Trade-offs between male fertility reduction and selected growth factors or the klotho response in a lipopolysaccharide-dependent mouse model

Abstract

The increasing number of depression cases leads to a greater need for new antidepressant treatment development. It is postulated that antidepressants may harm male fertility, but the cellular mechanism is still poorly understood. The role of growth factors and klotho protein in maintaining normal male reproductive function is well documented. Hence, the study aimed to investigate the effect of the antidepressant drug - imipramine (tricyclic AD), and other substances with antidepressant potential (ALS), administered in combination or in combination with LPS (an animal model of depression) on gene expression and protein synthesis of IGF-2 (insulin-like growth factor 2), TGF-β1 (transforming growth factor β1), NGF (nerve growth factor), KGF (keratinocyte growth factor) and protein synthesis of VEGF-A (vascular endothelial growth factor A), IGF-IR (insulin-like growth factor receptor 1), EGFR (epidermal growth factor receptor) and klotho in the testis of mice. Mice were injected intraperitoneally with selected ALS and LPS or 10% DMSO (controls) (n = 7/group) once a day for 14 days. Animals were decapitated and testes collected for RNA and protein purification. PCR and western blot methods were employed for the evaluation of growth factors and klotho expression. The results obtained indicated a decreased level of most of the analyzed genes and proteins, except KGF; its expression increased after treatment with MTEP and IMI administrated individually and after NS-398, and IMI in combination with LPS. Our results may suggest that the tested ALS and LPS can contribute to a reduction of male fertility, but NS-398, IMI, and IMI+NS-398 may also act as stimulants after LPS.

Keywords: Antidepressant-like substances; Growth factors; Klotho; LPS; Testis.

Fig. 1

mRNA expression of: A IGF-2, B TGF-β1, C NGF, D KGF analyzed by RT PCR technique in mouse (C57BL/6J) testis; Bars indicate mean ± SD, n = 6, asterisks indicate comparison between control group (C) and analysed groups after administration of: MTEP, NS-398, IMI, MTEP + NS-398 (M1), IMI + NS-398 (M2) with and without LPS; ^^^,***p < 0.001, ^^,**p < 0.01, ^,*p < 0.05; (*) indicate comparison between control group and used substances, (^) indicate comparison between group after the same substances used LPS-non-treated and treated, no indication – no statistical significance (p > 0.05) (one –way ANOVA with Dunnett’s a posteriori test). The results were normalized to ACTB

Fig. 2

Ethidium bromide staining of the results of RT PCR for IGF-2, TGF-β1, NGF, KGF mRNA in mouse (C57BL/J) testis; representative images of gels after administration of: vehicle (C) MTEP, NS-398, IMI, MTEP + NS-398 (M1), IMI + NS-398 (M2) with and without LPS

Fig. 3

A VEGF-A, B IGF-IRβ, C EGFR, D klotho membrane isoform, E klotho secreted isoform protein levels analyzed by Western blot technique in mouse (C57BL/6J) testis. Bars indicate mean ± SD, n = 6; asterisks indicate comparison between control group and analysed groups after administration of: MTEP, NS-398, IMI, MTEP + NS-398 (M1), IMI + NS-398 (M2) with and without LPS; ^^^,***p < 0.001, ^^,**p < 0.01, ^,*p < 0.05; (*) indicate comparison between control group and used substances, (^) indicate comparison between group after the same substances used LPS-non-treated and treated; no indication – no statistical significance (p > 0.05) (one –way ANOVA with Dunnett’s a posteriori test). The results were normalized to β-actin

Fig. 4

Representative immunoblots of: VEGF-A, IGF-IRβ, EGFR, KOTHO membrane isoform, KLOTHO secrete isoform and β-actin proteins in mouse (C57BL/6J) testis after administration of: vehicle (C), MTEP, NS-398, IMI, MTEP + NS-398 (M1), IMI + NS-398 (M2) with and without LPS

Fig. 5

Scheme of the effect of antidepressant-like substances on mouse testis