Contractile Behavior Research Articles

Entropy production rate is maximized in non-contractile actomyosin

The actin cytoskeleton is an active semi-flexible polymer network whose non-equilibrium properties coordinate both stable and contractile behaviors to maintain or change cell shape. While myosin motors drive the actin cytoskeleton out-of-equilibrium, the role of myosin-driven active stresses in the accumulation and dissipation of mechanical energy is unclear. To investigate this, we synthesize an actomyosin material in vitro whose active stress content can tune the network from stable to contractile. Each increment in activity determines a characteristic spectrum of actin filament fluctuations which is used to calculate the total mechanical work and the production of entropy in the material. We find that the balance of work and entropy does not increase monotonically and the entropy production rate is maximized in the non-contractile, stable state of actomyosin. Our study provides evidence that the origins of entropy production and activity-dependent dissipation relate to disorder in the molecular interactions between actin and myosin.

Contraction Dynamics and Respiration of Small Single-Osculum Explants of the Demosponge Halichondria panicea

: Sponges pump large amounts of seawater through their water canal system, providing both food and oxygen to the sponge body. Sponge pumping activity may show considerable variation as a consequence of contractile behavior, which includes contraction and expansion of the exhalant opening (osculum) in regular or irregular time intervals. The present study unravels short- and long-term effects of contraction-expansion events on the respiration rate of small single-osculum explants of the demosponge Halichondria panicea. Based on simultaneous video-microscopic time-lapse recordings of osculum cross-sectional area (OSA) and projected area (A), combined with respiration rate measurements, we further evaluate the role of pumping activity for oxygen uptake in the explants. Pumping dynamics were expressed by cyclic contraction-expansion events of the OSA and A, including osculum closure with a mean duration of 37.5 ± CI95% 13.7 min. The respiration rate of sponge explants remained relatively constant at 0.046 ± CI95% 0.014 µmol O2 h-1 (i.e., 7.41 µmol O2 h-1 g-1 DWsponge) during contraction-expansion cycles, but with a marginal decrease of 9.6 % during osculum closure. Periods of pumping cessation during osculum closure likely caused reduced oxygen levels in the sponge body, increasing the oxygen gradient between the environment and the sponge interior, allowing enhanced diffusion of oxygen across the explant surface. This is a key mechanism for balancing respiratory demands during sponge contractions. While contractile behavior is only marginally associated with decreased respiratory demands in small single-osculum H. panicea explants, it may control the degree of internal oxygen depletion and thereby ensure maintenance of sponge-associated microorganisms during non-pumping periods.

Contractile behaviors of cardiac muscle cells on mushroom-shaped micropillar arrays.

In this work we propose mushroom-shaped PDMS (Polydimethylsiloxane) μpillar arrays for enhancing the contractile force of cardiomyocytes during cell culturing. Conventional micropillar (μpillar) arrays with flat surfaces were employed as a standard sample to quantitatively recognize experimental data and to conclusively demonstrate the improved performance of mushroom-shaped PDMS μpillar arrays. Cardiomyocytes isolated from experimental animals were cultured on both of the fabricated μpillar arrays and then monitored over a growing period. Deflections of μpillars were precisely measured through a home-built analyzing system quantitatively representing the contractile force of cardiomyocytes. Mushroom-shaped PDMS μpillar arrays exhibited a 20% improved contractile force compared to conventional PDMS μpillar arrays due to their topographical dependency. Preliminary results also show that the proposed mushroom-shaped PDMS μpillar surface positively affects the Z-band width, actin filament polymerization and focal adhesion (FA) of cardiomyocytes. Further, the enhanced performance of mushroom-shaped PDMS μpillar arrays was confirmed by measuring the cardiac sarcomere α-actin length and myofilament width via ICC (immunocytochemistry) staining and western blot experiments.

Sarcolemmal Alterations in Unloaded Rat Heart after Heterotopic Transplantation.

