Contractile Effects Research Articles

Engineering Microgel Packing to Tailor the Physical and Biological Properties of Gelatin Methacryloyl Granular Hydrogel Scaffolds.

Granular hydrogel scaffolds (GHS) are fabricated via placing hydrogel microparticles (HMP) in close contact (packing), followed by physical and/or chemical interparticle bond formation. Gelatin methacryloyl (GelMA) GHS have recently emerged as a promising platform for biomedical applications; however, little is known about how the packing of building blocks, physically crosslinked soft GelMA HMP, affects the physical (pore microarchitecture and mechanical/rheological properties) and biological (in vitro and in vivo) attributes of GHS. Here, the GHS pore microarchitecture is engineered via the external (centrifugal) force-induced packing and deformation of GelMA HMP to regulate GHS mechanical and rheological properties, as well as biological responses in vitro and in vivo. Increasing the magnitude and duration of centrifugal force increases the HMP deformation/packing, decreases GHS void fraction and median pore diameter, and increases GHS compressive and storage moduli. MDA-MB-231 human triple negative breast adenocarcinoma cells spread and flatten on the GelMA HMP surface in loosely packed GHS, whereas they adopt an elongated morphology in highly packed GHS as a result of spatial confinement. Via culturing untreated or blebbistatin-treated cells in GHS, the effect of non-muscle myosin II-driven contractility on cell morphology is shown. In vivo subcutaneous implantation in mice confirms a significantly higher endothelial, fibroblast, and macrophage cell infiltration within the GHS with a lower packing density, which is in accordance with the in vitro cell migration outcome. These results indicate that the packing state of GelMA GHS may enable the engineering of cell response in vitro and tissue response in vivo. This research is a fundamental step forward in standardizing and engineering GelMA GHS microarchitecture for tissue engineering and regeneration.

Neurogenic-derived 6-nitrodopamine is the most potent endogenous modulator of the mouse urinary bladder relaxation

6-Nitrodopamine (6-ND) modulates vas deferens, seminal vesicles, and corpus cavernosum contractility; however, its role on the lower urinary tract organs has not been evaluated. Investigations of isolated urinary bladders from wild-type (WT) mice revealed 6-ND release was comparable to that of dopamine and adrenaline, whereas noradrenaline was hardly detected, as assessed by liquid chromatography coupled to tandem mass spectrometry. In vitro, 6-ND induced concentration-dependent relaxations in carbachol pre-contracted bladders with high potency (pEC50: 8.04 ± 0.86), independently of eNOS/sGC activity. Co-incubation of 6-ND (1–10 μM) antagonizes the contractile effects of acetylcholine (p < 0.05). Experiments using nitric oxide synthase (NOS) knockout mice demonstrated that 6-ND release from isolated urinary bladder was significantly reduced by neuronal NOS (nNOS−/−) deletion and abolished by triple NOSs deletion (n/i/eNOS−/−), while no significant changes were observed in endothelial (eNOS−/−) or inducible (iNOS−/−) knockout mice. Incubation with tetrodotoxin resulted in a significant decrease in 6-ND release in bladders obtained from WT, but not in nNOS−/− mice. The bladders from nNOS−/− and n/i/eNOS−/− mice exhibited significantly higher contractile responses to electric field stimulation (EFS), compared to eNOS−/−, iNOS−/−, or WT bladders. The hyperreactivity observed in triple NOS knockouts was reversed by the incubation with bladder mucosal layer obtained from a donor WT mice, but not with the muscular layer. These findings clearly demonstrate 6-ND is the most potent endogenous relaxing agent of urinary bladder, and inhibition of its release is associated with bladder hyperreactivity.

