Surface Membrane Alterations Research Articles

Mechanistically distinct steps in the mitochondrial death pathway triggered by oxidative stress in cardiac myocytes.

Oxidative stress plays an important role in the pathogenesis of cardiovascular diseases. In the present study, we characterize three distinct phases of the H2O2-induced response, which leads to loss of mitochondrial membrane potential (DeltaPsi(m)) and subsequent cell death in cultured cardiac myocytes. (1) Priming: After H2O2 exposure (100 micromol/L), cells maintain a constant DeltaPsi(m) for the cell-to-cell specific latency but at the same time undergo progressive changes in inner mitochondrial membrane structure (swelling and loss of cristae by electron microscopy). An increase of matrix calcium is required, but not sufficient, for this process. (2) Depolarization: Priming is followed by sudden depolarization of DeltaPsi(m), which is mediated by mitochondrial permeability transition pore opening, as evidenced by the concomitant release of calcein from mitochondria. This process is rapid (<4 minutes), complete, and irreversible. The duration of depolarization is constant and does not depend on the length of the priming process in any given cell. (3) Fragmentation: Along with massive mitochondrial swelling and release of cytochrome c into the cytoplasm, cells undergo surface membrane alterations, such as exposure of phosphatidylserine and eventual loss of membrane integrity and cellular fragmentation. Thus, oxidant stress elicits reproducible and stereotyped responses in cardiac cells. The priming phase, during which mitochondria undergo major ultrastructural alterations but remain functional, represents a particularly attractive target for intervention in the prevention of cell death.

Intracellular calcium dynamics and cerebral injury: modeling various insults in vitro

The magnitude and time course of intracellular [Ca 2+] i alterations were studied after excitatory amino acid challenge (EAA) or chemical energy depletion in mature spinal cultures. While either cytotoxic event led to prompt increases in [Ca 2+] i, the pattern of these changes before and after exposure to the toxin was different. EAA [Ca 2+] i changes seem primarily dependent on surface membrane alterations from which the cells rapidly recover while energy depletion effects release of [Ca 2+] i from intracellular stores and produces a lasting compromise in the ability of these neurons in culture to recover from the initial insult.

Flow‐cytometric detection of surface membrane alterations and concomitant changes in the cytoskeletal actin status of activated platelets

Occlusive vascular diseases are promoted by a "prethrombotic state" with increased platelet activity. Polymerization of cytoskeletal proteins and exposure of subcellular structures or rebinding of secreted proteins have been characterized as early reactions after platelet activation preceding adhesion and aggregation. Here, we demonstrate the kinetic increase in specific binding of monoclonal antibodies to thrombospondin (P10) and to platelet membrane activation markers CD63 (GP53, a 53 kD lysosomal protein) and CD62 (GMP140, a 140 kD alpha granule protein) by using a flow-cytometric bio-assay and the related change in the actin status by using the DNase-I inhibition assay after stimulation of normal human platelets with 0.2 U/ml thrombin. F-actin was raised from 41% to 51% of total platelet actin content 30 s after stimulation and remained thereafter constant (50% at 60 s). Simultaneously, the percentage of P10, CD63, and CD62 positive platelets was elevated from 5.4%, 24.4%, and 9.1% to 67.4%, 80.2%, and 82.3% respectively. The mean number of P10, CD63, and CD62 antibody binding sites increased from 3,300, 1,715, and 2,146 to 6,400, 6,800, and 9,016 per platelet. Conclusively, changes in the organization of the cytoskeletal protein "actin" and exposure of subcellular structures indicating platelet secretion can be regarded as markers of early platelet activation. Thus, the parallel response in both analytical systems provides further support for the diagnostic concept of flow-cytometric detection of preactivated platelets in the peripheral blood by using fluochrome staining procedures detecting activation dependent structural alterations directly at the cellular level.

Mechanisms of Psoralen Action in the Skin.

The combination of psoralens and ultraviolet light (UVA, 320-400 nm), also referred to as PUVA, is a potent modulator of epidermal cell growth and differentiation. Although it has been postulated that PUVA exerts its actions by binding to DNA, our laboratory has obtained evidence that a specific, saturable, high-affinity receptor site independent of the DNA mediates the biologic actions of these drugs. This receptor is a 22,000 molecular weight protein present in membrane and cytoplasmic fractions of responsive cell types. Treatment of cells with psoralens followed by UV light causes activation of the receptor. This leads to specific cell surface membrane alterations, in particular phosphorylation of the receptor for epidermal growth factor (EGF). The EGF receptor is a transmembrane glycoprotein possessing intrinsic tyrosine kinase activity. Modification of the EGF receptor leads to a loss in its ability to bind EGF, as well as an inhibition of its tyrosine kinase activity. These data indicate that the psoralens act at the level of the cell membrane and that their biologic effects in the skin may be mediated by the ability of these drugs to disrupt normal growth factor functions.

