The mystery of capsular vesicles: Preliminary studies

Abstract

An unexpected finding from a previous experiment was the finding of capsular vesicles in the uterine flush 48h after the transfer of a Day 8 embryo that had been punctured with an acupuncture needle. The authors of this study initially thought the vesicles originated from fragments of capsule, in the same way fragments of cultured trophoblast readily form vesicles. However, recovery of a capsule with the hole made by the acupuncture needle alongside the capsular vesicles showed that this was not the case. A preliminary study was undertaken to determine if trophoblast vesicles (TVs) could initiate capsule vesicle formation in recipient mares at different stages post ovulation when transferred for 24h. Day 11 embryos were recovered from donor mares and the capsule was removed before fragmenting the trophoblast and incubating the tissue in DMEMF12 (+ 5% FBS and pen/strep) for 24-48h. Between 20 and 50 TVs were transferred to the uteri of ten recipient mares. Recipient mares were divided into 3 groups; G1, Days 4 or 5 post ovulation (n = 2); G2, Days 6 to 9 (n = 5); and G3, Days 12 and 13 (n = 3). The uteri of the recipient mares were flushed 24h after transfer of the TVs and the recovered fluid searched. In all of the mares, TVs were recovered withvarying degrees of degeneration. In G1 no capsular vesicles were recovered, in G2 and G3 between 5 and 32 capsular vesicles were found in each flush. Although different numbers of vesicles were transferred to recipients the greatest number of capsular vesicles per transferred TV was in G2 compared to G3 (0.44 - 1.6 vs. 0.12 – 0.25 capsular vesicles/transferred TV, respectively). Proper formation of the capsule in the mare is believed to involve interaction of the embryo with the endometrium because in vitro produced embryos fail to produce a proper capsule until after transfer. In vivo, developmentally regulated changes in composition of the capsule have also been noted and the capsule increases in thickness up until at least Day 11. These preliminary results suggest that capsular vesicles can form in the uterus following transfer of TVs, providing that the uterus is at a stage when an embryo would normally be forming the capsular layer since transfers prior to Day 6 failed to produce capsular vesicles and reduced numbers were produced after Day 12. The mechanism of their formation remains a mystery as they are free in the uterus and did not envelop the trophoblastic vesicles.