Toll-like receptor (TLR) and cytokine-mediated signaling comprise major functional categories of up-regulated genes, demonstrating that both hypoxia and classic inflammatory pathways are involved in inflammatory responses of the testis.