Effects of Cold on the Adrenoreactivity of the Rat Superior Mesenteric Artery

In experiments on segments of the rat superior mesenteric artery (SMA) under isometric conditions, the effect of cold on the reactivity of this artery, estimated by the magnitude of the contractile reaction of its segments to noradrenaline (NA) at a concentration of 0.01–10.0 µM, as well as to electrical field stimulation (EFS) of perivascular nerves with a frequency of 3, 10 and 40 Hz in the absence and presence of NA in the medium and at a temperature of 37 oC or 25 oC was studied. Cooling was found not to change the tone of the SMA caused by NA at all the concentrations used. In the absence and presence of NA at a low concentration (0.01–0.10 µM), cooling leads to a significant decrease in the neurogenic response of the SMA at all frequencies of EFS, while in the presence of NA at a high concentration (1–10 µM), cooling has no statistically significant effect on this response. In the presence of NA at a low concentration, cooling at low EFS frequencies insignificantly, and at high frequency significantly reduces the potentiating effect of NA on neurogenic contraction of SMA. NA at a high concentration under normal temperature conditions, reduces the neurogenic contraction at all EFS frequencies, while under cooling conditions at low frequencies it has a potentiating effect, and at high EFS frequency it has a depressing effect. The results obtained show that the depressing effect of cold on the neurogenic constriction of the rat SMA, observed at low and disappearing at high concentrations of NA, as well as on the NA-evoked potentiation of the neurogenic vasoconstriction, is not associated with a decrease in the contractile effect of NA, which is the main neurotransmitter in this artery. These effects can contribute to the redistribution of blood from the surface deep into the body, thus reducing heat loss and improve thermoregulation.

Vascular smooth muscle BK channels limit ouabain-induced vasocontraction: Dual role of the Na/K-ATPase as a hub for Src-kinase and the Na/Ca-exchanger.

Large-conductance, calcium-activated potassium channels (BK channels) and the Na/K-ATPase are expressed universally in vascular smooth muscle. The Na/K-ATPase may act via changes in the intracellular Ca2+ concentration mediated by the Na/Ca exchanger (NCX) and via Src kinase. Both pathways are known to regulate BK channels. Whether BK channels functionally interact in vascular smooth muscle cells with the Na/K-ATPase remains to be elucidated. Thus, this study addressed the hypothesis that BK channels limit ouabain-induced vasocontraction. Rat mesenteric arteries were studied using isometric myography, FURA-2 fluorimetry and proximity ligation assay. The BK channel blocker iberiotoxin potentiated methoxamine-induced contractions. The cardiotonic steroid, ouabain (10-5 M), induced a contractile effect of IBTX at basal tension prior to methoxamine administration and enhanced the pro-contractile effect of IBTX on methoxamine-induced contractions. These facilitating effects of ouabain were prevented by the inhibition of either NCX or Src kinase. Furthermore, inhibition of NCX or Src kinase reduced the BK channel-mediated negative feedback regulation of arterial contraction. The effects of NCX and Src kinase inhibition were independent of each other. Co-localization of the Na/K-ATPase and the BK channel was evident. Our data suggest that BK channels limit ouabain-induced vasocontraction by a dual mechanism involving the NCX and Src kinase signaling. The data propose that the NCX and the Src kinase pathways, mediating the ouabain-induced activation of the BK channel, act in an independent manner.

Contractile Effects of Semaglutide in the Human Atrium.

Semaglutide is a glucagon-like peptide 1 receptor (GLP-1R) agonist. GLP-1R agonists are used to treat type 2 diabetes and obesity. It is currently unknown whether semaglutide can directly increase force of contraction (FOC) in the human heart. We tested the hypothesis that semaglutide might increase the FOC in the isolated human atrium. To this end, we conducted contraction experiments in isolated human right atrial muscle preparations (HAP). HAP were obtained during open-heart surgery. We detected a concentration- and time-dependent positive inotropic effect (PIE) of semaglutide in HAP. These PIEs were accompanied by increases in the rates of tension development and tension relaxation and a reduction in muscle relaxation time. The PIE of semaglutide in HAP was attenuated by H89, an inhibitor of the cyclic AMP-dependent protein kinase and by ryanodine, an inhibitor of sarcoplasmic Ca2+ release. Semaglutide up to 100 nM failed to exert a PIE in isolated electrically paced (1 Hz) wild-type mouse left atrial preparations studied for comparison. Our data suggest that semaglutide can increase the FOC in the atria of patients at therapeutic drug concentrations.

Opposite Contractile Effects of Amphetamine-Related Hallucinogenic Drugs in the Isolated Human Atrium.

