High Cholesterol/Low Cholesterol: Effects in Biological Membranes: A Review

Abstract

Lipid composition determines membrane properties, and cholesterol plays a major role in this determination as it regulates membrane fluidity and permeability, as well as induces the formation of coexisting phases and domains in the membrane. Biological membranes display a very diverse lipid composition, the lateral organization of which plays a crucial role in regulating a variety of membrane functions. We hypothesize that, during biological evolution, membranes with a particular cholesterol content were selected to perform certain functions in the cells of eukaryotic organisms. In this review, we discuss the major membrane properties induced by cholesterol, and their relationship to certain membrane functions.

Keywords: Cholesterol; EPR; Lipid spin label; Membrane domains; Oximetry.

Fig. 1

Phase diagram for DMPC/Chol mixtures is reproduced from Reference , Copyright 2013, with permission from American Chemical Society. The shaded area indicates the region where CBDs are detected, forming the structured l_o phase of DMPC membranes. Between 50 and 66 mol% Chol is a structured, one-phase region (dispersed phase region) where CBDs are supported by the DMPC bilayer saturated with Chol. The phase boundary at 66 mol% Chol separates the structured, one-phase region from the two-phase region (structured l_o phase of DMPC and Chol crystals). It should be noted that the phase rule has to be obeyed for all regions and phase boundaries: f = c − p + 1 (f, degree of freedom; c, number of components; p, number of phases, assuming that the pressure is constant for membranes). Schematic drawings of membrane structures (including phases, domains, and crystals) formed at a different Chol content in the DMPC/Chol mixture. l_d, liquid-disordered phase; l_o, liquid-ordered phase; s_o, solid-ordered phase; CBD, Chol bilayer domain; Chol crystals, monohydrated Chol crystals.

Fig. 2

Chemical structures of PL- (n-PC), stearic acid- (n-SASL), and Chol-analog spin labels (ASL and CSL) together with the structure of Chol (CHOL), DMPC, 1-palmitoyl-2-oleoylphosphatidylcholine (POPC), and sphingomyelin (PSM). POPC is a major PL of bioloical membranes and PSM is a major PL of the fiber cell plasma membranes in human eye lenses. Approximate locations of these molecules across the lipid bilayer are illustrated.

Fig. 3

Order parameter (amplitude of the wobbling of the acyl chain segment). (A) Representative EPR spectra from 5- and 14-PC in DMPC membranes in the absence and presence of 50 mol% Chol recorded at 27°C. Measured values for evaluating order parameters are indicated. The order parameter S was calculated according to Marsh [45] using the equation $S=0.5407\left(A_{\Vert }^{\prime }-A_{\perp }^{\prime }\right)/a_0$, where $a_0=\left(A_{\Vert }^{\prime }+A_{\perp }^{\prime }\right)/3$ (B) Profiles of the order parameter obtained with n-PCs at 27°C for DMPC membranes in the absence and presence of 50 mol% Chol. Approximate localizations of the nitroxide moieties of spin labels are indicated by arrows. (C) Order parameter of 5- and 14-PC in DMPC membranes plotted as a function of the mole fraction of Chol for measurements made at 27°C. Curves represent averages of the values obtained from various experiments and show qualitative dependence of the indicated parameter on the Chol bilayer content.

Fig. 4

Membrane fluidity (spin-lattice relaxation rate T₁⁻¹). (A) Representative SR signals with fitted curves from 5- and 14-PC in DMPC membranes in the absence and presence of 50 mol% Chol recorded at 27°C. Signals were recorded for deoxygenated samples (equilibrated with 100% nitrogen). The SR signals can be satisfactorily fitted only with a single exponential function (see details in [49]): for 5-PC in the absence and presence of 50 mol% Chol, with time constants (T₁) of 4.61 ± 0.01 μs and 5.73 ± 0.01 μs, respectively, and for 14-PC in the absence and presence of 50 mol% Chol, with time constants of 3.01 ± 0.01 μs and 2.73 ± 0.01 μs, respectively. (B) Profiles of T₁⁻¹ (the spin-lattice relaxation rate) obtained for n-PC spin labels at 27°C for DMPC membranes in the absence and presence of 50 mol% Chol. Approximate localizations of the nitroxide moieties of spin labels are indicated by arrows. (C) T₁⁻¹ of 5- and14-PC in DMPC membranes plotted as a function of the mole fraction of Chol for measurements made at 27°C. Curves represent averages of the values obtained from various experiments and show qualitative dependence of the indicated parameter on the Chol bilayer content.

