FAS RNA and Enzyme Analysis in PPARAlpha -/-SV-129 Murine Model Skin and Liver as an Indicator of Tumor Proliferation After Varied CLA Isotype Diet
Date of Award
Restricted Access Thesis
College of Theology, Arts, & Sciences
Dr. Rici Hallstrand
Chemoprevention is a pharmacological cancer intervention with the intent to slow or reverse carcinogenesis before invasion and metastasis occurs (Sporn 1991). While the field of dietary cancer research is still in its infancy, there is justified optimism surrounding chemoprevention and its potential to help control and reduce human cancers. Studies have shown reversal of abnormal cell differentiation, (a characteristic of premalignant cancer) blocked preneoplastic lesions, and induced apoptosis in cells that have undergone carcinogenic mutations (Hong 1997).
This project focuses on two isotypes of the fatty acid conjugated linoleic acid (CLA), which is a derivative of linoleic acid (LA), an essential fatty acid that must be supplemented through diet in humans. CLA acts as an exogenous activator of the peroxisome‐proliferator activated receptor (PPARα) nuclear receptor pathway, becoming a nuclear transcription factor upregulating target genes helpful for their anticarcinogenic effects. CLA is thus proposed to be a chemopreventive fatty acid via the PPARα pathway. Through use of in vivo, genetically altered models, the effects of 9‐11 and 10‐12 CLA isotypes can be examined for their specific role as dietary suppressors of early tumor promotion when compared to standard diet and wild type mice after varied diet conditions. Also, this study examines fatty acid synthase (FAS), a known marker of carcinogenesis which is involved in lipid metabolism along with PPARα. Through examining FAS activity in murine liver, and FAS RNA expression in the liver and skin, insight can be gained to better understand the mechanisms by which CLA affects murine models via the PPARα pathway.
This project uses an inbred strain of mice called SV‐129, which are transgenically modified to lack both alleles necessary for PPAR α expression. These mice were homozygous PPARα knockout (KO) mice, and a few wild type (WT), or non genetically altered SV‐129 mice were used as controls. One group of mice were on varied isotypes (9‐11, 10‐12 and mix of the two) of CLA and standard diets for one week. The second group of mice was on a 30 week CLA diet, having been painted with a tumor promoter at 6 weeks old, followed by weekly application of an inflammatory agent to aid in inducing tumor proliferation for this study. The third, and final group of mice, were part of the 30 week tumor study though were removed from their CLA diets one week before sample harvest and put on a regular diet. This was a result of insufficient diet to finish the study, but allowed the opportunity to examine changes in physiology after one week off of the CLA diets. All mice were painted with the pro‐inflammatory compound before harvesting and all mouse groups were harvested within the same time period after painting, allowing examination of FAS as an early indicator of carcinogenesis. FAS is examined through real time FAS mRNA expression in skin and both real time mRNA expression and FAS enzyme activity in the liver. Liver size and tumor incidence were also compared among selected specimens.
The results of this study, while not conclusive, suggest a possible role for the PPARα dependent pathway for the anticarcinogenic effects of CLA 9,11, as well as supporting the hypothesis that FAS can be an early carcinogenic indicator.