Protein Kinase Type Research Articles

A genetic variation associated with plasma erythropoietin and a non-coding transcript of PRKAR1A in sickle cell disease.

Blood erythropoietin (EPO) increases primarily to hypoxia. In sickle cell anaemia (homozygous HBBE6V; HbSS), plasma EPO is elevated due to hemolytic anaemia-related hypoxia. Hydroxyurea treatment reduces haemolysis and anaemia by increasing foetal haemoglobin, which leads to lower hypoxic transcriptional responses in blood mononuclear cells but paradoxically further increases EPO. To investigate this apparent hypoxia-independent EPO regulation, we assessed two sickle cell disease (SCD) cohorts for genetic associations with plasma EPO, by prioritizing 237,079 quantitative trait loci for expression level and/or transcript isoform variations of 12,727 genes derived from SCD blood mononuclear cells. We found an association between the T allele of SNP rs60684937 and increased plasma EPO (n = 567, combined P = 5.5 × 10 − 8 adjusted for haemoglobin and hydroxyurea) and validated it in independent SCD patients (n = 183, P = 0.018). The T allele of rs60684937 was associated with a relatively increased expression of a non-coding transcript of PRKAR1A (cAMP-dependent protein kinase type I-alpha regulatory subunit) in 58 SCD patients (P = 7.9 × 10 − 7) and 58 HapMap Yoruba samples (P = 0.0011). In conclusion, we demonstrate that plasma EPO elevation with hydroxyurea in SCD is independent of hypoxic responses and that genetic variation at SNP rs60684937 may contribute to EPO regulation through a cAMP-dependent protein kinase A pathway.

Genome-Wide Identification and Expression Analysis of the Mitogen-Activated Protein Kinase Gene Family in Cassava.

Mitogen-activated protein kinases (MAPKs) play central roles in plant developmental processes, hormone signaling transduction, and responses to abiotic stress. However, no data are currently available about the MAPK family in cassava, an important tropical crop. Herein, 21 MeMAPK genes were identified from cassava. Phylogenetic analysis indicated that MeMAPKs could be classified into four subfamilies. Gene structure analysis demonstrated that the number of introns in MeMAPK genes ranged from 1 to 10, suggesting large variation among cassava MAPK genes. Conserved motif analysis indicated that all MeMAPKs had typical protein kinase domains. Transcriptomic analysis suggested that MeMAPK genes showed differential expression patterns in distinct tissues and in response to drought stress between wild subspecies and cultivated varieties. Interaction networks and co-expression analyses revealed that crucial pathways controlled by MeMAPK networks may be involved in the differential response to drought stress in different accessions of cassava. Expression of nine selected MAPK genes showed that these genes could comprehensively respond to osmotic, salt, cold, oxidative stressors, and abscisic acid (ABA) signaling. These findings yield new insights into the transcriptional control of MAPK gene expression, provide an improved understanding of abiotic stress responses and signaling transduction in cassava, and lead to potential applications in the genetic improvement of cassava cultivars.

Adiponectin corrects premature cellular senescence and normalizes antimicrobial peptide levels in senescent keratinocytes

Stress-induced premature senescence or aging causes dysfunction in the human somatic system. Adiponectin (Acrp30) plays a role in functional recovery, especially with adenosine 3′,5′-monophosphate (AMP)-activated protein kinase (AMPK) and silent mating type information regulation 2 homolog 1 (SIRT1). Acrp30 stimulation reduced the premature senescence positive ratio induced by hydrogen peroxide (H2O2) and restituted human β-defensin 2 (hBD-2) levels in senescent keratinocytes. Acrp30 recovered AMPK activity in senescent keratinocytes and increased SIRT1 deacetylation activity. As a result, FoxO1 and FoxO3 transcription activity was recovered. Additionally, Acrp30 stimulation suppresses NFκB p65, which induces abnormal expression of hBD-2 induced by H2O2. In the present study, we have shown that Acrp30 reduces premature senescence and recovers cellular function in keratinocytes. These results suggest a role for Acrp30 as an anti-aging agent to improve impaired skin immune barriers.