Following heterotopic transplantation, the rat heart undergoes atrophy and exhibits delayed cardiac relaxation without any changes in contraction and systolic Ca 2+ transients. Furthermore, the sarcoplasmic reticular Ca 2+ uptake and release activities were reduced and Ca 2+ influx through L-type Ca 2+ channels was increased in the atrophied heart. Since Ca 2+ movements at sarcolemma are intimately involved in the regulation of intracellular Ca 2+ concentration, the present study was undertaken to test if sarcolemma plays any role to maintain cardiac function in the atrophied heart.The characteristics of sarcolemmal Ca 2+ pump and Na + -Ca 2+ exchange activities were examined in 8 weeks heterotopically isotransplanted rat hearts which did not support hemodynamic load and underwent atrophy. Sarcolemmal ATP (adenosine triphosphate)-dependent Ca 2+ uptake and Ca 2+ -stimulated ATPase (adenosine triphosphatase) activities were increased without any changes in Na + -K + ATPase activities in the transplanted hearts. Although no alterations in the Na + -dependent Ca 2+ uptake were evident, Na + -induced Ca 2+ release was increased in the transplanted heart sarcolemmal vesicles. The increase in Na + -induced Ca 2+ release was observed at different times of incubation as well as at 5, 20, and 40 mM Na + . The sarcolemma from transplanted hearts also showed higher contents of phosphatidic acid, sphingomyelin, and cholesterol.These results indicate that increases in the sarcolemmal, Ca 2+ transport activities in unloaded heart may provide an insight into adaptive mechanism to maintain normal contractile behavior of the atrophic heart.

Balance between Force Generation and Relaxation Leads to Pulsed Contraction of Actomyosin Networks

Actomyosin contractility regulates various biological processes, including cell migration and cytokinesis. The cell cortex underlying the membrane of eukaryote cells exhibits dynamic contractile behaviors facilitated by actomyosin contractility. Interestingly, the cell cortex shows reversible aggregation of actin and myosin called “pulsed contraction” in diverse cellular phenomena, such as embryogenesis and tissue morphogenesis. Although contractile behaviors of actomyosin machinery have been studied extensively in several in vitro experiments and computational studies, none of them successfully reproduced the pulsed contraction observed in vivo. Recent experiments have suggested the pulsed contraction is dependent upon the spatiotemporal expression of a small GTPase protein called RhoA. This only indicates the significance of biochemical signaling pathways during the pulsed contraction. In this study, we reproduced the pulsed contraction with only the mechanical and dynamic behaviors of cytoskeletal elements. First, we observed that small pulsed clusters or clusters with fluctuating sizes may appear when there is subtle balance between force generation from motors and force relaxation induced by actin turnover. However, the size and duration of these clusters differ from those of clusters observed during the cellular phenomena. We found that clusters with physiologically relevant size and duration can appear only with both actin turnover and angle-dependent F-actin severing resulting from buckling induced by motor activities. We showed how parameters governing F-actin severing events regulate the size and duration of pulsed clusters. Our study sheds light on the underestimated significance of F-actin severing for the pulsed contraction observed in physiological processes.

Dual excitatory and smooth muscle-relaxant effect of β-phenylethylamine on gastric fundus strips in rats.

β-Phenylethylamine (β-PEA) is a trace amine with chemical proximity to biogenic amines and amphetamines. It is an endogenous agonist of trace amine-associated receptors (TAARs) that acts as a neuromodulator of classic neurotransmitters in the central nervous system. At high concentrations, β-PEA contracts smooth muscle, and a role for TAARs in these responses has been postulated. The high dietary intake of trace amines has been associated with such symptoms as hypertension and migraine, especially after the intake of foods containing such compounds. In gastrointestinal tissues, TAAR expression was reported, although the effect of β-PEA on gastric contractile behaviour is unknown. Here, isolated strips that were obtained from the rat gastric fundus were stimulated with high micromolar concentrations of β-PEA. Under resting tonus, β-PEA induced contractions. In contrast, when the strips were previously contracted with KCl, a relaxant response to β-PEA was observed. The contractile effect of β-PEA was inhibited by 5-hydroxytryptamine (5-HT) receptor antagonists (i.e., cyproheptadine and ketanserin) but not by the TAAR1 antagonist EPPTB. In gastric fundus strips that were previously contracted with 80mmol/L KCl, the relaxant effect of β-PEA intensified in the presence of 5-HT receptor antagonists, which was inhibited by EPPTB and the adenylyl cyclase inhibitor MDL-12,330A. The guanylyl cyclase inhibitor ODQ did not alter the relaxant effects of β-PEA. In conclusion, β-PEA exerted dual contractile and relaxant effects on rat gastric fundus. The contractile effect appeared to involve the recruitment of 5-HT receptors, and the relaxant effect of β-PEA on KCl-elicited contractions likely involved TAAR1 .

