Universal DNA methylation age across mammalian tissues.

Aging, often considered a result of random cellular damage, can be accurately estimated using DNA methylation profiles, the foundation of pan-tissue epigenetic clocks. Here, we demonstrate the development of universal pan-mammalian clocks, using 11,754 methylation arrays from our Mammalian Methylation Consortium, which encompass 59 tissue types across 185 mammalian species. These predictive models estimate mammalian tissue age with high accuracy (r > 0.96). Age deviations correlate with human mortality risk, mouse somatotropic axis mutations and caloric restriction. We identified specific cytosines with methylation levels that change with age across numerous species. These sites, highly enriched in polycomb repressive complex 2-binding locations, are near genes implicated in mammalian development, cancer, obesity and longevity. Our findings offer new evidence suggesting that aging is evolutionarily conserved and intertwined with developmental processes across all mammals.

Genome-wide association study of monoamine metabolite levels in human cerebrospinal fluid.

Studying genetic determinants of intermediate phenotypes is a powerful tool to increase our understanding of genotype-phenotype correlations. Metabolic traits pertinent to the central nervous system (CNS) constitute a potentially informative target for genetic studies of intermediate phenotypes as their genetic underpinnings may elucidate etiological mechanisms. We therefore conducted a genome-wide association study (GWAS) of monoamine metabolite (MM) levels in cerebrospinal fluid (CSF) of 414 human subjects from the general population. In a linear model correcting for covariates, we identified one locus associated with MMs at a genome-wide significant level (standardized ß=0.32, P=4.92 × 10(-8)), located 20 kb from SSTR1, a gene involved with brain signal transduction and glutamate receptor signaling. By subsequent whole-genome expression quantitative trait locus (eQTL) analysis, we provide evidence that this variant controls expression of PDE9A (ß=0.21; P unadjusted=5.6 × 10(-7); P corrected=0.014), a gene previously implicated in monoaminergic transmission, major depressive disorder and antidepressant response. A post hoc analysis of loci significantly associated with psychiatric disorders suggested that genetic variation at CSMD1, a schizophrenia susceptibility locus, plays a role in the ratio between dopamine and serotonin metabolites in CSF. The presented DNA and mRNA analyses yielded genome-wide and suggestive associations in biologically plausible genes, two of which encode proteins involved with glutamate receptor functionality. These findings will hopefully contribute to an exploration of the functional impact of the highlighted genes on monoaminergic transmission and neuropsychiatric phenotypes.

Use of clopidogrel post-coronary artery bypass surgery in canadian patients with acute coronary syndromes.

BACKGROUND: Randomized trials have established the efficacy of clopidogrel in acute coronary syndromes (ACS). The benefit of clopidogrel has also been observed in the subgroup of ACS patients who subsequently undergo coronary artery bypass surgery (CABG); however, this therapy is discontinued preoperatively and the frequency with which clopidogrel is restarted post-CABG is unknown. METHODS: We examined the pattern of clopidogrel use in the Canadian Global Registry of Acute Coronary Events (GRACE), GRACE2, and CANRACE (2003-2008) post-CABG ACS patients. We stratified the patients according to whether they underwent CABG during their index hospitalization for ACS and whether they were prescribed clopidogrel at discharge. RESULTS: Among those patients in whom clopidogrel status at discharge was known, 5904 (60%) of 9841 were discharged from hospital on clopidogrel. Use of clopidogrel at discharge was observed in 2222 (40.8%) of 5443 patients who were medically managed (ie, did not undergo percutaneous coronary intervention [PCI] or CABG) and in 3585 (90.1%) of 3980 patients who underwent in-hospital PCI. Overall, 455 (3.3%) of 13,776 patients underwent CABG during the index hospitalization; 255 (56%) patients were started on clopidogrel during the first 24 hours, and 66 of these patients (25.9%) were discharged on clopidogrel. In contrast, 5681 (61.3%) of the 9262 patients who did not undergo in-hospital CABG were discharged on clopidogrel. CONCLUSIONS: Although current guidelines recommend the use of clopidogrel post-CABG in patients with ACS, our observations suggest that only 1 in 4 or 5 Canadian patients are discharged on this therapy.

Temporal trends in the use of invasive cardiac procedures for non-ST segment elevation acute coronary syndromes according to initial risk stratification.

BACKGROUND: Current guidelines support an early invasive strategy in the management of high-risk non-ST elevation acute coronary syndromes (NSTE-ACS). Although studies in the 1990s suggested that highrisk patients received less aggressive treatment, there are limited data on the contemporary management patterns of NSTE-ACS in Canada. OBJECTIVE: To examine the in-hospital use of coronary angiography and revascularization in relation to risk among less selected patients with NSTE-ACS. METHODS: Data from the prospective, multicentre Global Registry of Acute Coronary Events (main GRACE and expanded GRACE2) were used. Between June 1999 and September 2007, 7131 patients from across Canada with a final diagnosis of NSTE-ACS were included the study. The study population was stratified into low-, intermediate- and high-risk groups, based on their calculated GRACE risk score (a validated predictor of in-hospital mortality) and according to time of enrollment. RESULTS: While rates of in-hospital death and reinfarction were significantly (P<0.001) greater in higher-risk patients, the in-hospital use of cardiac catheterization in low- (64.7%), intermediate- (60.3%) and highrisk (42.3%) patients showed an inverse relationship (P<0.001). This trend persisted despite the increase in the overall rates of cardiac catheterization over time (47.9% in 1999 to 2003 versus 51.6% in 2004 to 2005 versus 63.8% in 2006 to 2007; P<0.001). After adjusting for confounders, intermediate-risk (adjusted OR 0.80 [95% CI 0.70 to 0.92], P=0.002) and high-risk (adjusted OR 0.38 [95% CI 0.29 to 0.48], P<0.001) patients remained less likely to undergo in-hospital cardiac catheterization. CONCLUSION: Despite the temporal increase in the use of invasive cardiac procedures, they remain paradoxically targeted toward low-risk patients with NSTE-ACS in contemporary practice. This treatment-risk paradox needs to be further addressed to maximize the benefits of invasive therapies in Canada.