Electroacupuncture improves cognitive ability following cerebral ischemia reperfusion injury via CaM-CaMKIV-CREB signaling in the rat hippocampus.

The aim of the present study was to investigate the effect of electroacupuncture (EA) on cognitive deficits, and the underlying mechanism following cerebral ischemia-reperfusion (I/R) via the calmodulin (CaM)-calmodulin-dependent protein kinase type IV (CaMKIV)-cyclic adenosine monophosphate response elements binding protein (CREB) intracellular signaling pathway in the hippocampus. In total, 45 adult female Sprague-Dawley rats were randomly divided into three groups, namely the sham group, the middle cerebral artery occlusion (MCAO) group and the MCAO + EA group. Rats in the MCAO and MCAO + EA groups were modeled for post-stroke cognitive impairment. EA was performed at the Baihui and Shenting acupuncture points for 30 min/day for one week in the MCAO + EA group. Behavioral testing was analyzed using a step-down apparatus, while 2,3,5-triphenyl tetrazolium chloride was used to detect the infarct volume and lesion size. In addition, CaM activity was assessed by cyclic nucleotide-dependent phosphodiesterase analysis, and the protein expression levels of CaM, CaMKIV, phosphorylated (p)-CaMKIV, CREB and p-CREB were analyzed by western blot analysis. The cerebral I/R injured rat model in the MCAO group was established successfully with regard to the infarct volume and neuronal lesion size, as compared with the sham group. EA was demonstrated to effectively improve the cognitive ability, as measured by the step-down apparatus test, and decrease the infarct volume when compared with the MCAO group (P<0.05). The step-down apparatus test for the EA-treated rats revealed improved learning and reduced memory impairment when compared with the MCAO group. Furthermore, CaM activity and CaM protein expression levels in the MCAO + EA group were lower compared with those in the MCAO group (P<0.05). By contrast, the protein expression levels of CaMKIV, p-CaMKIV, CREB and p-CREB were significantly reduced in the MCAO group when compared with the sham group (P<0.05), although the expression levels increased following EA treatment when compared with the MCAO group (P<0.05). Therefore, cognitive repair benefited from EA, and the main intracellular signaling pathway in the hippocampus was mediated by CaM-CaMKIV-CREB. EA effectively inhibited the expression and activity of CaM, while further enhancing the expression of CaMKIV and CREB, and their associated phosphorylated functions.

Cutaneous tissue damage induces long-lasting nociceptive sensitization and regulation of cellular stress- and nerve injury-associated genes in sensory neurons

Tissue damage is one of the major etiological factors in the emergence of chronic/persistent pain, although mechanisms remain enigmatic. Using incision of the back skin of adult rats as a model for tissue damage, we observed sensitization in a nociceptive reflex enduring to 28days post-incision (DPI). To determine if the enduring behavioral changes corresponded with a long-term impact of tissue damage on sensory neurons, we examined the temporal expression profile of injury-regulated genes and the electrophysiological properties of traced dorsal root ganglion (DRG) sensory neurons. The mRNA for the injury/stress-hub gene Activating Transcription Factor 3 (ATF3) was upregulated and peaked within 4 DPI, after which levels declined but remained significantly elevated out to 28 DPI, a time when the initial incision appears healed and tissue-inflammation largely resolved. Accordingly, stereological image analysis indicated that some neurons expressed ATF3 only transiently (mostly medium-large neurons), while in others it was sustained (mostly small neurons), suggesting cell-type-specific responses. In retrogradely-traced ATF3-expressing neurons, Calcium/calmodulin-dependent protein kinase type IV (CAMK4) protein levels and isolectin-B4 (IB4)-binding were suppressed whereas Growth Associated Protein-43 (GAP-43) and Neuropeptide Y (NPY) protein levels were enhanced. Electrophysiological recordings from DiI-traced sensory neurons 28 DPI showed a significant sensitization limited to ATF3-expressing neurons. Thus, ATF3 expression is revealed as a strong predictor of single cells displaying enduring pain-related electrophysiological properties. The cellular injury/stress response induced in sensory neurons by tissue damage and indicated by ATF3 expression is positioned to contribute to pain which can occur after tissue damage.

