Low KCl Concentrations Research Articles

Highly Elastic Hydrated Cellulosic Materials with Durable Compressibility and Tunable Conductivity.

Anisotropic cellular materials with direction-dependent structure and durable mechanical properties enable various applications (e.g., nanofluidics, biomedical devices, tissue engineering, and water purification), but their widespread use is often hindered by complex and scale-limited fabrication and unsatisfactory mechanical performance. Here, inspired by the anisotropic and hierarchical material structure of tendons, we demonstrate a facile, scalable top-down approach for fabricating a highly elastic, ionically conductive, anisotropic cellulosic material (named elastic wood) directly from natural wood via chemical treatment. The resulting elastic wood demonstrates good elasticity and durable compressibility, showing no sign of fatigue after 10 000 compression cycles. The chemical treatment not only softens the wood cell walls by partially removing lignin and hemicellulose but introduces an interconnected cellulose fibril network into the wood channels. Atomistic and continuum modeling further reveals that the absorbed water can freely and reversibly move inside the elastic wood and therefore helps the elastic wood accommodate large compressive deformation and recover to its original shape upon compression release. In addition, the elastic wood showed a high ionic conductivity of up to 0.5 mS cm-1 at a low KCl concentration of 10-4 M, which can be tuned by changing the compression ratio of the material. The demonstrated elastic, mechanically robust, and ionically conductive cellulosic material combining inherited anisotropic cellular structure from natural wood and a self-formed internal gel may find a variety of potential applications in ionic nanofluidics, sensors, soft robots, artificial muscle, environmental remediation, and energy storage.

Importance of soil interparticle forces and organic matter for aggregate stability in a temperate soil and a subtropical soil

Soil organic matter (SOM) and interparticle forces, including the electrostatic repulsive force (ERF), surface-hydration repulsive force (SHRF) and van der Waals attractive force (vDWAF), play crucial roles in aggregate stability. However, few studies investigated their coupled effects on aggregate stability in variably charged soils, in which soil particle interactions are more complex than in permanently charged soils due to the coexistence of positive and negative charges and the variable surface charge characteristics. Therefore, this study aims to: 1) investigate the combined effects of soil solution pH and the overlapping of electric double layers (EDLs) between positively charged Fe/Al (hydro)oxides and negatively charged SOM on the ERF and aggregate stability before (control) and after SOM removal and after straw incubation for 240 days; and 2) evaluate the importance of soil interparticle forces and SOM for aggregate stability in a permanently charged temperate soil (Vertisol) and a variably charged subtropical soil (Ultisol). Soil aggregate stability was determined by measuring the release of small particles (w(<d)%) after aggregate breakdown at different KCl concentrations (10−1 to 10−5 mol L−1). Soil solution pH was adjusted by varying KCl concentration. In both soils, the w(<d)% increased exponentially with the net force of the soil interparticle forces. However, in contrast to the Vertisol, the w(<d)% of the differently treated Ultisol showed almost no change before the KCl concentration decreased to 10−2 mol L−1 and continuous changes after the KCl concentration decreased to 10−2 mol L−1. In addition, the differences in the w(<d)% between the SOM removal treatment and the control and straw incubation treatments were much larger in the Ultisol than in the Vertisol. In the Ultisol, the ERF did exhibit little and continuous increase over the whole range of tested pH. When considering the impact of the overlapping of oppositely charged EDLs between Fe/Al (hydro)oxides and SOM, a much stronger reduction in ERF was observed in the control and straw incubation treatments and at lower KCl concentrations. Consequently, in both the Vertisol and Ultisol, aggregate stability is essentially controlled by the soil interparticle forces, whereas the effects of soil solution pH and the overlapping of oppositely charged EDLs between SOM and Fe/Al (hydro)oxides on the ERF must be considered in variably charged soils (i.e. Ultisol). Moreover, SOM can play a more important role in aggregate stability in subtropical soil than in temperate soil.

