Characterization of VDR and CYP27B1 expression in the endometrium during the menstrual cycle before embryo transfer: implications for endometrial receptivity

Abstract

Background: Molecular analyses of vitamin D in a typical cycling endometrium has received minimal research attention in the reproductive field. This study was designed to assess how expression of the endometrial vitamin D receptor (VDR) and CYP27B1, a vitamin D metabolizing enzyme, change during the menstrual cycle in women of reproductive age. In addition, this study explores the association between expression of vitamin D-VDR system and endometrial receptivity during the implantation window.

Methods: Sixteen patients underwent standardized in vitro fertilization (IVF) treatment and freeze-all techniques. Before embryo transfer, total serum 25(OH) D levels were determined through blood samples and VDR, CYP27B1, HOXA10, and CYP19 expression were determined through endometrial samples. Endometrial receptivity was also assessed using an electron microscope.

Results: We found that VDR protein expression was significantly lower throughout the endometrial secretory phase compared to the proliferative phase, while CYP27B1 expression remained constant during the menstrual cycle. During the implantation window, ultrastructural evaluation showed that higher serum vitamin D levels were associated with more mature pinopodes; VDR and HOXA10 protein expression were substantially elevated in pregnant women compared to non-pregnant women; and VDR protein levels were positively correlated with HOXA10 levels. In addition, serum vitamin D levels were positively correlated with VDR and HOXA10 protein levels in the endometrium.

Conclusions: Women with increased VDR expression in the endometrium, especially during the implantation window of the menstrual cycle, were significantly more likely to be pregnant than women with decreased expression. Our results support the hypothesis that the Vitamin D-VDR system performs a role during the development of endometrial receptivity.

Keywords: CYP27B1; Endometrial receptivity; In-vitro fertilization; Vitamin D; Vitamin D receptor.

Fig. 1

SEM and TEM microphotographs of the endometrium under the influence of different concentrations of vitamin D during the implantation window. SEM and TEM microphotographs of vitamin D deficient (a and b), insufficient (c and d), and replete (e and f) patients. (a) Pinopodes not yet fully formed, ×3000 magnification. (c) Pinopodes in the developmental stage with abundant expression, × 3000 magnification. (e) Clusters of mature pinopodes with abundant expression, × 3000 magnification. b Intracytoplasmic organelles abundant with a few vacuoles; The nucleus is irregularly shaped and locally concave. Heterochromatin increases and aggregates can be seen around the nuclear envelope (examples are indicated by black arrowheads); The mitochondria are abundant, some are dropsical and dilated (reddish brown); The rough endoplasmic reticulum is swollen and dilated (blue); The Golgi apparatus (orange); Intracellular autophagy are less distributed (purple). d Intracytoplasmic organelles are abundant with a few vacuoles; The nucleus is irregularly shaped and locally concave without heterochromatin (black); The mitochondria are abundant without obvious edema or distention (reddish brown); The rough endoplasmic reticulum (blue); Intracellular autophagy is obviously increased (purple). The lysosomal (green). f Intracytoplasmic organelles are abundant; The nucleus is irregularly shaped and some are accompanied by nuclear division. Heterochromatin levels are elevated and aggregated around the nuclear envelope (black); The mitochondria are abundant, some are dropsical and dilated (reddish); The rough endoplasmic reticulum is swollen and dilated (blue); Intracellular autophagy is obviously increased (purple)

Fig. 2

Immunohistochemistry localization of VDR (left column), HOXA10 (second column), CYP27B1 (third column), and CYP19 (right column) during proliferative phase of the menstrual cycle in vitamin D deficient (top row), insufficient (second row), and replete (third row) patients. No signal was detected in the negative control sections (bottom row). All parts: magnification × 400, scale bar 50 μm

Fig. 3

Immunohistochemistry localization of VDR (left column), HOXA10 (second column), CYP27B1 third column), and CYP19 (right column) during the secretory phase of the menstrual cycle in vitamin D deficient (top row), insufficient (second row), and replete (third row) patients. No signal was detected in the negative control sections (bottom row). All parts: magnification × 400, scale bar 50 μm

Fig. 4

Western blot for VDR, CYP27B1, HOXA10, and CYP19 during the proliferative (a) and secretory (b) phases of the menstrual cycle. Summarized data are presented as the mean ± SEM of eight observations in each group, ***P < 0.001 (c)

Fig. 5

Target protein expression according to clinical pregnancy status. Comparison of target protein expression between pregnant and non-pregnant patients during the proliferative phase (a) and during the secretory phase (b) of the menstrual cycle. Summarized data are presented as the mean ± SEM of three to four observations in each subgroup, *P < 0.05