Limitations to exercise- and maximal insulin-stimulated muscle glucose uptake

The hypothesis of this investigation was that insulin and muscle contraction, by increasing the rate of skeletal muscle glucose transport, would bias control so that glucose delivery to the sarcolemma (and t tubule) and phosphorylation of glucose intracellularly would exert more influence over glucose uptake. Because of the substantial increases in blood flow (and hence glucose delivery) that accompany exercise, we predicted that glucose phosphorylation would become more rate determining during exercise. The transsarcolemmal glucose gradient (TSGG; the glucose concentration difference across the membrane) is inversely related to the degree to which glucose transport determines the rate of glucose uptake. The TSGG was determined by using isotopic methods in conscious rats during euglycemic hyperinsulinemia [Ins; 20 mU/(kg. min); n = 7], during treadmill exercise (Ex, n = 6), and in sedentary, saline-infused rats (Bas, n = 13). Rats received primed, constant intravenous infusions of trace 3-O-[3H]methyl-D-glucose and [U-14C]mannitol. Then 2-deoxy-[3H]glucose was infused for the calculation of a glucose metabolic index (Rg). At the end of experiments, rats were anesthetized, and soleus muscles were excised. Total soleus glucose concentration and the steady-state ratio of intracellular to extracellular 3-O-[3H]methyl-D-glucose (which distributes on the basis of the TSGG) were used to calculate ranges of possible glucose concentrations ([G]) at the inner and outer sarcolemmal surfaces ([G]im and [G]om, respectively). Soleus Rg was increased in Ins and further increased in Ex. In Ins, total soleus glucose, [G]om, and the TSGG were decreased compared with Bas, while [G]im remained near 0. In Ex, total soleus glucose and [G]im were increased compared with Bas, and there was not a decrease in [G]om as was observed in Ins. In addition, accumulation of intracellular free 2-deoxy-[3H]glucose occurred in soleus in both Ex and Ins. Taken together, these data indicate that, in Ex, glucose phosphorylation becomes an important limitation to soleus glucose uptake. In Ins, both glucose delivery and glucose phosphorylation influence the rate of soleus glucose uptake more than under basal conditions.     

A G protein gamma subunit-like domain shared between RGS11 and other RGS proteins specifies binding to Gbeta5 subunits

Regulators of G protein signaling (RGS) proteins act as GTPase-activating proteins (GAPs) toward the alpha subunits of heterotrimeric, signal-transducing G proteins. RGS11 contains a G protein gamma subunit-like (GGL) domain between its Dishevelled/Egl-10/Pleckstrin and RGS domains. GGL domains are also found in RGS6, RGS7, RGS9, and the Caenorhabditis elegans protein EGL-10. Coexpression of RGS11 with different Gbeta subunits reveals specific interaction between RGS11 and Gbeta5. The expression of mRNA for RGS11 and Gbeta5 in human tissues overlaps. The Gbeta5/RGS11 heterodimer acts as a GAP on Galphao, apparently selectively. RGS proteins that contain GGL domains appear to act as GAPs for Galpha proteins and form complexes with specific Gbeta subunits, adding to the combinatorial complexity of G protein-mediated signaling pathways.     

Participation as a complicating mechanism: the effect of clinical professional and middle manager participation on hospital performance

One way hospitals complicate themselves is by increasing the participation of clinical professionals and middle managers in making strategic decisions. Using a survey methodology this article investigates the relationships between the participation of clinical professionals (MDs and RNs) and middle managers with hospital costs, as well as the possible moderating effect of strategic complexity.     

Human wound contraction: collagen organization, fibroblasts, and myofibroblasts

The closure of ungrafted sacrococcygeal pilonidal sinus excisional wounds was studied in 15 patients. Wound punch biopsies were taken on a regular basis, and histologic sections were made. To document changes, computer-assisted morphometric image analysis was employed. Initial average wound depth was 37.8 +/- 4.6 mm, and complete closure (0 wound depth) was reached by 68 days. Wound contraction contributed 88 percent to wound closure, whereas the deposition of scar only contributed 12 percent. Maximum cells density within granulation tissue was reached by day 18. Myofibroblasts, identified by alpha-smooth muscle actin immunostaining, first appeared on day 11. Unlike those observed in laboratory animals, myofibroblasts were a minor cell population of granulation tissue, never exceeding 10 percent of the cells. The pattern of collagen fiber organization was documented by polarized light microscopy of Sirius red-stained sections. Early granulation tissue collagen fibers demonstrated a fine greenish birefringence, whereas more mature granulation tissue collagen fibers were thicker, displaying orange-yellowish birefringence. Myofibroblasts were associated exclusively with thicker collagen fibers, whereas fibroblasts were associated with both fine and thick collagen fibers. It is proposed that human wound contraction involves a volume change whereby normal dermal and adipose tissues are pulled into the defect by forces generated within fibroblasts.     

