Cottonseed extracts and gossypol regulate diacylglycerol acyltransferase gene expression in mouse macrophages

Authors: Cao, Heping; Sethumadhavan, Kandan

Submitted to: Journal of Agricultural and Food Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/28/2018
Publication Date: 6/20/2018
Citation: Cao, H., Sethumadhavan, K. Cottonseed extracts and gossypol regulate diacylglycerol acyltransferase gene expression in mouse macrophages. Journal of Agricultural and Food Chemistry. 66(24):6022-6030. https://doi.org/10.1021/acs.jafc.8b01240
DOI: https://doi.org/10.1021/acs.jafc.8b01240

Interpretive Summary: Plant bioactive polyphenols have been used for the prevention and treatment of various diseases since ancient history. Cotton seeds are classified as glanded or glandless seeds depending on the presence or absence of pigment glands, which contain polyphenolic gossypol. Diacylglycerol acyltransferases (DGATs) are integral membrane proteins for the last step of triacylglycerol biosynthesis in eukaryotes. Understanding the regulation of DGATs will help to create transgenic plants and microbes with value-added properties and provide information for therapeutic intervention for obesity and related diseases. The objective of this study was to investigate the effects of cottonseed extracts and gossypol on the expression of DGAT genes in mouse macrophages. We have discovered that only DGAT2 gene expression is strongly regulated by gossypol and extract from glanded cottonseed kernel, that glandless seed extracts have only minimal effects on DGAT gene expression, and that LPS modestly regulates the expression of both genes in mouse macrophages. The coefficient correlation between DGAT2 mRNA levels in macrophages and the concentrations of glanded kernel extract, gossypol and LPS were 0.87-0.99. Our results suggest that Dgat2 is an inducible gene which may play important role in rapidly responding to stimulators. We conclude that gossypol and ethanol extract from glanded cottonseed kernel are strong stimulators of DGAT2 gene expression and that they may be novel agents for intervention of lipid-related dysfunction via increasing DGAT2 gene expression in target tissues. Our results provide further evidence for glandless seeds as safer food and/or feed source because of lacking gossypol which may cause weight gain due to DGAT activity by gossypol. Our results suggest that gossypol can be potentially used to increase TAGs in plants and animals.

Technical Abstract: Plant bioactive polyphenols have been used for the prevention and treatment of various diseases since ancient history. Cotton (Gossypium hirsutum L.) seeds are classified as glanded or glandless depending on the presence or absence of pigment glands, which contain polyphenolic gossypol. Diacylglycerol acyltransferases (DGATs) are integral membrane proteins that catalyze the last step of triacylglycerol biosynthesis in eukaryotes. Understanding the regulation of DGATs will provide information for therapeutic intervention for obesity and related diseases. However, little is known if DGAT gene expression is regulated by natural products. The objective of this study was to investigate the effects of cottonseed extracts and gossypol on the expression of DGAT genes in mouse macrophages. Mouse macrophages were treated with different concentrations of cottonseed extracts, gossypol, and lipopolysaccharides (LPS) for various times. Quantitative PCR assay showed that DGAT2 mRNA was 4-9 fold higher than DGAT1 mRNA in untreated mouse RAW264.7 macrophages. Coat extract of glanded seeds had modest effects on DGAT1 and DGAT2 mRNA levels. Kernel extract of glanded seeds also had a modest effect on DGAT1 but increased DGAT2 mRNA levels more than 20-fold. Coat extract of glandless seeds had no effect on DGAT1 and a modest effect on DGAT2 mRNA levels. Kernel extract of glandless seeds did not have much effect on DGAT1 and slightly increased DGAT2 mRNA levels. Gossypol increased DGAT1 and DGAT2 mRNA levels by 3-fold and more than 80-fold, respectively. LPS increased DGAT1 and DGAT2 mRNA levels by 2-6 fold. The coefficient correlation (R2) between DGAT2 mRNA levels in macrophages and the concentrations of glanded kernel extract and gossypol were 0.82-0.99. This study suggests Dgat2 is an inducible gene rapidly responding to stimulators such as polyphenols whose protein product DGAT2 play important roles in fat biosynthesis. We conclude that gossypol and ethanol extract from glanded cottonseed kernel are strong stimulators of DGAT2 gene expression and that they may be novel agents for intervention of lipid-related dysfunction via increasing DGAT2 gene expression in target tissues.