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Biomedical subjects

D F Gerson

Publications and source records attributed to D F Gerson.

At least 19 recordsLinked to original sources

Menstrum for culture preservation and medium for seed preparation in a tetanus toxin production process containing no animal or dairy products.

AIMS: To completely eliminate animal and dairy products from the lyophilization menstrum and the seed medium used to produce tetanus toxin with Clostridium tetani. METHODS AND RESULTS: Tetanus toxin production in a recently developed fermentation medium lacking animal and dairy products was studied with different seed media. It was found that soy peptone could completely replace the beef heart infusion plus animal peptone previously used as seed medium. In addition, we found that cells lyophilized in soy milk could replace the usual type of cells lyophilized in cow's milk. CONCLUSIONS: We have now developed a complete tetanus toxin production process containing no animal and dairy products. SIGNIFICANCE AND IMPACT OF THE STUDY: Toxoid preparations made from toxin produced with animal and dairy products can contain undesirable contaminants such as the prion causing bovine spongiform encephalopathy (Mad Cow's Disease) or antigenic peptides that stimulate anaphylactic reactions and other undesirable immune reactions in immunized hosts. The new vegetable-based process described here avoids such unfortunate possibilities.

Animals↗

Cell surface energy and membrane associated actin in lymphocytes.

We have shown previously that membrane associated actin correlates with the migratory abilities of lymphocytes during recirculation, and that cell surface energy correlates with the adhesiveness of lymphocytes to other cells. In this study, measurements of actin content and cell surface energy have been made for various lymphocyte subpopulations to examine the possibility that recirculation ability may be related to nonspecific adhesiveness. We have found that: both cell surface energy and actin content combine to provide a consistent explanation for the relative rates of recirculation of various lymphocyte subpopulations, and cell surface energies and actin contents vary independently in these lymphocyte subpopulations. Comparison of the actin contents and cell surface energies of metastatic and nonmetastatic lymphoma cell lines indicated that the differences in metastatic potential were more likely attributable to specific receptor-ligand interactions than to nonspecific adhesiveness. Cell surface energy and actin content are consistent with the greater adhesiveness of B cells than T cells to nylon wool, providing a physical basis for this common cell separation technique.

Actins↗

Interference of cyclosporin with lymphocyte activation maintenance of the cells in a non-cycling phase and with a low intracellular pH.

Cyclosporin can block T cell activation by mitogens as evidenced by inhibition of exogenous thymidine integration into DNA and by blockage of the cells in the G0/G1 phase of the cell cycle. The mitogenic activation sequence shows at least one cyclosporin sensitive step at or prior to the rise of cytoplasmic pH (pHi), which normally occurs after exposure to mitogen and which seems to be a permissive condition for initiation of DNA synthesis (S phase of the cell cycle).

Animals↗

Intracellular pH and the cell cycle of mitogen-stimulated murine lymphocytes.

Rapidly proliferating, polyclonally stimulated mouse spleen lymphocytes were separated by density-gradient unit-gravity sedimentation. The following measurements were made on each fraction: the average intracellular water volume, the distribution of DNA content by flow microfluorometry, the rate of 3H-thymidine incorporation, and the intracellular pH. Fractions of cells with a small average intracellular volume were predominantly in G0 or G1 phase of the cell cycle, while fractions of larger cells had higher proportions of cells in S or G2. Multiple regression analysis of the data for both T and B lymphocytes indicated that the intracellular pH of cells in G0, G1, or G2 is around pH 7.2, and that the intracellular pH of cells in S phase of the cell cycle is around pH 7.4.

Cell Cycle↗

Adhesion, phagocytosis and cell surface energy. The binding of fixed human erythrocytes to rat macrophages and polymethylpentene.

Fixed human erythrocytes were used as model particles for the study of adhesion and phagocytosis by rat peritoneal macrophages. Erythrocytes were fixed with various concentrations of glutaraldehyde or tannic acid, or were treated with neuraminidase. Adhesion and phagocytosis of these cells were measured. In addition, the surface energy of these erythrocytes and macrophages was estimated by the contact angle technique. Free energies of adhesion, based on the cell surface energies, were correlated with both adhesion and phagocytosis.

Animals↗

Intracellular pH of mitogen-stimulated lymphocytes.

Mitogenic stimulation of mouse lymphocytes results in two sequential intracellular alkalinizations. The first shift of intracellular pH from 7.18 to 7.35 coincides with early biochemical events following mitogenic stimulation. The second alkalinization begins 12 hours after stimulation and rises in parallel with the rate of thymidine incorporation. The results suggest that intracellular alkalinization following stimulation may play a key role in the enhancement of cellular activation and mitogenesis.

