Most physiological systems show daily variations in functional output, entrained to the day-night cycle. Humans exhibit a daily rhythm in urinary voiding (micturition), and disruption of this rhythm (nocturia) has significant clinical impact. However, the underlying mechanisms are not well-understood. Recently, a circadian rhythm in micturition was demonstrated in rodents, correlated with functional changes in urodynamics, providing the opportunity to address this issue in an animal model. Smooth muscle cells from mouse bladder have been proposed to express a functional and autonomous circadian clock at the molecular level. In this study, we addressed whether a semi-intact preparation of mouse urinary bladder smooth muscle (UBSM) exhibited measurable differences in contractility between day and night. UBSM tissue strips were harvested at four time points over the diurnal cycle, and spontaneous (phasic) and nerve-evoked contractions were assessed using isometric tension recordings. During the active period (ZT12-24) when micturition frequency is higher in rodents, UBSM strips had no significant differences in maximal- (high K(+)) or nerve-evoked contractions compared to strips harvested from the resting period (ZT0-12). However, a diurnal rhythm in phasic contraction was observed, with higher amplitudes at ZT10. Consistent with the enhanced phasic amplitudes, expression of the BK K(+) channel, a key suppressor of UBSM excitability, was lower at ZT8. Higher expression of BK at ZT20 was correlated with an enhanced effect of the BK antagonist paxilline (PAX) on phasic amplitude, but PAX had no significant time-of-day dependent effect on phasic frequency or nerve-evoked contractions. Overall, these results identify a diurnal difference for one contractile parameter of bladder muscle. Taken together, the results suggest that autonomous clocks in UBSM make only a limited contribution to the integrated control of diurnal micturition patterns.

Chronic volume overload induces multiple cardiac remodeling processes that finally result in eccentric cardiac hypertrophy and heart failure. We have hypothesized that chronic angiotensin-converting enzyme (ACE) inhibition by trandolapril might affect various remodeling processes differentially, thus allowing their dissociation. Cardiac remodeling due to chronic volume overload and the effects of trandolapril were investigated in rats with an aortocaval fistula (ACF rats). The aortocaval shunt was created using a needle technique and progression of cardiac remodeling to heart failure was followed for 24 weeks. In ACF rats, pronounced eccentric cardiac hypertrophy and contractile and proarrhythmic electrical remodeling were associated with increased mortality. Trandolapril substantially reduced the electrical proarrhythmic remodeling and mortality, whereas the effect on cardiac hypertrophy was less pronounced and significant eccentric hypertrophy was preserved. Effective suppression of electrical proarrhythmic remodeling and mortality but not hypertrophy indicates that the beneficial therapeutic effects of ACE inhibitor trandolapril in volume overload heart failure might be dissociated from pure antihypertrophic effects.

Vascular smooth muscle cell (VSMC) phenotypic transition represents the fundamental pathophysiological alteration in the vascular remodeling process during the initiation and progression of cardiovascular diseases. Recent studies have revealed that Icariside II (ICS-II), a flavonol glycoside derived from the traditional Chinese medicine Herba Epimedii, exhibited therapeutic effects in various cardiovascular diseases. However, the therapeutic efficacy and underlying mechanisms of ICS-II regarding VSMC phenotypic transition were unknown. In this study, we investigated the therapeutic effects of ICS-Ⅱ on vascular remodeling with a rat's balloon injury model in vivo. The label-free proteomic analysis was further implemented to identify the differentially expressed proteins (DEPs) after ICS-II intervention. Gene ontology and the pathway enrichment analysis were performed based on DEPs. Moreover, platelet-derived growth factor (PDGF-BB)-induced primary rat VSMC was implemented to verify the restoration effects of ICS-II on the VSMC contractile phenotype. Results showed that ICS-II could effectively attenuate the vascular remodeling process, promote SMA-α protein expression, and inhibit OPN expression in vivo. The proteomic analysis identified 145 differentially expressed proteins after ICS-II intervention. Further, the bioinformatics analysis indicated that the focal adhesion signaling pathway was enriched in the ICS-II group. In vitro studies showed that ICS-II suppressed VSMC proliferation and migration, and promoted VSMC contractile phenotype by modulating the focal adhesion signaling pathway. Taken together, our results suggest that ICS-II attenuates the vascular remodeling process and restores the VSMC contractile phenotype by promoting the focal adhesion pathway.

