Nutrient restriction, also known as caloric restriction, has been extensively examined for its positive impact on lifespan, immune system boost, and aging. In addition, nutrient restriction is implicated in decreasing cancer initiation and progression. Given the phenotypic changes associated with nutrient restriction, we hypothesized significant protein expression alterations must be associated with caloric restriction. To compare the molecular and phenotypic changes caused by glucose restriction and fetal bovine serum restriction there is need for an efficient model system. We establish 3-dimensional cell culture models, known as spheroids, in the HCT 116 colorectal cancer cell line as a high throughput model for studying the proteomic changes associated with nutrient restriction. Flow cytometry was used to assess apoptosis and autophagy levels in the spheroids under nutrient restriction. Isobaric tags for relative and absolute quantification and liquid chromatography tandem mass spectrometry were used to determine differential protein abundances between the nutrient restriction conditions. We identified specific proteins that have implications in cancer progression and metastasis that are differentially regulated by restriction of either glucose or serum. These proteins include the up-regulation of sirtuin 1 and protein inhibitor of activated STAT 1 and down-regulation of multi-drug resistance protein and Zinc finger and BTB domain-containing protein 7A. The results indicate nutrient restriction causes lower apoptotic and higher autophagy rates in HCT 116 spheroids. In addition, proteins shown to be differentially regulated by both glucose and serum restriction were similarly regulated.
Keywords: Apoptosis; Autophagy; Colorectal cancer; Nutrient restriction; Quantitative proteomics; Three dimensional cell culture.
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