Animals↗

High intracellular pH accompanies mitotic activity in murine lymphocytes.

Intracellular pH is crucial to the control of many cellular processes such as glycolysis, protein synthesis, and cell division. To study the relation between intracellular pH and mitotic activity in actively dividing Con A- or LPS-stimulated splenic lymphocytes, a method was developed to determine intracellular pH using a fluorescence-activated cell sorter. The new method uses the pH-sensitive fluorochrome, 4-methylumbelliferone. Results obtained with it not only correspond qualitatively with the results obtained using 14C-dimethyloxazolidinedione (DMO) but also clearly show the active and inactive subpopulations. The intracellular pH of mitogen-stimulated murine lymphocytes increases from pH 7.15 to pH 7.45 when the population has greatest mitotic activity. The intracellular pH of three virus-transformed lymphocyte cell lines is higher by approximately 0.5 pH units when the cells are in exponential growth compared to stationary phase.

Animals↗

Determination of cell/medium interfacial tensions from contact angles in aqueous polymer systems.

The contact angles on cell layers of a series of polymeric droplets from aqueous two-phase systems of dextran and poly(ethylene glycol) have been used to determine the critical or limiting interfacial tension for spreading on the cell layers. Test droplets of the denser dextran-rich phase were formed in the lighter poly(ethylene glycol)-rich phase. The interfacial tensions, gamma, between the phases were determined with the pendant drop method, and a linear relationship was found between gamma-1/2 and the cosine of the angle the droplets made with the cell layers (Good-Girifalco plot). We were thus able to determine the limiting or critical interfacial tension, gamma c, for spreading on the cell layers. The value of gamma c is a measure of the interfacial energy of the cell/bathing medium interface. Values of gamma c obtained by this method include the following: 0.65 and 0.84 microN . m-1 for human erythrocytes and neutrophils, respectively; 0.93 microN . m-1 for porcine pulmonary macrophages; 0.75--3.60 microM . m-1 for various transformed murine lymphoid cell lines, and 2.53 microN . m-1 for Balb/c murine spleen lymphocytes. Exposure to various agents has differing effects on gamma c. Concanavalin A reduces gamma c, and bacterial lipopolysaccharide increases gamma c of murine spleen lymphocytes. The calcium ionophore, A23187, increases gamma c of both porcine pulmonary macrophages and murine spleen lymphocytes. This new method provides a quantitative approach to the cell surface energy and hydrophobicity which are thought to play an important role in membrane-mediated phenomena and in cell adhesion.

Animals↗

Cell surface energy, contact angles and phase partition. III. Adhesion of bacterial cells to hydrophobic surfaces.

The densities of adhesion of Staphylococcus epidermidis, Staphylococcus aureus and Serratia marcescens to five types of plastics were studied in relation to interfacial free energies at the aqueous interfaces of both the bacteria and the plastics. The free energy of adhesion of bacteria to plastic in an aqueous medium is a linear function of partititon of the bacteria between the solid surface and the liquid phase. These results show that the thermodynamics of the partitioning of a suspended particle between two immiscible liquid phases also apply to partitioning between a liquid and a solid phase.

Bacterial Physiological Phenomena↗

Cell surface energy, contact angles and phase partition. I. Lymphocytic cell lines in biphasic aqueous mixtures.

The surface energy of cells is the quantity which dominates certain physical interactions of cells such as adhesion to hydrophobic surfaces and phagocytosis. A linear relationship is derived relating the equilibrium constant obtained from phase partition in liquid-liquid systems with the surface energy difference obtained from contact-angle measurements. Using biphasic mixtures of Dextran and poly(ethylene glycol) in a medium of constant salt composition the expression is confirmed for transformed lymphocytic cell lines. The results demonstrate the importance of van der Waals' interactions in the phase-partition process, that phase partition can be used as a direct measure of cell surface hydrophobicity, and that the equilibrium constant of phase partition is directly related to the difference in the surface energy of the partitioned particle between the two phases.

Animals↗

Cell surface energy, contact angles and phase partition. II. Bacterial cells in biphasic aqueous mixtures.

Partition coefficients in biphasic mixtures of poly(ethylene glycol) and Dextran are compared to cell surface energies obtained from contact angles of each liquid phase on cell layers. Linear relationships are observed between these two independent measurements for a variety of bacterial cells. The results demonstrate the importance of interfacial phenomena and contact angles in the phase-partition process.

