THE DIFFERENTIATING ABILITY OF RAT LENS EPITHELIAL CELLS IN CELL CULTURE

Dissociated cells of lens epithelia of adult rats were monolayerly cultured in vitro. After about 15–20 days' period of active cell growth, such characteristic structures that correspond to "lentoid bodies" described previously in chick cultures were formed. These structures consisted of elongated cells, ultrastructural profile of which was similar with lens fiber. The presence of gamma-crystallin, a marker molecule specific to mature lens fiber, was confirmed in these elongated cells by means of fluorescent antibody technique. The differentiation of lens fiber in vitro was also recognized in clones originating from single lens epithelial cells cultured at very low cell density.     

A new in vitro system for studying cell response to mechanical stimulation : Different effects of cyclic stretching and agitation on smooth muscle cell biosynthesis

An experimental model has been devised to permit morphologic and metabolic characterization of cells subjected to a range of cyclic mechanical stimuli similar to those which may prevail in blood vessel walls. A unique feature is the use of purified elastin membranes prepared from bovine aortas as extensible substrates for cell growth. Cells attached firmly to such membranes which could then be subjected to continuous stretching and relaxation or displacement without stretching by a motor coupled to a movable supporting frame. Various combinations of frequencies, amplitudes and rates of deformation have been used over extended periods with minimal fatigue or disruption of the elastin substrate. The effects of cyclic stretching on [¹⁴C]proline incorporation into protein and collagen and [³H]thymidine incorporation into DNA by rabbit aortic smooth muscle cells were distinct from those attributable to agitation without stretching, indicating that cells responded differently to these modes of stimulation. Increases in rate of protein or DNA synthesis induced by stretching were just as marked after 48 h of stimulation as they were at the outset of an experimental period. Since the system permits observations of cell response to independently variable components of pulsatile stress over extended periods and under a variety of culture conditions, it may be expected to provide new data concerning the interaction of mechanical with hormonal and genetic factors in the elaboration of connective tissue components.     

Isolation and characterization of lipopolysaccharides from cell walls of blue-green algae of the genus Phormidium.

Lipopolysaccharide (LPS) fractions were isolated from three species of blue-green algae of the genus Phormidium, namely, P. africanum, P. laminosum, and P. uncinatum, by using a phenol-water procedure followed by exhaustive extraction with ammonium oxalate. The materials obtained were shown to be closely related biochemically. Nearly 60% of the LPS consisted of the polysaccharides galactose, glucose, mannose, xylose, arabinose, and rhamnose and an unidentified, fast-moving sugar residue. In addition, glucosamine, galactosamine, and 2-keto-3-deoxyoctonate were detected. Oleic, stearic, and palmitic acids were found in the hydrolysate of the lipid component, which averaged 1.5% of the LPS. Concomitantly, the protein component (7 to 20%) was shown to contain the following amino acids: aspartic acid, threonine, serine glutamic acid, proline, glycine, alanine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, and arginine. Whole cells, as well as the LPS, of Phormidium possessed antigenic properties.     

Purification and properties of an acid phosphatase of Micrococcus denitrificans distinct from thiamine phosphate phosphatase.

To determine whether the acid phosphatase in Micrococcus denitrificans participates in hydrolysis of thiamine phosphate in the synthesis of thiamine pyrophosphate, acid phosphatase was purified 280-fold by conventional procedures, which removed thiamine phosphate phosphatase completely. Studies showed that this acid phosphatase is a different protein from thiamine phosphate phosphatase and that it has no binding site for thiamine phosphate on its active site.     

Inhibition by Mg2+ of the interaction of Ca2+ with spin-labeled g2 bound to myosin.

According to the measurement of ESR spectrum, Ca2+ induced conformational change of spin-labeled g2 bound to myosin in the presence of 1 mM Mg2+. The half-maximal changes were observed at pCa 6.8 and at pCa 3.7. Spin-labeled phosphorylated g2 bound to myosin showed one transition at pCa 4.5, which shifted to pCa 6.5 after the dephosphorylation with E. coli alkaliphosphatase.     

Microtubular structures in a stable staphylococcal L-form.

Microtubular structures, which were demonstrated as straight, dense-walled cylinders attached to cell membranes, were found in a stable staphylococcal L-form grown in the absence of antibiotic.     

A fluorescent probe response to the interaction of pyocin R1 with sensitive cells.

Additon of pyocin R1, a bacteriocin of Pseudomonas aeruginosa, to sensitive cells caused a fluorescence increase of 8-anilino-1-naphthalenesulfonate (ANS) in the cell suspension. The reaction was rapid, starting with a short time lag after adsorption of pyocin onto the cells and finishing within several minutes. The fluorescence response was attributed to the interaction of the cell body and ANS, not to that of the medium outside the cells and ANS. The maximal amplitude of fluorescence after pyocin addition was dependent on temperature, and the relation appeared to be biphasic. Similarly, Arrhenius plots of the initial rate of fluorescence change were biphasic. The transition of slopes in both cases occurred in the temperature range between 18 and 19 degrees. These results suggest that ANS interacts with lipids in the cell envelope and that pyocin causes a structural change of the cell envelope leading to increased fluorescence of ANS.     

Conformational relaxations of urea- and guanidine hydrochloride-unfolded ferricytochrome c.

Several recent studies of protein the unfolded proteins. In urea- and guanidine HCl-unfolded ferricytochrome c (horse heart), an acid-induced spin state transformation of the heme group has been detected by the heme absorptions, Trp-59 fluorescence, and the intrinsic viscosity of protein. Kinetics of this second conformational transition, by the temperature jump and stopped flow methods, are complex. One rapid reaction (tau1), pH-independent, occurs in a 50-mus range; the second reaction (tau2), in a 1-ms range, depends linearly upon pH and is faster at the alkaline side; a third reaction (tau3), in a 1-s range, shows a sigmoidal transition at pH 5.1 and is faster at the acidic side. The results are consistent with a kinetic scheme which involves protein conformational changes in the transformation of the heme coordination state. The kinetics, along with previous equilibrium studies, indicate that ligand or charge interactions within a protein molecule are not completely prohibited even in strongly denaturing conditions, such as in high concentrations of urea and guanidine HCl. Thus, local structures of peptide chain associated with these interactions can exist in the unfolded protein.