Proton-induced endocytosis is dependent on cell membrane fluidity, lipid-phase order and the membrane resting potential

AbstractRecently it has been shown that decreasing the extracellular pH of cells stimulates the formation of inward membrane invaginations and vesicles, accompanied by an enhanced uptake of macromolecules. This type of endocytosis was coined as proton-induced uptake (PIU). Though the initial induction of inward membrane curvature was rationalized in terms of proton-based increase of charge asymmetry across the membrane, the dependence of the phenomenon on plasma membrane characteristics is still unknown. The present study shows that depolarization of the membrane resting potential elevates PIU by 25%, while hyperpolarization attenuates it by 25%. Comparison of uptake in suspended and adherent cells implicates that the resting-potential affects PIU through remodeling the actin-cytoskeleton. The pH at the external interface of the cell membrane rather than the pH gradient across it determines the extent of PIU. PIU increases linearly upon temperature increase in the range of 4–36°C, in correlation with the membrane fluidity. The plasma membrane fluidity and the lipid phase order are modulated by enriching the cell's membrane with cholesterol, tergitol, dimethylsulfoxide, 6-ketocholestanol and phloretin and by cholesterol depletion. These treatments are shown to alter the extent of PIU and are better correlated with membrane fluidity than with the lipid phase order. We suggest that the lipid phase order and fluidity influence PIU by regulating the lipid order gradient across the perimeter of the lipid-condensed microdomains (rafts) and alter the characteristic tension line that separates the higher ordered lipid-domains from the lesser ordered ones

Leveraging Social Media to Promote EvidenceBased Continuing Medical Education

Importance New dissemination methods are needed to engage physicians in evidence-based continuing medical education (CME). Objective To examine the effectiveness of social media in engaging physicians in non-industry-sponsored CME. Design We tested the effect of different media platforms (e-mail, Facebook, paid Facebook and Twitter), CME topics, and different “hooks” (e.g., Q&A, clinical pearl and best evidence) on driving clicks to a landing site featuring non-industry sponsored CME. We modelled the effects of social media platform, CME topic, and hook using negative binomial regression on clicks to a single landing site. We used clicks to landing site adjusted for exposure and message number to calculate rate ratios. To understand how physicians interact with CME content on social media, we also conducted interviews with 10 physicians. Setting The National Physicians Alliance (NPA) membership. Participants NPA e-mail recipients, Facebook followers and friends, and Twitter followers. Main Outcomes and Measures Clicks to the NPA’s CME landing site. Results On average, 4,544 recipients received each message. Messages generated a total of 592 clicks to the landing site, for a rate of 5.4 clicks per 1000 recipients exposed. There were 5.4 clicks from e-mail, 11.9 clicks from Facebook, 5.5 clicks from paid Facebook, and 6.9 clicks from Twitter to the landing site for 1000 physicians exposed to each of 4 selected CME modules. A Facebook post generated 2.3x as many clicks to the landing site as did an e-mail after controlling for participant exposure, hook type and CME topic (p Conclusions Social media has a modest impact on driving traffic to evidence-based CME options. Facebook had a superior effect on driving physician web traffic to evidence-based CME compared to other social media platforms and email

Correlation between local cell membrane displacements and filterability of human red blood cells

AbstractLocal mechanical fluctuations of the cell membrane of human erythrocytes were shown to involve MgATP- and Mg2+-driven fast membrane displacements. We propose that these local bending deformations of the cell membrane are important for cell passage through capillaries. In order to verify this hypothesis, we examined cell membrane fluctuations and filterability of erythrocytes over a wide range of medium osmolalities (180–675 mosmol/kg H2O). The results indicate the existence of a positive correlation between the amplitude of local cell membrane displacements and cell filterability. We suggest that the occurrence of metabolically driven membrane displacements on the side surface of the red blood cell diminishes its bending stiffness and enables it to fold more efficiently upon entrance into blood capillaries. Thus, local cell membrane displacements seem to play an important role in microcirculation

Regulation of the Na+/H+ exchanger under conditions of abolished proton gradient: Isosmotic and hyperosmotic stimulation

AbstractActivation of the Na+/H+ exchanger following isosmotic and hyperosmotic stimuli was investigated in an osteoblast cell line (RCJ 1.20). The pH dependence of the transporter activity was studied under conditions of abolished proton gradient (pHi = PH*) across the membrane. The isotonic response is Na+* dependent, increases towards higher pH. values, displaying a sigmoidal dependence on pH1** (Hill coefficient ≈ 1.8) and is controlled by pH*. The greater than first order dependence of pH suggest that H+* inhibits the exchange beyond the rate expected from competition with the Na+* alone. This may be due to the existence of an external H+ regulatory site with a negative cooperative effect on the intra- or extracellular transport site. The hyperosmotic activation is Na+* independent, parallels the sigmoidal pH dependence of the isosmotic stimulus (Hill coefficient ≈ 2.0) and is mediated through an increase of the Vmax without a change in the intracellular proton sensitivity