Ascomycota↗

Cell surface measurements in hydrocarbon and carbohydrate fermentations.

Acinetobacter calcoaceticus was grown in 11-liter batch fermentations with hexadecane or sodium citrate as the sole source of carbon. Surface and interfacial tension measurements of the microbial broth indicated that surface-active compounds were being produced only during growth on the hydrocarbon substrate. Contact angle measurements of an aqueous drop on a smooth lawn of cells in a hexadecane bath indicated a highly hydrophobic surface of the cells in the initial stages of the hydrocarbon fermentation (120 degrees contact angle). At this stage, the entire cell population was bound to the hydrocarbon-aqueous interface. The contact angle dropped rapidly to approximately 45 degrees after 14 h into the fermentation. This coincided with a shift of the cell population to the aqueous phase. Thus, the cells demonstrated more hydrophilic characteristics in the later stages of the fermentation. Contact angles on cells grown on sodium citrate ranged from 18 to 24 degrees throughout the fermentation. The cells appear to be highly hydrophilic during growth on a soluble substrate. From the contact angle and aqueous-hydrocarbon interfacial tension, the surface free energy of the cells was calculated along with the cell-aqueous and cell-hydrocarbon interfacial tension. The results of these measurements were useful in quantitatively evaluating the hydrophobic nature of the cell surface during growth on hydrocarbons and comparing it with the hydrophilic nature of the cell surface during growth on a soluble substrate.

Journal Article↗

The effect of substrate on inhibition of Corynebacterium lepus by isonicotinic acid hydrazide (Isoniazid).

Isonicotinic acid hydrazide (INH) inhibits the growth of Corynebacterium lepus on hexadecane but has no effect on its growth on fructose. INH also inhibits the production of the mycolic acid containing lipopeptide bioemulsifier normally produced by C. lepus in response to an insoluble substate. The primary effect of INH appears to be inhibition of mycolic acid synthesis, which limits the growth of C. lepus on hexadecane by reducing the concentration of bioemulsifier.

Alkanes↗

Comparison of surfactant production from kerosene by four species of Corynebacterium.

Four species of Corynebacterium-C. fasciens, C. hydrocarboclastus, C. lepus and C. xerosis, were compared for growth and surfactant production from kerosene. Nitrate as sole nitrogen source resulted in a sharp peak in surfactant concentration during mid-exponential growth phase. Ammonium plus nitrate resulted in growth-associated surfactant production. Surfactant was produced at a concentration of 50-80 times the critical micelle concentration by these species; greatest concentrations were produced by C. lepus. All species were pleomorphic, with greatest cell length occurring in early exponential phase. C. hydrocarboclastus had distinctly longer cells (5 mu) than the other species (3 mu) when grown in shake-flask culture. Ammonium sulfate at concentrations greater than 0.1% (w/v) increased cell length in stationary phase.

Ammonium Sulfate↗

Production of surface-active lipids by Corynebacterium lepus.

Corynebacterium lepus was grown in 20-liter batch fermentations with kerosene as the sole carbon source. Critical micelle concentration measurements indicated the production of appreciable quantities of biosurfactants. This surface activity of the culture medium was due to lipids, which were extracted and identified. Samples of C. lepus whole broth were taken during a fermentation and monitored for surface tension, amount of surfactant present, and lipid content. The changes in the surfactant measured correlated with concentration changes of several surface-active lipids. An early dramatic increase in surfactant concentration was attributed to the production of a mixture of corynomycolic acids (beta-hydroxy alpha-branched fatty acids). Surface activity at the end of the fermentation was due to a lipopeptide containing corynomycolic acids plus small amounts of several phospholipids and neutral lipids which were identified by thin-layer chromatography.

Amino Acids↗

Emulsifying and surface active agents from Corynebacterium hydrocarboclastus.

A Corynebacterium hydrocarboclastus culture isolated in our laboratory (see, Zajic and Knettig, Developments in Industrial Microbiology, 1971, p. 87) has been shown to produce an extracellular biopolymer with emulsifying properties when grown on a mixture of linear hydrocarbons. This microorganism was found to grow well on a variety of carbohydrates and hydrocarbons. However, the best substrates were pure linear hydrocarbons and particularly, n-C12, n-C13, and n-C14. The substrates supporting good growth gave good polymer production. Maximum cell mass of 10-11 g/liter and a maximum amount of polymer of 5-6 g/liter were recorded. The polymers recovered from the different substrates were found to be complex molecules or mixtures with a protein, a lipid, and a carbohydrate moiety. All the polymers are surface active and have two critical micelle concentrations.

Bacterial Proteins↗