Sea cucumber plasmalogen enhance lipophagy to alleviate abnormal lipid accumulation induced by high-fat diet.

Image of an orange-footed sea cucumber (Cucumaria frondosa).

Wang Z, Liu Y, Wang X, Wang X, Wu Y, Song Y, Xu J, and Xue C. (2024) Sea cucumber plasmalogen enhance lipophagy to alleviate abnormal lipid accumulation induced by high-fat diet. Molecular and Cell Biology of Lipids.

Autophagy is a process used by cells to prevent cell damage and maintain metabolic homeostasis by lysosomes degrading proteins and damaged organelles. This process can occur in stress-induced conditions, but also in basal conditions where it is a defense mechanism to promote cell growth and protect against metabolic and oxidative stress. There are many types of autophagy which are classified based on the specificity of what is degraded. Lipophagy, is the selective degradation of lipid droplets and if this mechanism is not working efficiently in the cell, lipid accumulation can occur. Plasmalogens, a unique class of phospholipid that contain a vinyl-ether bond at the sn-1 position have been associated with the improvement of several chronic diseases related to metabolic remodeling of lipids. Because of this, Wang et al wanted to compare treatment with plasmalogens or plasmanylcholine isolated from sea cucumbers, Cucumaria frondosa, for lipophagy-activating efficacy.

To determine how lipid metabolism is improved by plasmalogens through lipophagy, HepG2 cells (a human liver cancer cell line which is commonly used for drug metabolism and hepatoxicity studies) were treated with palmitic acid (PA), often used to produce a high-fat diet (HFD) model. The cells treated with PA had high levels of triglycerides and total cholesterol, similar to what is seen in mice fed a high-fat diet. Oil Red O kit was utilized to stain lipid droplets to compare lipid accumulation in the cells after the PA treatment and with either plasmalogens or plasmanylcholine treatment. It was demonstrated that plasmalogens significantly reduced the accumulation of the lipids and phospholipids in cells, indicating an ability to inhibit lipid accumulation compared to the cells only treated with PA or to those treated with PA and plasmanylcholine.

To measure the changes in lipophagy, HepG2 cells were transfected so that they would express mRFP-GFP-LC3 protein. RFP (red fluorescent protein) and GFP (green fluorescent protein) are fluorescent tags, and LC3 is a protein in the autophagy pathway. During the autophagosome stage, LC3 will express the two fluorescences at the same time, but after autophagosome and lysosome fusion, the acidic lysosome quenches the green GFP and only allows LC3 to express the red fluorescence of RFP. Because of this, if both fluorescence colours can be detected, this would indicate a change in lipophagy. To further understand this change, clearing an autophagy cargo adaptor, p62, corresponds to completing current autophagy function and therefore levels of p62 were measured. It was found that p62 increased after PA treatment which suggested that PA resulted in lipophagy disorder. Interestingly, treatment with plasmalogens reduced the abnormal p62 accumulation. This is also seen when detecting fluorescence where both GFP and RFP are expressed the highest in the cells treated with plasmalogens. Specifically, when the fluorescence was merged, the yellow spots (where both fluorescences were expressed at the same time) increased and these represent the autophagosome stage, which suggests that PA induced lipophagy activation and promotion of autophagosome formation.

Wang et al were interested in determining the effect of plasmalogens on activating lipophagy and wanted to investigate if the double bond in plasmalogens was necessary for this effect or whether any ether lipid, such as plasmanylcholine, could also elicit the same effect. HepG2 cells that express mRFG-GFP-LC3 protein were induced with PA to create a HFD cell model then were treated with either plasmalogens or plasmanylcholine. They found that plasmalogens were able to significantly reduce the accumulation of lipids, however this was not seen in cells treated with plasmanylcholine. In addition, when studying the fluorescent tags attached to LC3, they found an increase in yellow, signifying the autophagosome stage, which supported that PA induced lipophagy activation and there was a decrease in green fluorescence which represented weak autophagosome and lysosome fusion. However, plasmalogen treatment restored expression of green fluorescence, indicating stronger autophagosome and lysosome fusion, and the treatment caused a decrease in abnormal p62 accumulation, which suggests it can relieve abnormal lipid accumulation. Both plasmalogens and plasmanylcholine, the latter of which does not have the vinyl bond at sn-1, were evaluated and the authors suggest that the double bond was necessary for these effects to be seen since PlsEtn and PakCho have EPA at sn-2 so a different side chain cannot explain these differences. This could indicate that plasmalogens are a lipophagy enhancer and could explain their role in many lipid metabolism disorders. This group has also done work in mice and that will be discussed in the next blog.