The present study examined three hallucinogenic amphetamine derivatives, namely, 2,5-dimethoxy-4-iodoamphetamine (DOI) as well as 2,5-dimethoxy-4-methylamphetamine (DOM) and 4-methylmethcathinone (mephedrone). The objective of this study was to test the hypothesis that DOI, DOM, and mephedrone would increase the contractile force in isolated human atrial preparations in a manner similar to amphetamine. To this end, we measured contractile force under isometric conditions in electrically stimulated (1 Hz) human atrial preparations obtained during open surgery. DOI and DOM alone or in the presence of isoprenaline reduced the contractile force concentration-dependently in human atrial preparations. These negative inotropic effects of DOM and DOI were not attenuated by 10 µM atropine. However, mephedrone increased the contractile force in human atrial preparations in a concentration- and time-dependent manner. Furthermore, these effects were attenuated by the subsequent addition of 10 µM propranolol or pretreatment with 10 µM cocaine in the organ bath. Therefore, it can be concluded that amphetamine derivatives may exert opposing effects on cardiac contractile force. The precise mechanism by which DOI and DOM exert their negative inotropic effects remains unknown at present. The cardiac effects of mephedrone are probably due to the release of cardiac noradrenaline.

Contractile effects of stimulation of D1-dopamine receptors in the isolated human atrium.

Dopamine receptors have been claimed not to directly increase contractility in the human heart. Therefore, we performed contraction experiments in isolated electrically driven human atrial preparations (HAP). For comparison, we performed contraction experiments with left atrial preparations of transgenic mice which harbor a cardiac overexpression of human D1-dopamine receptors (D1-TG). In D1-TG, first we noted that dopamine (10nM-10µM cumulatively applied) in the presence of propranolol exerted a concentration- and time-dependent positive inotropic effect in D1-TG. In a similar fashion, dopamine increased force of contraction in the presence of 0.4µM propranolol in HAP and these effects were amplified by pre-treatment with inhibitor of phosphodiesterase III (1µM) cilostamide. Moreover, contractile effects of dopamine in the presence of propranolol 0.4µM in HAP were antagonized by odapipam, haloperidol, or raclopride. Ten micromolars of fenoldopam in the presence of cilostamide increased force of contraction in HAP and this effect was antagonized by SCH23390. We conclude that stimulation of human D1-dopamine receptors can increase force of contraction in the HAP.

Effect of impaired myocardial contractility on coronary flow reserve and inflammatory processes in chronic coronary syndrome

Aim. To study the relationship between myocardial contractility impairment in chronic coronary syndrome and the state of coronary flow reserve, the degree of systemic inflammation and endothelial dysfunction. Materials and methods. We examined 120 patients with stable angina pectoris (SAP) of functional class (FC) II–III, who were assigned into two groups: group 1 comprised 65 patients with left ventricular ejection fraction (LVEF) ≥55 %, and group 2 was composed of 55 patients with LVEF &lt;55 %. Diagnostic methods included clinical and instrumental examination data, analyses of lipid profile, inflammation biomarkers, endothelial functional state, hemogram data, leukocyte inflammation markers, and questionnaires. Results. Compared to group 1, group 2 patients showed an increase in the left atrium (p &lt; 0.001) and right ventricle (p = 0.027) sizes; significantly lower LVEF (50.16 ± 0.42 % vs 58.77 ± 0.41 %; p &lt; 0.001); thickening of the intima-media complex in the right (p = 0.003) and left (p = 0.017) common carotid arteries; significantly lower load threshold values (p = 0.008), exercise tolerance (p = 0.004) and heart rate variability indicators – SDNN (standard deviation of normal-to-normal intervals over 24 hours; p = 0.046). Group 2 patients were characterized by more active low-grade chronic inflammation, as indicated by increased levels of leukocytes (p = 0.024), neutrophils (p &lt; 0.001), and decreased lymphocyte levels (p = 0.021); significantly higher levels of leukocyte inflammation markers, in particular, the neutrophil-to-lymphocyte ratio (p &lt; 0.001), platelet-to-lymphocyte ratio (p = 0.004), systemic immune-inflammation index (p &lt; 0.001), systemic inflammation response index (p &lt; 0.001), and aggregate index of systemic inflammation (p &lt; 0.001) as compared to group 1 individuals. Patients with LVEF &lt;55 % had higher levels of fibrinogen (p &lt; 0.001), uric acid (p = 0.002), high-sensitivity C-reactive protein (p = 0.007), and endothelin-1 (p &lt; 0.001) compared to those with LVEF ≥55 %. Conclusions. Group 2 patients need a more thorough monitoring and a more intensive treatment aimed at reducing the inflammatory process.