The main adverse effect of tyrosine kinase inhibitors, such as sunitinib, is cardiac contractile dysfunction; however, the molecular mechanisms of this effect remain largely obscure. MicroRNAs (miRNAs) are key regulatory factors in both cardiovascular diseases and the tyrosine kinase pathway. Therefore, we analyzed the differential expression of miRNAs in the myocardium in mice after exposure to sunitinib using miRNA microarray. A significant downregulation of miR-146a was observed in the myocardium of sunitinib-treated mice, along with a 20% decrease in left ventricle ejection fraction (LVEF). The downregulation of miR-146a was further validated by RT-qPCR. Among the potential targets of miR-146a, we focused on Pln and Ank2, which are closely related to cardiac contractile dysfunction. Results of luciferase reporter assay confirmed that miR-146a directly targeted the 3' untranslated region of Pln and Ank2. Significant upregulation of PLN and ANK2 at the mRNA and protein levels was observed in the myocardium of sunitinib-treated mice. Cardiac-specific overexpression of miR-146a prevented the deteriorate effect of SNT on calcium transients, thereby alleviating the decreased contractility of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). SiRNA knockdown of PLN or ANK2 prevented sunitinib-induced suppression of contractility in hiPSC-CMs. Therefore, our in vivo and in vitro results showed that sunitinib downregulated miR-146a, which contributes to cardiac contractile dysfunction by regulating the downstream targets PLN and ANK2, and that upregulation of miR-146a alleviated the inhibitory effect of SNT on cardiac contractility. Thus, miR-146a could be a useful protective agent against sunitinib-induced cardiac dysfunction.

Inflammatory conditions of the urinary bladder have been shown to be associated with urothelial damage and loss of function. The purinergic P2X7 receptor has been implicated in several inflammatory conditions. The aim of this study was to investigate the role of the P2X7 receptor in acrolein-induced inflammatory damage using the porcine urinary bladder. For this purpose, an ex-vivo model of porcine urothelial damage induced by direct instillation of acrolein into the whole bladder lumen was used. To determine the role of the P2X7 receptor, the bladders were pre-incubated with a selective P2X7 receptor antagonist, A804598 (10 μM), for 1 h. The effects of the acrolein-induced urothelial damage on the bladder's function were assessed by examining the bladder wall contractile response, structure changes, apoptosis, and oxidative stress in the bladder tissues. The acrolein treatment led to significant damage to the urothelium histology, tight junction expression, and contractile responses. Acrolein also induced apoptosis in the mucosa layer. All these acrolein-induced responses were attenuated by pre-treatment with the P2X7 receptor antagonist A804598. Acrolein also significantly induced DNA oxidation in the submucosal layer; however, the P2X7 receptor antagonism did not show any protective effect towards the acrolein-induced oxidative stress. These findings suggested that the P2X7 receptor is involved in the acrolein-induced damage to the urothelium; therefore, the P2X7 receptor antagonists may be a new therapeutic option for the treatment of bladder inflammation.

