Pulse‐chase experiments in conjunction with subcellular fractionation and quantitative immunoprecipitation have been used to study the intracellular transport of four secretory proteins, albumin, transferrin, prealbumin and retinol‐binding protein, in isolated rat hepatocytes. The proteins were found to be transported from the endoplasmic reticulum (ER) to the Golgi complex (GC) at greatly different rates (t1/2 = 14‐137 min), indicating that transport of secretory proteins between these organelles is effected by a selective, possibly receptor‐mediated process and not through bulk phase transfers. The transport from the Golgi complex to the medium was rapid for all proteins (t1/2 approximately 15 min) and possibly occurred at the same rate. Consistent with these kinetic data, the amount of a rapidly transported protein (albumin) in the GC fraction was found to be high (relative to its amount in the ER fraction) whereas the amount of a slowly transported protein (transferrin) in the GC fraction was found to be low, as determined by radioimmunoassays.
Spider silk has recently become a material of high interest for a large number of biomedical applications. Previous work on structuring of silk has resulted in particles (0D), fibers (1D), films (2D), and foams, gels, capsules, or microspheres (3D). However, the manufacturing process of these structures is complex and involves posttreatment of chemicals unsuitable for biological applications. In this work, the self-assembly of recombinant spider silk on micropatterned superhydrophobic surfaces is studied. For the first time, structuring of recombinant spider silk is achieved using superhydrophobic surfaces under conditions that retain the bioactivity of the functionalized silk. By tuning the superhydrophobic surface geometry and the silk solution handling parameters, this approach allows controlled generation of silk coatings, nanowires, and sheets. The underlying mechanisms and governing parameters are discussed. It is believed that the results of this work pave the way for fabrication of silk formations for applications including vehicles for drug delivery, optical sensing, antimicrobial coatings, and cell culture scaffolds.
Biologically compatible membranes are of high interest for several biological and medical applications. Tissue engineering, for example, would greatly benefit from ultrathin, yet easy‐to‐handle, biodegradable membranes that are permeable to proteins and support cell growth. In this work, nanomembranes are formed by self‐assembly of a recombinant spider silk protein into a nanofibrillar network at the interface of a standing aqueous solution. The membranes are cm‐sized, free‐standing, bioactive and as thin as 250 nm. Despite their nanoscale thickness, the membranes feature an ultimate engineering strain of over 220% and a toughness of 5.2 MPa. Moreover, they are permeable to human blood plasma proteins and promote cell adherence and proliferation. Human keratinocytes seeded on either side of the membrane form a confluent monolayer within three days. The significance of these results lays in the unique combination of nanoscale thickness, elasticity, toughness, biodegradability, protein permeability and support for cell growth, as this may enable new applications in tissue engineering including bi‐layered in vitro tissue models and support for clinical transplantation of coherent cell layers.
The effect of acutely induced polycythemia on blood flow and viscosity in the vasodilated vascular bed of working and non-working skeletal muscle was studied. In 12 mongrel dogs anesthetized with thiopental sodium the calf muscle of one hind limb was isolated. Vasodilation was induced either by sciatic stimulation setting the muscle at maximal work or by i.a. infusion of papaverine. Blood flow was measured at different perfusion pressure before and after infusion of 300 ml packed homologous red cells. Blood viscosity in vitro was determined in a coneplate viscometer. Apparent viscosity in vivo was analyzed by comparing pressure-flow relationships for blood and a reference solution. Polycythemia decreased blood flow by 35% in the non-working muscle but less than 10% in the working muscle at comparable perfusion pressures. Blood viscosity in vitro increased by 35% at low shear rates. Apparent viscosity in vivo increased by 35% in the non-working muscle but less than 10% in the working muscle. The flow impairment induced by polycythemia was far more pronounced in the non-working skeletal muscle indicating a flow facilitation by the rhythmic muscle contractions. Erythrocyte flow in fact increased in the working muscle after induced polycythemia while decreased in the non-working muscle.
The effects of increased plasma viscosity and induced red blood cell (RBC) aggregation on apparent viscosity of blood in vivo in the skeletal muscle of the dog were studied. Apparent viscosity in vivo was determined in the isolated and vasodilated calf muscles of one hindlimb by comparing pressure-flow relationships for RBC suspensions with pressure-flow relationships for a Newtonian solution of known viscosity. RBC suspensions of increased plasma viscosity with and without RBC aggregation were obtained by substituting plasma with isoviscous solutions of high- and low-molecular-weight dextran in saline. Hematocrits of the suspensions were adjusted to either 45 or 60%. The viscosities of the suspensions in vitro were determined in a Wells-Brookfield viscometer. Apparent viscosity of blood in vivo was found to be mainly dependent on the viscosity of plasma. RBC aggregation had no significant influence on the viscosity in vivo.
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