Numerical investigation on flow-induced wall shear stress variation of metastatic cancer cells in lymphatics with elastic valves

Hematogenous metastasis occurs when cancer cells detach from the extracellular matrix in the primary tumor into the bloodstream or lymphatic system. Elucidating the response of metastatic tumor cells in suspension to the flow conditions in lymphatics with valves from a mechanical/fluidic perspective is necessary. A physiologically relevant computational model of a lymphatic vessel with valves was constructed using fully coupled fluid–cell–vessel interactions to investigate the effects of lymphatic vessel contractility, valve properties, and cell size and stiffness on the variations in magnitude and gradient of the flow-induced wall shear stress (WSS) experienced by suspended tumor cells. Results indicated that the maximum WSSmax increased with the increments in cell diameter, vessel contraction amplitude, and valve stiffness. The decrease in vessel contraction period and valve aspect ratio also increased the maximum WSSmax. The influence of the properties of the valve on the WSS was more significant among the factors mentioned above. The maximum WSSmax acting on the cancer cell when the cell reversed the direction of its motion in the valve region increased by 0.5–1.4 times that before the cell entered the valve region. The maximum change in WSS was in the range of 0.004–0.028 Pa/µm depending on the factors studied. They slightly exceeded the values associated with breast cancer cell apoptosis. The results of this study provide biofluid mechanics-based support for mechanobiological research on the metastasis of metastatic cancer cells in suspension within the lymphatics.

Toxicology of milrinone by zebrafish embryotoxicity test.

Milrinone, a phosphodiesterase III inhibitor with contractile and vasodilatory effects, is widely used in acute decompensated heart failure and medically refractory end-stage heart failure (HF). The adverse reactions of milrinone have been extensively explored clinically, but its possible toxicities and underlying molecular mechanisms in embryo development need further understanding as its clinical applications increase. Herein, we assessed the milrinone toxicity using the zebrafish embryotoxicity test (ZET), with a view of providing evidence and guidancefor gravidas medicine. We carried out ZET by exposing embryos to a milrinone culture with a series concentration gradients since 1.5 hours post fertilization (hpf) and observed and assessed mortality and hatching rates of drug-treated zebrafishes at 24, 48, 72, and 96 hpf. No significant lethal effect was found in milrinone-treated zebrafish, but hatching rate of eggs at 48 hpf was up-regulated with the increase of milrinone concentration. The impact of milrinone on embryogenesis was assessed through body length, eye area, yolk sac area, swim bladder inflation area, pericardial area and venous congestion area at 96hpf. 150 μg/mL or higher milrinone treatment showed significant effects in the indicators. Organ disorders including enlarged pericardium, liver atrophy and decreased blood vessels were observed in dysplasia individuals versus controls. TUNEL assay suggested the ability of milrinone to induce apoptosis in malformation embryos. Quantitative real-time PCR showed aberrant expressions of transcription factors associated with heartdevelopment and genes related to liver development and apoptosis regulation. Therefore, ZET is feasible for the milrinone toxicity test, and high-dose milrinone causes harm to the embryonic development of zebrafish, especially in embryonic carcinogenesis, vasculogenesis, and hepatogenesis.

Effect of high-fat diet on isometric, concentric and eccentric contractile performance of skeletal muscle isolated from female CD-1 mice.