The etiology of diarrhea-predominant irritable bowel syndrome (IBS-D) is complicated and closely related to neurotransmission in the gastrointestinal (GI) tract. Developing new strategies for treating this disease is a major challenge for IBS-D research. Berberine hydrochloride (BBH), the derivative of berberine, is a herbal constituent used to treat IBS. Previous studies have shown that BBH has potential anti-inflammatory, antibacterial, analgesic, and antidiarrheal effects and a wide range of biological activities, especially in regulating the release of some neurotransmitters. A modified IBS-D rat model induced by chronic restraint stress was used in all experiments to study the effects of BBH on the GI tract. This study measured the abdominal withdrawal reflex (AWR) response to graded colorectal distention (CRD; 20, 40, 60, and 80 mmHg) and observed the fecal areas of stress-induced IBS-D model. Experiments were conducted using organ bath techniques, which were performed in vitro using strips of colonic longitudinal smooth muscle. Inhibitory and excitatory neurotransmitter agents were added to each organ bath to observe contractile responses on the strips and the treatment effect exerted by BBH. The IBS-D rat model was successfully induced by chronic restraint stress, which resulted in an increased defecation frequency and visceral hypersensitivity similar to that of humans. BBH could reduce 4-h fecal areas and AWR response to CRD in IBS-D. The stress-induced IBS-D model showed upregulated colonic mRNA expression levels of 5-hydroxytryptamine-3A receptor and downregulated expression levels of neuronal nitric oxide synthase. Meanwhile, BBH could reverse this outcome. The responses of substances that regulate the contraction induced by related neurotransmission in the longitudinal smooth muscle of IBS-D colon (including the agonist of acetylcholine, carbachol; NOS inhibitor, L-NAME; and P2Y1 receptor antagonist, MRS2500) can be inhibited by BBH. In summary, BBH promotes defecation frequency and visceral hypersensitivity in IBS-D and exerts inhibitory effects on contractile responses in colonic longitudinal smooth muscle. Thus, BBH may represent a new therapeutic approach for treating IBS-D.

Smooth muscle (SM) contraction is one of the important physiological functions of the human body, and SM abnormal contraction will induce many diseases. The phosphorylated regulatory light chains (p-RLC) play a decisive role in SM contraction, and dephosphorylation of p-RLC is an effective way to relax SM. Our previous study showed that the novel benzylisoquinoline alkaloid, neoliensinine (Neo), could relax microvascular SM contracted by KCl hyperpolarization. In this study, mesenteric capillaries isolated from 45 mice were divided into normal tension group (Control), 124 mM KCl induced contraction model group (Model), and KCl and Neo-treatment group (Drug). The dephosphorylation levels of RLC in the three groups were measured. Compared with the model group, the phosphorylation of RLC in the drug group was decreased dramatically as expected, suggesting that the relaxation effect of Neo was caused by downregulating p-RLC of microvessel SM. In order to fully understand its fundamental mechanism, our research focused on the identification of target proteins in mice with KCl-induced contractile mesenteric capillary. Isobaric tags for relative and absolute quantification (ITRAQ) tagging was carried out by nanospray liquid chromatography-tandem mass spectrometry. The results allowed the upregulation of 164 differential abundance proteins (DAPs) among the 3,474 protein abundance disturbances identified from the model/control samples. Further comparison showed that there were 16 DAP convergences associated with vascular SM contraction between the drug/model and the drug/control samples. Among them, two proteins with known function, PLCβ and RhoGEF12, were selected as target proteins of the relaxation effect of Neo. The two selective target DAPs were verified by Western blot at protein level. The results suggested that changes of the two proteins were consistent with that of the iTRAQ results. Our present work reveals that Neo relaxes vascular smooth muscle via inhibition of RLC phosphorylation, and PLCβ and RhoGEF12 may be potential biomarkers for evaluating the effects mediated by Neo.

Introduction: Corydalis yanhusuo total alkaloids (CYTA) are the primary active ingredients in yanhusuo, known for their analgesic and cardioprotective effects. However, the mechanisms underlying the treatment of Myocardial ischemia (MI) with CYTA have not been reported. The purpose of this study was to explore the protective effect of CYTA on MI and its related mechanisms. Methods: A network pharmacology was employed to shed light on the targets and mechanisms of CYTA's action on MI. The protective effect of CYTA against hypoxia damage was evaluated in H9c2 cells. Furthermore, the effects of CYTA on L-type Ca2+ current (ICa-L), contractile force, and Ca2+ transient in cardiomyocytes isolated from rats were investigated using the patch clamp technique and IonOptix system. The network pharmacology revealed that CYTA could regulate oxidative stress, apoptosis, and calcium signaling. Cellular experiments demonstrated that CYTA decreased levels of CK, LDH, and MDA, as well as ROS production and Ca2+ concentration. Additionally, CYTA improved apoptosis and increased the activities of SOD, CAT, and GSH-Px, along with the levels of ATP and Ca2+-ATPase content and mitochondrial membrane potential. Moreover, CYTA inhibited ICa-L, cell contraction, and Ca2+ transient in cardiomyocytes. Results: These findings suggest that CYTA has a protective effect on MI by inhibiting oxidative stress, mitochondrial damage, apoptosis and Ca2+ overload. Discussion: The results prove that CYTA might be a potential natural compound in the field of MI treatment, and also provide a new scientific basis for the its utilization.