The molecular, temporal and region-specific requirements of the beta isoform of Calcium/Calmodulin-dependent protein kinase type 2 (CAMK2B) in mouse locomotion.

Genetic approaches using temporal and brain region-specific restricted gene deletions have provided a wealth of insight in the brain regions and temporal aspects underlying spatial and associative learning. However, for locomotion such extensive studies are still scarce. Previous studies demonstrated that Camk2b–/– mice, which lack the β isoform of Calcium/Calmodulin-dependent protein kinase 2 (CAMK2B), show very severe locomotion deficits. However, where these locomotion deficits originate is unknown. Here we made use of novel Camk2b mutants (Camk2bf/f and Camk2bT287A), to explore the molecular, temporal and brain region-specific requirements of CAMK2B for locomotion. At the molecular level we found that normal locomotion requires Calcium/Calmodulin mediated activation of CAMK2B, but CAMK2B autonomous activity is largely dispensable. At a systems level, we found that global deletion of Camk2b in the adult mouse causes only mild locomotion deficits, suggesting that the severe locomotion deficits of Camk2b–/– mice are largely of developmental origin. However, early onset deletion of Camk2b in cerebellum, striatum or forebrain did not recapitulate the locomotion deficits, suggesting that these deficits cannot be attributed to a single brain area. Taken together, these results provide the first insights into the molecular, temporal and region-specific role of CAMK2B in locomotion.

Structure of the Dictyostelium Myosin-II Heavy Chain Kinase A (MHCK-A) α-kinase domain apoenzyme reveals a novel autoinhibited conformation.

The α-kinases are a family of a typical protein kinases present in organisms ranging from protozoa to mammals. Here we report an autoinhibited conformation for the α-kinase domain of Dictyostelium myosin-II heavy chain kinase A (MHCK-A) in which nucleotide binding to the catalytic cleft, located at the interface between an N-terminal and C-terminal lobe, is sterically blocked by the side chain of a conserved arginine residue (Arg592). Previous α-kinase structures have shown that an invariant catalytic aspartic acid residue (Asp766) is phosphorylated. Unexpectedly, in the autoinhibited conformation the phosphoryl group is transferred to the adjacent Asp663, creating an interaction network that stabilizes the autoinhibited state. The results suggest that Asp766 phosphorylation may play both catalytic and regulatory roles. The autoinhibited structure also provides the first view of a phosphothreonine residue docked into the phospho-specific allosteric binding site (Pi-pocket) in the C-lobe of the α-kinase domain.

Lateral habenula as a link between thyroid and serotoninergic system modiates depressive symptoms in hypothyroidism rats.

Depression-like behavior is observed in both rats and people with hypothyroidism, which suggests that altered thyroid hormone levels are closely associated with mental illness. Furthermore, decreased serotonin (5-hydroxytryptamine, 5-HT) levels are found in some brain regions of hypothyroid rats with depression-like behavior. However, the mechanism underlying the effects of hypothyroidism on the central serotonin system is unclear. The lateral habenula (LHb) is related to both the serotonin and thyroid systems and also plays an important role in the pathogenesis of depression. Our study aimed to disclose the role of the LHb in the onset of depression-like behavior in thyroidectomy (TD) rats. Forced swimming (FST) and open-field tests (OFT) were performed to measure behavioral changes in TD rats. The expression of β calmodulin-dependent protein kinase type II (β CaMKII) in the LHb, cytochrome C oxidase (COX) activity in the LHb and dorsal raphe nucleus (DRN), and 5-HT levels in the DRN were assayed. We found that TD rats exhibited depression-like behavior in the FST and OFT. Compared with the sham group, neural activity and the expression of β CaMKII in TD rats were higher in the LHb, and neural activity and 5-HT levels were lower in the DRN. Depressive behavior and decreased 5-HT levels in the DRN in TD rats were reversed by LHb lesioning. Our study indicates that depression-like behavior in TD rats can be attributed to decreased 5-HT levels in the DRN resulting from inhibition by an overactive LHb. The LHb mediates the effect of the thyroid system on 5-HT function in the DRN.