Despite evidence inferring muscle and contractile mode-specific effects of high-fat diet (HFD), no study has yet considered the impact of HFD directly on eccentric muscle function. The present work uniquely examined the effect of 20-week HFD on the isometric, concentric and eccentric muscle function of isolated mouse soleus (SOL) and extensor digitorum longus (EDL) muscles. CD-1 female mice were randomly split into a control (n=16) or HFD (n=17) group and for 20 weeks consumed standard lab chow or HFD. Following this period, SOL and EDL muscles were isolated and assessments of maximal isometric force and concentric work loop (WL) power were performed. Each muscle was then subjected to either multiple concentric or eccentric WL activations. Post-fatigue recovery, as an indicator of incurred damage, was measured via assessment of concentric WL power. In the EDL, absolute concentric power and concentric power normalised to muscle mass were reduced in the HFD group (P<0.038). HFD resulted in faster concentric fatigue and reduced eccentric activity-induced muscle damage (P<0.05). For the SOL, maximal isometric force was increased, and maximal eccentric power normalised to muscle mass and concentric fatigue were reduced in the HFD group (P<0.05). HFD effects on eccentric muscle function are muscle-specific and have little relationship with changes in isometric or concentric function. HFD has the potential to negatively affect the intrinsic concentric and eccentric power-producing capacity of skeletal muscle, but a lack of a within-muscle uniform response indicates disparate mechanisms of action which require further investigation.

The Enigma of Norbormide, a Rattus-Selective Toxicant.

Norbormide (NRB) is a Rattus-selective toxicant, which was serendipitously discovered in 1964 and formerly marketed as an eco-friendly rodenticide that was deemed harmless to non-Rattus species. However, due to inconsistent efficacy and the emergence of second-generation anticoagulants, its usage declined, with registration lapsing in 2003. NRBs' lethal action in rats entails irreversible vasoconstriction of peripheral arteries, likely inducing cardiac damage: however, the precise chain of events leading to fatality and the target organs involved remain elusive. This unique contractile effect is exclusive to rat arteries and is induced solely by the endo isomers of NRB, hinting at a specific receptor involvement. Understanding NRB's mechanism of action is crucial for developing species-selective toxicants as alternatives to the broad-spectrum ones currently in use. Recent research efforts have focused on elucidating its cellular mechanisms and sites of action using novel NRB derivatives. The key findings are as follows: NRB selectively opens the rat mitochondrial permeability transition pore, which may be a factor that contributes to its lethal effect; it inhibits rat vascular KATP channels, which potentially controls its Rattus-selective vasoconstricting activity; and it possesses intracellular binding sites in both sensitive and insensitive cells, as revealed by fluorescent derivatives. These studies have led to the development of a prodrug with enhanced pharmacokinetic and toxicological profiles, which is currently undergoing registration as a novel efficacious eco-sustainable Rattus-selective toxicant. The NRB-fluorescent derivatives also show promise as non-toxic probes for intracellular organelle labelling. This review documents in more detail these developments and their implications.

Real-time regurgitation estimation in percutaneous left ventricular assist device fully supported condition using an unscented Kalman filter

Objective. Significant aortic regurgitation is a common complication following left ventricular assist device (LVAD) intervention, and existing studies have not attempted to monitor regurgitation signals and undertake preventive measures during full support. Regurgitation is an adverse event that can lead to inadequate left ventricular unloading, insufficient peripheral perfusion, and repeated episodes of heart failure. Moreover, regurgitation occurring during full support due to pump position offset cannot be directly controlled through control algorithms. Therefore, accurate estimation of regurgitation during percutaneous left ventricular assist device (PLVAD) full support is critical for clinical management and patient safety. Approach. An estimation system based on the regurgitation model is built in this paper, and the unscented Kalman filter estimator (UKF) is introduced as an estimation approach. Three offset degrees and three heart failure states are considered in the investigation. Using the mock circulatory loop experimental platform, compare the regurgitation estimated by the UKF algorithm with the actual measured regurgitation; the errors are analyzed using standard confidence intervals of ±2 SDs, and the effectiveness of the mentioned algorithms is thus assessed. The generalization ability of the proposed algorithm is verified by setting different heart failure conditions and different rotational speeds. The root mean square error and correlation coefficient between the estimated and actual values are quantified and the statistical significance of accuracy differences in estimation is illustrated using one-way analysis of variance (One-Way ANOVA), which in turn assessed the accuracy and stability of the UKF algorithm. Main results. The research findings demonstrate that the regurgitation estimation system based on the regurgitation model and UKF can relatively accurately estimate the regurgitation status of patients during PLVAD full support, but the effect of myocardial contractility on the estimation accuracy still needs to be taken into account. Significance. The proposed estimation method in this study provides essential reference information for clinical practitioners, enabling them to promptly manage potential complications arising from regurgitation. By sensitively detecting LVAD adverse events, valuable insights into the performance and reliability of the LVAD device can be obtained, offering crucial feedback and data support for device improvement and optimization.