Effect of Potassium Concentration on Triplex Stability under Molecular Crowding Conditions.

The properties of non-canonical DNA structures, like G-quadruplexes and triplexes, change under cell-mimicking molecular crowding conditions relative to dilute aqueous solutions. The analysis of environmental effects on their stability is crucial since they play important roles in gene expression and regulation. In this study, three intramolecular and intermolecular triplex-forming sequences of different C+*G-C triplet content (*: Hoogsteen base pair; - : Watson–Crick base pair) were designed and their stability measured in the absence and presence of a crowding agent with different K+ concentrations. In dilute solution, the stability of the triplexes was reduced by decreasing the concentration of KCl. This reduction became smaller as the number of C+*G-C triplets increased. Under molecular crowding conditions, Watson–Crick base pairs and Hoogsteen base pairs were destabilized and stabilized, respectively. Interestingly, with lower KCl concentrations (≤1 M), the destabilization of the triplexes due to reduction of KCl concentration was significantly smaller than in dilute solutions. In addition, the C+*G-C content had greater influence on triplex stability under molecular crowding conditions. Our work provides quantitative information about the effects of K+ concentration on triplex stability under molecular crowding conditions and should further our understanding of the function and regulation of triplexes in bioprocesses.

A Highly Conductive Cationic Wood Membrane

AbstractHere, a highly conductive cationic membrane is developed directly from natural wood via a two‐step process, involving etherification and densification. Etherification bonds the cationic functional group ((CH3)3N+Cl−) to the cellulose backbone, converting negatively charged (ξ‐potential of −27.9 mV) wood into positively charged wood (+37.7 mV). Densification eliminates the large pores of the natural wood, leading to a highly laminated structure with the oriented cellulose nanofiber and a high mechanical tensile strength of ≈350 MPa under dry conditions (20 times higher than the untreated counterpart) and ≈98 MPa under wet conditions (×5.5 increase compared to the untreated counterpart). The nanoscale gaps between the cellulose nanofibers act as narrow nanochannels with diameters smaller than the Debye length, which facilitates rapid ion transport that is 25 times higher than the ion conductance of the natural wood at a low KCl concentration of 10 × 10−3 m. The demonstrated cationic wood membrane exhibits enhanced mechanical strength and excellent nanofluidic ion‐transport properties, representing a promising direction for developing high‐performance nanofluidic material from renewable, and abundant nature‐based materials.

Exercise training‐induced adaptations of vascular smooth muscle vasodilatory function are absent in muscle arterioles from rats lacking the nuclear factor (erythroid‐derived 2)‐like 2 (Nrf2) gene