Hypertension is a major risk factor for stroke and cardiovascular events in clinic, which is accompanied by the abnormality of vascular tone and endothelial dysfunction of small artery. Here we report that Kang Le Xin (KLX), a novel anthraquinones compound, could reduce blood pressure and the underlying mechanisms involves that KLX induces endothelium-dependent vasodilation. KLX significantly decreases the arterial blood pressure of spontaneous hypertensive rats (SHR), decreases the contractile reactivity of superior mesenteric artery to phenylephrine and increases the vasodilatory reactivity of superior mesenteric artery to carbachol in a dose-dependent manner. Besides, KLX reduces vascular tension of endothelium-intact mesenteric artery pre-constricted with phenylephrine in a dose-dependent manner, while this effect is inhibited by depriving vascular endothelium or pretreating vascular rings with L-NAME (endothelial nitric oxide synthase inhibitor) or compound C (AMP-activated protein kinase inhibitor). Moreover, KLX increases nitric oxide (NO) generation, endothelial nitric oxide synthase (eNOS), AKT and AMP-activated protein kinase (AMPK) phosphorylation in cultured human umbilical vein endothelial cells (HUVECs), while these effects are inhibited by pretreating cells with compound C. In conclusion, KLX is a new compound with the pharmacological action of reducing arterial blood pressure. The underlying mechanism involves KLX induces endothelium-dependent vasodilation through activating AMPK-AKT-eNOS signaling pathway.

Post-cardiac arrest myocardial dysfunction significantly contributes to early mortality after the return of spontaneous circulation. However, no effective therapy is available now. Aldehyde dehydrogenase 2 (ALDH2) enzyme has been shown to protect the heart from aldehyde toxicity such as 4-hydroxy-2-nonenal (4-HNE) and oxidative stress. In this study, we evaluated the effect of enhanced activity or expression of ALDH2 on post-cardiac arrest myocardial dysfunction and survival in a rat cardiac arrest model. Furthermore, we elucidated the underlying mechanisms with a focus on mitochondrial reactive oxygen species (ROS) production in a cell hypoxia/reoxygenation model. A total of 126 rats were used for the ALDH2 activation or cardiac overexpression of ALDH2 studies. Randomization was done 10 min before the respective agonist injection or in vivo gene delivery. We showed that enhanced activity or expression of ALDH2 significantly improved contractile function of the left ventricle and survival rate in rats subjected to cardiac arrest-cardiopulmonary resuscitation procedure. Moreover, ALDH2 prevented cardiac arrest-induced cardiomyocyte death from apoptosis and mitochondrial damage. Mechanistically, 4-HNE, a representative substrate of ALDH2, was dominantly increased in the hypoxia/reoxygenation-exposed cardiomyocytes. Direct addition of 4-HNE led to significantly augmented succinate accumulation and mitochondrial ROS production. Through metabolizing 4-HNE, ALDH2 significantly inhibited mitochondrial ROS production. Our findings provide compelling evidence of the cardioprotective effects of ALDH2 and therapeutic targeting this enzyme would provide an important approach for treating post-cardiac arrest myocardial dysfunction.

