Jurgen Hannig

Surfactant Sealing of Membranes Permeabilized by Ionizing Radiation

Abstract Hannig, J., Zhang, D., Canaday, D. J., Beckett, M. A., Astumian, R. D., Weichselbaum, R. R. and Lee, R. C. Surfactant Sealing of Membranes Permeabilized by Ionizing Radiation. Acute tissue injury and subsequent inflammation, including tissue edema and erythema, can be caused by sufficiently high levels of exposure to γ radiation. The mechanism of this tissue injury is related to the generation of reactive oxygen intermediates (ROI) which chemically alter biological molecules and cell physiology. Cell membrane lipids are vulnerable to ROI-mediated lipid peroxidation that then leads to many of the acute tissue effects. We hypothesize that increased cell membrane permeability leading to osmotic swelling and vascular transudation is one of these effects. Thus we used adult postmitotic rhabdomyocytes in culture and microscopic fluorescence techniques to quantify radiation-induced changes in cell membrane permeability. Based on time-resolved dye flux measurements, a characteristic lag time of 34 ± 3 min was determined between exposure to 160 Gy of γ radiation and the decrease in membrane permeability. Administration of 0.1 mM nonionic surfactant Poloxamer 188 added to the cell medium after irradiation completely inhibited the dye loss over the time course of 2 h. Thus a reproducible model was developed for studying the mechanism of acute radiation injury and the efficacy of membrane-sealing agents. As only supportive measures now exist for treating the acute, nonlethal injuries from high-dose radiation exposure, agents that can restore cell membrane function after radiation damage may offer an important tool for therapy.

Pharmaceutical Therapies for Sealing of Permeabilized Cell Membranes in Electrical Injuriesa

ABSTRACT: Several years ago, we proposed that loss of cell membrane structural integrity by electroporation is a substantial cause of tissue necrosis in victims of electrical trauma. 1 Specifically, this involves the permeabilization of the lipid bilayer by thermal and electrical forces. We further suggested that certain mild surfactants in low concentration could induce sealing of permeabilized lipid bilayers and salvage of cells that had not been extensively heat‐damaged. Successful restoration of membrane transport properties using the surfactant poloxamer 188 was reported in 1992. 2 The purpose of this study is to further examine the response of electroporated rat skeletal muscle membranes to poloxamer 188 (P188) therapy by direct assay of membrane transport properties. Experimental evidence accumulated to date suggests that P188 is effective in sealing permeabilized cell membranes both in vitro and in vivo.

Poloxamine 1107 sealing of radiopermeabilized erythrocyte membranes

PURPOSE Lipid peroxidation-mediated permeabilization of cell membranes following intense ionizing irradiation is well documented. This form of membrane radiopermeabilization leads to rapid exhaustion of cellular high-energy compounds, resulting in the acute onset of cellular necrosis. Strategies to reverse the process of necrosis and preserve cell viability require membrane sealing. This report documents the relative efficacy of Poloxamine 1107, a non-ionic surfactant, compared with other polymers, in sealing radiopermeabilized cell membranes. MATERIALS AND METHODS Isolated erythrocytes were exposed to 600 Gy 60Co irradiation at a dose rate of 1.3 Gy/s. Different polymer compounds were added 10 min later to the irradiated cell suspensions. At 2 h later the haemoglobin content in the supernatants was determined spectrophotometrically. RESULTS Compared with the non-treated irradiated control, Poloxamine 1107 significantly reduced the leakage of haemoglobin from irradiated erythrocytes. Poloxamer 188 and dextran at equal concentrations had no significant reverse effect on the irradiation-mediated increased membrane permeability. The amount of haemoglobin released from irradiated erythrocytes was inversely related to the Poloxamine 1107 concentration. CONCLUSIONS This study demonstrates the capability of Poloxamine 1107 to seal radiopermeabilized cell membranes. Thus, surfactants such as Poloxamine 1107 might be useful as a therapeutic agent in the treatment of high-dose radiation injuries since cellular necrosis due to metabolic exhaustion following radiopermeabilization of their membranes might be prevented.

Oxidative Cell Membrane Alteration: Evidence for Surfactant-Mediated Sealinga

ABSTRACT: Exposure to very intense ionizing irradiation produces acute tissue sequelae including inflammation, pain, and swelling that often results in tissue fibrosis and/or necrosis. Acute tissue necrosis occurs in hours when sufficiently rapid damage to membrane lipids and proteins leads to altered membrane structure, disrupting the vital electrochemical diffusion barrier necessary for cell survival. 1,2 This damage mechanism is thought to underlie the interphase death of lethally irradiated postmitotic cells such as neurons, but it has also been implicated in the rapid cell death of lymphocytes and acute vascular changes due to capillary epithelium dysfunction. 3,4 It is not known whether sealing of radiation‐permeabilized cell membranes will prolong survival of lethally irradiated cells or perhaps lead to repair of damaged nucleic acids. The purpose of this study is to begin to address the first question.

