The Journal of Membrane Biology

We used specific binding of phlorizin to the intact intestinal mucosa in order to measure glucose transport site density in intestines of mice fed a high-carbohydrate or no-carbohydrate diet. Nonspecific binding varied with intestinal position but showed only modest dependence on diet. Specific binding to glucose transporters was 1.9 times greater in jejunum of high-carbohydrate mice than of no-carbohydrate mice; this ratio was the same as the ratio for Vmax values of actived-glucose uptake between the two diet groups. The gradient in specific binding of phlorizin along the intestine paralleled the gradient in Vmax of glucose transport. These results directly demonstrate that the increase in intestinal glucose transport caused by a high-carbohydrate diet is due to induction of glucose transporter. They also indicate that the normal positional graident in glucose transport along the intestine arises from a gradient in transporters, induced by the normal gradient in luminal glucose concentration.

Connexins are proteins that form gap junctions. Perturbations in the cell membrane reportedly promote changes in the expression profile of connexins. Electroporation promotes destabilization by applying electrical pulses, and this procedure is used in electrochemotherapy and gene therapy, among others. This in vitro work aimed to study the interference of electroporation on the expression profile of GJB2 (Cx26 gene) and Connexin 26 in melanoma cell line B16/BL6. The techniques of immunocytochemistry, Western blot, and real-time PCR were used. After electroporation, cells showed a transient decrease in GJB2 mRNA. The immunostaining of Cx26 showed no noticeable change after electroporation at different time points. However, Western blot showed a significant reduction in Cx26 30 min after electroporation. Our results showed that electroporation interferes transiently in the expression of Connexin 26 in melanoma and are consistent with the idea that electroporation is a process of intense stress that promotes cell homeostatic imbalance and results in disruption of cell physiological processes such as transcription and translation.

Optogenetics is the integration of genetics and optics to achieve gain or loss of function of well-defined events in specific cells of living tissue. As a versatile tool, upon light illumination, it allows fast control of precisely defined events in biological systems from single cell to different parts of whole tissue in freely moving animals. Taking advantage of this method, a multitude of studies have been published to understand brain functions and dysfunctions. Although from the beginning, it has been used to target neurons within the neural networks and to understand how specific neurons contribute to brain function, it gradually has been extended to other fields of biology such as stem cell research and therapy. With a combination of optogenetics and stem cells, new opportunities were opened up in stem cell biology and also its integration in new circuit as a cell-based treatment strategy for more common disorders like neurodegenerative and cardiovascular one. Recently, some studies showed that engineered stem cells expressing exogenous light-activated opsins can be used in stem cell biology including tracking the differentiation of stem cells, functional analysis of embryonic stem cell-derived graft, and testing the functional integration of induced pluripotent stem cell-derived neurons. With the advent of non-invasive approach, such as transcranial excitation or inhibition, optogenetics also holds promise for non-invasive control of engineered stem cell.

The effect of biliary salts and fatty acids on the bilayer structure of rabbit intestinal brush-border membranes was studied using the nonperturbing probe31P NMR. The broad. asymmetric lineshape of the31P NMR spectrum of isolated brush-border vesicles demostrates that their component phospholipids are organized in extended bilayers. These membranes are not significantly perturbed by incubation with physiological concentrations of biliary salts (3, 9, 18mm), demonstrating that the vesicles are highly stable, corresponding to their biological function. However, the emergence of a narrow peak superimposed on the broad lineshape indicates that a small proportion of the membrane phospholipids has reached isotropic motion, which may correspond to external or internal micellar structures. Incubation with mixed micelles of fatty acids and taurochlorate show that long-chain fatty acids enhance the membrane-perturbing effect of taurocholate while short-chain, watersoluble fatty acids do not, suggesting a difference in the absorption mechanisms.

Five major membrane glycoproteins of the BHK-B4 hamster fibroblast plasma membrane have been identified by binding specific rabbit antibodies to the cell surface and by recovering the detergent solubilized immunocomplexes with Protein A-Sepharose immunoadsorption. These glycoproteins, designated as gp45, gp65, gp95, gp130 and gp140, are exposed at the cell surface since: (i) they were accessible to antibodies in intact viable cells; (ii) they were radioiodinated by the lactoperoxidase-glucose oxidase procedure; and (iii) they were cleaved by proteolytic enzymes in conditions affecting only the cell surface. Among these glycoproteins the gp130 is the predominant component and its exposed portion is characterized by lack of sensitivity to trypsin cleavage. Glycoproteins of different molecular weight, but immunologically related to the major hamster membrane glycoproteins, have been detected at the surface of both rat and mouse fibroblasts.

An assay has been developed for quantitating the reassociation of cortical secretory vesicles (CVs) with fragments of sea urchin egg plasma membrane attached to glass slides (PM lawns). Binding ofS. pupuratus CVs to homologous PM lawns increased with time and CV concentration. The observation that CV binding was blocked by chymotrypsin digestion of the PM fragments suggested that a PM protein(s) is required for reassociation. The possibility that the extent of CV lysis that occurred during CV preparation (15.4±3.8% as assessed by ovoperoxidase assay) influenced reassociation was investigated by determining the effect of CV content proteins (isolated as fertilization product) on binding. Various concentrations of fertilization product (up to equivalent amounts of fertilization product and CV protein) had no effect on CV binding. The specificity of binding was investigated by assessing the ability of CVs to bind to PM lawns prepared from human red blood cells, and by determining the ability of heterologous vesicles to bind to egg PM fragments. PM lawns from HRBCs did not support CV binding; however, PM lawns prepared from the eggs of several species of sea urchin did bindS. pupuratus CVs. Vesicles from a partially purified preparation of yolk platelets bound to egg PM lawns with low efficiency (1/7 that of CVs), but immunofluorescence analysis with an anti-hyalin monoclonal antibody demonstrated that 74±9% of the bound vesicles were CVs that contaminated the yolk platelet preparation. Dioleoylphosphatidyl choline liposomes were also unable to bind to egg PM lawns. These results are consistent with hypothesis that CV binding to egg PM lawns is a specific, protein-mediated event.