Galectin-1 overexpression in endometriosis and its regulation by neuropeptides (CRH, UCN) indicating its important role in reproduction and inflammation

Abstract

Endometriosis is an inflammatory disease of women of reproductive age featured by the presence of ectopic endometrium and is strongly related to infertility. Galectins, carbonhydrate-binding proteins, have been found to have pro- or anti-inflammatory roles in the reproductive tract and in pathological conditions concerning infertility. Galectin-1, which is expressed at endometrium and decidua, plays a major role in implantation and trophoblast invasion. Also, the neuropeptides, corticotropin releasing hormone (CRH) and urocortin (UCN) and their receptors are expressed in eutopic and ectopic endometrium showing a differential expression pattern in endometriotic women compared to healthy ones. The aim of this study was to examine the galectin-1 expression in endometriotic lesions and compare its expression in eutopic endometrium of endometriotic and healthy women. Furthermore, we examined the effect of CRH and UCN in galectin-1 expression in Ishikawa cell line and macrophages and investigated the implication of CRHR1 in these responses. Eutopic and ectopic endometrium specimens, Ishikawa cell line and mice macrophages were used. Immunohistochemistry and western blot analysis were performed in order to identify galectin-1 expression in ectopic and eutopic endometrium of women with and without endometriosis and the regulatory effect of CRH and UCN on galectin-1 expression. This study presents for the first time that galectin-1 is overexpressed in endometriotic lesions compared to eutopic endometrium of endometriotic women and is more abundantly expressed in eutopic endometrium of disease women compared to healthy ones. Furthermore, it is shown that CRH and UCN upregulate galectin-1 expression in Ishikawa cell line and macrophages and this effect is mediated through CRHR1. These results suggest that galectin-1 might play an important role in endometriosis pathology and infertility profile of women suffering from endometriosis by being at the same time regulated by CRH and UCN interfering in the immune disequilibrium which characterizes this pathological condition.

Figure 1. Galectin-1 overexpression in late secretory phase of eutopic endometrium and in endometriotic sites.

1a: Immunohistochemical expression of galectin-1 expression through the different phases of the menstrual cycle of the endometrium (A: proliferative, B: early secretory phase, C: late secretory phase) showing a higher expression pattern at the late secretory phase of the endometrium(C), *p<0.05. 1b: Western blot immunodetection of galectin-1 (14 kDa) protein expression in eutopic and ectopic endometrium. Galectin-1 is overexpressed in eutopic endometrium of endometriotic women (ee) when compared to eutopic endometrium of healthy women (he), *p<0.05 and galectin-1 shows an abundant expression in ectopic endometrium (e) when compared to eutopic endometrium of the same endometriotic women (ee), **p<0.01. GAPDH (37 KDa) was used as a house keeping gene.

Figure 2. Galectin-1 is upregulated in Ishikawa cell line upon CRH and UCN stimulation and this is mediated through CRHR1.

Western blot immunodetection of galectin-1 (14 kDa) protein expression in Ishikawa cell line, regulated by CRH and UCN neuropeptides. GAPDH (37 KDa) was used as a house keeping gene. 2a: In Ishikawa cell line, galectin-1 expression is upregulated by CRH, showing a higher expression at 8 hrs and 24 hrs of stimulation. When antalarmin is added, galectin-1 shows an impaired expression mostly at 24 hrs of stimulation. 2b: In Ishikawa cell line, galectin-1 is upregulated by UCN, showing a higher expression at 8 hrs of stimulation.

Figure 3. Galectin-1 is upregulated in macrophages upon CRH and UCN stimulation and this is mediated through CRHR1.

Western blot immunodetection of galectin-1 (14 kDa) protein expression in macrophages, regulated by CRH and UCN neuropeptides. GAPDH (37 KDa) was used as a house keeping gene. 3a: In mice macrophages, galectin-1 expression is upregulated by CRH mostly at 24 hrs and Antalarmin effect is stronger at 24 hrs. 3b: In mice macrophages, galectin-1 expression is mostly upregulated by UCN at 24 hrs.