The oxytocin antagonist cligosiban reduces human prostate contractility: Implications for the treatment of benign prostatic hyperplasia.

With increasing life expectancy, benign prostatic hyperplasia (BPH) consequently affects more ageing men, illustrating the urgent need for advancements in BPH therapy. One emerging possibility may be the use of oxytocin antagonists to relax smooth muscle cells in the prostate, similar to the currently used (although often associated with side effects) α1-adrenoceptor blockers. For the first time we used live-imaging, combined with a novel image analysis method, to investigate the multidirectional contractions of the human prostate and determine their changes in response to oxytocin and the oxytocin antagonists atosiban and cligosiban. Human prostate samples were obtained and compared from patients undergoing prostatectomy due to prostate cancer as well as from patients with transurethral resection of prostate tissue due to severe BPH. The two cohorts of tissue samples showed spontaneous multidirectional contractions, which significantly increased after the addition of oxytocin. Different to atosiban, which showed ambiguous effects of short duration, only long-acting cligosiban reliably prevented, as well as counteracted, any contractile oxytocin effect. Furthermore, cligosiban visibly reduced not only oxytocin-induced contractions, but also showed intrinsic activity to relax prostatic tissue. Thus, the oxytocin antagonist cligosiban could be an interesting candidate in the search for novel BPH treatment options.

Compartment specific responses to contractility in the small intestinal epithelium.

Tissues are subject to multiple mechanical inputs at the cellular level that influence their overall shape and function. In the small intestine, actomyosin contractility can be induced by many physiological and pathological inputs. However, we have little understanding of how contractility impacts the intestinal epithelium on a cellular and tissue level. In this study, we probed the cell and tissue-level effects of contractility by using mouse models to genetically increase the level of myosin activity in the two distinct morphologic compartments of the intestinal epithelium, the crypts and villi. We found that increased contractility in the villar compartment caused shape changes in the cells that expressed the transgene and their immediate neighbors. While there were no discernable effects on villar architecture or cell polarity, even low levels of transgene induction in the villi caused non-cell autonomous hyperproliferation of the transit amplifying cells in the crypt, driving increased cell flux through the crypt-villar axis. In contrast, induction of increased contractility in the proliferating cells of the crypts resulted in nuclear deformations, DNA damage, and apoptosis. This study reveals the complex and diverse responses of different intestinal epithelial cells to contractility and provides important insight into mechanical regulation of intestinal physiology.

Spironolactone Induces Vasodilation by Endothelium-Dependent Mechanisms Involving NO and by Endothelium-Independent Mechanisms Blocking Ca2+ Channels.

Spironolactone (SPI) is a diuretic widely used to treat cardiovascular diseases (CVD) and is non-specific for mineralocorticoid receptors (MR) and with an affinity for progesterone (PR) and androgen (AR) receptors. Since 2009, it has been suggested that pharmaceuticals are emerging contaminants (called EDC), and recently, it was reported that most EDC are AR and MR antagonists and estrogen receptors (ER) agonists. Concerning SPI, endocrine-disrupting effects were observed in female western mosquitofish, but there are still no data regarding the SPI effects as a possible human EDC. In this work, aortic rings were used to analyze the contractility effects of SPI and the mode of action concerning the involvement of Ca2+ channels and endothelial pathways. Moreover, cytotoxic effects were analyzed by MTT assays. SPI induces vasodilation in the rat aorta by endothelium-dependent mechanisms involving NO and by endothelium-independent mechanisms blocking Ca2+ channels. Moreover, a non-monotonic effect characteristic of EDC was observed for SPI-induced decrease in cell viability. Our findings suggest that SPI may act as an EDC at a human level. However, ex vivo studies with human arteries should be carried out to better understand this drug's implications for human health and future generations.

