The effect of skin allografting on the equine endometrial cup reaction

Abstract

This research tested the hypothesis that immunological sensitization of mares by skin allografting, followed by the establishment of pregnancy using semen from the skin-graft donor, would give rise to secondary immune responses to the developing horse conceptus, resulting in an earlier demise of the fetally derived endometrial cups. Maiden mares received skin allografts from a stallion homozygous for Major Histocompatibility Complex (MHC) antigens and/or equivalent autografts and were subsequently mated to the skin-graft donor stallion during the next two breeding seasons. Mares that had been immunologically primed to the foreign MHC class I antigens of the skin-graft donor stallion developed strong secondary antibody responses early in their first pregnancies, whereas autografted mares made weak primary antibody responses in their first pregnancies and strong secondary responses in their second pregnancies. In contrast, histological examination of the endometrial cups after surgical pregnancy termination at Day 60 of gestation revealed no discernible differences between allografted and autografted mares, and there were no significant differences in the concentrations and/or duration of secretion of the endometrial cup-specific hormone, equine chorionic gonadotrophin (eCG), between allografted and autografted mares, nor in either group between first and second pregnancies. The vigorous antibody response observed in the pregnant allografted mares supported the first part of our hypothesis, providing evidence of systemic immunological priming. However, there was a lack of an equivalent heightened cellular response to the endometrial cups. These findings provided strong evidence for an asymmetric immune response to the conceptus, characterized by strong humoral immunity and a dampened cellular response.

Fig. 1

Histological evaluation of the cellular response to equine skin allografts. Hematoxylin and eosin staining was performed on biopsies of formalin-fixed samples of skin autografts and allografts taken on Day 8 after grafting. (A) Low magnification view of normal skin. (B) Higher magnification view of the boxed area in (A). (C) Low magnification view of a skin autograft on Day 8 after grafting. (D) Higher magnification view of the boxed area in (C). Note the scarcity of mononuclear cells (arrow) at the interface between the viable skin autograft (au) and recipient skin (r). (E) Low magnification view of a skin allograft on Day 8 after grafting. (F) Higher magnification view of the boxed area in (E). Note the large number of mononuclear cells (arrow) at the interface between the dying skin allograft (al) and recipient skin (r). Bar = 400 micrometers (A, C, and E) and 100 micrometers (B, D, and F).

Fig. 2

Time course of cytotoxic anti-paternal antibody to donor antigens in horses receiving skin allografts. Skin allografts (solid symbols) elicited strong primary cytotoxic antibody responses against the MHC antigens of the skin graft donor, but no cytotoxic antibodies were detected in sera from autografted recipients (open symbols).

Fig. 3

Anamnestic cytotoxic anti-paternal antibody responses in mares following skin allografting. Allografted mares produced strong secondary cytotoxic antibody responses when bred to the skin graft donor sire (A, solid symbols). In contrast, autografted mares produced weak primary antibody responses when bred to the same sire (A, open symbols). During the second pregnancy to the skin graft donor sire, allografted mares made strong tertiary antibody responses (B, solid symbols), whereas autografted mares made strong secondary antibody responses (B, open symbols).

Fig. 4

Histology of the cellular response to endometrial cups in mares during first and second pregnancy following either a skin autograft or a skin allograft from the mating stallion. Hematoxylin and eosin staining was performed on formalin-fixed endometrial cup tissues from Day 60 of pregnancy. Cellular responses to endometrial cups were compared between the first (A, C) and second (B, D) pregnancies of autografted (A, B) or allografted (C, D) mares. Note the large number of mononuclear cells (arrow) at the interface between the endometrial cup (ec) and the normal endometrium (em). Bar = 200 micrometers (A-D) and 500 micrometers (insets of A-D).

Fig. 5

Serum eCG concentrations measured in mares during their first and second consecutive pregnancies following either a skin autograft or a skin allograft from the mating stallion. Serial serum samples were collected during the first (A) and second (B) pregnancies and analyzed for eCG concentrations using an amplified enzyme-linked immunoassay [39]. There were no significant differences in peak serum eCG concentrations between allografted (solid symbols) and autografted (open symbols) animals during first (A) and second (B) pregnancies, nor in either group between first (A) and second (B) pregnancies.