The Serial Cultivation of Suspended BHK-21/13 Cells in Serum-Free Waymouth Medium

Guskey, Louis E.; Jenkin, Howard M.

doi:10.3181/00379727-151-39178

Cited by 7 publications

(5 citation statements)

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“…Cells suspended in growth medium were treated with polyene antibiotic and incubated as described above. After incubation for 30 min, 4 h, 8 h, 12 h, 20 h, and 24 h post-treatment, duplicate samples were removed from each cell suspension. Intracellular potassium was determined after washing the cells twice with K+-free maintenance medium (as described above).…”

Section: Methodsmentioning

confidence: 99%

Dissociation between the induction of potassium efflux and cytostatic activity of polyene macrolides in mammalian cells

Malewicz¹,

Jenkin²,

Borowski³

1980

Antimicrob Agents Chemother

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The paper contains data on the induction of K+ efflux and viability of baby hamster kidney (BHK-21) cells after their treatment with macrolide antibiotics inducing specific pores in membrane. New water-soluble semisynthetic derivatives of amphotericin B and aureofacin (N-glycosyl and trimethylammonium methyl ester derivatives) as well as the parent compounds were used to compare the concentration of antibiotics inducing permeabilizing and cytostatic effects. We found that a two-to eight-times-higher concentration of polyene antibiotic was required to observe a cytostatic effect than for release of 50% of the cellular potassium (Kr5o concentration) from BHK-21 cells. These differences were larger for water-soluble derivatives than for the parent compounds. The amount of intracellular potassium in treated cells incubated under optimal growth conditions was higher than that in cells which had been further washed with K+-free maintenance medium. The membrane permeability changes induced by low concentrations of specific polyenes were observed to be reversible. BHK-21 cells were able to repair polyene-induced membrane permeability within 3 to 12 h under optimal growth conditions, after cell treatment with K5w concentration of specific macrolide antibiotics. The repair phenomenon is postulated as an explanation for the dissociation observed between permeabilizing and cytostatic effect of specific polyenes in BHK-21 cells.

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Section: Methodsmentioning

confidence: 99%

Dissociation between the induction of potassium efflux and cytostatic activity of polyene macrolides in mammalian cells

Malewicz¹,

Jenkin²,

Borowski³

1980

Antimicrob Agents Chemother

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“…Our results extended the use of L cells grown in a serum-free medium in a spinner culture (17) to their successful use in a shaker culture (10). With this culture system we were able to obtain equal or higher titers of C. trachomatis LGV-404L than those observed with other methods of cultivation, as well as organisms free of serum contamination and adventitious agents from serum.…”

Section: Discussionmentioning

confidence: 56%

“…Cx-treated L cells grown in modified Waymouth 752/1 (WO5) medium (17) without serum or undefined exogenous lipids provides an ideal system for studying chlamydial lipid synthesis. The shaker culture system provides a means of obtaining high yields of cells and chlamydiae with a minimal chance of contamination (10).…”

mentioning

confidence: 99%

Use of cycloheximide to study independent lipid metabolism of Chlamydia trachomatis cultivated in mouse L cells grown in serum-free medium

Reed¹,

Anderson²,

Jenkin³

1981

Infect Immun

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A system for measuring chlamydial lipid synthesis was developed with mouse L cells grown in serum-free modified Waymouth 752/l medium in a shaker culture. Host lipid synthesis was reduced approximately 90% when cells were incubated for 24 h in medium containing cycloheximide (2 micrograms/ml). Lipid metabolism was monitored by measuring the incorporation of [3H]isoleucine into the total lipid of normal and infected cells. The results suggested that lipid synthesis of Chlamydia trachomatis lymphogranuloma venereum (LGV-404L) was not inhibited by cycloheximide treatment when the chlamydiae were grown in L cells, whereas host lipid synthesis was inhibited. Chlamydial lipid metabolism began about 6 to 12 h after infection when the noninfectious reticulate body was found and continually increased until the beginning of the appearance of intracellular infectious elementary bodies at 24 to 30 h.

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“…Additional components, in mg/liter, included: galactose, 1,890; ammonium sulfate, 250; biotin, 0.1; vitamin B12, 1.25; a-lipoic acid, 1.0; and folinic acid, 1.0. Fatty acids were incorporated into the medium as the sodium salts completed with BSA (12). Palmitic (16:0), stearic (18:0), oleic (cis 9-18:1), and elaidic (trans 9-18:1) acids were sterilized in tightly capped tubes at 121'C for 20 min and then converted to the sodium salts by slowly heating over a flame with a 20% molar excess of sterile 0.1 N NaOH.…”

mentioning

confidence: 99%

“…A second biological system was also used to assess the toxicity of the two oleate preparations. In serum-free cultures of BHK 21 cells routinely cultivated with oleate as fatty acid source (12), typical cell growth occurred with freshly prepared oleate whereas inhibition of growth and Voi.. 19, 1978 A, palmitic plus stearic (20 ,ug/ml each); U, oleic plus linoleic (20 pg/ml each); 0, combination ofpalmitic (22 pig/ml), stearic (10 pg/ml), oleic (17.4 ptg/ml), and linoleic (14.3 pg/ml). The fatty acid concentrations in the combination offour were based on analysis of the fatty acid composition of rabbit testes infected with T. pallidum (unpublished data).…”

mentioning

confidence: 99%

Effects of fatty acids on motility retention by Treponema pallidum in vitro

Matthews¹,

Jenkin²,

Crilly³

et al. 1978

Infect Immun

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Treponema pallidum (Nichols virulent strain) was incubated under 75% N2 + 20% H2 + 5% CO2 in prereduced serum-free modified Eagle-Richter medium supplemented with different concentrations of various long-chain fatty acids complexed with fatty acid-free bovine serum albumin. Motility retention was greater in medium with oleic acid containing 15 rather than 2 mg of albumin per ml. Palmitic, stearic, oleic, or linoleic acid alone caused rapid loss of motility at concentrations as low as 5 microgram/ml. Elaidic acid (92 microgram/ml) alone had no effect on motility. Various combinations of saturated plus unsaturated fatty acids did not inhibit motility retention or were less inhibitory than either of the individual fatty acid components. The combination of palmitic plus oleic acids was least toxic. Rapid loss of motility occurred with pairs of unsaturated or saturated fatty acids, or with Tween 40, 60, or 80, alone or combined. Autoxidation of oleic acid resulted in decreased toxicity for T. pallidum but increased toxicity for baby hamster kidney cells.

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The Serial Cultivation of Suspended BHK-21/13 Cells in Serum-Free Waymouth Medium

Cited by 7 publications

References 0 publications

Dissociation between the induction of potassium efflux and cytostatic activity of polyene macrolides in mammalian cells

Dissociation between the induction of potassium efflux and cytostatic activity of polyene macrolides in mammalian cells

Use of cycloheximide to study independent lipid metabolism of Chlamydia trachomatis cultivated in mouse L cells grown in serum-free medium

Effects of fatty acids on motility retention by Treponema pallidum in vitro

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