A new oxime synthesized from Senecio nutans SCh. Bip (chachacoma) reduces calcium influx in the vascular contractile response in rat aorta.

Senecio nutans Sch. Bip is an endemic plant commonly employed in the Andes culture to counteract the effects of mountain sickness, and its bioactive molecules could provide new drugs for treating hypertension. The purpose was to determine whether the vascular response of the plant bioactive molecules, such as (5-acetyl-6-hydroxy-2-isopropenyl-2,3-dihydrobenzofurane; Sn-I), could be improved by a simple structural modification to synthesize oximes (Ox-Sn-I). We characterized both compounds using IR and NMR spectroscopy and Heteronuclear Multiple Quantum Coherence (HMQC). We investigated vascular relaxation mechanisms in response to Sn-I and Ox-Sn-I using rat aorta and vascular smooth muscle cells (A7r5) as experimental models. Preincubation of aortic rings with Sn-I (10-5 M) significantly (p < 0.001) decreased the contractile effect in response to phenylephrine (PE) and potassium chloride (KCl). The sensitivity (EC50) to PE significantly (p < 0.01) decreased in the presence of Sn-I (10-5 M), but not with Ox-Sn-I. Sn-I significantly (p < 0.001) reduced the PE-induced contraction under calcium-free conditions. When A7r5 cells were preincubated with Sn-I and Ox-Sn-I (10-5 M), both compounds blunted the increase in intracellular Ca2+ induced by KCl. 2,3-Dihydrobenzofurane derived from S. nutans (Sn-I) reduces the contractile response probably by blocking Ca2+ entry through voltage-gated Ca2+ channels (VGCC) in vascular smooth cells. This effect also causes relaxation in rat aorta mediated by reduction of intracellular Ca2+ concentration, rather than an increase of NO generation in endothelial vascular cells.

Contractile Effects of Amphetamine, Pseudoephedrine, Nor-pseudoephedrine (Cathine), and Cathinone on Atrial Preparations of Mice and Humans.

Amphetamine derivatives are used worldwide legally or illegally and intoxications may be accompanied by cardiac arrhythmias. Here, we tested contractile effects of cumulative applied (±)-amphetamine, pseudoephedrine, nor-pseudoephedrine (cathine), and cathinone in electrically stimulated (1 Hz) human right atrial preparations (HAP) and mouse left atrial preparations and in spontaneously beating mouse right atrial preparations. In mouse atrial preparations, amphetamine increased force of contraction and beating rate in a concentration- and time-dependent manner, starting at 1 µM in left atrial preparations to 157.1% ± 3.0% and right atrial preparations to 146.6% ± 9.8% at 10 µM, respectively [mean ± standard error of the mean (SEM); n = 5; P < 0.05]. Pseudoephedrine, cathine, or cathinone alone were ineffective in mouse atrial preparations but after pre-incubation with the phosphodiesterase IV inhibitor rolipram (0.1 µM), a positive inotropic effect was noted (mean ± SEM: pseudoephedrine: 112.3% ± 9.8%; cathine: 109.0% ± 4.3%; cathinone: 138.3% ± 21.2%). The effects of all drugs were greatly attenuated by 10 µM cocaine or 10 µM propranolol treatments. However, In HAP, not only amphetamine (to a mean ± SEM of 208% ± 32%) but also pseudoephedrine (to a mean ± SEM of 287% ± 60%), cathine (to a mean ± SEM of 234% ± 52%), and cathinone (to a mean ± SEM of 217% ± 65%) increased force of contraction without the need of phosphodiesterase inhibition. The contractile effects in HAP were attenuated by 10 µM cocaine and antagonized by 10 µM propranolol. We conclude that amphetamine, pseudoephedrine, cathine, and cathinone act probably via release of noradrenaline from cardiac stores as indirect sympathomimetic agents in mouse and more pronounced in human atrial preparations.

PGF2α induces a pro-labour phenotypical switch in human myometrial cells that can be inhibited with PGF2α receptor antagonists.

