Substructure of inner mitochondrial membranes of myocardial cells as shown by cryo-ultramicrotomy

Summary The substructure of the inner mitochondrial membranes has been studied by cryo-ultramicrotomy under conditions during which denaturation of proteins by treatment with chemical solutes has been totally avoided. In such preparations, the inner membrane has a substructure consisting of globular subunits. These subunits have an average diameter of ca. 20Å–ca. 62Å and are fairly regularly spaced. Intracristal space is absent in the unstained, freeze-dried preparations, whereas a space of ca 40Å is seen in preparations lightly treated by OsO₄-vapour. It is concluded that the subunits of the inner mitochondrial membranes probably consist either of single protein molecules or of complexes of protein molecules.This work was supported by grants from The Norwegian Council on Cardiovascular Disease and from The Norwegian Research Council for Science and the Humanities     

On the origin of Z-band material and myofilaments in myoblasts from the human atrial wall

Summary The origin of cardiac myofibrils in cells from the atrial wall in human embryos was studied. Z-band substance appears throughout the cytoplasm as irregular electron dense patches in a network of thin filaments. The thin and thick filaments are synthesized as separate units in the sarcoplasm and are later aggregated into myofibrils. Complexes of Z substance and thin filaments occur numerously at different stages of myofibrillar organisation. Thick filaments are formed in close proximity to free ribosomes and are later incorporated in an hexagonal pattern into the Z-band/thin filament complex.This work was supported by grants from The Norwegian Council on Cardiovascular Disease and from The Norwegian Research Council for Science and the Humanities     

Some molecular properties of NAD(P)H dehydrogenase from rat liver

NAD(P)H dehydrogenase (EC 1.6.99.2) purified from rat liver cytosol revealed three discrete bands, of mol.wts. about 27000, 18000 and 9000, when subjected to polyacrylamidegel electrophoresis in the presence of sodium dodecyl sulphate. Elution of the bands from the gel and individual re-electrophoresis on separate gels showed that the 27000-mol.wt. band yielded three bands similar to those obtained with the intact enzyme, whereas the 18000-mol.wt. band retained its characteristic mobility. Amino acid analysis of native enzyme and protein extracted from each of the three bands from sodium dodecyl sulphate/polyacrylamide gels suggests that the native enzyme is composed of two subunits and that each subunit consists of two dissimilar non-covalently bound polypeptides, so that altogether the enzyme is composed of four polypeptides, two of mol.wt. 18000 and two of mol.wt. 9000. NAD(P)H dehydrogenase was active over a wide pH range with no sharp optimum. The same K(m) value for NADH but different values for V(max.) were obtained for the enzyme purified from Sprague-Dawley and Wistar rats. In immunodiffusion, however, the enzymes from the two rat strains showed a reaction of complete identity. NAD(P)H dehydrogenase was effectively inhibited by thiol-blocking reagents, indicating that the activity is dependent on free thiol group(s). By amino acid analysis six cysteine residues were found per mol of enzyme. Guanidino-group- and amino-group-selective reagents had only moderate inactivating effects on the enzyme activity.     

Ultrastructure of the heart of the sturgeon Acipenser stellatus (Chondrostei)

Summary The ultrastructure of atrial and ventricular myocardial cells from Acipenser stellatus is described. The cells of the atrium are more loosely connected than those of the ventricle. Cell contact is by simple intercalated discs and by desmosomes. The cells are flattened, with peripheral myofibrils and a central region of mitochondria and the nucleus. The sarcoplasmic reticulum consists of subsarcolemmal tubules, that frequently extend towards the central mitochondria. Dyads are small and positioned at any sarcomeric level. No T-tubules are present. Specific granules are restricted to the atrial cell, and are sometimes present within the SR tubules.     

