Expression of growth differentiation factor 9 (GDF-9), an apparent regulator of follicular development, reportedly differs between compartments of the rodent (oocytes) and human (oocytes and granulosa cells) ovary. To further characterize GDF-9 expression and action in the primate periovulatory follicle, adult female rhesus monkeys received recombinant human gonadotropins to promote multiple follicular development. Whole ovaries or follicular aspirates were obtained before and at various times after administration of an ovulatory dose of hCG; time points for tissue collection spanned the 40-h periovulatory interval. GDF-9 mRNA was detected by reverse transcription polymerase chain reaction assay in each oocyte and every granulosa cell sample examined, but granulosa cell GDF-9 mRNA levels did not change across the periovulatory interval. GDF-9 was also detected in follicular fluid using Western blotting; GDF-9 protein concentration in follicular fluid did not change across the periovulatory interval. Immunocytochemical staining for GDF-9 indicated that oocytes of both small and large antral follicles were positive for GDF-9. GDF-9 immunoreactivity was also present in cumulus granulosa cells and mural granulosa cells near the cumulus stalk. When granulosa cells from preovulatory follicles were exposed to recombinant GDF-9 in culture, GDF-9 increased vascular endothelial growth factor levels in culture medium. These data demonstrate that the cells of the primate periovulatory follicle both produce and respond to GDF-9. However, GDF-9 expression and action differ between rodent and primate follicles, suggesting a possible regulatory role for GDF-9 that is unique to the primate follicle.

Bioprosthetic heart valves (BHVs) are commonly used to replace severely diseased heart valves but their susceptibility to structural valve degeneration (SVD) limits their use in young patients. We hypothesized that antibodies against immunogenic glycans present on BHVs, particularly antibodies against the xenoantigens galactose-α1,3-galactose (αGal) andN-glycolylneuraminic acid (Neu5Gc), could mediate their deterioration through calcification. We established a large longitudinal prospective international cohort of patients (n = 1668, 34 ± 43 months of follow-up (0.1–182); 4,998 blood samples) to investigate the hemodynamics and immune responses associated with BHVs up to 15 years after aortic valve replacement. Early signs of SVD appeared in <5% of BHV recipients within 2 years. The levels of both anti-αGal and anti-Neu5Gc IgGs significantly increased one month after BHV implantation. The levels of these IgGs declined thereafter but anti-αGal IgG levels declined significantly faster in control patients compared to BHV recipients. Neu5Gc, anti-Neu5Gc IgG and complement deposition were found in calcified BHVs at much higher levels than in calcified native aortic valves. Moreover, in mice, anti-Neu5Gc antibodies were unable to promote calcium deposition on subcutaneously implanted BHV tissue engineered to lack αGal and Neu5Gc antigens. These results indicate that BHVs manufactured using donor tissues deficient in αGal and Neu5Gc could be less prone to immune-mediated deterioration and have improved durability.

Programmed cell death represents an important pathogenic mechanism in various autoimmune diseases. Type I diabetes mellitus (IDDM) is a T cell-dependent autoimmune disease resulting in selective destruction of the β cells of the islets of Langerhans. β cell apoptosis has been associated with IDDM onset in both animal models and newly diagnosed diabetic patients. Several apoptotic pathways have been implicated in β cell destruction, including Fas, perforin, and TNF-α. Evidence for Fas-mediated lysis of β cells in the pathogenesis of IDDM in nonobese diabetic (NOD) mice includes: 1) Fas-deficient NOD mice bearing the lpr mutation (NOD-lpr/lpr) fail to develop IDDM; 2) transgenic expression of Fas ligand (FasL) on β cells in NOD mice may result in accelerated IDDM; and 3) irradiated NOD-lpr/lpr mice are resistant to adoptive transfer of diabetes by cells from NOD mice. However, the interpretation of these results is complicated by the abnormal immune phenotype of NOD-lpr/lpr mice. Here we present novel evidence for the role of Fas/FasL interactions in the progression of NOD diabetes using two newly derived mouse strains. We show that NOD mice heterozygous for the FasL mutation gld, which have reduced functional FasL expression on T cells but no lymphadenopathy, fail to develop IDDM. Further, we show that NOD-lpr/lpr mice bearing the scid mutation (NOD-lpr/lpr-scid/scid), which eliminates the enhanced FasL-mediated lytic activity induced by Fas deficiency, still have delayed onset and reduced incidence of IDDM after adoptive transfer of diabetogenic NOD spleen cells. These results provide evidence that Fas/FasL-mediated programmed cell death plays a significant role in the pathogenesis of autoimmune diabetes.

Background: The two common sialic acids (Sias) in mammals are N‐acetylneuraminic acid (Neu5Ac) and its hydroxylated form N‐glycolylneuraminic acid (Neu5Gc). Unlike most mammals, humans cannot synthesize Neu5Gc that is considered foreign and recognized by circulating antibodies. Thus, Neu5Gc is a potential xenogenic carbohydrate antigen in bioprosthetic heart valves (BHV) that tend to deteriorate in time within human patients.Methods: We investigated Neu5Gc expression in non‐engineered animal‐derived cardiac tissues and in clinically used commercial BHV, and evaluated Neu5Gc immunogenicity on BHV through recognition by human anti‐Neu5Gc IgG.Results: Neu5Gc was detected by immunohistochemistry in porcine aortic valves and in porcine and bovine pericardium. Qualitative analysis of Sia linkages revealed Siaα2‐3>Siaα2‐6 on porcine/bovine pericardium while the opposite in porcine aortic/pulmonary valve cusps. Similarly, six commercial BHV containing either porcine aortic valve or porcine/bovine/equine pericardium revealed Siaα2‐3>Siaα2‐6 expression. Quantitative analysis of Sia by HPLC showed porcine/bovine pericardium express 4‐fold higher Neu5Gc levels compared to the porcine aortic/pulmonary valves, with Neu5Ac at 6‐fold over Neu5Gc. Likewise, Neu5Gc was expressed on commercial BHV (186.3±16.9 pmol Sia/μg protein), with Neu5Ac at 8‐fold over Neu5Gc. Affinity‐purified human anti‐Neu5Gc IgG showing high specificity toward Neu5Gc‐glycans (with no binding to Neu5Ac‐glycans) on a glycan microarray, strongly bound to all tested commercial BHV, demonstrating Neu5Gc immune recognition in cardiac xenografts.Conclusions: We conclusively demonstrated Neu5Gc expression in native cardiac tissues, as well as in six commercial BHV. These Neu5Gc xeno‐antigens were recognized by human anti‐Neu5Gc IgG, supporting their immunogenicity. Altogether, these findings suggest BHV‐Neu5Gc/anti‐Neu5Gc may play a role in valve deterioration warranting further investigation.