Nuclear factor (erythroid‐derived 2)‐like 2 (Nrf2) is a transcription factor that is crucial in maintaining cellular redox homeostasis. Nrf2 protein increases with exercise training in skeletal muscle. Adaptations of the vasculature are often linked to changes in the metabolic profile of skeletal muscle. We hypothesized that deletion of Nrf2 would negate or reduce exercise training‐induced adaptations of arterioles from the soleus muscle. To test this hypothesis, we exercise trained wild type Sprague‐Dawley (WT) rats, and their littermates that were lacking the Nrf2 gene (Nrf2KO). Rats were either exercise trained (EX) on a motor‐driven treadmill 5 days/wk or remained sedentary (SED) in their cages for 10–12 weeks. A mixed moderate aerobic and high‐intensity interval training protocol was utilized. At the end of the training period, soleus muscle arterioles were isolated for assessment of vascular reactivity. Responsiveness to potassium chloride (KCl), phenylephrine, and the NO donor, Diethylamine‐NONO‐ate were evaluated. In resistance soleus muscle arterioles from WT rats, constriction to high concentrations of KCl (80–100 mM) was reduced (WT EX vs WT SED; P&lt;0.05) by exercise training. In contrast, vasodilatory responsiveness to low concentrations of KCl (10–20 mM) was increased (WT EX vs WT SED; P&lt;0.05) by exercise training in resistance arterioles from the soleus muscle of WT rats. Exercise training also increased vasodilatory responsiveness (WT EX vs WT SED; P=0.02) to DEA‐NONO‐ate in soleus muscle arterioles from WT rats. These exercise training‐induced adaptations were absent in soleus muscle arterioles from Nrf2KO rats. Vasoconstrictor responses to phenylephrine were not changed by exercise training in soleus muscle arterioles from WT rats, but tended to increase with exercise training in arterioles from Nrf2KO rats. These data suggest that exercise training enhances vasodilatory responsiveness of the vascular smooth muscle in soleus muscle arterioles from WT rats; however, exercise training‐induced enhancement of vasodilation in smooth muscle is absent in soleus muscle arterioles from Nrf2KO rats. This lack of vasodilatory adaptation may contribute to greater vasoconstrictor responses in arterioles from exercise‐trained Nrf2KO rats. Further experiments in denuded arterioles are needed to definitively determine whether these exercise training‐induced adaptations occur in the vascular smooth muscle or in the endothelium of soleus muscle arterioles from WT and Nrf2KO rats.Support or Funding InformationNIH Grant Number R15AG055029This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Deficiency of the transcription factor, Nrf2, impairs vasomotor responses and limits exercise training‐induced adaptations of coronary arterioles

Nuclear factor (erythroid‐derived 2)‐like 2 (Nrf2) is a transcription factor that is crucial in maintaining cellular redox homeostasis. Vasomotor responses of the coronary resistance vasculature and regulation of coronary blood flow are closely linked to myocardial metabolism and redox status of the myocardium. We hypothesized that deletion of Nrf2 would alter vascular responses of coronary arterioles and limit exercise training‐induced adaptations of vasomotor function. We exercise trained wild type (WT) Sprague‐Dawley rats, and their littermates that were lacking the Nrf2 gene (Nrf2KO). Rats were either exercise trained (EX) on a motor‐driven treadmill 5 days/wk or remained sedentary (SED) in their cages for 10–12 weeks. A mixed moderate aerobic and high‐intensity interval training protocol was utilized. At the end of the training period, coronary arterioles were isolated for assessment of vasomotor responses to potassium chloride (KCl) and endothelin, as both are considered potential endogenous regulators of coronary vascular resistance. We also evaluated responsiveness of coronary arterioles to increasing intraluminal pressure (the myogenic response). Contractile responses of coronary arterioles to KCl and endothelin were reduced in arterioles from sedentary Nrf2KO rats as compared to arterioles from sedentary WT rats. Low concentrations of KCl induced vasodilation in coronary arterioles from exercise trained WT rats but not in arterioles from sedentary WT rats. Constriction to high concentrations of KCl was reduced in arterioles from exercise trained WT rats as compared to those from sedentary WT (WT EX vs WT SED; P&lt;0.01); however, these adaptations to exercise training were absent in arterioles from Nrf2KO rats (KO EX vs KO SED; P=0.94). Exercise training altered vasoconstrictor responses to endothelin in coronary arterioles from both WT and Nrf2KO rats; however, the effects of exercise training were directionally opposite in arterioles from WT and Nrf2KO rats. In arterioles from WT rats, exercise training reduced vasoconstrictor responses to endothelin, whereas in arterioles from Nrf2KO rats, vasoconstrictor responses to endothelin were increased by exercise training. Myogenic responsiveness was decreased in coronary arterioles from Nrf2KO rats as compared to arterioles from WT rats, but exercise training did not alter myogenic responsiveness in arterioles from either WT or Nrf2KO rats. These data suggest that the contractile responsiveness of coronary arterioles is decreased in the absence of Nrf2. The absence of Nrf2 also limits adaptation of coronary arterioles in response to exercise training. These changes in arteriolar function may be linked to alterations of redox homeostasis and metabolite production in cardiac muscle that is deficient in Nrf2.Support or Funding InformationNIH R15AG055029This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Reverse micelle Extraction of Antibiotics using an Eco-friendly Sophorolipids Biosurfactant

