The Gly206Ala mutation was found in 8 of 19 unrelated Caribbean Hispanic families with early-onset familial AD. This genetic change may be a prevalent cause of early-onset familial AD in the Caribbean Hispanic population.

Apoptosis of myocardial cells occurs during cardiac allograft rejection. Apoptosis during rejection parallels the expression of iNOS, which suggests that apoptosis may be triggered by NO and peroxynitrite.

Background-The hypothesis that cyclooxygenase-2 (COX-2) is involved in the myocardial inflammatory response during cardiac allograft rejection was investigated using a rat heterotopic abdominal cardiac transplantation model. Methods and Results-COX-2 mRNA and protein in the myocardium of rejecting cardiac allografts were significantly elevated 3 to 5 days after transplantation compared with syngeneic controls (nϭ3, PϽ0.05). COX-2 upregulation paralleled in time and extent the upregulation of iNOS mRNA, protein, and enzyme activity in this model. COX-2 immunostaining was prominent in macrophages infiltrating the rejecting allografts and in damaged cardiac myocytes. Prostaglandin (PG) levels in rejecting allografts were also higher than in native hearts. Because NO has been reported to modulate PG synthesis by COX-2, additional transplants were performed using animals treated with a selective COX-2 inhibitor (SC-58125) and a selective inhibitor of the inducible nitric oxide synthase (iNOS) N-aminomethyl-L-lysine. At posttransplant day 5, inhibitor administration resulted in a significant reduction of COX-2 mRNA expression (3764Ϯ337 versus 5110Ϯ141 arbitrary units, nϭ3, PϽ0.05) and iNOS enzymatic activity (1.7Ϯ0.4 versus 22.8Ϯ14.4 nmol/mg protein, nϭ3, PϽ0.01) compared with vehicle-treated allogeneic transplants. Allograft survival in treated animals was increased modestly from 5.4 to 6.4 days (PϽ0.05). However, apoptosis of cardiac myocytes (TUNNEL method) was only marginally reduced relative to vehicle controls in treated graft recipients. The intensity of allograft rejection was also similar in the treated and untreated allografts. Conclusions-The data indicates that COX-2 expression is enhanced in parallel with iNOS in the myocardium during cardiac allograft rejection. (Circulation. 2000;101:430-438.)

Excessive nitric oxide (NO) production within the heart is implicated in the pathogenesis of myocyte death, but the mechanism whereby NO kills cardiac myocytes is not known. To determine whether NO may trigger programmed cell death (apoptosis) of adult rat ventricular myocytes in culture, the NO donor S-nitroso- N-acetylpenicillamine (SNAP) was shown to kill purified cardiac myocytes in a dose-dependent fashion. In situ analysis of ventricular myocytes plated on chamber slides using nick-end labeling of DNA demonstrated that SNAP induces cardiac myocyte apoptosis, which was confirmed by the identification of oligonucleosomal DNA fragmentation on agarose gel electrophoresis. Similarly, treatment of cardiac myocytes with cytokines that induce inducible NO synthase was shown to cause an NO-dependent induction of apoptosis. Addition of reduced hemoglobin to scavenge NO liberated by SNAP extinguished both the increase in percentage of apoptotic cells and the appearance of DNA ladders. Treatment with SNAP (but not with N-acetylpenicillamine or SNAP + hemoglobin) not only induced apoptosis but resulted in a marked increase in p53 expression in cardiac myocytes detected by Western blotting and immunohistochemistry. These data indicate that NO has the capacity to kill cardiac myocytes by triggering apoptosis and suggest the involvement of p53 in this process.

BackgroundThe p73 protein, a paralogue of the p53 tumor suppressor, is essential for normal development and survival of neurons. TP73 is therefore of interest as a candidate gene for Alzheimer's disease (AD) susceptibility. TP73 mRNA is transcribed from three promoters, termed P1 – P3, and there is evidence for an additional complexity in its regulation, namely, a variable allelic expression bias in some human tissues.MethodsWe utilized RT-PCR/RFLP and direct cDNA sequencing to measure allele-specific expression of TP73 mRNA, SNP genotyping to assess genetic associations with AD, and promoter-reporter assays to assess allele-specific TP73 promoter activity.ResultsUsing a coding-neutral BanI polymorphism in TP73 exon 5 as an allelic marker, we found a pronounced allelic expression bias in one adult brain hippocampus, while 3 other brains (two adult; one fetal) showed approximately equal expression from both alleles. In a tri-ethnic elderly population of African-Americans, Caribbean Hispanics and Caucasians, a G/A single nucleotide polymorphism (SNP) at -386 in the TP73 P3 promoter was weakly but significantly associated with AD (crude O.R. for AD given any -386G allele 1.7; C.I. 1.2–2.5; after adjusting for age and education O.R. 1.5; C.I. 1.1–2.3, N= 1191). The frequency of the -386G allele varied by ethnicity and was highest in African-Americans and lowest in Caucasians. No significant differences in basal P3 promoter activity were detected comparing -386G vs. -386A promoter-luciferase constructs in human SK-NSH-N neuroblastoma cells.ConclusionsThere is a reproducible allelic expression bias in mRNA expression from the TP73 gene in some, though not all, adult human brains, and inter-individual variation in regulatory sequences of the TP73 locus may affect susceptibility to AD. However, additional studies will be necessary to exclude genetic admixture as an alternative explanation for the observed associations.