Testosterone increases the virulence traits of uropathogenic Escherichia coli

Abstract

Uropathogenic Escherichia coli (UPEC) is the most common cause of urinary tract infections (UTIs) in humans. Testosterone negatively impacts UTIs by affecting the immune response, leading to higher susceptibility of chronic cystitis in individuals with elevated testosterone levels, regardless of gender. Current research is mostly focused on how testosterone affects the host response to UPEC, but not so much is known about how testosterone directly affect UPEC virulence. The aim of the present study was to investigate the impact of testosterone exposure on the virulence of UPEC. We found that testosterone directly increases UPEC growth, endotoxin release and biofilm formation. We also found that testosterone-stimulated CFT073 increased colonization and invasion of bladder epithelial cells. Testosterone-stimulated CFT073 also increased the release of IL-1β and LDH from bladder epithelial cells. Additionally, by using a Caenorhabditis elegans survival assay we also showed that testosterone decreased the survival of CFT073 infected C. elegans worms. Taken together, our findings show that testosterone directly increases the virulence traits of UPEC.

Keywords: cross-kingdom interaction; growth; testosterone; uropathogenic Escherichia coli; virulence.

Figure 1

CFT073 growth with or without the presence of testosterone (100 pg/mL, 2 ng/mL and 60 ng/mL) during 24 h (A) and at 8 (B), 10 (C) and 12 (D) hours. Data are presented as mean (A) and mean ± SD (B) of n = 3 independent experiments. The asterisk distinguishes statistical significance: * = p < 0.05 vs. CFT073.

Figure 2

Biofilm formation (A), endotoxin release (B) and hemolysis (C) with or without the presence of testosterone (100 pg/mL, 2 ng/mL and 60 ng/mL) after 4–8 (C) or 24 h (A–B). Data are presented as mean ± SD of n = 3–5 independent experiments. The asterisk distinguishes statistical significance: * = p < 0.05 and ** = p < 0.01 vs. CFT073.

Figure 3

CFT073 colonization (A,C 4 h) or invasion (B, 2 h) of bladder epithelial cells after CFT073 pre-treated with or without testosterone (100 pg/mL, 2 ng/mL and 60 ng/mL). CFT073 (carrying a GFP-expressing plasmid) colonization was measured as mean fluorescence intensity (MFI) (A) and imaged (C). Data are presented as mean ± SD of n = 3–8 independent experiments. The asterisk distinguishes statistical significance: * = p < 0.05, ** = p < 0.01, and *** = p < 0.001 vs. CFT073. Scale bar: 500 μm.

Figure 4

IL-1β (A), IL-8 (B) and LDH (C) release from bladder epithelial cells stimulated with CFT073 MOI10 pre-treated with or without testosterone (100 pg/mL, 2 ng/mL and 60 ng/mL) at 4 h. Data are presented as mean ± SD of n = 6–8 independent experiments. The asterisks distinguish statistical significance: * = p < 0.05 and **** = p < 0.0001.

Figure 5

RNASE7 (A), LL37 (B), beta-defensin 1 (C) and beta-defensin 2 (D) release from bladder epithelial cells stimulated with CFT073 MOI10 pre-treated with or without testosterone (100 pg/mL, 2 ng/mL and 60 ng/mL) at 4 h. Data are presented as mean ± SD of n = 3 independent experiments. The asterisks distinguish statistical significance: * = p < 0.05 and **** = p < 0.0001.

Figure 6

Caenorhabditis elegans survival after stimulation with CFT073 pre-treated with or without testosterone (100 pg/mL, 2 ng/mL and 60 ng/mL) for 4 (B), 5 (C), 6 (D) and 12 h (A). Data are presented as mean ± SD of n = 9 independent experiment. Statistical significance is denoted with asterisks: ** = p < 0.01, *** = p < 0.001, and **** = p < 0.0001 vs. CFT073.