Reverse micelles extraction of erythromycin and amoxicillin were carried out using the novel Sophorolipids biosurfactant. By replacing commonly used chemical surfactants with biosurfactant, reverse micelle extraction can be further improved in terms of environmental friendliness and sustainability. A central composite experimental design was used to investigate the effects of solution pH, KCl concentration, and sophorolipids concentration on the reverse micelle extraction of antibiotics. The most significant factor identified during the reverse micelle extraction of both antibiotics is the pH of aqueous solutions. Best forward extraction performance for erythromycin was found at feed phase pH of approximately 8.0 with low KCl and sophorolipids concentrations. Optimum recovery of erythromycin was obtained at stripping phase pH around 10.0 and with low KCl concentration. On the other hand, best forward extraction performance for amoxicillin was found at feed phase pH around 3.5 with low KCl concentration and high sophorolipids concentration. Optimum recovery of erythromycin was obtained at stripping phase pH around 6.0 with low KCl concentration. Both erythromycin and amoxicillin were found to be very sensitive toaqueous phase pH and can be easily degraded outside of their stable pH ranges.

The effect of kappa carrageenan and salt on thermoreversible gelation of methylcellulose

Methylcellulose (MC) and kappa carrageenan (KCG) are widely used in food and pharmaceutical industries as a viscosity modifier, a gelling aid, and a film former due to their reversible thermal gelation properties. Thermoreversible gelation of MC/salt, MC/KCG/water, and MC/KCG/salt mixtures was investigated utilizing dynamic and steady shear rheological measurements. It was found that for the MC/salt mixture, gelation temperatures decreased linearly with increasing salt concentrations independent of valences of cations and molar concentrations of anions. For the MC/KCG blend, double gelation was not observed, and KCG is not influenced or disturbed the gelation properties of MC. Double gelation was observed for the mixture of MC/KCG/KCl for the low concentration of salts of 0.01 M KCl and 0.04 M KCl with the maximum moduli values for the mixture of MC/KCG/0.04 M KCl and then gradually decreased with increasing KCl salt concentration and eventually became similar to the gelation of MC solution. Therefore, KCl concentration played a major role in double gelation properties of MC/KCG/KCl mixture. It was also found that for the MC/KCG/KCl system, gelation transition matrices are linearly depending on salt concentration and independent of KCG and salt type. It was shown that for MC/salt mixture, solution rheology follows the principle of time–temperature superposition (TTS) below the gelation temperature. However, TTS failed above the gelation temperature. TTS also failed for MC/KCG and MC/KCG with low KCl concentration mixtures.

Possible involvement of transient receptor potential ankyrin 1 in Ca2+ signaling via T-type Ca2+ channel in mouse sensory neurons.

T-type Ca2+ channels and TRPA1 are expressed in sensory neurons and both are associated with pain transmission, but their functional interaction is unclear. Here we demonstrate that pharmacological evidence of the functional relation between T-type Ca2+ channels and TRPA1 in mouse sensory neurons. Low concentration of KCl at 15 mM (15K) evoked increases of intracellular Ca2+ concentration ([Ca2+ ]i ), which were suppressed by selective T-type Ca2+ channel blockers. RT-PCR showed that mouse sensory neurons expressed all subtypes of T-type Ca2+ channel. The magnitude of 15K-induced [Ca2+ ]i increase was significantly larger in neurons sensitive to allylisothiocyanate (AITC, a TRPA1 agonist) than in those insensitive to it, and in TRPA1-/- mouse sensory neurons. TRPA1 blockers diminished the [Ca2+ ]i responses to 15K in neurons sensitive to AITC, but failed to inhibit 40 mM KCl-induced [Ca2+ ]i increases even in AITC-sensitive neurons. TRPV1 blockers did not inhibit the 15K-induced [Ca2+ ]i increase regardless of the sensitivity to capsaicin. [Ca2+ ]i responses to TRPA1 agonist were enhanced by co-application with 15K. These pharmacological data suggest the possibility of functional interaction between T-type Ca2+ channels and TRPA1 in sensory neurons. Since TRPA1 channel is activated by intracellular Ca2+ , we hypothesize that Ca2+ entered via T-type Ca2+ channel activation may further stimulate TRPA1, resulting in an enhancement of nociceptive signaling. Thus, T-type Ca2+ channel may be a potential target for TRPA1-related pain.

