Keratinocyte growth factor protects against elastase-induced pulmonary emphysema in mice

Pulmonary emphysema is characterized by persistent inflammation and progressive alveolar destruction. The keratinocyte growth factor (KGF) favorably influences alveolar maintenance and repair and possesses anti-inflammatory properties. We aimed to determine whether exogenous KGF prevented or corrected elastase-induced pulmonary emphysema in vivo. Treatment with 5 mg x kg(-1) x day(-1) KGF before elastase instillation prevented pulmonary emphysema. This effect was associated with 1) a sharp reduction in bronchoalveolar lavage fluid total protein and inflammatory cell recruitment, 2) a reduction in the pulmonary expression of the chemokines CCL2 (or monocyte chemoattractant protein-1) and CXCL2 (or macrophage inflammatory protein-2alpha) and of the adhesion molecules ICAM-1 and VCAM-1, 3) a reduction in matrix metalloproteinase (MMP)-2 and MMP-9 activity at day 3, and 4) a major reduction in DNA damage detected by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) in alveolar cells at day 7. Treatment with KGF after elastase instillation had no effect on elastase-induced emphysema despite the conserved expression of the KGF receptor in the lungs of elastase-instilled animals as determined by immunohistochemistry. In vitro, KGF abolished the elastase-induced increase in CCL2, CXCL2, and ICAM-1 mRNA in the MLE-12 murine alveolar epithelial cell line. We conclude that KGF pretreatment protected against elastase-induced pulmonary inflammation, activation of MMPs, alveolar cell DNA damage, and subsequent emphysema in mice.     

Automatic morphological sieving: comparison between different methods, application to DNA ploidy measurements.

The aim of the present study is to propose alternative automatic methods to time consuming interactive sorting of elements for DNA ploidy measurements. One archival brain tumour and two archival breast carcinoma were studied, corresponding to 7120 elements (3764 nuclei, 3356 debris and aggregates). Three automatic classification methods were tested to eliminate debris and aggregates from DNA ploidy measurements (mathematical morphology (MM), multiparametric analysis (MA) and neural network (NN)). Performances were evaluated by reference to interactive sorting. The results obtained for the three methods concerning the percentage of debris and aggregates automatically removed reach 63, 75 and 85% for MM, MA and NN methods, respectively, with false positive rates of 6, 21 and 25%. Information about DNA ploidy abnormalities were globally preserved after automatic elimination of debris and aggregates by MM and MA methods as opposed to NN method, showing that automatic classification methods can offer alternatives to tedious interactive elimination of debris and aggregates, for DNA ploidy measurements of archival tumours.     

Dielectric breakdowm in the membranes of itValonia utricularis. The role of energy dissipation

The electrical properties of the membranes of Valoniautricularis were investigated using intracellular electrodes. Using short (0.5–1.0 ms) current pulses it was found that at a critical membrane potential difference of 0.85 V there was a large and discontinuous decrease in the membrane impedance and the slope resistance beyond this potential was virtually zero.The electrical breakdown of the membranes did not lead to global damage of the cells and after a resealing time of approx. 5 s could be repeated with identical results.Experiments with long current pulses and long bursts of pulses repeated at 1 kHz are described which show that the electrical breakdown is not due to thermal damage arising from localized heating in the membrane. Thus a dissipation of some 10³–10⁵ times the energy normally dissipated during the onset of breakdown did not lead to breakdown itself unless the critical membrane potential was exceeded.The results also show that punch-through and avalanche ionization are not likely to be important in the breakdown mechanism. The results are consitent, however, with there being a critical instability in the electro-mechanical stresses set up in the membrane at large electric field strengths.     

Electromechanical stresses and the effect of pH on membranes structure

The presence of an electric field in a membrane creates stresses which lead to a mechanical compression of the membrane material.It is shown that considerations of this effect yield results consistent with the observed differential effect of pH on the plasma membrane substructure if the central electron-lucent layer in OsO₄-fixed membranes is identified with the ionic depletion in the double-fixed charge model of cell membranes.     

