Effect of porcine zonae pellucidae immunisation on ovarian follicular development and endocrine function in domestic ewes (Ovis aries).

Domestic ewes (Ovis aries) were immunised with porcine zonae pellucidae (pZP) or pZP conjugated to keyhole limpet haemocyanin (KLH) in adjuvant(s) to examine the feasibility of the species to serve as a model for further development of pZP-based vaccines in ungulates. Two immunisation groups were employed, with a third group receiving only adjuvant (n = 5 per group). Early in the study, oestrous activity was monitored by the use of a vasectomised ram fitted with a marking harness. Eventually, ewes were exposed to an intact ram for breeding. In addition, weekly serum and every-other-day faecal samples were collected to measure pZP antibodies and progesterone metabolite concentrations respectively. At the conclusion of the study, fecundity was established, and ovarian tissue was examined. Ewes immunised against pZP : KLH with adjuvant produced minimal antibody absorbance levels, displayed normal oestrous cycles, became pregnant upon introduction of the intact ram and exhibited normal ovarian histopathology. Ewes immunised against pZP with adjuvant produced high antibody absorbance levels, were acyclic following primary immunisation and were infertile. Examination of the ovarian tissue revealed atrophic changes that included: (1) the absence of growing follicles; (2) significant reduction in the number of primordial follicles; and (3) the presence of abnormal granulosa cell clusters lacking oocytes. Antisera displayed immunoreactivity to the major components of pZP, and immunohistochemical labelling of ovarian tissue showed specificity to the ZP. These data are the first generated in an ungulate species showing deleterious effects of pZP immunisation on folliculogenesis and oestrous cyclicity.

Persistence of anti-zonae pellucidae antibodies following a single inoculation of porcine zonae pellucidae in the domestic equine.

In this study of equids, we investigated the antibody response and the effect on the estrous cycle following a single inoculation of porcine zonae pellucidae (pZP) employing controlled-release methodology. We also investigated the use of two different water-soluble adjuvants as an alternative to oil-based adjuvants. Twenty-seven domestic mares were inoculated with various formulations of pZP and adjuvant. We showed that the anti-pZP antibodies generated as a result of the inoculations persisted for at least 43 weeks (length of the study). Of the various formulations used in the study, pZP and QS-21 water-soluble adjuvant, administered in combination with an emulsified preparation of pZP and Freund's Complete Adjuvant generated a significantly (P < 0.05) higher titer of anti-pZP antibodies when compared with other formulations employing the water-soluble adjuvant, Carbopol. Hormone analyses for cyclicity indicated a high incidence and extended duration of persistent corpora lutea among the treated mares. The positive control group of mares receiving two standard inoculations of pZP and Freund's Complete and Incomplete Adjuvants, as well as the placebo group of mares injected with QS-21 only, also exhibited high incidences of persistent corpora lutea. However, all mares eventually returned to normal cyclicity. The basis for the high incidence and extended duration of persistent corpora lutea was unexplained. The results demonstrate for the first time the persistent generation of anti-pZP antibodies following a single inoculation of pZP incorporated into a controlled-released preparation in the horse. This study further suggests that a single inoculation of pZP sequestered in a controlled-release lactide-glycolide polymer may serve as an alternative to traditional two-inoculation protocols for contraception investigations in the equine.