Transfer of amphotericin B from gel state vesicles to mycoplasma cells: biphasic action on potassium transport and permeability

Abstract

The action of amphotericin B on the K+ permeability of Mycoplasma mycoides var. capr cells, containing either cholesterol or ergosterol in their membranes, was studied. When the drug, solubilized in dimethyl sulfoxide, was added directly to the cell suspension, a slightly greater sensitivity to permeabilization was observed for ergosterol-containing cells, confirming the data reported in the literature. When amphotericin B bound to gel state phospholipid vesicles was added to the cell suspension, two effects on cholesterol-containing cells were observed. First, the K+ active transport rates increased; membrane permeabilization and K+ leakage were subsequently detected. For ergosterol containing cells these sequential events were observed only at amphotericin B concentrations below 10-6 M. At higher concentrations only K+ leakage was observed. The second permeabilization effect varied with the amphotericin B concentration in different ways in the two types of cells. The permeabilization of ergosterol-containing membranes depended on the amphotericin B/phospholipid molar ratio, whereas the permeabilization of cholesterol-containing membranes did not. In general, the latter remained fairly constant when the total amphotericin B concentration in the medium varied.The antibiotic activity of amphotericin B (AmB) is related to the channels this drug forms in cell membranes (for a review, see reference 11). AmB and related compounds are used in antifungal therapy, since ergosterol-containing membranes are more sensitive to their action than cholesterolcontaining ohes (for a review, see reference 15). Although numerous studies have been carried out, leading to the development of elaborate models (9, 17), little is known about the complex process which occurs during the interaction of AmB with membranes and which leads to pore formation.The poor water solubility of AmB, which results in its aggregation in aqueous medium, has been a major impediment in studying this process. The interaction of these aggregates with membranes is difficult to analyze in terms of the successive molecular events involved and thus has been dealt with as an all or nothing process.This impediment can be overcome by making use of the ability of AmB to exchange between membranes: instead of introducing into the cell suspension AmB as a dimethyl formamide or dimethyl sulfoxide (DMSO) solution, which results in its aggregation, AmB is first incorporated into lipid vesicles which are then introduced into the membrane suspension. By this method, which completely eliminates the presence of AmB in its aggregated form, it has been shown that the affinity of AmB for membranes is a function of the physical state of the membrane (6). More recently, a study of the interaction of AmB with cholesterol and ergosterol-containing phospholipid vesicles revealed that the selectivity of AmB for the latter was much greater than the selectivity currently admitted on the basis of the results obtained by direct application to these membranes of the antibiotic dissolved i...

Carrier effects on biological activity of amphotericin B

“…VOL. 9,1996 CARRIER EFFECTS ON AMPHOTERICIN B EFFICACY 517 A similar difference in selectivity was observed with ergosterolor cholesterol-containing Mycoplasma cells (191). Since the affinity of AmB to DPPC vesicles is greater than the affinity to EPC vesicles (106), these data support a role for stability of the complex in AmB sterol selectivity.…”

“…Therefore, with the discovery that AmB incorporated into liposome or lipid complexes was less toxic to animals than was Fungizone but at the same time was equally therapeutic (134), experiments were designed to compare various toxic and antifungal activities of AmB alone and ABLC in vitro. Studies on carrier effects were extended to detergents other than deoxycholate (82,165,179) and to various lipids added to AmB-cell suspensions, e.g., lipoproteins (22), triglycerides (172), preformed phospholipid vesicles (190,191), preformed mixed micelles of egg lecithin and glycocholate (32), and lecithin-stabilized triglyceride emulsions (117). The techniques used for measuring damage in these in vitro assays can be grouped into short-and long-term assays (101).…”

Amphotericin B and its new formulations: pharmacologic characteristics, clinical efficacy, and tolerability

“…AmB has strong affinity for ergosterol, the principal sterol of fungal membranes, while exhibiting less affinity for cholesterol, the principal sterol of mammalian cell membranes ( 1). AmB and ergosterol form micelles that combine to form transmembrane channels that modify the permeability of the fungal membrane, with leakage of ions and other cellular components ( 2, 3). Other mechanisms of action include proton pump inhibition in Candida albicans , and induction of membrane lipid peroxidation ( 4, 5).…”

Recommendations for the treatment of fungal pneumonias

“…Polyenes act by absorbing the ergosterol of the fungal membrane and disinfecting it by increasing the permeability of the membrane, allowing the ions to leak out of the cytoplasm [99,100]. On the contrary, amphotericin B exhibits a lower affinity for cholesterol, the principal sterol of mammalian cell membrane [101].…”