Structural changes in cell membranes after ionizing electromagnetic field exposure

Ionizing radiation damages chemical structures in biological tissues mainly through the generation of reactive oxygen intermediates (ROI). Consequences on the cells transcriptional mechanisms are widely investigated but cannot account for the rapid lethal effects in victims of accidental high-dose radiation exposure. ROI-mediated structural damage of cell membranes through lipid peroxidation can lead to increased ion permeability followed by the loss of ion-homeostasis and cell death. We monitored radiation-induced changes in the morphology of isolated skeletal muscle cells. During the first two hours after gamma irradiation large blebs were formed on the surface of muscle cells. These blebs are the initiation point for a successive total collapse of the cells over a time period of about 15 min similar to Ca/sup 2+/-influx mediated cell contraction. Acute radiation-induced death as well as possible relations to lipid peroxidation, cell membrane permeability increases, irradiation-induced apoptosis are discussed. Our ongoing research on ionizing electromagnetic field effects on cell membranes and the induced structural phenomena are closely linked to pertinent membrane damage by some nonionizing electromagnetic fields and ionized gases.

25 – Membrane Biology and Biophysics

The structural integrity of the lipid bilayer component of the cell membrane is essential for facilitating the maintenance of physiological transmembrane ionic concentration gradients at a metabolic energy cost that is affordable. Despite the effectiveness of the lipid bilayer in restricting diffusive ionic transport, approximately 85%–95% of cellular metabolic energy is used to maintain the gradients by specialized ion pumps. The sodium–potassium pump may serve as an example. It simultaneously transports potassium into and sodium out of the cell, against the transmembrane ionic concentration gradients, with ATP as the energy source. It is ironic that despite its critical role in supporting life, the lipid bilayer is quite fragile in comparison to other biologic macromolecular structures. Permeabilization and disruption of the membrane lipid bilayer is a major component of massive radiation injury, reperfusion injury, thermal burns, frostbite, electrical shock, and many other forms of trauma-mediated tissue injury. Thus, the biology and biophysics of cell membrane damage and subsequent sealing are central to the science of surgery.

Surfactant therapy after electroporation mediated muscle injury in vivo

High voltage electrical shock can produce extensive muscle and nerve tissue damage due to poration of cell membranes by electric field gradients in addition to thermal denaturation of biomolecules. The authors studied the therapeutic membrane sealing effect of the surfactant Poloxamer 188 on muscle tissue damage in vivo following non-thermal electrical injury in their rat hind limb model (DC-pulses, 150 V/cm) by radiotracer imaging. Poloxamer 188 surfactant injections (50 mg/kg) showed a significant decrease in radiotracer retention compared to control injections post-shock. The results indicate the importance of membrane permeabilization and the therapeutic potential of surfactant sealing in electrical shock victims.

Physiological imaging of electrical trauma and therapeutic responses

In victims of electrical trauma, electroporation of cell membrane, in which lipid bilayer is permeabilized by thermal and electrical forces, is thought to be a substantial cause of tissue damage. It has been suggested that certain mild surfactant in low concentration could induce sealing of permeabilized lipid bilayers, thus repairing cell membranes that had not been extensively damaged. With an animal model of electrically injured hind limb of rats, we have demonstrated and validated the use of radiotracer imaging technique to assess the physiology of the damaged tissues after electrical shock and of their repairs after applying surfactant as a therapeutic strategy. For example, using Tc-99m labeled pyrophosphate (PYP), which follows calcium in cellular function and is known to accumulate in damaged tissues, we have established a physiological imaging approach for assessment of the extent of tissue injury for diagnosis and surgical planning, as well as for evaluation of responses to therapy. With the use of a small, hand-held, miniature gamma camera, this physiological imaging method can be employed at patients bedside and even in the field, for example, at accident site or during transfer for emergency care, rapid diagnosis, and prompt treatment in order to maximize the chance for tissue survival.

Histological manifestations of muscle electroporation injury

Joule heating has long been considered the principal component of tissue damage in electrical injury. In the past decade it has been shown that electroporation, a nonthermally mediated mechanism of cell membrane damage, is also a major factor. The authors investigated the electric field dose dependent extent of electroporation-mediated muscle necrosis histopathologically in vivo. Pulsed electric fields approximately 150 V/cm were produced in hind limbs of anesthetized rats. Histopathologic examination revealed prominent hypercontraction band degeneration of myofibers. The observed skeletal muscle damage was strongly dose dependent. These results support the role of non-thermal electroporation-mediated tissue damage in the extremities of high-voltage electrical injury victims.

Therapeutic effect of Vitamin C on electroporation mediated muscle injury in vivo

High voltage electrical shock can produce extensive muscle and nerve tissue damage due to thermal denaturation of proteins and poration of cell membranes by electric field gradients. To study the therapeutic effect of Vitamin C on the neurophysiologic performance of motor nerves following electrical injury, the authors used an in vivo model exposing an anesthetized rats hindlimb to thermally neutral electrical shocks. The applied pulsed electrical fields had a strength of approximately 150 V/cm. A standard EMG test was used to monitor the damage and recovery of the motor nerve function. Intravenously administered Vitamin C at concentrations of 5.8 mg/kg and 340 mg/kg showed a dose-dependent, statistically significant improvement in the recovery of the muscle action potential amplitude.