Research ethics training in Peru: a case study

With the rapidly increasing number of health care professionals seeking international research experience, comes an urgent need for enhanced capacity of host country institutional review boards (IRB) to review research proposals and ensure research activities are both ethical and relevant to the host country customs and needs. A successful combination of distance learning, interactive courses and expert course instructors has been applied in Peru since 2004 through collaborations between the U.S. Naval Medical Research Center Detachment, the University of Washington and the Department of Clinical Bioethics of the National Institutes of Health to provide training in ethical conduct of research to IRB members and researchers from Peru and other Latin American countries. All training activities were conducted under the auspices of the Peruvian National Institute of Health (INS), Ministry of Health. To date, 927 people from 12 different Latin American countries have participated in several of these training activities. In this article we describe our training model.     

Magnetic signature of environmental change reflected by Pleistocene lacustrine sediments from the Nihewan Basin, North China

A mineral magnetic study has been conducted on Pleistocene lacustrine sediments from the Xujiayao Paleolithic site within the Nihewan Basin, North China. The magnetic signal is characterized by a dominance of high-coercivity (hematite) minerals that is modulated by low-coercivity magnetite–maghemite assemblages during interglacials. A high-amplitude record of the composition dependent magnetic S_− 0.3 T-parameter varies in accord with composite marine oxygen isotope records, suggesting climate modulation of magnetic composition. The Xujiayao site is situated at the northern outskirts of the Chinese Loess Plateau (CLP) and has neither received significant eolian input nor been exposed to pedogenic processes comparable to those acting on CLP. Mineral magnetic analyses suggest that variations in both magnetite and hematite are primarily controlled by detrital fluxes rather than by authigenic production of magnetic minerals. Here we propose that the fluctuations of magnetic signals at Xujiayao are not derived from varying compositions of eolian flux with glacial/interglacial periods or minute eolian influx during interglacials, but is likely dominated by runoff-related processes, by which some weathered/pedogenized loess and soils with stronger magnetic signals have been eroded and transported to the catchment during interglacials resulting in magnetic enhancement. The inferred continuous deposition from the Brunhes/Matuyama boundary to the paleosol S₁ (ca. 129 ka) gives the age of ca. 500 ka for the sediments carrying the Xujiayao Paleolithic site, suggesting an allogenic origin of the Paleolithic artifacts and mammalian bones dated by U–Th method to ca. 100 ka.     

Disulfide-dependent protein folding is linked to operation of the vitamin K cycle in the endoplasmic reticulum. A protein disulfide isomerase-VKORC1 redox enzyme complex appears to be responsible for vitamin K1 2,3-epoxide reduction

Gamma-carboxylation of vitamin K-dependent proteins is dependent on formation of reduced vitamin K1 (Vit.K1H2) in the endoplasmic reticulum (ER), where it works as an essential cofactor for gamma-carboxylase in post-translational gamma-carboxylation of vitamin K-dependent proteins. Vit.K1H2 is produced by the warfarin-sensitive enzyme vitamin K 2,3-epoxide reductase (VKOR) of the vitamin K cycle that has been shown to harbor a thioredoxin-like CXXC center involved in reduction of vitamin K1 2,3-epoxide (Vit.K>O). However, the cellular system providing electrons to the center is unknown. Here data are presented that demonstrate that reduction is linked to dithiol-dependent oxidative folding of proteins in the ER by protein disulfide isomerase (PDI). Oxidative folding of reduced RNase is shown to trigger reduction of Vit.K>O and gamma-carboxylation of the synthetic gamma-carboxylase peptide substrate FLEEL. In liver microsomes, reduced RNase-triggered gamma-carboxylation is inhibited by the PDI inhibitor bacitracin and also by small interfering RNA silencing of PDI in HEK 293 cells. Immunoprecipitation and two-dimensional SDS-PAGE of microsomal membrane proteins demonstrate the existence of a VKOR enzyme complex where PDI and VKORC1 appear to be tightly associated subunits. We propose that the PDI subunit of the complex provides electrons for reduction of the thioredoxin-like CXXC center in VKORC1. We can conclude that the energy required for gamma-carboxylation of proteins is provided by dithiol-dependent oxidative protein folding in the ER and thus is linked to de novo protein synthesis.