Plasmodium falciparum GPCR-like receptor SR25 mediates extracellular K+ sensing coupled to Ca2+ signaling and stress survival

The malaria parasite Plasmodium falciparum is exposed, during its development, to major changes of ionic composition in its surrounding medium. We demonstrate that the P. falciparum serpentine-like receptor PfSR25 is a monovalent cation sensor capable of modulating Ca2+ signaling in the parasites. Changing from high (140 mM) to low (5.4 mM) KCl concentration triggers [Ca2+]cyt increase in isolated parasites and this Ca2+ rise is blocked either by phospholipase C (PLC) inhibition or by depleting the parasite’s internal Ca2+ pools. This response persists even in the absence of free extracellular Ca2+ and cannot be elicited by addition of Na+, Mg2+ or Ca2+. However, when the PfSR25 gene was deleted, no effect on [Ca2+]cyt was observed in response to changing KCl concentration in the knocked out (PfSR25−) parasite. Finally, we also demonstrate that: i) PfSR25 plays a role in parasite volume regulation, as hyperosmotic stress induces a significant decrease in parasite volume in wild type (wt), but not in PfSR25− parasites; ii) parasites lacking PfSR25 show decreased parasitemia and metacaspase gene expression on exposure to the nitric oxide donor sodium nitroprusside (SNP) and iii), compared to PfSR25− parasites, wt parasites showed a better survival in albumax-deprived condition.

Alignment of actin filament streams driven by myosin motors in crowded environments

BackgroundCellular dynamics depend on cytoskeletal filaments and motor proteins. Collective movements of filaments driven by motor proteins are observed in the presence of dense filaments in in vitro systems. As multiple macromolecules exist within cells and the physiological ionic conditions affect their interactions, crowding might contribute to ordered cytoskeletal architecture because of collective behavior. MethodsUsing an in vitro reconstituted system, we observed the emergence of stripe patterns resulting from collective actin filament streaming driven by myosin motors in the presence of the crowding agent, methylcellulose (MC). ResultsAlthough at high KCl concentrations (150mM), actin filaments tended to dissociate from a myosin-coated surface, 1% MC prevented this dissociation and enabled filament movement on myosin molecules. At concentrations of actin filaments above 0.2mg/mL, the moving filaments accumulated and progressively formed long, dense bands. The bands were spaced at about 10-μm intervals. Increasing the KCl concentration up to 300mM resulted in narrowing of the spacing between the aligned bands. On the other hand, low KCl concentrations (≤25mM) induced broad streams, where actin filaments exhibited bidirectional movement. ConclusionsThese results suggest that crowded environments can promote spatial patterning of the actin cytoskeleton, depending on the intensity of the myosin driving force and filament velocity, both modulated by the ionic strength. General significanceThe mutual contribution of packing and driving forces provides insight into cytoskeleton organization in living cells, in which various macromolecules mingle.