Electroactive graphene composite scaffolds for cardiac tissue engineering.

Contractile behavior of cardiomyocytes relies on directed signal propagation through the electroconductive networks that exist within the native myocardium. Due to their unique electrical properties, electroactive materials, such as graphene, have recently emerged as promising candidate materials for cardiac tissue engineering applications. In this work, the potential of three-dimensional (3D) nanofibrous graphene and poly(caprolactone) (PCL + G) composite scaffold for cardiac tissue engineering has been explored for the first time. The addition of graphene into PCL led to an increased volume conductivity of scaffolds with an even distribution of graphene particles throughout the matrix. Graphene particles provided local conductive sites within the PCL matrix, which enabled application of external electrical stimulation throughout the scaffold using a custom point stimulation device. When mouse embryonic stem cell derived cardiomyocytes (mES-CM) were seeded on PCL + G scaffolds, cells adhered well, contracted spontaneously, and exhibited classical cardiomyocyte phenotype confirming the biocompatibility of electroactive PCL + G scaffolds. Further functional characterization demonstrated that graphene especially affected Ca2+ handling properties of mES-CM compared to that of cardiomyocytes cultured in 2D culture, highlighting the potential of PCL + G for in vitro cardiac tissue engineering. © 2018 Wiley Periodicals, Inc. J. Biomed. Mater. Res. Part A: 106A: 2923-2933, 2018.

Specialized Living Wound Dressing Based on the Self-Assembly Approach of Tissue Engineering

There is a high incidence of failure and recurrence for chronic skin wounds following conventional therapies. To promote healing, the use of skin substitutes containing living cells as wound dressings has been proposed. The aim of this study was to produce a scaffold-free cell-based bilayered tissue-engineered skin substitute (TES) containing living fibroblasts and keratinocytes suitable for use as wound dressing, while considering production time, handling effort during the manufacturing process, and stability of the final product. The self-assembly method, which relies on the ability of mesenchymal cells to secrete and organize connective tissue sheet sustaining keratinocyte growth, was used to produce TESs. Three fibroblast-seeding densities were tested to produce tissue sheets. At day 17, keratinocytes were added onto 1 or 3 (reference method) stacked tissue sheets. Four days later, TESs were subjected either to 4, 10, or 17 days of culture at the air–liquid interface (A/L). All resulting TESs were comparable in terms of their histological aspect, protein expression profile and contractile behavior in vitro. However, signs of extracellular matrix (ECM) digestion that progressed over culture time were noted in TESs produced with only one fibroblast-derived tissue sheet. With lower fibroblast density, the ECM of TESs was almost completely digested after 10 days A/L and lost histological integrity after grafting in athymic mice. Increasing the fibroblast seeding density 5 to 10 times solved this problem. We conclude that the proposed method allows for a 25-day production of a living TES, which retains its histological characteristics in vitro for at least two weeks.

Spatial and temporal analysis of PCP protein dynamics during neural tube closure.

Planar cell polarity (PCP) controls convergent extension and axis elongation in all vertebrates. Although asymmetric localization of PCP proteins is central to their function, we understand little about PCP protein localization during convergent extension. Here, we use quantitative live imaging to simultaneously monitor cell intercalation behaviors and PCP protein dynamics in the Xenopus laevis neural plate epithelium. We observed asymmetric enrichment of PCP proteins, but more interestingly, we observed tight correlation of PCP protein enrichment with actomyosin-driven contractile behavior of cell-cell junctions. Moreover, we found that the turnover rates of junctional PCP proteins also correlated with the contractile behavior of individual junctions. All these dynamic relationships were disrupted when PCP signaling was manipulated. Together, these results provide a dynamic and quantitative view of PCP protein localization during convergent extension and suggest a complex and intimate link between the dynamic localization of core PCP proteins, actomyosin assembly, and polarized junction shrinking during cell intercalation in the closing vertebrate neural tube.