Role of CaMK II in primary somatosensory area and hippocampi in reduction of remifentanil-induced hyperalgesia by lidocaine in rats

Objective To investigate the role of calcium/calmodulin-dependent protein kinase Ⅱ (CaMKⅡ) in the primary somatosensory area (S1 area) and hippocampi in reduction of remifentanil-induced hyperalgesia by lidocaine in rats. Methods One hundred fifty-six male Sprague-Dawley rats, aged 8-10 weeks, weighing 240-260 g, were randomly allocated into 4 groups using a radom number table: control group (group C, n=6), remifentanil group (group R, n=50), lidocaine group (group L, n=50), and remifentanil+ lidocaine group (group RL, n=50). Remifentanil was given as a bolus of 6 mg/kg followed by an 2 h infusion of 2.4 μg·kg-1·min-1 in group R. Lidocaine was given as a bolus of 6 mg/kg followed by an infusion of 200 μg·kg-1·min-1 for 2 h in group L. In group RL, drug administration was similar to those previously described in R and L groups.The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured before administration and at 0.5, 2, 5 and 24 h after the end of administration.The rats were then sacrificed immediately after administration and at 0.5, 2, 5 and 24 h after the end of administration in R, L and RL groups, or at the corresponding time point in group C. The S1 area and hippocampi were isolated for determination of phosphorylated CaMKⅡ (p-CaMKⅡ) expression by Western blot. Results Compared with the value before administration, the MWT was significantly decreased at 0.5 and 2 h after the end of administration (P 0.05). Compared with group C, p-CaMKⅡ expression in the S1 area and hippocampi was significantly up-regulated immediately after administration and at 0.5 and 2 h after the end of administration in group R (P 0.05). Conclusion The mechanism by which lidocaine mitigates remifentanil-induced hyperalgesia is associated with inhibited activity of CaMKII in the S1 area and hippocampi of rats. Key words: Piperidines; Lidocaine; Hyperalgesia; Calcium; calmodulin dependent protein kinase type 2; Hippocampus; Somatosensory cortex

Oxidative Stress-Activated NHE1 Is Involved in High Glucose-Induced Apoptosis in Renal Tubular Epithelial Cells.

PurposeDiabetic nephropathy (DN) is a prevalent chronic microvascular complication of diabetes mellitus involving disturbances in electrolytes and the acid-base balance caused by a disorder of glucose metabolism. NHE1 is a Na+/H+ exchanger responsible for keeping intracellular pH (pHi) balance and cell growth. Our study aimed to investigate roles of NHE1 in high glucose (HG)-induced apoptosis in renal tubular epithelial cells.Materials and MethodsRenal epithelial tubular cell line HK-2 was cultured in medium containing 5 mM or 30 mM glucose. Then, cell apoptosis, oxidative stress, NHE1 expression, and pHi were evaluated. NHE1 siRNA and inhibitor were used to evaluate its role in cell apoptosis.ResultsHG significantly increased cell apoptosis and the production of reactive oxygen species (ROS) and 8-OHdG (p<0.05). Meanwhile, we found that HG induced the expression of NHE1 and increased the pHi from 7.0 to 7.6 after 48 h of incubation. However, inhibiting NHE1 using its specific siRNA or antagonist DMA markedly reduced cell apoptosis stimulated by HG. In addition, suppressing cellular oxidative stress using antioxidants, such as glutathione and N-acetyl cysteine, significantly reduced the production of ROS, accompanied by a decrease in NHE1. We also found that activated cyclic GMP-Dependent Protein Kinase Type I (PKG) signaling promoted the production of ROS, which contributed to the regulation of NHE1 functions.ConclusionOur study indicated that HG activates PKG signaling and elevates the production of ROS, which was responsible for the induction of NHE1 expression and dysfunction, as well as subsequent cell apoptosis, in renal tubular epithelial cells.

