Background: Cardiac hypertrophy is a key biological response to injurious stresses such as pressure overload and when excessive can lead to heart failure. Innate immune activation by danger signals, through intracellular pattern recognition receptors such as nucleotide-binding oligomerization domain-containing protein 1(Nod1) and its adaptor receptor-interacting protein 2 (RIP2), might play a major role in cardiac remodeling and progression to heart failure. We hypothesize that Nod1/RIP2 are major contributors to cardiac hypertrophy, but may not be sufficient to fully express the phenotype alone. Methods: To elucidate the contribution of Nod1/RIP2 signaling to cardiac hypertrophy, we randomized Nod1 -/- , RIP2 -/- or wild-type (WT) mice to transverse aortic constriction (TAC) or sham operations. Cardiac hypertrophy, fibrosis, and cardiac function were examined in these mice. Results: Nod1 and RIP2 proteins were up-regulated in the heart after TAC, and this was paralleled by increased expression of mitochondrial proteins, including mitochondrial antiviral signaling protein (MAVS). Nod1 -/- and RIP2 -/- mice subjected to TAC exhibited better survival, improved cardiac function and decreased cardiac hypertrophy. Downstream signal transduction pathways that regulate inflammation and fibrosis including NF-κB and MAPK-GATA4/p300, were reduced in both Nod1 -/- and RIP2 -/- mice after TAC compared with WT mice. Co-immunoprecipitation of extracted cardiac proteins and confocal immunofluorescence microscopy showed that Nod1/ RIP2 interaction was robust and that this complex also included MAVS as an essential component. Suppression of MAVS expression attenuated the complex formation, NF-κB signalling and myocyte hypertrophy. Interrogation of mitochondrial function compared in the presence or ablation of MAVS revealed that MAVS serves to suppress mitochondrial energy output and mediate fission/fusion related dynamic changes. The latter is possibly linked to mitophagy during cardiomyocytes stress, which may provide an intriguing link between innate immune activation and mitochondrial energy balance under stress or injury conditions. Conclusions: We have identified that innate immune Nod1/RIP2 signaling is a major contributor to cardiac remodeling following stress. This process is critically joined by and regulated through the mitochondrial danger signal adapter MAVS. This novel complex coordinates remodeling, inflammatory response and mitochondrial energy metabolism in stressed cardiomyocytes. Thus Nod1/RIP2/MAVS signaling complex may represent an attractive new therapeutic approach toward heart failure.
BackgroundCarbapenem-resistant K. pneumoniae 2297, isolated from a patient treated with tigecycline for pneumonia, developed tigecycline resistance, in contrast to carbapenem-resistant isolate 1215, which was collected four months prior to the 2297 isolate. Mechanisms underlying tigecycline resistance were elucidated for the clinical isolates.MethodsThe tigecycline minimum inhibitory concentration (MIC) was determined using the broth microdilution method, with or without phenylalanine-arginine β-naphthylamide (PABN), and whole-genome sequencing was carried out by single-molecule real-time sequencing. The expression levels of the genes acrA, oqxA, ramA, rarA, and rpoB were determined by reverse-transcription quantitative PCR.ResultsBoth isolates presented identical antibiograms, except for tigecycline, which showed an MIC of 0.5 mg/L in 1215 and 2 mg/L in 2297. The addition of PABN to tigecycline-resistant 2297 caused a four-fold decrease in the tigecycline MIC to 0.5 mg/L, although acrA expression (encoding the AcrAB efflux pump) was upregulated by 2.5 fold and ramA expression (encoding the pump activator RamA) was upregulated by 1.4 fold. We identified a 6,096-bp fragment insertion flanking direct TATAT repeats that disrupted the romA gene located upstream of ramA in the chromosome of K. pneumoniae 2297; the insertion led the ramA gene promoter replacement resulting in stronger activation of the gene.ConclusionsThe K. pneumoniae isolate developed tigecycline resistance during tigecycline treatment. It was related to the overexpression of the AcrAB resistance-nodulation-cell division efflux system due to promoter replacement.
The fast transient outward potassium current (I to,f ) plays a critical role in the electrical and contractile properties of the myocardium. I to,f channels are formed by the co-assembly of the pore-forming ␣-subunits, Kv4.2 and Kv4.3, together with the accessory -subunit KChIP2. Reductions of I to,f are common in the diseased heart, which is also associated with enhanced stimulation of -adrenergic receptors (-ARs). We used cultured neonatal rat ventricular myocytes to examine how chronic -AR stimulation decreases I to,f . To determine which downstream pathways mediate these I to,f changes, adenoviral infections were used to inhibit CaMKII␦c, CaMKII␦b, calcineurin, or nuclear factor B (NF-B). We observed that chronic -AR stimulation with isoproterenol (ISO) for 48 h reduced I to,f along with mRNA expression of all three of its subunits (Kv4.2, Kv4.3, and KChIP2). Inhibiting either CaMKII␦c nor CaMKII␦b did not prevent the ISO-mediated I to,f reductions, even though CaMKII␦c and CaMKII␦b clearly regulated I to,f and the mRNA expression of its subunits. Likewise, calcineurin inhibition did not prevent the I to,f reductions induced by -AR stimulation despite strongly modulating I to,f and subunit mRNA expression. In contrast, NF-B inhibition partly rescued the ISO-mediated I to,f reductions in association with restoration of KChIP2 mRNA expression. Consistent with these observations, KChIP2 promoter activity was reduced by p65 as well as -AR stimulation. In conclusion, NF-B, and not CaMKII␦ or calcineurin, partly mediates the I to,f reductions induced by chronic -AR stimulation. Both mRNA and KChIP2 promoter data suggest that the ISO-induced I to,f reductions are, in part, mediated through reduced KChIP2 transcription caused by NF-B activation.Fast cardiac transient outward potassium currents are generated by channels comprised of voltage-gated ␣-pore-forming subunits (which in humans/canines is predominantly Kv4.3, and in rodents is Kv4.2 and Kv4.3) and the accessory -subunit KChIP2 (1). These currents play a critical role in early cardiac repolarization (1), excitation contraction-coupling (2, 3), and arrhythmias (4). I to,f and its molecular constituents are invariably reduced in cardiac hypertrophy and disease (1), and heart disease is also characterized by elevations in catecholamines and consequently enhanced activation of -adrenergic receptors (-AR) 6 (5). Although chronic -AR stimulation has been shown to decrease I to,f (6), the molecular mechanisms underlying regulation of I to,f by -ARs remain unclear.I to,f is regulated by several pathways activated in the diseased myocardium, such as calcineurin, nuclear factor-activated T-cells (NFAT) (6, 7) and mitogen activated protein kinases (8). Although calcineurin/NFAT signaling regulates I to,f in a manner that is model dependent (6, 9, 10), this pathway does not mediate ␣-AR-induced changes in I to,f or its molecular subunits (9,10). We previously showed that ␣-AR stimulation activates the transcription factor nuclear factor B (NF-B), which media...
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