Production of human normal adult and fetal hemoglobins in Escherichia coli

A hemoglobin expression system in Escherichia coli is described. In order to produce authentic human hemoglobin, we need to co-express both methionine aminopeptidase and globin genes under the control of a strong promoter. We have constructed three plasmids, pHE2, pHE4 and pHE7, for the expression of human normal adult hemoglobin and a plasmid, pHE9, for the expression of human fetal hemoglobin, in high yields. The globin genes can be derived from either synthetic genes or human globin cDNAs. The extra amino-terminal methionine residues of the expressed globins can be removed by the co-expressed methionine aminopeptidase. The heme is inserted correctly into the expressed alpha- globin from our expression plasmids. A fraction (approximately 25%) of the heme is not inserted correctly into the expressed beta- or gamma- globin. However, the incorrectly inserted hemes can be converted into the correct conformation by carrying out a simple oxidation-reduction process on the purified hemoglobin molecule. We have investigated the functional properties of the expressed hemoglobins by measuring their oxygen-binding properties and their structural features by obtaining their 1H-NMR spectra. Our results show that authentic human normal adult and fetal hemoglobins can be produced from our expression plasmids in E. coli and in high yields. Our expression system allows us to design and to produce any recombinant hemoglobins needed for our research on the structure-function relationship in hemoglobin.      

Electrosynthesis in systems of two immiscible liquids and a phase transfer catalyst. V. The anodic chlorination of naphthalene

The anodic chlorination of naphthalene in a water/methylene chloride emulsion and using tetrabutylammonium ion as the phase transfer catalyst is demonstrated; in conditions where the aqueous phase is saturated NaCl, the organic yield of 1-chloronaphthalene is 56% and the current yield is 33% after the passage of 2.33 F mol^–1 of naphthalene. It is shown, however, that when the aqueous phase also contains zinc chloride so that the species transferred is [(C₄H₉)₄N⁺]₂ZnCl ₄ ^–– , the yields can be increased to 92% and 49% respectively. The mechanism of these chlorinations is discussed.     

Autoimmunity, immunodeficiency and mucosal infections: chronic intestinal inflammation as a sensitive indicator of immunoregulatory defects in response to normal luminal microflora

Despite the fact that target antigens and the genetic basis of several autoimmune diseases are now better understood, the initial events leading to a loss of tolerance towards self-components remain unknown. One of the most attractive explanations for autoimmune phenomena involves various infections as possible natural events capable of initiating the process in genetically predisposed individuals. The most accepted explanation of how infection causes autoimmunity is based on the concept of "molecular mimicry" (similarity between the epitopes of an autoantigen and the epitopes in the environmental antigen). Infectious stimuli may also participate in the development of autoimmunity by inducing an increased expression of stress proteins (hsp), chaperones and transplantation antigens, which leads to abnormal processing and presentation of self antigens. Superantigens are considered to be one of the most effective bacterial components to induce inflammatory reactions and to take part in the development and course of autoimmune mechanisms. It has long been known that defects in the host defense mechanism render the individual susceptible to infections caused by certain microorganisms. Impaired exclusion of microbial antigens can lead to chronic immunological activation which can affect the tolerance to self components. Defects in certain components of the immune system are associated with a higher risk of a development of autoimmune disease. The use of animal models for the studies of human diseases with immunological pathogenesis has provided new insights into the influence of immunoregulatory factors and the lymphocyte subsets involved in the development of disease. One of the most striking conclusion arising from work with genetically engineered immunodeficient mouse models is the existence of a high level of redundancy of the components of the immune system. However, when genes encoding molecules involved in T cell immunoregulatory functions are deleted, spontaneous chronic inflammation of the gut mucosa (similar to human inflammatory bowel disease) develops. Surprisingly, when such immunocompromised animals were placed into germfree environment, intestinal inflammation did not develop. Impairment of the mucosal immune response to the normal bacterial flora has been proposed to play a crucial role in the pathogenesis of chronic intestinal inflammation. The use of immunodeficient models colonized with defined microflora for the analysis of immune reactivity will shed light on the mode of action of different immunologically important molecules responsible for the delicate balance between luminal commensals, nonspecific and specific components of the mucosal immune system.     

Comparison of bond strengths of three denture base resins to treated nickel-chromium-beryllium alloy

The success of radiotherapy in eradicating tumours depends on the total radiation dose, but what limits this dose is the tolerance of the normal tissues within the treatment volume. Studies involving fibroblast survival have demonstrated the theoretical feasibility of a predictive assay of radiation sensitivity, but such an assay is still far from clinical application. Using pulsed-field gel electrophoresis (PFGE), we have quantified the initial "apparent" number of DNA double-strand breaks (dsb) induced by the radiation as an alternative measure of sensitivity in 2 different normal cell types from the same patients, epidermal skin cells and lymphocytes. We found significant inter-individual variation in the measured dsb (1-5 dsb/Gy/DNA unit). We also found a linear correlation between molecular damage in lymphocytes and skin samples from the same patient (slope = 0.83; r = 0.694; p = 0.0001). These results suggest that the initial number of dsb could be used as an indicator of the in vivo response to radiation.