Terpolymer Gel System Formed by Resorcinol–Hexamethylenetetramine for Water Management in Extremely High-Temperature Reservoirs

An in situ terpolymer gel system formed by resorcinol–hexamethylenetetramine (HMTA) was developed and systematically evaluated for water management in extremely high-temperature reservoirs. Suitable gelation time and favorable gelation performance were obtained by adjusting terpolymer and/or cross-linker system concentrations. With the increase of the terpolymer, the resorcinol–HMTA concentration increased both the gelation time and gelation performance. The gel system was prepared by deionized water and maintained good thermostability in a high-salinity environment. Very low concentrations of NaCl, KCl, and CaCl2 can delay gelation time. After the critical concentrations are reached, these inorganic ions will boost the cross-linking reaction; however, the presence of MgCl2 shortens gelation time. The gel system in ampules was kept stable at 150 °C for 5 months, and the differential scanning calorimetry measurement indicated that the gel system could be used for water management at temperatures up to 240 ...

Substitutions in Spodoptera exigua topoisomerase I modulate its relaxation activity and camptothecin sensitivity.

Topoisomerase I (Top I) is referred as the cellular target of the camptothecins (CPTs) which are now being explored as potential pesticides for insect control. Three amino acid substitutions, including L530P, A653T and S729T, in Top Is of insects were found in our previous studies. In order to investigate the effect of these three substitutions, a comparative analysis was conducted between the wild-type and mutant Top Is in Spodoptera exigua Hübner. The optimal salt concentration of A653T and S729T was 150 mm, which is consistent with that of the wild-type Top I, while the mutant L530P showed maximum relaxation activity at a lower KCl concentration (100 mm). The mutated L530P and A653T Top Is showed a higher relaxation efficiency owing to an increased relaxation velocity toward the negatively supercoiled plasmid pBR322 DNA, which rendered L530P and A653T resistant to CPTs, whereas mutant S729T exhibited sensitivity to CPTs as a result of a decreased relaxation activity toward plasmid pBR322 DNA. These results suggested that the polymorphism in Top I of insects was related to the biological activity of CPTs, which provided the basic information for reasonable usage of CPTs to control insect pests. © 2016 Society of Chemical Industry.

Folding and misfolding pathways of G-quadruplex DNA.

G-quadruplexes adopt various folding topologies, but information on their folding pathways remains scarce. Here, we used electrospray mass spectrometry to detect and quantify the specifically bound potassium ions, and circular dichroism to characterize the stacking topology of each ensemble. For human telomeric (hTel) sequences containing the d((GGGTTA)3GGG) core, K+ binding affinity and cooperativity strongly depends on the chosen construct. The shortest sequences bind only one K+ at low KCl concentration, and this 2-quartet G-quadruplex is antiparallel. Flanking bases increase the K+ binding cooperativity. To decipher the folding pathways, we investigated the kinetics of K+ binding to telomeric (hybrid) and c-myc (parallel) G-quadruplexes. G-quadruplexes fold via branched pathways with multiple parallel reactions. Up to six states (one ensemble without K+, two ensembles with 1-K+ and three ensembles with 2-K+) are separated based on their formation rates and ion mobility spectrometry. All G-quadruplexes first form long-lived misfolded structures (off-pathway compared to the most stable structures) containing one K+ and two quartets in an antiparallel stacking arrangement. The results highlight the particular ruggedness of G-quadruplex nucleic acid folding landscapes. Misfolded structures can play important roles for designing artificial G-quadruplex based structures, and for conformational selection by ligands or proteins in a biological context.

Conducted dilatation to ATP and K+ in rat skeletal muscle arterioles.

