Hyaluronan (HA) content, the ratio of HA fragments and the expression of CD44 in the ovine cervix vary with the stage of the oestrous cycle

Abstract

The complex anatomy of the ovine cervix limits the success of trans-cervical artificial insemination in sheep. However, there is a degree of natural relaxation of cervix at oestrus that is accompanied by an increase in the water content. As hyaluronan (HA) has a high affinity for water molecules, in this study, we tested the hypothesis that the HA content of the cervix, the proportion of different size fragments of HA and expression of its receptor CD44 vary with the stage of the oestrous cycle. Oestrous was synchronized in 25 Welsh mountain ewes, and their cervices were collected either during luteal phase (n=8) or pre-LH (n=8) or post-LH (n=9) surge stage of the oestrous cycle. The pre-LH surge group had the highest HA content (2.96 ng/mg of cervical tissue), which was significantly (P≤0.05) higher than that observed for the post-LH surge (2.04 ng/mg) group. The luteal phase group had a mean HA content intermediate between the pre- and post-LH surge groups, and was significantly different from either. The frequency of cervical samples containing both sizes of HA fragments (small and large) was significantly higher (P≤0.05) in the pre-LH surge group compared with the luteal and the post-LH surge groups, whereas that in post-LH surge group was significantly (P≤0.05) higher than that in the luteal group. The number of cervical samples that contained only small HA fragments was significantly (P≤0.05) higher in the luteal group compared with both the pre- and post-LH surge groups, whereas the number of samples containing only large HA fragments was significantly (P≤0.05) higher in the post-LH surge group compared with the luteal or pre-LH surge groups. Overall mean expression of CD44 in the vaginal and mid regions was significantly (P≤0.001) higher than that in the uterine region, with no difference between the vaginal and mid regions of the cervix. Pattern of CD44 expression depended on the stage of the oestrous cycle. At the luteal stage, CD44 expression did not vary among epithelial, sub-epithelial, circular and longitudinal smooth muscle layers, whereas at the pre- and post-LH surge stages, the expression in the epithelial layer was significantly (P≤0.001) higher than that in the other three layers. In general, CD44 expression in the transverse smooth muscle layer was significantly (P≤0.05) lower than the expression in all the other layers at all the stages of the oestrous cycle. The results indicated that the HA varied with the steroid status. Higher HA values at a time when cervical relaxation is naturally higher may indicate its involvement in remodelling of the cervix at oestrus.