Preterm birth is the leading cause of infant morbidity and mortality. There has been an interest in developing prostaglandin F2α (PGF2α) antagonists as a new treatment for preterm birth, although much of the rationale for their use is based on studies in rodents where PGF2α initiates labour by regressing the corpus luteum and reducing systemic progesterone concentrations. How PGF2α antagonism would act in humans who do not have a fall in systemic progesterone remains unclear. One possibility, in addition to an acute stimulation of contractions, is a direct alteration of the myometrial smooth muscle cell state towards a pro-labour phenotype. In this study, we developed an immortalised myometrial cell line, MYLA, derived from myometrial tissue obtained from a pregnant, non-labouring patient, as well as a novel class of PGF2α receptor (FP) antagonist. We verified the functionality of the cell line by stimulation with PGF2α, resulting in Gαq-specific coupling and Ca2+ release, which were inhibited by FP antagonism. Compared to four published FP receptor antagonists, the novel FP antagonist N582707 was the most potent compound [Fmax 7.67 ± 0.63 (IC50 21.26nM), AUC 7.30 ± 0.32 (IC50 50.43nM), and frequency of Ca2+ oscillations 7.66 ± 0.41 (IC50 22.15nM)]. RNA-sequencing of the MYLA cell line at 1, 3, 6, 12, 24, and 48h post PGF2α treatment revealed a transforming phenotype from a fibroblastic to smooth muscle mRNA profile. PGF2α treatment increased the expression of MYLK, CALD1, and CNN1 as well as the pro-labour genes OXTR, IL6, and IL11, which were inhibited by FP antagonism. Concomitant with the inhibition of a smooth muscle, pro-labour transition, FP antagonism increased the expression of the fibroblast marker genes DCN, FBLN1, and PDGFRA. Our findings suggest that in addition to the well-described acute contractile effect, PGF2α transforms myometrial smooth muscle cells from a myofibroblast to a smooth muscle, pro-labour-like state and that the novel compound N582707 has the potential for prophylactic use in preterm labour management beyond its use as an acute tocolytic drug.

Effects of Cirsium palustre Extracts and Their Main Flavonoids on Colon Motility-An Ex Vivo Study.

For centuries, various species from the genus Cirsium have been utilized in traditional medicine worldwide. A number of ethnopharmacological reports have pointed out that Cirsium plants can be applied to diminish digestive problems. Among them, Cirsium palustre (L.) Scop. (Asteraceae) stands out as a promising herbal drug candidate because its constituents exhibit antimicrobial and antioxidant potential, as evidenced by ethnopharmacological reports. As a result, the species is particularly intriguing as an adjunctive therapy for functional gastrointestinal and motility disorders. Our research goal was to verify how the extracts, fractions, and main flavonoids of C. palustre affect colon contractility under ex vivo conditions. An alternative model with porcine-isolated colon specimens was used to identify the effects of C. palustre preparations and their primary flavonoids. LC-ESI-MS was utilized to evaluate the impacts of methanol (CP1), methanolic 50% (CP2), and aqueous (CP3) extracts as well as diethyl ether (CP4), ethyl acetate (CP5), and n-butanol (CP6) fractions. Additionally, the impacts of four flavonoids, apigenin (API), luteolin (LUT), apigenin 7-O-glucuronide (A7GLC), and chrysoeriol (CHRY), on spontaneous and acetylcholine-induced motility were assessed under isometric conditions. The results showed that C. palustre extracts, fractions, and their flavonoids exhibit potent motility-regulating effects on colonic smooth muscle. The motility-regulating effect was observed on spontaneous and acetylcholine-induced contractility. All extracts and fractions exhibited an enhancement of the spontaneous contractility of colonic smooth muscle. For acetylcholine-induced activity, CP1, CP2, and CP4 caused a spasmolytic effect, and CP5 and CP6 had a spasmodic effect. LUT and CHRY showed a spasmolytic effect in the case of spontaneous and acetylcholine-induced activity. In contrast, API and A7GLC showed a contractile effect in the case of spontaneous and pharmacologically induced activity. Considering the results obtained from the study, C. palustre could potentially provide benefits in the treatment of functional gastrointestinal disorders characterized by hypomotility and hypermotility.