Background: Norbormide (NRB) is a selective rat toxicant endowed with vasoconstrictor activity confined to the rat peripheral arteries. In a recent work we used a fluorescent derivative of NRB (NRB-AF12), obtained by coupling the NBD fluorophore to the parent molecule via a linker, in order to gain information about the possible site of action of the unlabeled compound. We found that NRB-AF12 labeled intracellular organelles in both NRB-sensitive and -insensitive cells and we accordingly proposed its use as a scaffold for the development of a new class of fluorescent probes. In this study, we examined the fluorescent properties of a BODIPY FL-conjugated NRB probe (MC009) developed: (A) to verify if NRB distribution could be influenced by the attached fluorophore; (B) to improve the fluorescent performance of NRB-AF12. Methods: MC009 characteristics were investigated by confocal fluorescence microscopy, in freshly isolated rat caudal artery myocytes (FIRCAM) and in LX2 cells, representative of NRB-sensitive and insensitive cells, respectively. Main results: In both FIRCAM and LX2 cells MC009 stained endoplasmic reticulum, mitochondria, Golgi apparatus and lipid droplets, revealing the same intracellular distribution as NRB-AF12, and, at the same time, had both improved photostability and gave a more intense fluorescent signal at lower concentrations than was possible with NRB-AF12, which resulted in a better and finer visualization of intracellular structures. Furthermore, MC009 was effective in cellular labeling in both living and fixed cells. At the concentration used to stain the cells, MC009 did not show any cytotoxic effect and did not affect the regular progression of cell cycle and division. Conclusions: This study demonstrates that the distribution of fluorescently labeled NRB is not affected by the type of fluorophore attached to the parent compound, supporting the idea that the localization of the fluorescent derivatives may reasonably reflect that of the parent compound. In addition, we observed a marked improvement in the fluorescent properties of BODIPY FL-conjugated NRB (MC009) over its NBD-derived counterpart (NRB-AF12), confirming NRB as a scaffold for the development of new, high performance, non-toxic fluorescent probes for the labeling of intracellular structures in both living and fixed cells.

Myocardial contractile dysfunction in diabetic cardiomyocytes is a significant promoter of heart failure. Herein, we investigated the effect of icariin, a flavonoid monomer isolated from Epimedium, on diabetic cardiomyopathy (DCM) and explored the mechanisms underlying its unique pharmacological cardioprotective functions. High glucose (HG) conditions were simulated in vitro using cardiomyocytes isolated from neonatal C57 mice, while DCM was stimulated in vivo in db/db mice. Mice and cardiomyocytes were treated with icariin, with or without overexpression or silencing of Apelin and Sirt3 via transfection with adenoviral vectors (Ad-RNA) and specific small hairpin RNAs (Ad-sh-RNA), respectively. Icariin markedly improved mitochondrial function both in vivo and in vitro, as evidenced by an increased level of mitochondrial-related proteins via western blot analysis (PGC-1α, Mfn2, and Cyt-b) and an increased mitochondrial membrane potential, as observed via JC-1 staining. Further, icariin treatment decreased cardiac fibrogenesis (Masson staining), and inhibited apoptosis (TUNEL staining). Together, these changes improved cardiac function, according to multiple transthoracic echocardiography parameters, including LVEF, LVSF, LVESD, and LVEDD. Moreover, icariin significantly activated Apelin and Sirt3, which were inhibited by HG and DCM. Importantly, when Ad-sh-Apelin and Ad-sh-Sirt3 were transfected in cardiomyocytes or injected into the heart of db/db mice, the cardioprotective effects of icariin were abolished and mitochondrial homeostasis was disrupted. Further, it was postulated that since Ad-Apelin induced different results following increased Sirt3 expression, icariin may have attenuated DCM development by preventing mitochondrial dysfunction through the Apelin/Sirt3 pathway. Hence, protection against mitochondrial dysfunction using icariin may prove to be a promising therapeutic strategy against DCM in diabetes.