AimDuring exercise in humans, circulating levels of ATP and K+ increase at a time when blood flow increases to satisfy metabolic demand. Both molecules can activate arteriolar K+ channels to stimulate vasodilatation; here, it is established whether conducted dilatation is observed in a skeletal muscle bed.MethodsIsolated and cannulated rat cremaster arterioles were used to assess both local and conducted responses. Agents were either added to the bath, focally pulse‐ejected to the downstream end of arterioles, or in triple‐cannulated arterioles, luminally perfused into the downstream branches to assess both local and conducted responses.ResultsThe endothelium‐dependent agonist ACh and the KATP channel opener levcromakalim each stimulated both local and conducted vasodilatation. Focal, bolus delivery of ATP (10 μ m) or KCl (33 mm) to the outside of arterioles stimulated a biphasic vasomotor response: rapid vasoconstriction followed by dilatation as each washed away. At lower concentrations of KCl (19 mm), constriction was avoided, and instead, Ba2+‐sensitive local dilatation and conducted dilatation were both observed. Luminal perfusion of ATP avoided constriction and activated P2Y1 receptors stimulating vasodilatation secondary to opening of KC a channels. In triple‐cannulated arterioles, either ATP (10 μ m) or K+ (15 mm) luminally perfused into daughter branches of a bifurcation stimulated local dilatation which conducted into the parent arteriole.ConclusionThe recognized physiological autocrine and paracrine mediators ATP and K+ each act to evoke both local and conducted vasodilatation in rat cremaster arterioles. Therefore, in situations when circulating levels are raised, such as during exercise, these agents can act as important regulators of blood flow.

The calcium channel agonist BAY K8644 enhances the responsiveness of human airway muscle to KCl and histamine but not to carbachol.

The role of calcium channel antagonists in asthma remains controversial. A new compound BAY K8644 is an agonist at the calcium voltage-dependent channel (VDC). Therefore, we studied the effect of BAY K8644 on contraction to KCl, histamine, and carbachol in human bronchus in vitro to assess the role of the VDC in this tissue. Cumulative concentration response curves were obtained to the agonists in bronchial spiral strips in the absence and in the presence of BAY K8644. Contractile responses to histamine were significantly (p less than 0.05) enhanced by BAY K8644 (10(-6) M) at concentrations between 10(-7) M and 3 X 10(-6) M and there was a significant decrease in the histamine EC50 (the concentration producing half the maximal response). Responses to low concentrations of KCl (10(-4) M to 3 X 10(-3) M) were also potentiated. BAY K8644 did not affect carbachol-induced contractions. We conclude that calcium influx through VDC occurs in response to KCl and histamine but not to carbachol. Such findings could explain the variability in efficacy of calcium channel antagonists in inhibiting various forms of bronchospasm.

Comparison of Activation Energy and Pore Dynamics in Liquid and Gel Phases of Electroporated Lipid Bilayers using Temperature Dependent MD Simulations

The molecular level understanding of electroporation has been studied by few research groups [1, 2, 3] over the last decade. We have performed molecular dynamics simulation(MDS) of electroporation at different temperatures to find activation energy as well as pore dynamics in the gel and liquid phases of POPC and DPPC lipid bilayers.The MDS of bilayers were performed using NAMD, the Particle mesh Ewald(PME) method, the all-atom CHARMM force field and an integrated time-step of 2 fs.The bilayers were composed of 256 lipids which were solvated with TIP3 water molecules with a low KCl concentration. The MD simulations were performed in temperature range from 250 K to 350 K with varying electric fields (0.02 to 1 V/nm).A plot of pore initiation rate as a function of inverse temperature showed Arrhenius type behaviour. The activation energy was determined to be 25.5 and 21.5 kJ/mol for the liquid phase of POPC and DPPC lipids respectively for an electric field of 0.3 V/nm, and reduces at higher fields. The activation energy in the gel phase of POPC increases to 28.8 kJ/mol at the same field. The pore closing time after the field is switched off was found to be longer in the gel phase than in the liquid phase. Remarkably, pores of radii ∼0.7nm in the gel phase of POPC did not close even after 50ns, whereas they close completely within 10ns in the liquid phase.[1]M Tarek, Biophysical. J., 88 (2005) 4045-4053.[2]PT Vernier, MJ Ziegler, Y Sun, WV Chang, MA Gundersen and DP Tieleman, J. Am. Chem. Soc., 128 (2006) 6288-6289.[3]WFD Bennett, N Sapay and DP Tieleman, Biophysical. J., 106 (2014) 210-219.