A 3D diffusional-compartmental model of the calcium dynamics in cytosol, sarcoplasmic reticulum and mitochondria of murine skeletal muscle fibers.

Variations of free calcium concentration ([Ca2+]) are powerful intracellular signals, controlling contraction as well as metabolism in muscle cells. To fully understand the role of calcium redistribution upon excitation and contraction in skeletal muscle cells, the local [Ca2+] in different compartments needs to be taken into consideration. Fluorescent probes allow the determination of [Ca2+] in the cytosol where myofibrils are embedded, the lumen of the sarcoplasmic reticulum (SR) and the mitochondrial matrix. Previously, models have been developed describing intracellular calcium handling in skeletal and cardiac muscle cells. However, a comprehensive model describing the kinetics of the changes in free calcium concentration in these three compartments is lacking. We designed a new 3D compartmental model of the half sarcomere with radial symmetry, which accounts for diffusion of Ca2+ into the three compartments and simulates its dynamics at rest and at various rates of stimulation in mice skeletal muscle fibers. This model satisfactorily reproduces both the amplitude and time course of the variations of [Ca2+] in the three compartments in mouse fast fibers. As an illustration of the applicability of the model, we investigated the effects of Calsequestrin (CSQ) ablation. CSQ is the main Ca2+ buffer in the SR, localized in close proximity of its calcium release sites and near to the mitochondria. CSQ knock-out mice muscles still preserve a near-normal contractile behavior, but it is unclear whether this is caused by additional SR calcium buffering or a significant contribution of calcium entry from extracellular space, via stored-operated calcium entry (SOCE). The model enabled quantitative assessment of these two scenarios by comparison to measurements of local calcium in the cytosol, the SR and the mitochondria. In conclusion, the model represents a useful tool to investigate the impact of protein ablation and of pharmacological interventions on intracellular calcium dynamics in mice skeletal muscle.

ECIS based wounding and reorganization of cardiomyocytes and fibroblasts in co-cultures

The crosstalk of two major heart cell groups, cardiomyocytes and fibroblasts, relies on direct electromechanical cellular coupling as well as indirect mechanical signal transmission through the surrounding viscoelastic extracellular matrix. Upon injury of cardiac tissue, this communication becomes unbalanced: fibrosis is initiated leading to increased collagen deposition, accompanied by an activation of fibroblasts – the key players of fibrosis. They undergo a reorganization or partial transformation to myofibroblasts during this process, which precedes scar formation within the infarcted heart in vivo. Here, we induce wound formation in an in vitro system as a model for these fibrotic conditions: we assessed the dynamics of wound healing in co-cultures of fibroblasts and myocytes upon targeted wound initiation using Electric Cell Substrate Impedance Sensing (ECIS) under optical control. We discovered distinct wound closure dynamics for mono- and co-cultures of myocytes and fibroblasts and observed a cessation of the contractile behavior for recovering cardiomyocyte cultures. We furthermore identified a change of cellular impedance for recovering fibroblasts and the presence of α-SMA, suggesting a partial transformation into myofibroblasts. This was concomitant with a modulation of connectivity, cell-substrate dynamics and membrane capacitance of all wounded cell cultures. Qualitatively, connexin 43 observation confirmed the ECIS trend found for cell-cell connectivity. Finally, we were able to validate the ECIS based wounding approach against an ECIS based barrier assay – the so-called electric fence. In particular the cell-cell connectivity and thus cell layer integrity dominates the healing dynamics within the two intrinsically different assays.

Generation and primary characterization of iAM-1, a versatile new line of conditionally immortalized atrial myocytes with preserved cardiomyogenic differentiation capacity.