The number of people taking statins is set to increase across the globe due to recent changes in prescription guidelines. For example, half the US population over 40 is now eligible for these drugs, whether they have high serum cholesterol or not. With such development in policy comes a stronger need for understanding statins' myriad of effects. Surprisingly little is known about possible direct actions of statins on cardiac myocytes, although claims of a direct myocardial toxicity have been made. Here, we determine the impact of simvastatin administration (40 mg/kg/day) for 2 weeks in normocholesterolemic rats on cardiac myocyte contractile function and identify an underlying mechanism. Under basal conditions, statin treatment increased the time to half (t0.5) relaxation without any effect on the magnitude of shortening, or the magnitude/kinetics of the [Ca2+]i transient. Enhanced myocyte lusitropy could be explained by a corresponding increase in phosphorylation of troponin I (TnI) at Ser23,24. Statin treatment increased expression of eNOS and Ser1177 phosphorylated eNOS, decreased expression of the NOS-inhibitory proteins caveolins 1 and 3, and increased (P = 0.06) NO metabolites, consistent with enhanced NO production. It is well-established that NO stimulates protein kinase G, one of the effectors of TnI phosphorylation at Ser23,24. Trends for parallel changes in phospho-TnI, phospho-eNOS and caveolin 1 expression were seen in atrial muscle from patients taking statins. Our data are consistent with a mechanism whereby chronic statin treatment enhances TnI phosphorylation and myocyte lusitropy through increased NO bioavailability. We see no evidence of impaired function with statin treatment; the changes we document at the level of the cardiac myocyte should facilitate diastolic filling and cardiac performance.

Palmitoylethanolamide (PEA), an endogenous lipid mediator, is emerging as a promising pharmacological agent in multiple neurodegenerative disorders for its anti-inflammatory and neuroprotective properties. However, its effects on enteric inflammation and colonic dysmotility associated with Alzheimer's disease (AD) are lacking. This study was designed to investigate the beneficial effect of PEA administration in counteracting the enteric inflammation and relieving the bowel motor dysfunctions in an AD mouse model, SAMP8 mice. In addition, the ability of PEA in modulating the activation of enteric glial cells (EGCs), pivotally involved in the pathophysiology of bowel dysfunctions associated with inflammatory conditions, has also been examined. SAMP8 mice at 4 months of age were treated orally with PEA (5 mg/kg/day) for 2 months. SAMR1 animals were employed as controls. At the end of treatment, parameters dealing with colonic motility, inflammation, barrier integrity and AD protein accumulation were evaluated. The effect of PEA on EGCs was tested in cultured cells treated with lipopolysaccharide (LPS) plus β-amyloid 1-42 (Aβ). SAMP8 treated with PEA displayed: 1) an improvement of in vitro colonic motor activity, citrate synthase activity and intestinal epithelial barrier integrity and 2) a decrease in colonic Aβ and α-synuclein (α-syn) accumulation, S100-β expression as well as enteric IL-1β and circulating LPS levels, as compared with untreated SAMP8 mice. In EGCs, treatment with PEA counteracted the increment of S100-β, TLR-4, NF-κB p65 and IL-1β release induced by LPS and Aβ. These results suggest that PEA, under a condition of cognitive decline, prevents the enteric glial hyperactivation, reduces AD protein accumulation and counteracts the onset and progression of colonic inflammatory condition, as well as relieves intestinal motor dysfunctions and improves the intestinal epithelial barrier integrity. Therefore, PEA represents a viable approach for the management of the enteric inflammation and motor contractile abnormalities associated with AD.

Parturition involves the transformation of the quiescent myometrium into a highly excitable and contractile state, a process that is driven by changes in myometrial gene expression. This study aimed to identify myometrial transcriptomic signatures and potential novel hub genes in parturition, which have great significance for understanding the underlying mechanisms of successful parturition and treating labor-associated pathologies such as preterm birth. In our study, comparative transcriptome analysis was carried out on human myometrial tissues collected from women undergoing caesarean section at term in the presence (TL = 8) and absence of labor (TNL = 8). A total of 582 differentially expressed genes (DEGs) between TL and TNL tissues were identified. Gene ontology (GO), Kyoto encyclopedia of genes and genomes (KEGG) and gene set enrichment analysis (GSEA) revealed that the DEGs were enriched in signal transduction, regulation of signaling receptor activity, inflammatory response, cytokine-cytokine receptor interaction, IL-17 signaling pathway, TNF signaling pathway, among others. Thus, transcriptome analysis of the myometrium during term labor revealed that labor onset was associated with an inflammatory response. Moreover, protein-protein interactions network analysis identified FPR1, CXCL8, CXCL1, BDKRB2, BDKRB1, and CXCL2 as the hub genes associated with onset of labor. Formyl peptide receptor 1 (FPR1) was highly expressed in laboring myometrial tissues, with the activation of FPR1 in vitro experiments resulting in increased myometrial contraction. Our findings demonstrate the novel role of FPR1 as a modulator of myometrial contraction.

