Fig. 2. Effects of C18:1 or C22:1 on VLCFA-induced apoptosis in peroxisome-deficient CHO cells.CHO-zp102 (peroxisome-deficient) cells were cultured in serum-free medium with various concentrations of VLCFAs for 72 h in the presence or absence of C18:1, C22:1, or their combinations (A, B). Cells were harvested by trypsin-EDTA treatment, and subjected to the trypan blue dye exclusion test. The data shown are the percentages of the values of the vehicle-treated control cells. Values represent the mean± S.D. of three independent experiments. Statistical significance was determined using one-way analysis of variance (ANOVA), followed by Tukey’s post-hoc test. *p < 0.05, indicating a significant difference between indicated groups.ER stress involvement in VLCFA toxicity and its alleviation by C18:1We further investigated whether ER stress contributes to VLCFA-induced apoptosis. Treatment with C24:0 or C22:1 significantly upregulated ER stress marker (XBP1s, ATF3, CHOP) at 24 h. Co-treatment with C18:1 completely suppressed these responses at 24 and 48 hours. Electron microscopy revealed that C24:0 caused disruption and collapse of ER membranes, while C18:1 preserved normal ER structure and spacing. These results suggest that C18:1 protects peroxisome-deficient cells by stabilizing ER membranes, thereby mitigating ER stress and preventing VLCFA-induced apoptosis.Mechanism of protection by C18:1Lipidomics revealed that C18:1 promotes redistribution of C24:0 from toxic phospholipids, such as phosphatidylcholine (PC) and lysophosphatidylcholine (LPC), into less harmful lipid species triacylglycerol (TAG) and sphingomyelin (SM). This redistribution was associated with improved cell survival and restored levels of protective C18:1-containing phospholipids such as PC 36:2 and PE 36:2. VLCFA exposure depleted these lipids, leading to membrane instability and endoplasmic reticulum (ER) stress. Supplementation with C18:1 effectively restored these phospholipid levels, alleviating membrane stress and protecting cells from apoptosis. Notably, phospholipids with 18:1/18:1 acyl chains play a key role in stabilizing cellular membranes compromised by VLCFA accumulation. This membrane-stabilizing effect of C18:1 explains its ability to rescue cells without altering the levels of accumulated VLCFAs. A well-ordered and flexible membrane is essential for maintaining proper vesicle trafficking, enzyme function, and overall cellular homeostasis.PerspectivesIn this study, we aimed to develop a medicinal oil for treatment of ALD. Our study demonstrates that C18:1 effectively suppresses the VLCFA-induced ER stress response, restores membrane integrity, and facilitates ― 268 ―
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