Time-dependent evolution of functional vs. remodeling signaling in induced pluripotent stem cell-derived cardiomyocytes and induced maturation with biomechanical stimulation.

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are a powerful platform for uncovering disease mechanisms and assessing drugs for efficacy/toxicity. However, the accuracy with which hiPSC-CMs recapitulate the contractile and remodeling signaling of adult cardiomyocytes is not fully known. We used β-adrenergic receptor (β-AR) signaling as a prototype to determine the evolution of signaling component expression and function during hiPSC-CM maturation. In "early" hiPSC-CMs (less than or equal to d 30), β2-ARs are a primary source of cAMP/PKA signaling. With longer culture, β1-AR signaling increases: from 0% of cAMP generation at d 30 to 56.8 ± 6.6% by d 60. PKA signaling shows a similar increase: 15.7 ± 5.2% (d 30), 49.8 ± 0.5% (d 60), and 71.0 ± 6.1% (d 90). cAMP generation increases 9-fold from d 30 to 60, with enhanced coupling to remodeling pathways (e.g., Akt and Ca(2+)/calmodulin-dependent protein kinase type II) and development of caveolin-mediated signaling compartmentalization. By contrast, cardiotoxicity induced by chronic β-AR stimulation, a major component of heart failure, develops much later: 5% cell death at d 30vs 55% at d 90. Moreover, β-AR maturation can be accelerated by biomechanical stimulation. The differential maturation of β-AR functionalvs remodeling signaling in hiPSC-CMs has important implications for their use in disease modeling and drug testing. We propose that assessment of signaling be added to the indices of phenotypic maturation of hiPSC-CMs.-Jung, G., Fajardo, G., Ribeiro, A. J. S., Kooiker, K. B., Coronado, M., Zhao, M., Hu, D.-Q., Reddy, S., Kodo, K., Sriram, K., Insel, P. A., Wu, J. C., Pruitt, B. L., Bernstein, D. Time-dependent evolution of functionalvs remodeling signaling in induced pluripotent stem cell-derived cardiomyocytes and induced maturation with biomechanical stimulation.

Insulin Does Not Target CamkIIα Neurones to Critically Regulate the Neuroendocrine Reproductive Axis in Mice.

Insulin signalling in the brain plays an important role in the central regulation of energy homeostasis and fertility, such that mice exhibiting widespread deletion of insulin receptors (InsR) throughout the brain and peripheral nervous system display diet sensitive obesity and hypothalamic hypogonadism. However, the specific cell types mediating the central effects of insulin on fertility remain largely unidentified. To date, the targeted deletion of InsR from individual neuronal populations implicated in the metabolic control of fertility has failed to recapitulate the hypogonadic and subfertile phenotype observed in brain-specific InsR knockout mice. Because insulin and leptin share similar roles as centrally-acting metabolic regulators of fertility, we used the Cre-loxP system to generate mice with a selective inactivation of the Insr gene from the same widespread neuronal population previously shown to mediate the central effects of leptin on fertility by crossing Insr-flox mice with calcium/calmodulin-dependent protein kinase type IIα (CamkIIα)-Cre mice. Multiple reproductive and metabolic parameters were then compared between male and female Insr-flox/Cre-positive (CamK-IRKO) and Insr-flox/Cre-negative control mice. Consistent with brain-specific InsR knockout mice, CamK-IRKO mice exhibited a mild but significant obesogenic phenotype. Unexpectedly, CamK-IRKO mice exhibited normal reproductive maturation and function compared to controls. No differences in the age of puberty onset, oestrous cyclicity or fecundity were observed between CamK-IRKO and control mice. We conclude that the central effects of insulin on the neuroendocrine reproductive axis are not critically mediated via the same neuronal populations targeted by leptin.