Increasing neuropeptide Y (NPY) has been shown to be a risk factor for cardiovascular diseases. However, its role and mechanism in myocardial infarction (MI) have not yet been fully understood. H9c2 cells and neonatal rat ventricular myocytes with loss of function of NPY and rats with global knockout were used in this study. MI model of rats was induced by the ligation of left coronary artery, and the extent of MI was analyzed through echocardiographic, pathological, and molecular analyses. Our data demonstrated that NPY expression was significantly increased in MI rats and hypoxia/hydrogen peroxide (H2O2)-treated cardiomyocytes. At the same time, NPY-knockout rats exhibited a remarkable decrease in infarct size, serum lactate dehydrogenase activity, cardiomyocyte apoptosis, and caspase-3 expression and activity and a strong improvement in heart contractile function compared with MI rats. Meanwhile, NPY small interfering RNA (siRNA) inhibited the cell apoptosis in H2O2-treated H9c2 cells and hypoxia-treated neonatal rat ventricular myocytes. NPY deletion increased miR-499 expression and decreased FoxO4 expression in MI in vivo and in vitro. Moreover, NPY type 1 receptor antagonist BIBO3304 can reverse miR-499 decrease and FoxO4 increase in H2O2-induced cardiomyocytes. NPY siRNA inhibited cell apoptosis in H2O2-treated H9c2 cells that were reversed by miR-499 inhibitor. Additionally, FoxO4 was validated as the direct target of miR-499. Moreover, BIBO3304 and FoxO4 siRNA significantly increased the cell activity, inhibited the cell apoptosis, and decreased caspase-3 expression and activity in H2O2-treated cardiomyocytes that NPY presented the opposite effect. Collectively, deletion of NPY reduced myocardial ischemia, improved cardiac function, and inhibited cardiomyocyte apoptosis by NPY type 1 receptor-miR-499-FoxO4 axis, which provides a new treatment for MI.

Norbormide is a toxicant selective for rats to which it induces a widespread vasoconstriction. In a recent paper, we hypothesized a role of ATP-sensitive potassium (KATP) channels in norbormide-induced vasoconstriction. The current study was undertaken to verify this hypothesis by comparing the effects of norbormide with those of glibenclamide, a known KATP channel blocker. The whole-cell patch-clamp method was used to record KATP currents in myocytes freshly isolated from the rat and mouse caudal artery and from the rat gastric fundus, as well as in insulin-secreting pancreatic beta cells (INS-1 cells). Smooth muscle contractile function was assessed on either rat caudal artery rings or gastric fundus strips. Molecular modeling and docking simulation to KATP channel proteins were investigated in silico. Both norbormide (a racemic mixture of endo and exo isomers) and glibenclamide inhibited KATP currents in rat and mouse caudal artery myocytes, as well as in gastric fundus smooth muscle cells. In rat INS-1 cells, only glibenclamide blocked KATP channels, whereas norbormide was ineffective. The inhibitory effect of norbormide in rat caudal artery myocytes was not stereo-specific as both the endo isomers (active as vasoconstrictor) and the exo isomers (inactive as vasoconstrictor) had similar inhibitory activity. In rat caudal artery rings, norbormide-induced contraction was partially reverted by the KATP channel opener pinacidil. Computational approaches indicated the SUR subunit of KATP channels as the binding site for norbormide. KATP channel inhibition may play a role in norbormide-induced vasoconstriction, but does not explain the species selectivity, tissue selectivity, and stereoselectivity of its constricting activity. The lack of effect in INS-1 cells suggests a potential selectivity of norbormide for smooth muscle KATP channels.