AimsThe generation of homogeneous cardiomyocyte populations from fresh tissue or stem cells is laborious and costly. A potential solution to this problem would be to establish lines of immortalized cardiomyocytes. However, as proliferation and (terminal) differentiation of cardiomyocytes are mutually exclusive processes, their permanent immortalization causes loss of electrical and mechanical functions. We therefore aimed at developing conditionally immortalized atrial myocyte (iAM) lines allowing toggling between proliferative and contractile phenotypes by a single-component change in culture medium composition.Methods and resultsFreshly isolated neonatal rat atrial cardiomyocytes (AMs) were transduced with a lentiviral vector conferring doxycycline (dox)-controlled expression of simian virus 40 large T antigen. Under proliferative conditions (i.e. in the presence of dox), the resulting cells lost most cardiomyocyte traits and doubled every 38 h. Under differentiation conditions (i.e. in the absence of dox), the cells stopped dividing and spontaneously reacquired a phenotype very similar to that of primary AMs (pAMs) in gene expression profile, sarcomeric organization, contractile behaviour, electrical properties, and response to ion channel-modulating compounds (as assessed by patch-clamp and optical voltage mapping). Moreover, differentiated iAMs had much narrower action potentials and propagated them at >10-fold higher speeds than the widely used murine atrial HL-1 cells. High-frequency electrical stimulation of confluent monolayers of differentiated iAMs resulted in re-entrant conduction resembling atrial fibrillation, which could be prevented by tertiapin treatment, just like in monolayers of pAMs.ConclusionThrough controlled expansion and differentiation of AMs, large numbers of functional cardiomyocytes were generated with properties superior to the differentiated progeny of existing cardiomyocyte lines. iAMs provide an attractive new model system for studying cardiomyocyte proliferation, differentiation, metabolism, and (electro)physiology as well as to investigate cardiac diseases and drug responses, without using animals.

Spontaneous and α-adrenoceptor-induced contractility in human collecting lymphatic vessels require chloride.

Human lymphatic vessels are myogenically active and respond to sympathetic stimulation. The role of various cations in this behavior has recently been investigated, but whether the anion Cl- is essential is unclear. With ethical approval and informed consent, human thoracic duct and mesenteric lymphatic vessels were obtained from surgical patients. Spontaneous or norepinephrine-induced isometric force production from isolated vessels was measured by wire myography; the transmembrane Cl- gradient and Cl- channels were investigated by substitution of extracellular Cl- with the impermeant anion aspartate and inhibition of Cl- transport and channels with the clinical diuretics furosemide and bendroflumethiazide as well as DIDS and 5-nitro-2-(3-phenylpropylamino)benzoic acid. The molecular expression of Ca2+-activated Cl- channels was investigated by RT-PCR, and proteins were localized using immunoreactivity. Spontaneous and norepinephrine-induced contractility in human lymphatic vessels was highly abrogated after Cl- substitution with aspartate. About 100-300 µM DIDS or 5-nitro-2-(3-phenylpropylamino)benzoic acid inhibited spontaneous contractile behavior. Norepinephrine-stimulated tone was furthermore markedly abrogated by 200 µM DIDS. Furosemide lowered only spontaneous constrictions, whereas bendroflumethiazide had nonspecific inhibitory effects. Consistent expression of transmembrane member 16A [TMEM16A (anoctamin-1)] was found in both the thoracic duct and mesenteric lymphatic vessels, and immunoreactivity with different antibodies localized TMEM16A to lymphatic smooth muscle cells and interstitial cells. The significant change in contractile function observed with inhibitors and anion substitution suggests that Cl- movement over the plasma membrane of lymphatic myocytes is integral for spontaneous and α-adrenoceptor-evoked contractility in human collecting lymphatic vessels. Consistent detection and localization of TMEM16A to myocytes suggests that this channel could play a major functional role. NEW & NOTEWORTHY In this study, we report the first observations of Cl- being a critical ionic component of spontaneous and agonist-evoked contractility in human lymphatics. The most consistently expressed Ca2+-activated Cl- channel gene in the human thoracic duct and mesenteric lymphatic vessels appears to be transmembrane member 16A, suggesting that this channel plays a major role.

A multiscale active structural model of the arterial wall accounting for smooth muscle dynamics.