Temperature and Electrolyte Optimization of the α-Hemolysin Latch Sensing Zone for Detection of Base Modification in Double-Stranded DNA

The latch region of the wild-type protein pore α-hemolysin (α-HL) constitutes a sensing zone for individual abasic sites (and furan analogs) in double-stranded DNA (dsDNA). The presence of an abasic site or furan within a DNA duplex, electrophoretically captured in the α-HL vestibule and positioned at the latch region, can be detected based on the current blockage prior to duplex unzipping. We investigated variations in blockage current as a function of temperature (12–35°C) and KCl concentration (0.15–1.0 M) to understand the origin of the current signature and to optimize conditions for identifying the base modification. In 1 M KCl solution, substitution of a furan for a cytosine base in the latch region results in an ∼8 kJ mol−1 decrease in the activation energy for ion transport through the protein pore. This corresponds to a readily measured ∼2 pA increase in current at room temperature. Optimal resolution for detecting the presence of a furan in the latch region is achieved at lower KCl concentrations, where the noise in the measured blockage current is significantly lower. The noise associated with the blockage current also depends on the stability of the duplex (as measured from the melting temperature), where a greater noise in the measured blockage current is observed for less stable duplexes.

The influence of gap junction network complexity on pulmonary artery smooth muscle reactivity in normoxic and chronically hypoxic conditions

Experiments on intrapulmonary arteries (IPAs) isolated from rats maintained in normoxia and chronic hypobaric hypoxia showed that in normoxia, the IPA contractile sensitivity to KCl was not modified by gap junction inhibition. In contrast, chronic hypoxia induced an endothelium-independent hypersensitivity, which was suppressed by gap junction inhibition. For the theoretical analysis of these results, we developed a model of interconnected myocytes. Given that smooth muscle cells in IPAs are known to communicate via gap junctions, we regard the cytoarchitecture of the IPA as a spatial network, in which nodes represent individual smooth muscle cells and the links signify intercellular communication. A single-cell model that drives the dynamics of individual nodes includes the major elements of voltage-dependent Ca(2+) signalling. In addition, interindividual variability of SMCs is introduced by distributing the reversal potentials for K(+). Cell-to-cell connection consists of passive Ca(2+) diffusion and electrical coupling, and connection between cells is determined by the topology of the intercellular network. Model predictions indicate that the experimental results can be explained by topological modifications and not by changes in the number of gap junctions. According to the model, in normoxia the myocytes are connected in a complex network, whereas chronic hypoxia is related to loss of complexity, leading to hypersensitivity. Our results thus indicate that chronic hypoxia entails gap junction network rearrangements, leading to disturbances in the intercellular communication pathways.

Smooth Muscle Relaxation Activity of an Aqueous Extract of Dried Immature Fruit of Poncirus Trifoliata (PF-W) on an Isolated Strip of Rat Ileum

We demonstrated that an aqueous extract of dried immature fruit of Poncirus trifoliate (PF-W) produces relaxation of intestinal smooth muscle using the ileac strips of a rat. Furthermore, the underlying mechanism of its relaxant activity was investigated. PF-W was prepared using the standard extraction protocol. A 1.5 - 2 cm long rat ileac strip was placed in an organ bath with Tyrode's solution and smooth muscle contractility was recorded by connecting it to a force transducer. Various compounds were added to the organ baths, and changes in muscular contractility were measured. PF-W concentration-dependently induced relaxation of rat ileac strips that were contracted both spontaneously and via acetylcholine treatment. Various potassium channel blockers did not inhibit the relaxation by PF-W. No difference in the effect of PF-W was observed between ileac strips treated with low (20 mM) and high concentrations (60 mM) of KCl. PF-W inhibited the contraction of rat ileac strips induced by extracellular calcium. PF-W acts as a potent smooth muscle relaxant, implicating its possible action as a rapid acting reliever for abdominal pains and a cure for intestinal convulsion. Considering that PF-W also exhibits prokinetic activity, its use in various gastrointestinal disorders seems promising.