8 research outputs found
Single-Cell Transcriptional Analysis of Neuronal Progenitors
AbstractThe extraordinary cellular heterogeneity of the mammalian nervous system has largely hindered the molecular analysis of neuronal identity and diversity. In order to uncover mechanisms involved in neuronal differentiation and diversification, we have monitored the expression profiles of individual neurons and progenitor cells collected from dissociated tissue or captured from intact slices. We demonstrate that this technique provides a sensitive and reproducible representation of the single-cell transcriptome. In the olfactory system, hundreds of transcriptional differences were identified between olfactory progenitors and mature sensory neurons, enabling us to define the large variety of signaling pathways expressed by individual progenitors at a precise developmental stage. Finally, we show that regional differences in gene expression can be predicted from transcriptional analysis of single neuronal precursors isolated by laser capture from defined areas of the developing brain
Disruption of Protein Kinase A Interaction with A-kinase-anchoring Proteins in the Heart in Vivo: EFFECTS ON CARDIAC CONTRACTILITY, PROTEIN KINASE A PHOSPHORYLATION, AND TROPONIN I PROTEOLYSIS*
Protein kinase A (PKA)-dependent phosphorylation is regulated by targeting
of PKA to its substrate as a result of binding of regulatory subunit, R, to
A-kinase-anchoring proteins (AKAPs). We investigated the effects of disrupting
PKA targeting to AKAPs in the heart by expressing the 24-amino acid regulatory
subunit RII-binding peptide, Ht31, its inactive analog, Ht31P, or enhanced
green fluorescent protein by adenoviral gene transfer into rat hearts in
vivo. Ht31 expression resulted in loss of the striated staining pattern
of type II PKA (RII), indicating loss of PKA from binding sites on endogenous
AKAPs. In the absence of isoproterenol stimulation, Ht31-expressing hearts had
decreased +dP/dtmax and
-dP/dtmin but no change in left ventricular
ejection fraction or stroke volume and decreased end diastolic pressure
versus controls. This suggests that cardiac output is unchanged
despite decreased +dP/dt and -dP/dt. There
was also no difference in PKA phosphorylation of cardiac troponin I (cTnI),
phospholamban, or ryanodine receptor (RyR2). Upon isoproterenol
infusion, +dP/dtmax and
-dP/dtmin did not differ between Ht31 hearts and
controls. At higher doses of isoproterenol, left ventricular ejection fraction
and stroke volume increased versus isoproterenol-stimulated controls.
This occurred in the context of decreased PKA phosphorylation of cTnI,
RyR2, and phospholamban versus controls. We previously
showed that expression of N-terminal-cleaved cTnI (cTnI-ND) in transgenic mice
improves cardiac function. Increased cTnI N-terminal truncation was also
observed in Ht31-expressing hearts versus controls. Increased cTnI-ND
may help compensate for reduced PKA phosphorylation as occurs in heart
failure
Removal of the N-terminal Extension of Cardiac Troponin I as a Functional Compensation for Impaired Myocardial β-Adrenergic Signaling*
Although β-adrenergic stimuli are essential for myocardial
contractility, β-blockers have a proven beneficial effect on the
treatment of heart failure, but the mechanism is not fully understood. The
stimulatory G protein α-subunit (Gsα) couples the
β-adrenoreceptor to adenylyl cyclase and the intracellular cAMP response.
In a mouse model of conditional Gsα deficiency in the cardiac
muscle (Gsα-DF), we demonstrated heart failure phenotypes
accompanied by increases in the level of a truncated cardiac troponin I
(cTnI-ND) from restricted removal of the cTnI-specific N-terminal extension.