Arterial wall dynamics arise from the synergy of passive mechano-elastic properties of the vascular tissue and the active contractile behaviour of smooth muscle cells (SMCs) that form the media layer of vessels. We have developed a computational framework that incorporates both these components to account for vascular responses to mechanical and pharmacological stimuli. To validate the proposed framework and demonstrate its potential for testing hypotheses on the pathogenesis of vascular disease, we have employed a number of pharmacological probes that modulate the arterial wall contractile machinery by selectively inhibiting a range of intracellular signalling pathways. Experimental probes used on ring segments from the rabbit central ear artery are: phenylephrine, a selective α1-adrenergic receptor agonist that induces vasoconstriction; cyclopiazonic acid (CPA), a specific inhibitor of sarcoplasmic/endoplasmic reticulum Ca2+-ATPase; and ryanodine, a diterpenoid that modulates Ca2+ release from the sarcoplasmic reticulum. These interventions were able to delineate the role of membrane versus intracellular signalling, previously identified as main factors in smooth muscle contraction and the generation of vessel tone. Each SMC was modelled by a system of nonlinear differential equations that account for intracellular ionic signalling, and in particular Ca2+ dynamics. Cytosolic Ca2+ concentrations formed the catalytic input to a cross-bridge kinetics model. Contractile output from these cellular components forms the input to the finite-element model of the arterial rings under isometric conditions that reproduces the experimental conditions. The model does not account for the role of the endothelium, as the nitric oxide production was suppressed by the action of L-NAME, and also due to the absence of shear stress on the arterial ring, as the experimental set-up did not involve flow. Simulations generated by the integrated model closely matched experimental observations qualitatively, as well as quantitatively within a range of physiological parametric values. The model also illustrated how increased intercellular coupling led to smooth muscle coordination and the genesis of vascular tone.

Activation of Fibroblast Contractility via Cell-Cell Interactions and Soluble Signals.

The collagen contraction assay is an in vitro, three-dimensional method to determine the factor(s) affecting the contractile behavior of activated cells such as fibroblasts in either physiological or pathological scenarios. The collagen lattices/hydrogels are seeded with fibroblasts to mimic the interactions between these cells and their surrounding extracellular matrix proteins in the connective tissue. This method is an important platform to assess components as potential therapeutic targets to prevent pathologies such as fibrosis, which are manifestations of hyperactivated fibroblasts. We have described a basic version of this collagen contraction assay, which is amenable to customization using different cell types under diverse experimental conditions.

Dynamic Model for Characterizing Contractile Behaviors and Mechanical Properties of a Cardiomyocyte

Studies on the contractile dynamics of heart cells have attracted broad attention for the development of both heart disease therapies and cardiomyocyte-actuated micro-robotics. In this study, a linear dynamic model of a single cardiomyocyte cell was proposed at the subcellular scale to characterize the contractile behaviors of heart cells, with system parameters representing the mechanical properties of the subcellular components of living cardiomyocytes. The system parameters of the dynamic model were identified with the cellular beating pattern measured by a scanning ion conductance microscope. The experiments were implemented with cardiomyocytes in one control group and two experimental groups with the drugs cytochalasin-D or nocodazole, to identify the system parameters of the model based on scanning ion conductance microscope measurements, measurement of the cellular Young’s modulus with atomic force microscopy indentation, measurement of cellular contraction forces using the micro-pillar technique, and immunofluorescence staining and imaging of the cytoskeleton. The proposed mathematical model was both indirectly and qualitatively verified by the variation in cytoskeleton, beating amplitude, and contractility of cardiomyocytes among the control and the experimental groups, as well as directly and quantitatively validated by the simulation and the significant consistency of 90.5% in the comparison between the ratios of the Young’s modulus and the equivalent comprehensive cellular elasticities of cells in the experimental groups to those in the control group. Apart from mechanical properties (mass, elasticity, and viscosity) of subcellular structures, other properties of cardiomyocytes have also been studied, such as the properties of the relative action potential pattern and cellular beating frequency. This work has potential implications for research on cytobiology, drug screening, mechanisms of the heart, and cardiomyocyte-based bio-syncretic robotics.