Tetrandrine prevents monocrotaline-induced pulmonary arterial hypertension in rats through regulation of the protein expression of inducible nitric oxide synthase and cyclic guanosine monophosphate-dependent protein kinase type 1

Pulmonary arterial hypertension (PAH) is a fatal disease characterized by a persistent elevation of pulmonary artery pressure and ventricular hypertrophy. Tetrandrine is a bisbenzylisoquinoline alkaloid that can decrease blood pressure, inhibit the proliferation of vascular smooth muscle cells, and block cardiac hypertrophy, but whether it has a therapeutic effect on PAH remains poorly defined. This study was undertaken to investigate the efficacy of tetrandrine on PAH. Forty-eight male Sprague-Dawley rats were randomly and equally divided into four groups. The control group was injected with normal saline; the others were injected with monocrotaline (MCT) to induce PAH, then treated with saline, tetrandrine, and vardenafil, respectively, from day 21 to day 42. On day 43, we measured the mean pulmonary artery pressure under general anesthesia, dissected the rat, and calculated the right ventricular hypertrophy index [right ventricle/(left ventricle plus septum)]. Later we observed the changes in the pulmonary vascular wall; measured the expression of cyclic guanosine monophosphate-dependent protein kinase type 1 and inducible nitric oxide synthase; measured the levels of superoxide dismutase, glutathione, malondialdehyde, and catalase; and then compared the results among groups. Compared with the MCT group, rats treated with tetrandrine had attenuated mean pulmonary artery pressure (20.48± 1.49 vs 30.07± 1.51; P< .01) and right ventricular hypertrophy index (49.19± 2.45 vs 68.50± 1.95; P<.01), inhibited proliferation of pulmonary artery smooth muscle cells, and improved endothelial function. Tetrandrine also upregulated the expression of protein kinase type 1 (90.86± 1.95 vs 67.34± 1.50; P< .01); downregulated the expression of inducible nitric oxide synthase (74.76± 1.48 vs 80.19± 0.28; P< .01); increased levels of superoxide dismutase (245.54± 12.98 vs 166.16± 21.42; P< .01), glutathione (0.699± 0.032 vs 0.514± 0.056; P< .01), and catalase (32.13± 2.33 vs 27.19± 2.72; P< .01); and decreased malondialdehyde (1.027± 0.039 vs 1.462± 0.055; P< .01). Tetrandrine alleviated MCT-induced PAH through regulation of nitric oxide signaling pathway and antioxidant and antiproliferation effects.

Cyclic GMP/cGMP-dependent protein kinase type I signaling and cysteine-rich LIM-only protein 4 in AngII/AT1R-induced heart hypertrophy and fibrosis

Background Cardiac hypertrophy is an adaptive response of the heart to many cardio-vascular disorders including hypertension, infarction and defects of the valves. Elevated levels of cardiac cyclic guanosine-3',5'-monophosphate (cGMP) activate cGMP-dependent protein kinase I (cGKI), which reportedly exhibited either anti-fibrotic and anti-hypertrophic effects or did not change the cardiac remodeling response. Based on these conflicting results, we and others suggested that the ability of natriuretic peptides (NP) to oppose detrimental changes via cGMP/cGKI might be strongly influenced by the stress stimuli that actually trigger the harmful events i.e. Angiotensin II (Ang II) modulating multiple reno-vascular and cardiac functions via G q-coupled Ang II type 1 receptors (AT1Rs). It is the overall aim of this study to dissect the molecular details underlying the cross-talk between the NP/cGMP/cGKI pathway and AngII/AT1R signaling in the heart.

A CNP-cGMP-cGKI-MAPK pathway promotes melanoma growth in vitro and in vivo in mice