Fig. 5

Hydrophobicity (measured as the hyperfine interaction, 2A_Z). (A) Representative EPR spectra of 5- and 14-PC from DMPC membranes in the absence and presence of 50 mol% Chol, recorded at −165°C to cancel motional effects. The measured 2A_Z value is indicated. (For more details, see [59]). (B) Profiles of the hydrophobicity (2A_Z) obtained with n-PCs and 9-SASL for DMPC membranes in the absence and presence of 50 mol% Chol. Approximate localizations of the nitroxide moieties of spin labels are indicated by arrows. (C) 2A_Z of 5- and 14-PC in DMPC membranes plotted as a function of the mole fraction of Chol. Curves represent averages of the values obtained from various experiments and show qualitative dependence of the indicated parameter on the Chol bilayer content. Fig. 5B is reproduced from Reference , Copyright 1994, with permission from American Chemical Society.

Fig. 6

Oxygen diffusion-concentration product (oxygen transport parameter). (A) Representative SR signals with fitted curves from 14-PC in DMPC membranes in the presence of 50 mol% Chol. Signals were recorded at 27°C for samples equilibrated with 100% nitrogen and a gas mixture of 30% air/70% nitrogen. These SR signals can be satisfactorily fitted with a single exponential function: for 14-PC in pure DMPC, with time constants T₁(N₂) of 3.01 ± 0.01 μs and T₁(30% air) 0.79 ± 0.01 μs. The OTP W(x) was calculated according to Kusumi et al. [61] using the equation W(x) = T₁⁻¹(Air,x) − T₁⁻¹(N₂,x) ~ D(x)C(x). Here, x is the “depth” in the membrane. Note that the OTP is defined for samples equilibrated with air, so appropriate extrapolation should be done. Representative SR signals with fitted curves from 5-PC in DMPC membranes in the absence and presence of 50 mol% Chol. Signals were recorded at 27°C for samples equilibrated with a 30% air/70% nitrogen gas mixture. SR signals can be satisfactorily fitted with a single exponential function: for 5-PC in pure DMPC, with a time constant T₁(30% air) of 1.36 ± 0.01 μs and for 5-PC in DMPC containing 50 mol% Chol, with a time constant T₁(30% air) of 4.01 ± 0.01 μs. Appropriate SR signals for 5-PC for deoxygenated samples are presented in Fig. 4A. (B) Profiles of the OTP obtained with n-PCs and 9-SASL for DMPC membranes in the absence and presence of 50 mol% Chol. Approximate localizations of the nitroxide moieties of spin labels are indicated by arrows. (C) OTP of 5- and 14-PC in DMPC membranes plotted as a function of the mole fraction of Chol for measurements made at 27°C indicate the region (between ~10 and ~30 mol% Chol) where l_d and l_o phases coexist. Data obtained with ASL indicate the region (>50 mol% Chol) where the l_o phase contains CBDs. Curves represent averages of the values obtained from various experiments and show qualitative dependence of the indicated parameter on the Chol bilayer content. Broken lines indicate that the amount of the phase or the domain change from zero to the maximal value. Fig. 6B is reproduced from Reference , Copyright 2007, with permission from Elsevier.

Fig. 7

Profiles of different membrane properties across lens lipid membranes of eye lenses from human donors of different age groups obtained using EPR spin labeling and approaches described in Figs. 3–6. All profiles were obtained at 37°C with the PL-analog spin labels. Data obtained with Chol-analog spin labels are included only into the profiles of the OTP. As indicated by profiles of the OTP (data obtained with ASL and CSL), the CBDs are present in membranes of all age groups, indicating that the surrounding PL bilayer is always saturated with Chol. Data for all age groups are reproduced from Reference , Copyright 2017, with permission from Taylor & Francis.