Faculty Bibliography

The regular application of hydrogen peroxide to gingival tissues is becoming widely used as part of dental hygiene. Since hydrogen peroxide can be toxic under certain experimental conditions, proof of both safety and efficacy of its chronic administration should be available before prescription to the general public.

Actin constitutes a major component of the cytoskeleton of human polymorphonuclear leukocytes (PMNs). In this study, we present a comprehensive view of the organization of actin in various PMN regions and functional states. Transmission electron microscopic observations were made on whole mount, migrating, and phagocytizing PMNs. Positive identification of actin filaments was made through S-1 myosin subfragment labeling. In all PMNs studied, actin filaments were primarily organized as a three-dimensional meshwork. The density of this meshwork was greatest within the cell cortex. At peripheral regions of nonpolarized (viz., no distinct head or tail region) and polarized PMNs, actin filaments organized into parallel bundles or overlapping arcs. These bundles or arcs were oriented either perpendicular or parallel to the cell periphery. At the base of the PMN, actin filaments converged upon dense, plaquelike condensations. This latter pattern of actin organization was also observed in some pseudopods at the cell front and in phagocytic processes engulfing bacteria. In areas of internalized bacteria, the surrounding actin appeared as a loose meshwork. Treatment of PMNs with the antiactin drug, cytochalasin B, revealed shearing of the peripheral actin meshwork, condensation of the meshwork around the nuclear region, and dissolution of the basal plaquelike condensations.

Purified muscle actin and mixtures of actin and actin-binding protein were examined in the transmission electron microscope after fixation, critical point drying, and rotary shadowing. The three-dimensional structure of the protein assemblies was analyzed by a computer-assisted graphic analysis applicable to generalized filament networks. This analysis yielded information concerning the frequency of filament intersections, the filament length between these intersections, the angle at which filaments branch at these intersections, and the concentration of filaments within a defined volume. Purified actin at a concentration of 1 mg/ml assembled into a uniform mass of long filaments which overlap at random angles between 0 degrees and 90 degrees. Actin in the presence of macrophage actin-binding protein assembled into short, straight filaments, organized in a perpendicular branching network. The distance between branch points was inversely related to the molar ratio of actin-binding protein to actin. This distance was what would be predicted if actin filaments grew at right angles off of nucleation sites on the two ends of actin-binding protein dimers, and then annealed. The results suggest that actin in combination with actin-binding protein self-assembles to form a three-dimensional network resembling the peripheral cytoskeleton of motile cells.

Native myosin filaments from rabbit psoas muscle are always 1.5 micrometer long. The regulated assembly of these filaments is generally considered to occur by an initial antiparallel and subsequent parallel aggregation of identical myosin subunits. In this schema myosin filament length is controlled by either a self-assembly or a Vernier process. We present evidence which refines these ideas. Namely, that the intact myosin bare zone assemblage nucleates myosin filament assembly. This suggestion is based on the following experimental evidence. (1) A native bare zone assemblage about 0.3 micrometer long can be formed by dialysis of native myosin filaments to either a pH 8 or a 0.2 M-KCl solution. (2) Upon dialysis back to 0.1 M-KCl, bare zone assemblages and distal myosin molecules recombine to form 1.5 micrometer long bipolar filaments. (3) The bare zone assemblage can be separated from the distal myosin molecules by column chromatography in 0.2 M-KCl. Upon dialysis of the fractionated subsets back to 0.1 M-KCl, the bare zone assemblage retains its length of about 0.3 micrometer. However, the distal molecules reassemble to form filaments about 5 micrometers long. (4) Filaments are formed from mixes of the isolated subsets. The lengths of these filaments vary with the amount of distal myosin present. (5) When native filaments, isolated bare zone assemblages or distal myosin molecules are moved sequentially to 0.6 M-KCl and then to 0.1 M-KCl, the final filament lengths are all about 5 micrometers. The capacity of the bare zone assemblage to nucleate filament assembly may be due to the bare zone myosin molecules, the associated M band components or both.