Several important effects of nitric oxide (NO) and natriuretic peptides (ANP, BNP, & CNP) are mediated by cyclic guanosine monophosphate (cGMP) and cGMP-dependent protein kinase type I (cGKI). Recent studies indicated that the cGMP pathway might also play a role in tumorigenesis. However, the cellular and molecular mechanisms of cGMP's effects on tumor growth are not well understood. In the present study, the expression and function of components of the cGMP pathway in melanoma cell lines of murine and human origin were investigated. We found that murine B16 melanoma cells express cGKI and treatment of the cells with the cell-permeable cGMP analog 8-bromo-cGMP induced the phosphorylation of cGKI substrate proteins VASP and PDE5. CNP, a ligand of the particulate guanylyl cyclase GC-B, but not NO or ANP, induced robust cGMP signals in melanoma cells. Interestingly, activation of the CNP-cGMP-cGKI pathway increased the growth and migration of B16 cells through a crosstalk with p44/42 MAPK signaling. The PDE5 inhibitor sildenafil enhanced CNP-mediated cGMP signaling and the growth of murine melanoma cells. Similar results were obtained with human melanoma cell lines. In line with the in vitro data, we found that cGKI overexpression in melanoma cells or administration of sildenafil promoted melanoma growth in vivo in mice. Finally, expression of cGKI was also detected in cancer cells of human melanoma patients. Taken together, we have identified a CNP-cGMP-cGKI-MAPK pathway in melanoma cells, which stimulates melanoma cell growth in vitro and in vivo and might be novel therapeutic target for treatment of melanoma.

Cyclic GMP signaling and mitochondrial BK channels in cardioprotection against ischemia/reperfusion injury

Background By studying hearts obtained from global BK-null mice (BK-KO) in an ex vivo Langendorff perfusion setup we and others previously found evidence for mitochondrial BK channels (mitoBKs) in cardiomyocytes as infarctlimiting factors [1,2]. It is well established that canonical BK channels usually present at the plasma membrane of cells are directly stimulated by the cyclic guanosine3’,5’-monophosphate (cGMP)/cGMP-dependent protein kinase type I pathway; however, it is unclear whether cardioprotection afforded by cardiomyocyte (CM) cGMP in vivo requires mitoBK.

The role of particulate guanylyl cyclase B (GC-B) in auditory function in adult mice

Background cGMP signaling triggered by the binding of C-type natriuretic peptide (CNP) to its receptor guanylyl cyclase B (GC-B; NPR2; NPRB) has been linked by genetic evidence to a remarkable variety of physiological functions like skeletal bone growth, female fertility, cardiac growth, fat metabolism and gastrointestinal function. For the nervous system it has been recently demonstrated that the CNP/GC-B/cGMP/cGMP-dependent protein kinase type I (cGKI) signaling pathway is essential for sensory axon branching at the dorsal root entry zone of the spinal cord and at the rhombomeres of the hindbrain during embryonic development [1]. Also in the auditory system, distinct auditory nerve fiber (ANF) types that differ in their discharge rate and sound sensitivity bifurcate in the cochlear nucleus (CN), sending collaterals to the anteroventral, posteroventral, and dorsal subdivisions. The lack of GC-B has been shown to lead to a central phenotype [2].

GYY4137 attenuates remodeling, preserves cardiac function and modulates the natriuretic peptide response to ischemia

AimsMyocardial infarction followed by adverse left ventricular (LV) remodeling is the most frequent proximate cause of heart failure. Hydrogen sulfide (H2S) is an important endogenous modulator of diverse physiological and pathophysiological processes. Its role in post-ischemic ventricular remodeling and the associated neurohormonal responses has not been defined. Here, we aimed at evaluating whether the slow-releasing water-soluble H2S donor GYY4137 (GYY) exerts cardioprotective effects and modulates the neurohormonal response to cardiac ischemic injury. Methods and resultsTreatment for 2 or 7days with GYY (100mg/Kg/48h, IP) after acute myocardial infarction (MI) in rats preserved LV dimensions and function in vivo, compared to untreated infarcted (MI), placebo- and dl-propargylglycine- (PAG, an inhibitor of endogenous H2S synthesis) treated animals (n=9/group/time-point). LV dimensions and function in GYY-treated animals were comparable to healthy sham-operated rats. GYY-treated hearts had significantly less LV fibrosis than MI, placebo and PAG hearts. A higher density of blood vessels was found in the LV scar area of GYY-treated animals compared to all other infarcted groups. Despite preserved LV structure and function, treatment with GYY increased the levels of the natriuretic peptides ANP and BNP in association with enhanced cyclic GMP levels, paralleled by higher cGMP-dependent protein kinase type I (cGKI) protein levels. ConclusionsOur data suggest that the slow-releasing H2S donor, GYY4137, preserves cardiac function, attenuates adverse remodeling and may exert post-ischemic cardioprotective (pro-angiogenic, anti-apoptotic, anti-hypertrophic and anti-fibrotic) effects in part through enhanced early post-ischemic endogenous natriuretic peptide activation.

