Effect of Host Cholesterol on the Membrane Dynamics of Outer Membrane Lipids of Mycobacteria.

The ability of Mycobacterium tuberculosis to remain dormant after primary infection represents the prime cause of new TB cases throughout the world. Hence, diagnosis and treatment of individuals hosting dormant mycobacterium is one of the crucial strategies to be adopted for the prevention of Tuberculosis. Among many strategies unleashed by the latent bacterium, one of them is scavenging host cholesterol for carbon source. Cholesterol modifies lipid membranes over many scales and here, its effect on mycobacterial membrane biophysics and the subsequent effect on partitioning of antibiotics into cholesterol- enriched mycobacterial membranes was investigated. Our research showed that cholesterol alters the phase state behavior of mycobacterial outer membrane lipids by enhancing the overall membrane order at the headgroup and acyl chain region and is integrated into both ordered and disordered domains/phases, with a preference for the latter. Exogenous cholesterol further alters the drug partitioning behavior of structurally different drugs, pointing to a larger clinical potential of using more hydrophobic medications to target dormant bacteria.

Aspirin locally disrupts the liquid-ordered phase.

Local structure and dynamics of lipid membranes play an important role in membrane function. The diffusion of small molecules, the curvature of lipids around a protein and the existence of cholesterol-rich lipid domains (rafts) are examples for the membrane to serve as a functional interface. The collective fluctuations of lipid tails, in particular, are relevant for diffusion of membrane constituents and small molecules in and across membranes, and for structure and formation of membrane domains. We studied the effect of aspirin (acetylsalicylic acid, ASA) on local structure and dynamics of membranes composed of dimyristoylphosphocholine (DMPC) and cholesterol. Aspirin is a common analgesic, but is also used in the treatment of cholesterol. Using coherent inelastic neutron scattering experiments and molecular dynamics (MD) simulations, we present evidence that ASA binds to liquid-ordered, raft-like domains and disturbs domain organization and dampens collective fluctuations. By hydrogen-bonding to lipid molecules, ASA forms 'superfluid' complexes with lipid molecules that can organize laterally in superlattices and suppress cholesterol's ordering effect.

Chlorhexidine: beta-cyclodextrin inhibits yeast growth by extraction of ergosterol

Chlorhexidine (Cx) augmented with beta-cyclodextrin (β-cd) inclusion compounds, termed Cx:β-cd complexes, have been developed for use as antiseptic agents. The aim of this study was to examine the interactions of Cx:β-cd complexes, prepared at different molecular ratios, with sterol and yeast membranes. The Minimal Inhibitory Concentration (MIC) against the yeast Candida albicans (C.a.) was determined for each complex; the MICs were found to range from 0.5 to 2 µg/mL. To confirm the MIC data, quantitative analysis of viable cells was performed using trypan blue staining. Mechanistic characterization of the interactions that the Cx:β-cd complexes have with the yeast membrane and assessment of membrane morphology following exposure to Cx:β-cd complexes were performed using Sterol Quantification Method analysis (SQM) and scanning electron microscopy (SEM). SQM revealed that sterol extraction increased with increasing β-cd concentrations (1.71 × 10³; 1.4 × 10³; 3.45 × 10³, and 3.74 × 10³ CFU for 1:1, 1:2, 1:3, and 1:4, respectively), likely as a consequence of membrane ergosterol solubilization. SEM images demonstrated that cell membrane damage is a visible and significant mechanism that contributes to the antimicrobial effects of Cx:β-cd complexes. Cell disorganization increased significantly as the proportion of β-cyclodextrin present in the complex increased. Morphology of cells exposed to complexes with 1:3 and 1:4 molar ratios of Cx:β-cd were observed to have large aggregates mixed with yeast remains, representing more membrane disruption than that observed in cells treated with Cx alone. In conclusion, nanoaggregates of Cx:β-cd complexes block yeast growth via ergosterol extraction, permeabilizing the membrane by creating cluster-like structures within the cell membrane, possibly due to high amounts of hydrogen bonding.

Chlorhexidine: beta-cyclodextrin inhibits yeast growth by extraction of ergosterol.

Chlorhexidine (Cx) augmented with beta-cyclodextrin (ß-cd) inclusion compounds, termed Cx:ß-cd complexes, have been developed for use as antiseptic agents. The aim of this study was to examine the interactions of Cx:ß-cd complexes, prepared at different molecular ratios, with sterol and yeast membranes. The Minimal Inhibitory Concentration (MIC) against the yeast Candida albicans (C.a.) was determined for each complex; the MICs were found to range from 0.5 to 2 µg/mL. To confirm the MIC data, quantitative analysis of viable cells was performed using trypan blue staining. Mechanistic characterization of the interactions that the Cx:ß-cd complexes have with the yeast membrane and assessment of membrane morphology following exposure to Cx:ß-cd complexes were performed using Sterol Quantification Method analysis (SQM) and scanning electron microscopy (SEM). SQM revealed that sterol extraction increased with increasing ß-cd concentrations (1.71 ×10(3); 1.4 ×10(3); 3.45 ×10(3), and 3.74 ×10(3) CFU for 1:1, 1:2, 1:3, and 1:4, respectively), likely as a consequence of membrane ergosterol solubilization. SEM images demonstrated that cell membrane damage is a visible and significant mechanism that contributes to the antimicrobial effects of Cx:ß-cd complexes. Cell disorganization increased significantly as the proportion of ß-cyclodextrin present in the complex increased. Morphology of cells exposed to complexes with 1:3 and 1:4 molar ratios of Cx:ß-cd were observed to have large aggregates mixed with yeast remains, representing more membrane disruption than that observed in cells treated with Cx alone. In conclusion, nanoaggregates of Cx:ß-cd complexes block yeast growth via ergosterol extraction, permeabilizing the membrane by creating cluster-like structures within the cell membrane, possibly due to high amounts of hydrogen bonding.