Distinction of viral and host-derived glycopolypeptides induced by "early" functions of human cytomegalovirus.

Human cytomegalovirus is shown to induce in phosphonoacetic acid-treated human fibroblasts glycosylation of five polypeptides with approximate molecular weights of 200-250, 150, 135, 130 and 100 kilodaltons (kd). Except for the 130 kd product, these glycopolypeptides (gp) separate with the cytoplasmic fraction, only one (200-250 kd) with the chromatin fraction as well. The gp of 135 and 100 kd were found to be virus-specified as determined by immunoblotting and immunoprecipitation. The gp of 200-250 kd exhibited an immunological relatedness to fibronectin and are therefore considered host-specific products. Both subsets of gp participate in virus-induced surface membrane alterations as documented by living cell immunofluorescence.

Surface Features of Leukaemic Megakaryocytic Precursors

Megakaryoblasts and maturing megakaryocytic precursor cells from 5 patients with megakaryoblastic leukaemia were studied by scanning electron microscopy (SEM). The diagnosis in all cases had been established by ultrastructural cytochemistry on the basis of a positive platelet peroxidase reaction with negative staining for myeloperoxidase. 1 case presented as acute myelofibrosis and 4 as acute megakaryoblastic transformation of chronic granulocytic leukaemia. Under the SEM, megakaryoblasts and maturing megakaryocytic precursors showed typical surface features including the presence of rounded and irregular blebs, broad folds and pseudopodia. The nature of these surface blebs is still unclear but they probably represent surface membrane alterations relating to imminent platelet shedding at least in the more mature precursors. These surface microprojections are distinctly different from those encountered on leukaemic lymphoblasts, myeloblasts and monoblasts. It is suggested that SEM may be used in conjunction with the PPO reaction as in aid in the diagnosis of megakaryocytic leukaemias.

Disturbance of redistribution of surface membrane receptors on peripheral mononuclear cells of patients with Down's syndrome and of aged individuals.

Redistribution of surface membrane immunoglobulins (SmIg) and concanavalin A (Con-A) receptor sites were studied in the peripheral mononuclear cells of institutionalized patients with Down's syndrome (DS), non-DS patients from the same institution, normal hospital staff members, and a group of healthy-appearing older volunteers averaging over 85 years of age. Two ligands were chosen: fluorescein-labeled polyvalant goat anti-human immunoglobulin for T-depleted mononuclear cells, and fluoresceinated Con-A for unfractionated peripheral mononuclear cells. The percentage of cells showing capping of SmIg and Con-A receptor sites was significantly lower in aged persons and in DS patients than in non-DS patients and normal staff members. Such a disturbance of mobility of cell membrane receptors in aged persons indicates the existence of alterations in surface membrane and associated structures of peripheral mononuclear (lymphoid) cells with aging in humans. The finding of a lower degree of capping in DS than in controls of similar age supports the supposition that DS shows features of accelerated aging.

Membrane damage caused by 8-MOP and UVA-treatment of cultivated cells

Treatment of red blood cells with a combination of 8-MOP and UVA did not cause any significant hemolysis under the experimental conditions used, and the deuterium test for identification of singlet oxygen mediated reactions was negative. However, similar experiments performed on in vitro cultivated human glia cells caused conspicuous surface membrane alterations, as revealed by scanning electron microscopy. The alterations were seen at higher concentration levels than was expected and might be secondary to earlier appearing intracellular events seen at lower concentrations. The clinical significance of membrane alterations for the results obtained with PUVA therapy require further investigation.

Utilization of exogenous linolenic and oleic acids for plasma membrane phosphoglyceride systhesis in L-Fibroblasts

The inability of the strain L-fibroblast to synthesize quatitatively significant amounts of polyenoic fatty acid and the apparent lack of turnover of their phosphoglyceride acyl groups under the usual conditions of cell culture makes them especially well suited for studies concerning the effect of fatty acid unsaturation on biological membranes. Such cells grown in the absence of exogenous lipid sources have in their phosphoglycerides only traces of polyenoic fatty acid. By infusing fatty acid supplements into suspension cultures of logarithmically growing cultures of L-fibroblasts it is possible to increase singnificantly their phosphoglyceride polyenoic fatty content to as much as 50% of the total lipid phosphoglyceride fatty acids. The infusion of fatty acid supplements at a constant rate over a 48 h time period diminishes the toxic effects which may accompany single doses of unesterified fatty acid and reduces considerably the accumulation of cytoplasmic lipid droplets. Cultures supplemented in this way have virtually the same generation times as non-supplemented control cultures. The data show that alterations in surface membrane and homogenate polyenoic fatty acid composition are minimal when oleic acid is supplied to the culture. During exposure to large amounts of polyenoic fatty acid, however, the unsaturation of plasma membrane total phosphoglyceride fraction is less than that of the cell homogenate. This effect is more pronounced in the phosphatidylethanolamine than in the phosphatidylcholine fraction.