Characterization of primary cell cultures as potential target cells for analysis of bovine cytotoxic T lymphocytes.

In domestic animal species, assessment of cell-mediated immune responses to virus infection is hampered by the requirement for class I MHC compatibility between target and effector cells. Additional complicating factors can include an inability to infect target cells in vitro, or virus-induced lysis of infected target cells. One way to circumvent these problems is to use virus-mediated gene transfer to deliver individual viral genes to autologous primary target cells. Several primary bovine cell cultures were assessed as potential target cells for cytotoxic T lymphocyte (CTL) assays by measuring their levels of class I MHC expression and susceptibilities to retroviral gene delivery. High levels in both class I MHC expression and susceptibility to gene delivery were seen in adherent cell cultures isolated from peripheral blood (PBAC). PBAC, which arose as an outgrowth of adherent peripheral blood mononuclear cell cultures, had morphology, protein expression patterns, and response to functional assays characteristic of high endothelial cells. Expression of viral vector-delivered genes in PBAC cells was confirmed with a recombinant retrovirus carrying the green fluorescent protein (GFP) gene. The use of vector-mediated delivery of viral genes to bovine high endothelial cells is a promising method for assessment of cell-mediated immunity in cattle.

A gene encoding a P-type ATPase mutated in two forms of hereditary cholestasis.

Cholestasis, or impaired bile flow, is an important but poorly understood manifestation of liver disease. Two clinically distinct forms of inherited cholestasis, benign recurrent intrahepatic cholestasis (BRIC) and progressive familial intrahepatic cholestasis type 1 (PFIC1), were previously mapped to 18q21. Haplotype analysis narrowed the candidate region for both diseases to the same interval of less than 1 cM, in which we identified a gene mutated in BRIC and PFIC1 patients. This gene (called FIC1) is the first identified human member of a recently described subfamily of P-type ATPases; ATP-dependent aminophospholipid transport is the previously described function of members of this subfamily. FIC1 is expressed in several epithelial tissues and, surprisingly, more strongly in small intestine than in liver. Its protein product is likely to play an essential role in enterohepatic circulation of bile acids; further characterization of FIC1 will facilitate understanding of normal bile formation and cholestasis.

Genetic and morphological findings in progressive familial intrahepatic cholestasis (Byler disease [PFIC-1] and Byler syndrome): evidence for heterogeneity.

Byler disease (ByD) is an autosomal recessive disorder in which cholestasis of onset in infancy leads to hepatic fibrosis and death. Children who have a clinically similar disorder, but are not members of the Amish kindred in which ByD was described, are said to have Byler syndrome (ByS). Controversy exists as to whether ByD and ByS (subtypes of progressive familial intrahepatic cholestasis [PFIC]) represent one clinicopathological entity. The gene for ByD has been mapped to a 19-cM region of 18q21-q22. PFIC caused by a lesion in this region, including ByD, can be designated PFIC-1. Examination of haplotypes in siblings with ByS in two unrelated non-Amish families showed that the gene(s) responsible for their disorder(s) did not lie in the PFIC-1 candidate region. On light microscopy and transmission electron microscopy (TEM), liver tissue differed between Amish children with PFIC-1, who had coarsely granular bile and at presentation had bland intracanalicular cholestasis, and the children with ByS in the two non-Amish families, who had amorphous or finely filamentous bile and at presentation had neonatal hepatitis. Bile acid composition of bile also differed: In the Amish children with PFIC-1 and in one ByS family, the proportional concentration of chenodeoxycholic acid (CDCA) in bile was low compared with normal bile; in the other ByS family, it was only slightly reduced. Genetic analysis and light microscopy and TEM of liver may help distinguish PFIC-1 from other forms of ByS.

Development of pancreatic enzyme microsphere technology and US findings with Pancrease in the treatment of chronic pancreatitis.

Enzyme replacement therapy is a vital aspect of the management of patients with chronic pancreatic insufficiency. However, pharmaceutical technology in the manufacture of enzyme products has advanced significantly only in the last decade. In the late 1970s, Johnson & Johnson scientists developed novel pH-sensitive enteric-coated microspheres of pancrelipase (Pancrease) that could be encapsulated for convenient administration. The increased efficiency of the formulation allowed lower daily doses than had been required with conventional enzyme products. In vitro studies indicate that the microspheres disintegrate at a pH appropriate for patients with cystic fibrosis and chronic pancreatitis. Clinical studies of Pancrease in chronic pancreatitis demonstrate a significant improvement in fecal fat excretion, fat utilization, stool weight, and stool frequency, as well as significant weight gain and improved quality of life of patients. Pancrease represents a major advance in the clinical management of chronic pancreatic insufficiency.

Tetracycline instillation for recurrent cystic thyroid nodules.

Ten patients, 5 with recurrent pure cystic and 5 with recurrent mixed cystic-solid thyroid nodules, were studied. When repeated needle aspirations failed to decrease reaccumulation of fluid, tetracycline was injected directly into the cysts. As a result, all the pure cysts resolved completely, and four of the five mixed cystic-solid nodules diminished in size and fluid did not reaccumulate. Follow-up ranged from 12 to 20 months (mean 15 months). The procedure was generally well tolerated. It is concluded that instillation of tetracycline into both recurrent pure cystic and mixed cystic-solid thyroid lesions is a simple, safe and effective treatment.