Controlling human corneal stromal stem cell contraction to mediate rapid cell and matrix organization of real architecture for 3-dimensional tissue equivalents.

For the first time, we have demonstrated that human CSSC exhibit the phenomenon of cellular self-alignment in tethered collagen gels. We found this mediated rapid co-alignment of collagen fibrils and thus subsequently exploited this property in vitro to improve the architecture of engineered RAFT tissue equivalents of the corneal stroma. Existing techniques are extremely lengthy and carry significant risk and cost for GMP manufacture. This rapid and tunable technique takes just 8 h of culture and is therefore ideal for clinical manufacture, creating biomimetic tissue equivalents with both cellular and ECM organization. Thus, cellular self-alignment can be a useful bioengineering tool for the development of organized tissue equivalents in a variety of applications.

Effects of Anticancer Drugs on Contractile Behaviors of Human-Induced Pluripotent Stem Cell-Derived Cardiomyocyte Monolayers Evaluated with an Image-based Analysis Using Motion Field Imaging Technique

Effects of Anticancer Drugs on Contractile Behaviors of Human-Induced Pluripotent Stem Cell-Derived Cardiomyocyte Monolayers Evaluated with an Image-based Analysis Using Motion Field Imaging Technique

A comparison of the effects of lindane and FeCl3/ADP on spontaneous contractions in isolated rat or human term myometrium

Oxidative stress affects the contractile behavior of smooth muscle resulting in complications during labor. Toxicants such as lindane and ferric chloride (FeCl3)/adenosine diphosphate (ADP) cause oxidative stress and have previously been shown to inhibit smooth muscle contraction. In this study we examined the effects of the oxygen species scavengers, ascorbic acid and N-acetylcysteine on lindane and FeCl3/ADP’s inhibition of spontaneous myometrial contractions in rat and human myometrium. Lindane and FeCl3/ADP gave rise to concentration-dependent reductions in rat (EC50 11.8×10−6M and 0.9×10−3M) and human myometrial contractions (EC50 16.3×10−6M and 1.1×10−3M, respectively). Pre-treatment with N-acetylcysteine significantly increased the EC50 for the effects of lindane on motility index of human tissue and reduced the maximum inhibitory effect of FeCl3/ADP on contractions in both rat and human myometrium. Ascorbic acid reduced the effects of FeCl3/ADP in rat tissue only. In conclusion pre-treatment with specific antioxidants may protect both rat and human myometrium from the inhibitory effects of lindane and FeCl3/ADP.

Myofilament Remodeling and Function Is More Impaired in Peripartum Cardiomyopathy Compared with Dilated Cardiomyopathy and Ischemic Heart Disease

Peripartum cardiomyopathy (PPCM) and dilated cardiomyopathy (DCM) show similarities in clinical presentation. However, although DCM patients do not recover and slowly deteriorate further, PPCM patients show either a fast cardiac deterioration or complete recovery. The aim of this study was to assess if underlying cellular changes can explain the clinical similarities and differences in the two diseases. We, therefore, assessed sarcomeric protein expression, modification, titin isoform shift, and contractile behavior of cardiomyocytes in heart tissue of PPCM and DCM patients and compared these with nonfailing controls. Heart samples from ischemic heart disease (ISHD) patients served as heart failure control samples. Passive force was only increased in PPCM samples compared with controls, whereas PPCM, DCM, and ISHD samples all showed increased myofilament Ca2+ sensitivity. Length-dependent activation was significantly impaired in PPCM compared with controls, no impairment was observed in ISHD samples, and DCM samples showed an intermediate response. Contractile impairments were caused by impaired protein kinase A (PKA)-mediated phosphorylation because exogenous PKA restored all parameters to control levels. Although DCM samples showed reexpression of EH-myomesin, an isoform usually only expressed in the heart before birth, PPCM and ISHD did not. The lack of EH-myomesin, combined with low PKA-mediated phosphorylation of myofilament proteins and increased compliant titin isoform, may explain the increase in passive force and blunted length-dependent activation of myofilaments in PPCM samples.