To investigate the functional significance of the increase of cTnI-ND in
Gsα-DF cardiac muscle, we generated double transgenic mice to
overexpress cTnI-ND in Gsα-DF hearts. The overexpression of
cTnI-ND in Gsα-DF failing hearts increased relaxation velocity and left
ventricular end diastolic volume to produce higher left ventricle maximum
pressure and stroke volume. Supporting the hypothesis that up-regulation of
cTnI-ND is a compensatory rather than a destructive myocardial response to
impaired β-adrenergic signaling, the aberrant expression of β-myosin
heavy chain in adult Gsα-DF but not control mouse hearts was
reversed by cTnI overexpression. These data indicate that the up-regulation of
cTnI-ND may partially compensate for the cardiac inefficiency in impaired
β-adrenergic signaling
Members of the miRNA-200 Family Regulate Olfactory Neurogenesis
MicroRNAs (miRNAs) are highly expressed in vertebrate neural tissues, but the contribution of specific miRNAs to the development and function of different neuronal populations is still largely unknown. We report that miRNAs are required for terminal differentiation of olfactory precursors in both mouse and zebrafish but are dispensable for proper function of mature olfactory neurons. The repertoire of miRNAs expressed in olfactory tissues contains over 100 distinct miRNAs. A subset, including the miR-200 family, shows high olfactory enrichment and expression patterns consistent with a role during olfactory neurogenesis. Loss of function of the miR-200 family phenocopies the terminal differentiation defect observed in absence of all miRNA activity in olfactory progenitors. Our data support the notion that vertebrate tissue differentiation is controlled by conserved subsets of organ-specific miRNAs in both mouse and zebrafish and provide insights into control mechanisms underlying olfactory differentiation in vertebrates
Gene regulation, alternative splicing, and posttranslational modification of troponin subunits in cardiac development and adaptation: a focused review
Ticagrelor in patients with diabetes and stable coronary artery disease with a history of previous percutaneous coronary intervention (THEMIS-PCI) : a phase 3, placebo-controlled, randomised trial
Background:
Patients with stable coronary artery disease and diabetes with previous percutaneous coronary intervention (PCI), particularly those with previous stenting, are at high risk of ischaemic events. These patients are generally treated with aspirin. In this trial, we aimed to investigate if these patients would benefit from treatment with aspirin plus ticagrelor.
Methods:
The Effect of Ticagrelor on Health Outcomes in diabEtes Mellitus patients Intervention Study (THEMIS) was a phase 3 randomised, double-blinded, placebo-controlled trial, done in 1315 sites in 42 countries. Patients were eligible if 50 years or older, with type 2 diabetes, receiving anti-hyperglycaemic drugs for at least 6 months, with stable coronary artery disease, and one of three other mutually non-exclusive criteria: a history of previous PCI or of coronary artery bypass grafting, or documentation of angiographic stenosis of 50% or more in at least one coronary artery. Eligible patients were randomly assigned (1:1) to either ticagrelor or placebo, by use of an interactive voice-response or web-response system. The THEMIS-PCI trial comprised a prespecified subgroup of patients with previous PCI. The primary efficacy outcome was a composite of cardiovascular death, myocardial infarction, or stroke (measured in the intention-to-treat population).
Findings:
Between Feb 17, 2014, and May 24, 2016, 11 154 patients (58% of the overall THEMIS trial) with a history of previous PCI were enrolled in the THEMIS-PCI trial. Median follow-up was 3·3 years (IQR 2·8–3·8). In the previous PCI group, fewer patients receiving ticagrelor had a primary efficacy outcome event than in the placebo group (404 [7·3%] of 5558 vs 480 [8·6%] of 5596; HR 0·85 [95% CI 0·74–0·97], p=0·013). The same effect was not observed in patients without PCI (p=0·76, p interaction=0·16). The proportion of patients with cardiovascular death was similar in both treatment groups (174 [3·1%] with ticagrelor vs 183 (3·3%) with placebo; HR 0·96 [95% CI 0·78–1·18], p=0·68), as well as all-cause death (282 [5·1%] vs 323 [5·8%]; 0·88 [0·75–1·03], p=0·11). TIMI major bleeding occurred in 111 (2·0%) of 5536 patients receiving ticagrelor and 62 (1·1%) of 5564 patients receiving placebo (HR 2·03 [95% CI 1·48–2·76], p<0·0001), and fatal bleeding in 6 (0·1%) of 5536 patients with ticagrelor and 6 (0·1%) of 5564 with placebo (1·13 [0·36–3·50], p=0·83). Intracranial haemorrhage occurred in 33 (0·6%) and 31 (0·6%) patients (1·21 [0·74–1·97], p=0·45). Ticagrelor improved net clinical benefit: 519/5558 (9·3%) versus 617/5596 (11·0%), HR=0·85, 95% CI 0·75–0·95, p=0·005, in contrast to patients without PCI where it did not, p interaction=0·012. Benefit was present irrespective of time from most recent PCI.
Interpretation:
In patients with diabetes, stable coronary artery disease, and previous PCI, ticagrelor added to aspirin reduced cardiovascular death, myocardial infarction, and stroke, although with increased major bleeding. In that large, easily identified population, ticagrelor provided a favourable net clinical benefit (more than in patients without history of PCI). This effect shows that long-term therapy with ticagrelor in addition to aspirin should be considered in patients with diabetes and a history of PCI who have tolerated antiplatelet therapy, have high ischaemic risk, and low bleeding risk