Association between miR-181b and PKG 1 in myocardial hypertrophy and its clinical implications.

The aim of this study was to explore the microRNA (miR)-181b expression in myocardial hypertrophy and to investigate its association with cGMP-dependent protein kinase type I (PKG 1) in an in vitro model. The miR-181b level in the peripheral blood was determined in patients with myocardial hypertrophy, and an in vitro model was established via phenylephrine (PE) treatment. Reverse transcription-quantitative polymerase chain reaction analysis and western blotting were performed to detect the expression levels of miR-181b, PKG 1 and hypertrophy-related genes. The results revealed that the expression of miR-181b was elevated in the peripheral blood of patients with myocardial hypertrophy, and this may have contributed to the pathology and progression of the disease. When the primary myocardial cells were treated with PE, microscopic observation and flow cytometry revealed significant hypertrophy. Furthermore, upregulation of myocardial hypertrophy-related genes, including β-myosin heavy chain, α-sarcomeric actinin and atrial natriuretic peptide, was observed. The miR-181b expression level in the PE-treated cells was elevated, while the mRNA and protein expression levels of PKG 1 were decreased, indicating a negative correlation between miR-181b and PKG 1 in myocardial hypertrophy. In addition, when the PE-treated primary myocardial cells were transfected with miR-181b inhibitor, the reduced PKG 1 expression was restored and the myocardial hypertrophy alleviated, as indicated by the reduced cellular sizes and decreased expression levels of the myocardial hypertrophy-related genes. In conclusion, miR-181b expression has been shown to be upregulated in myocardial hypertrophy, and this may play a role in the pathogenesis of the disease by regulating the expression of PKG 1. The present findings suggest that miR-181b is a promising molecular indicator for the clinical diagnosis and treatment of cardiac hypertrophy.

Sildenafil Does Not Prevent Heart Hypertrophy and Fibrosis Induced by Cardiomyocyte Angiotensin II Type 1 Receptor Signaling.

Analyses of several mouse models imply that the phosphodiesterase 5 (PDE5) inhibitor sildenafil (SIL), via increasing cGMP, affords protection against angiotensin II (Ang II)-stimulated cardiac remodeling. However, it is unclear which cell types are involved in these beneficial effects, because Ang II may exert its adverse effects by modulating multiple renovascular and cardiac functions via Ang II type 1 receptors (AT1Rs). To test the hypothesis that SIL/cGMP inhibit cardiac stress provoked by amplified Ang II/AT1R directly in cardiomyocytes (CMs), we studied transgenic mice with CM-specific overexpression of the AT1R under the control of the α-myosin heavy chain promoter (αMHC-AT1R(tg/+)). The extent of cardiac growth was assessed in the absence or presence of SIL and defined by referring changes in heart weight to body weight or tibia length. Hypertrophic marker genes, extracellular matrix-regulating factors, and expression patterns of fibrosis markers were examined in αMHC-AT1R(tg/+) ventricles (with or without SIL) and corroborated by investigating different components of the natriuretic peptide/PDE5/cGMP pathway as well as cardiac functions. cGMP levels in heart lysates and intact CMs were measured by competitive immunoassays and Förster resonance energy transfer. We found higher cardiac and CM cGMP levels and upregulation of the cGMP-dependent protein kinase type I with AT1R overexpression. However, even a prolonged SIL treatment regimen did not limit the progressive CM growth, fibrosis, or decline in cardiac functions in the αMHC-AT1R(tg/+) model, suggesting that SIL does not interfere with the pathogenic actions of amplified AT1R signaling in CMs. Hence, the cardiac/noncardiac cells involved in the cross-talk between SIL-sensitive PDE activity and Ang II/AT1R still need to be identified.