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• W4313461522 abstract "Abstract Background: Increased rates of cholesterol synthesis have been recognized as an important aspect of the metabolism of transformed cells. However, precisely how cholesterol dysmetabolism affects membrane homeostasis is not yet fully understood. Since free cholesterol can be transported from the plasma membrane to other organelles in a dynamic and bidirectional fashion, there is an urgent need to determine to what extent endomembranes are affected by driver oncogene-mediated distortions in cholesterol homeostasis. Aim: Determine if cholesterol is increased in endomembranes, relative to the plasma membrane in Apc mutant cell lines and primary Apc null mouse colonocytes amd CRC samples. Methods: Three mouse epithelial isogenic wild type and mutant Apc colonic cell lines (YAMC (Apc +/+), IMCE (Apc +/-), IMCE βcat (Apc +/- + ΔN89 βcat) were used. In addition, primary colonocytes from Apc wildtype and null mice were examined. mRNA was sequenced using a TruSeq Illumina Stranded mRNA kit. Data analysis was conducted as follows: EdgeR was used to identify differentially expressed genes. Human CRC RNAseq data was obtained from a public reservoir and analyzed. A gene target list (~ 290 genes) related to organelle cholesterol transport was subsequently queried. Membranes from the endoplasmic reticulum (ER), mitochondria and lysosomes were assessed using fluorescent probes: ERTracker, MitoTracker and LysoTracker, respectively. Cells were fixed and stained with Filipin III and imaged using confocal microscopy. Fluorescence colocalization was calculated and statistical analysis was performed using a one-way ANOVA. Results: Apc mutant colonocyte cell lines and Apc null mouse models and human CRC samples exhibited differential expression of cholesterol trafficking genes including Acaa1b, Acaa2, Pcsk9, Stard5. Genes related to mitochondrial cholesterol metabolism, transport, and accumulation, e.g., Tspo, Ppargc1a and Cyp27a, were also found differentially expressed in the three models. Imaging experiments revealed that mitochondrial cholesterol was 17% higher in IMCE and IMCE βcat cell lines. Lysosomal cholesterol was 21% higher in IMCE cells. ER cholesterol levels were unaffected. Conclusions: Our preliminary findings indicate dysregulation of mitochondrial and lysosomal unesterified cholesterol levels in colonocytes expressing mutant APC. Citation Format: Monica Munoz-Vega, Alfredo Erazo-Oliveras, Michael L. Salinas, Xiaoli Wang, Jennifer S. S Goldsby, Robert S. Chapkin. Endomembranes accumulate unesterified cholesterol in Apc mutant models and humans with CRC as assessed by RNAseq and confocal microscopy. [abstract]. In: Proceedings of the AACR Special Conference: Precision Prevention, Early Detection, and Interception of Cancer; 2022 Nov 17-19; Austin, TX. Philadelphia (PA): AACR; Can Prev Res 2023;16(1 Suppl): Abstract nr P016." @default.
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• W4313461522 date "2023-01-01" @default.
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• W4313461522 title "Abstract P016: Endomembranes accumulate unesterified cholesterol in Apc mutant models and humans with CRC as assessed by RNAseq and confocal microscopy" @default.
• W4313461522 doi "https://doi.org/10.1158/1940-6215.precprev22-p016" @default.
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