Re-examination of the lymphocytotoxic crossmatch in liver transplantation: can C4d stains help in monitoring?

C4d-assisted recognition of antibody-mediated rejection (AMR) in formalin-fixed paraffin-embedded tissues (FFPE) from donor-specific antibody-positive (DSA+) renal allograft recipients prompted study of DSA+ liver allograft recipients as measured by lymphocytotoxic crossmatch (XM) and/or Luminex. XM results did not influence patient or allograft survival, or cellular rejection rates, but XM+ recipients received significantly more prophylactic steroids. Endothelial C4d staining strongly correlates with XM+ (<3 weeks posttransplantation) and DSA+ status and cellular rejection, but not with worse Banff grading or treatment response. Diffuse C4d staining, XM+, DSA+ and ABO- incompatibility status, histopathology and clinical-serologic profile helped establish an isolated AMR diagnosis in 5 of 100 (5%) XM+ and one ABO-incompatible, recipients. C4d staining later after transplantation was associated with rejection and nonrejection-related causes of allograft dysfunction in DSA- and DSA+ recipients, some of whom had good outcomes without additional therapy. Liver allograft FFPE C4d staining: (a) can help classify liver allograft dysfunction; (b) substantiates antibody contribution to rejection; (c) probably represents nonalloantibody insults and/or complete absorption in DSA- recipients and (d) alone, is an imperfect AMR marker needing correlation with routine histopathology, clinical and serologic profiles. Further study in late biopsies and other tissue markers of liver AMR with simultaneous DSA measurements are needed.

Role of prostaglandins in the testosterone-dependent wolffian duct differentiation of the fetal mouse.

Recent observations from this laboratory indicated a role of prostaglandin E2 (PGE2) in masculine differentiation of the external genitalia of the fetal mouse, induced by fetal testosterone. In this communication, we further investigated the role played by PGE2 in the testosterone-induced differentiation of the internal genital tract (Wolffian duct) of the fetal mouse. Using in vitro organ culture bioassay of Wolffian duct differentiation, we determined the effect of a PG-depleting agent, namely, anti-PGE2 antibody, and of inhibitors of PG synthesis for their ability to prevent Wolffian duct differentiation in the presence of testosterone. We demonstrated that anti-PGE2 antibody inhibited Wolffian duct differentiation in a dose-dependent manner in embryonic male explants containing fetal testes. At 1:10 dilution, the antibody inhibited the appearance of the entire Wolffian duct as well as growth of the specimen. At 1:100 dilution, however, only development of the Wolffian duct was prevented, as indicated by the absence of regions of the Wolffian duct or by the presence of epithelial disintegration throughout the ductal lumen. The antibody at 1:1000 dilution produced no significant effect on the appearance of the Wolffian duct. PGE2 (10 micrograms/ml) replacement in the medium prevented Wolffian duct disintegration induced by anti-PGE2. We next determined whether the testosterone-dependent Wolffian duct differentiation requires ongoing PG synthesis within the reproductive tract and analyzed the effects of the compounds inhibiting PG synthesis at the level of phospholipase A2 (PLA2) namely, cortisone and dexamethasone-and of those inhibiting at the level of cyclooxygenase-namely, aspirin and indomethacin. We have demonstrated that both PLA2 and cyclooxygenase inhibitors inhibited Wolffian duct differentiation in the male explant, i.e., in the presence of testis. These compounds also prevented the appearance of the Wolffian duct in female explants induced by exogenous testosterone. PGE2 added in the medium blocked the anti-masculinizing effects of PG synthesis inhibitors both in the male and female specimens. Thus, it appears that PG synthesis plays a role in the testosterone-induced masculine differentiation of the Wolffian duct.