Generation of Cytotoxic T Lymphocytes during Coxsackievirus B-3 Infection

Abstract The production of cytotoxic cells in the spleens of adult male BALB/c mice infected with Coxsackievirus B-3 has been examined. An in vitro 51Cr release assay was used to measure cytotoxic activity against virus-infected and uninfected neonatal syngeneic fibroblasts. Cytotoxicity of immune spleen cells against virus-infected targets was detected on the 3rd day after infection, reached a peak on day 7, and then declined to low levels by days 12 and 14. Spleen cells obtained 3 and 5 days after infection also exerted cytotoxicity against uninfected fibroblasts, but by the 7th day there was little or no reactivity against uninfected target cells, although activity against infected fibroblasts was maximal at this time. Reciprocal assays performed by using Coxsackie and vaccinia viruses provided evidence of virus specificity of the cytotoxic reaction. When spleen cells were obtained 7 days after infection, the Coxsackievirus-immune population was not cytotoxic for vaccinia-infected fibroblasts, and the vaccinia-immune population was not cytotoxic for Coxsackievirus-infected targets, although each immune cell preparation caused significant lysis of fibroblasts infected with the homologous virus. Additional studies showed that primary mouse or hyperimmune rabbit anti-Coxsackieviral serum could not block immune spleen cell cytotoxicity or induce complement-mediated lysis of in fected targets. The findings indicate that Coxsackievirus infection results in surface membrane alterations, but no evidence was obtained that antiviral antibody could react with the infected cells.

Erythrocyte Surface Membrane Alterations

Electrophoretic mobility measurements were made of red blood cells obtained from patients with Duchenne and myotonic muscular dystrophy, from dystrophic mice and chickens, and from corresponding controls. Alterations in the erythrocyte surface electrokinetic properties were found in dystrophic mice and chickens and in many, but not all, patients with muscular dystrophy. The results are consistent with the concept of muscular dystrophy as a systemic membrane disease not limited to muscle.

Surface membrane alterations in somatic cell hybrids of neuroblastoma and glioma cells.

SOMATIC cell hybrids have been used to discern many properties of parental cells. Genes for neuronal properties can be activated in hybrids formed by fusion of mouse neuroblastoma and rat glioma cells (refs 1 and 2, and B. Hamprecht, T. Amano and M. Nirenberg, personal communication). These cell hybrids resemble the parent neuroblastoma cells in many of their properties as they can be induced to extend long neurites and possess electrically excitable membranes1. In addition the cell hybrids express other neuronal characteristics that are less pronounced in the parental cells, such as choline acetyltransferase1, and morphine receptors2. Many of the above properties suggest alterations in the surface membrane. We have shown previously that a particular group of glycopeptides is more prominently expressed on the cell surface in clones of mouse neuroblastoma which have the ability to undergo morphological differentiation than in clones that are unable to extend neurites3. In the present report, we show that this group of glycopeptides is abundant in the surface membranes of the hybrid cells.

Desmosomal structures in the cytoplasm of normal and abnormal keratinocytes (author's transl)

The occurrence of intracytoplasmic desmosomes in normal, hyperplastic, and hyperkeratotic epithelia, in carcinoma-in-situ and in invasive carcinoma of the human oral cavity is demonstrated by electron microscopy. The mechanism for formation of these structures by invagination, separation and by intracytoplasmic incorporation of plasma membrane-desmosome-complexes are described in various oral epithelia, and other possible mechanisms are discussed. Intracytoplasmic desmosomes may occur in normal and pathological keratinocytes of all layers of the oral epithelium. Their ultrastructure in the peripheral cytoplasm is similar to that of the regular desmosomes on the cell surface. However, as they migrate centripetally, they show signs of degeneration, suggesting dissolution by lysosomal enzyme systems. Various surface membrane alterations involved in the formation of intracytoplasmic desmosomes may lead to a reduction of plasma membrane material and of desmosome structures and to defective intercellular adhesion. The intracytoplasmic incorporation of desmosome structures is a ubiquitous phenomenon exhibited by epithelial keratinocytes under certain physiological or pathological conditions.

The SV40 "S" antigen and other papovavirus-induced surface antigens.

Although a variety of properties which characterize virus-transformed animal cells (e.g., morphological changes, loss of growth control, agglutination by plant lectins) have been linked to surface membrane alterations (Black et al., 1971), perhaps the most important surface alteration which occurs in the transformed cell is that involving antigenic changes, since the definitive feature of the neoplastic cell is the ability to produce a tumor in an appropriate host. Whether a transformed cell will produce a tumor in the autochthonous or syngeneic host involves many factors, but of great significance is the presence of “new” surface membrane antigens associated with the transformation process and the immunological response they evoke.KeywordsSialic AcidSurface AntigenSimian VirusPolyoma VirusTransplantation AntigenThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.