DHA-fluorescent probe is sensitive to membrane order and reveals molecular adaptation of DHA in ordered lipid microdomains

Abstract

Docosahexaenoic acid (DHA) disrupts the size and order of plasma membrane lipid microdomains in vitro and in vivo. However, it is unknown how the highly disordered structure of DHA mechanistically adapts to increase the order of tightly packed lipid microdomains. Therefore, we studied a novel DHA-Bodipy fluorescent probe to address this issue. We first determined if the DHA-Bodipy probe localized to the plasma membrane of primary B and immortal EL4 cells. Image analysis revealed that DHA-Bodipy localized into the plasma membrane of primary B cells more efficiently than EL4 cells. We then determined if the probe detected changes in plasma membrane order. Quantitative analysis of time-lapse movies established that DHA-Bodipy was sensitive to membrane molecular order. This allowed us to investigate how DHA-Bodipy physically adapted to ordered lipid microdomains. To accomplish this, we employed steady-state and time-resolved fluorescence anisotropy measurements in lipid vesicles of varying composition. Similar to cell culture studies, the probe was highly sensitive to membrane order in lipid vesicles. Moreover, these experiments revealed, relative to controls, that upon incorporation into highly ordered microdomains, DHA-Bodipy underwent an increase in its fluorescence lifetime and molecular order. In addition, the probe displayed a significant reduction in its rotational diffusion compared to controls. Altogether, DHA-Bodipy was highly sensitive to membrane order and revealed for the first time that DHA, despite its flexibility, could become ordered with less rotational motion inside ordered lipid microdomains. Mechanistically, this explains how DHA acyl chains can increase order upon formation of lipid microdomains in vivo.

Eicosapentaenoic and docosahexaenoic acid ethyl esters differentially enhance B-cell activity in murine obesity

“…The discrepancy between our fi ndings and the two aforementioned studies may be due to differences between the composition and the levels of the fat source ( 43,44 ). This study relied on a moderate dose of milkfat (45% of total kilocalories), modeling human suggest that when these fatty acids interact with ordered domains, they increase their molecular order and can adapt to the ordered environment ( 58,59 ). This may be driven by several variables, which include the loss of AA, a highly disordered fatty acid, and a displacement of cholesterol between disordered and ordered domains ( 58,60 ).…”

Docosahexaenoic acid promotes micron scale liquid-ordered domains. A comparison study of docosahexaenoic versus oleic acid containing phosphatidylcholine in raft-like mixtures

“…These differences have been attributed to the saturated nature of sphingolipid acyl chains that packs well with the rigid cholesterol backbone and separate from DHA containing phospholipids. DHA chains are disordered and enhance the segregation of cholesterol into SM-rich/sterolrich rafts, favoring DHA-rich domains via cholesterol exclusion but DHA chains can also partition into rafts [55].…”

The targeting mechanism of DHA ligand and its conjugate with Gemcitabine for the enhanced tumor therapy

“…Teague et al reported that a DHA-fluorescent probe was highly uptaken by EL4 cells at 37°C than at 23°C, which could be attributed to the high fluidity of the cell membrane at 37°C, while it is more ordered at 23°C [28]. Robinson et al found that DHA was initially incorporated into phosphatidylethanolamine (PE) accompanied by a less amounts into phosphatidylcholine (PC) or other phospholipid classes [29-30].…”