Location and effect of procaine on lecithin/cholesterol membranes using X-ray diffraction methods

X-ray diffraction studies were made on lecithin/cholesterol multilayers with very high water content and containing the local anaesthetic procaine. Narrow-angle diffraction experiments show that the procaine molecules are located with the uncharged aromatic amine group approx. 10 Å from the centre of the bilayer. The polar tertiary amine group of these molecules is almost certainly located in the polar head-group region of the membrane. Wide-angle diffraction experiments show that the incorporation of procaine molecules into such lipid membranes produces an approx. 30% increase in the spread of acyl chain separation, although the average spacing between the chains is slightly reduced.     

The Double Fixed Charge Membrane: Low Frequency Dielectric Dispersion

An analysis is made of the AC characteristics of a membrane consisting of two fixed charge regions of opposite sign, in contact. It is shown that the equivalent parallel capacitance and conductance of such a membrane undergo a strong dispersion at low frequencies. The dielectric dispersion is a result of polarization effects in the diffusion of coions in each of the two fixed charge lattices. This, at low frequencies, gives rise to a very large diffusion capacitance. The form of the dispersion characteristics is very similar to those observed for synthetic-fused anion-cation membranes and various cellular membranes.     

Cubyl amides: novel P2X7 receptor antagonists

Polycyclic amides 2 and 5-9 were successfully synthesised and their lipophilicity profiles were evaluated using reverse-phase HPLC. All synthesised compounds possessed P2X7R antagonistic properties when tested on rat spinal cord microglia cells. Extensive screening for binding to other neuroreceptor subtypes demonstrated their P2X7 selectivity.     

Combined enzymatic complex I and III deficiency associated with mutations in the nuclear encoded NDUFS4 gene

Combined OXPHOS-system enzyme deficiencies are observed in approximately 25% of all OXPHOS-system disturbances. Of these, combined complex I and III deficiency is relatively scarce. So far, only mtDNA and thymidine phosphorylase (TP) mutations have been associated with combined OXPHOS-system disturbances. In this report we show, for the first time, that a nuclear gene mutation in a structural, nuclear encoded complex I gene is associated with combined complex I and III deficiency. After our initial report we describe mutations in the NDUFS4 gene of complex I in two additional patients. The first mutation is a deletion of G at position 289 or 290. Amino acid 96 changes from a tryptophan to a stop codon. The mutation was found homozygous in the patient; both parents are heterozygous for the mutation. The second mutation is a transition from C to T at cDNA position 316. Codon is changed from CGA (arginine) to TGA (stop). The patient is homozygous for the mutation; both parents are heterozygous. Both mutations in the NDUFS4 gene led to a premature stop in Leigh-like patients with an early lethal phenotype. We hypothesise that the structural integrity of the OXPHOS system, in mammal supermolecular structures, may be responsible for the observed biochemical features.     

Insulin increases cell surface GLUT4 levels by dose dependently discharging GLUT4 into a cell surface recycling pathway

The insulin-responsive glucose transporter GLUT4 plays an essential role in glucose homeostasis. A novel assay was used to study GLUT4 trafficking in 3T3-L1 fibroblasts/preadipocytes and adipocytes. Whereas insulin stimulated GLUT4 translocation to the plasma membrane in both cell types, in nonstimulated fibroblasts GLUT4 readily cycled between endosomes and the plasma membrane, while this was not the case in adipocytes. This efficient retention in basal adipocytes was mediated in part by a C-terminal targeting motif in GLUT4. Insulin caused a sevenfold increase in the amount of GLUT4 molecules present in a trafficking cycle that included the plasma membrane. Strikingly, the magnitude of this increase correlated with the insulin dose, indicating that the insulin-induced appearance of GLUT4 at the plasma membrane cannot be explained solely by a kinetic change in the recycling of a fixed intracellular GLUT4 pool. These data are consistent with a model in which GLUT4 is present in a storage compartment, from where it is released in a graded or quantal manner upon insulin stimulation and in which released GLUT4 continuously cycles between intracellular compartments and the cell surface independently of the nonreleased pool.