Myocardial ischemia (MI) is an escalating public health care burden worldwide. Baoyuan decoction (BYD) is a traditional Chinese medicine formula with cardioprotective activity; however, its pharmacological characteristics and mechanisms are obscured. Herein, a multi-omics strategy via incorporating the metabonomics, transcriptomics, and pharmacodynamics was adopted to investigate the effects and molecular mechanisms of BYD for treating MI in a rat model of left anterior descending coronary artery (LADCA) ligation. The results indicated that BYD has a significantly cardioprotective role against MI by decreasing the infarct size, converting the echocardiographic abnormalities and myocardial enzyme markers, and reversing the serum metabolic disorders and myocardial transcriptional perturbations resulting from MI. Integrated bioinformatics analysis and literature reports constructed the interaction network based on the changes of the key MI targeted-metabolites and transcripts after BYD treatment and disclosed that the cardioprotection of BYD is mainly involved in the regulation of energy homeostasis, oxidative stress, apoptosis, inflammation, cardiac contractile dysfunction, and extracellular matrix remodeling. The results of histopathological examination, quantitative RT-PCR assay, cardiac energy synthesis, and serum antioxidant assessment complemented the multi-omics findings, and indicated the multi-pathway modulation mechanisms of BYD. Our investigation demonstrated that the multi-omics approach could achieve a complementary and verified view for the comprehensive evaluation of therapeutic effects and complex mechanisms of TCMF like BYD.

Vincristine is an effective anticancer agent for treating leukemias, lymphomas, and other solid tumors. Vincristine's better-known severe side effects include bone marrow depression, hyponatremia, peripheral neuropathy, and gastrointestinal distress. In recent years, cardiovascular damage also has been described during vincristine treatments. However, the vascular toxicity induced by vincristine is little studied. The aim of the present is to evaluate whether these alterations remain after the suspension of chemotherapy treatment (sequelae) and the possible mechanisms involved in this vascular damage. Adult male Wistar rats were used. The animals were divided into four treatment groups: two groups of saline (0.9% NaCl; saline, sequelae saline) and two groups of vincristine (100 μg/kg; vincristine, sequelae vincristine). Saline or vincristine was administered intraperitoneally in two cycles of 5 days each, leaving a rest period between cycles of 2 days. The final cumulative vincristine dose administered was 1 mg/kg. Sequelae groups correspond to 2 weeks after stopping treatment with the antitumor agent. At the end of the different experimental protocols, cardiac and vascular functions were analyzed. Alterations in the expression of different proteins in the cardiovascular tissues were also investigated. Chronic treatment with vincristine did not produce significant changes in basal cardiac function but provoked significant endothelial dysfunction in the aorta and a significant decrease in the mesenteric contractile function. These cardiovascular functional alterations disappeared 2 weeks after the suspension of chemotherapy treatment. Vincristine treatment caused a significant increase in the expression of tumor necrosis factor-alpha (TNFα), endothelial and inducible nitric oxide synthases (eNOS and iNOS), and connexin 43 in cardiac tissue. In the aorta, the chronic treatment with vincristine caused a slight non-significant increase in TNFα expression, a significant increase in eNOS and iNOS, and a significant decrease in connexin 43. After 2 weeks of vincristine treatment (sequelae group), the expression of TNFα increased and eNOS and iNOS expressions disappeared, but a significant decrease in the expression of connexin 43 was still observed in the aorta. In mesenteric arteries, similar data to those found in the aorta were observed. In conclusion, chronic treatment with vincristine causes functional alterations in the vascular function of both conductance and resistance vessels and changes in the expressions of TNFα, eNOS, iNOS, and connexin 43 in cardiovascular tissues, implicating direct toxicity during its treatment. These functional alterations are transitory and disappear after the suspension of its treatment.