Fast cell membrane displacements in B lymphocytes Modulation by dihydrocytochalasin B and colchicine

AbstractA novel type of cell membrane movement was characterized in B lymphocytes. Local submicron cell membrane displacements, within the frequency range 0.3–15 Hz, were registered in a murine lymphoma B cell line by a novel optical method based on point dark field microscopy. The cell membrane displacements were measured by monitoring changes in light scattering from very small illuminated areas (0.25 μm2) at the edge of the cell surface. B lymphocytes manifest a relative change in light scattering of 7.7 ± 1.3% (mean ± SD) which corresponds to cell membrane transverse displacement of 131 ± 22 nm. The confinement of cell membrane displacements to microdomains (≤0.2 μm2) emerged from the observed dependence of the displacement amplitude on the area size from which it is monitored. Colchicine (1 μM) decreased membrane fluctuations down to a value of 88 ± 14 nm, whereas dihydrocytochalasin B (2 μM) increased the amplitude of membrane displacements up to 184 ± 31 nm. These findings demonstrate the existence of a dynamic mechanical interaction between the cytoskeleton and the cell membrane in the frequency range of 0.3–15 Hz. The modulation of these interactions by the disruption of microfilaments or microtubules is explained in terms of the induced strain changes imposed on the cell membrane

Oxygenation-deoxygenation cycle of erythrocytes modulates submicron cell membrane fluctuations

Low frequency submicron fluctuations of the cell membrane were recently shown to be characteristic for different cell types, nevertheless their physiological role is yet unknown. Point dark-field microscopy based recordings of these local displacements of cell membrane in human erythrocytes, subjected to cyclic oxygenation and deoxygenation, reveals a reversible decrease of displacement amplitudes from 290 +/- 49 to 160 +/- 32 nm, respectively. A higher rate of RBC adhesion to a glass substratum is observed upon deoxygenation, probably due to a low level of fluctuation amplitudes. The variation in the amplitude of these displacements were reconstituted in open RBC ghosts by perfusing them with composite solutions of 2,3 diphosphoglycerate, Mg+2, and MgATP, which mimic the intracellular metabolite concentrations in oxygenated and deoxygenated erythrocytes. The mere change in intracellular Mg+2 during oxygenation-deoxygenation cycle is sufficient to explain these findings. The results imply that the magnitude of fluctuations amplitude is directly connected with cell deformability. This study suggests that the physiological cycle of oxygenation-deoxygenation provides a dynamic control of the bending deformability and adhesiveness characteristics of the RBC via a Mg+2-dependent reversible assembly of membrane-skeleton proteins. The existing coupling between oxygenation-deoxygenation of the RBC and its mechanical properties is expected to play a key role in blood microcirculation and may constitute an example of a general situation for other circulating blood cells, where the metabolic control of cytoskeleton dynamics may modulate their dynamic mechanical properties

Multifrequency Analysis of Single Inductive Coil Measurements Across a Gel Phantom Simulation of Internal Bleeding in the Brain

The present study is part of an ongoing effort to develop a simple diagnostic technology for detecting internal bleeding in the brain, which can be used in lieu or in support of medical imaging and thereby reduce the cost of diagnostics in general, and in particular, would make diagnostics accessible to economically disadvantaged populations. The study deals with a single coil inductive device to be used for detecting cerebral hemorrhage. It presents a first‐order experimental study that examines the predictions of our recently published theoretical study. The experimental model employs a homogeneous cylindrical phantom in which internal head bleeding was simulated by way of a fluid inclusion. We measured the changes in amplitude and phase across the coil with a network vector analyzer as a function of frequency (100–1,000 MHz), volume of blood simulating fluid, and the site of the fluid injection. We have developed a new mathematical model to statistically analyze the complex data produced in this experiment. We determined that the resolution for the fluid volume increase following fluid injection is strongly dependent on frequency as well as the location of liquid accumulation. The experimental data obtained in this study supports the predictions of our previous theoretical study, and the statistical analysis shows that the simple single coil device is sensitive enough to detect changes due to fluid volume alteration of two milliliters. Bioelectromagnetics. 2020;41:21–33. © 2019 Bioelectromagnetics SocietyThis work is based on a portion of a dissertation to be submitted by Moshe Oziel in partial fulfillment of the requirements for a PhD degree to Tel‐Aviv University