Effects of dietary methionine restriction on age-related changes in perivascular and beiging adipose tissues in the mouse.

OBJECTIVE: Perivascular adipose tissue (PVAT) regulates vascular health. Dietary methionine restriction (MetR) impacts age-related adiposity, and this study addresses its effects in PVAT. METHODS: Male C57BL/6 mice at 8, 52, and 102 weeks of age were fed a standard (0.86%) or low-methionine (0.12%) diet for 52 weeks in 8-week-old and 52-week-old mice and for 15 weeks in 102-week-old mice. RESULTS: Mice with dietary MetR were resistant to weight gain and maintained a healthy blood profile. Aging increased lipid accumulation, and MetR reversed this phenotype. Notch signaling in inguinal white adipose tissue (iWAT) was decreased by MetR but increased in gonadal white adipose tissue. However, the Notch phenotype of brown adipose tissue (BAT) was not affected by MetR. Uncoupling protein 1 (UCP1) was increased in PVAT, iWAT, and BAT by MetR when initiated in young mice, but this effect was lost in middle-aged mice. CONCLUSIONS: Lipid in mouse PVAT peaked at 1 year of age, consistent with peak body mass. MetR reduced body weight, normalized metabolic parameters, and decreased lipid in PVAT in all age cohorts. Mice fed a MetR diet from early maturity to 1 year of age displayed an increased thermogenic adipocyte phenotype in iWAT, PVAT, and BAT, all tissues with thermogenic capacity.

Quantification of Lipid Area within Thermogenic Mouse Perivascular Adipose Tissue Using Standardized Image Analysis in FIJI.

Quantification of adipocyte size and number is routinely performed for white adipose tissues using existing image analysis software. However, thermogenic adipose tissue has multilocular adipocytes, making it difficult to distinguish adipocyte cell borders and to analyze lipid proportion using existing methods. We developed a simple, standardized method to quantify lipid content of mouse thermogenic adipose tissue. This method, using FIJI analysis of hematoxylin/eosin stained sections, was highly objective and highly reproducible, with ∼99% inter-rater reliability. The method was compared to direct lipid staining of adipose tissue, with comparable results. We used our method to analyze perivascular adipose tissue (PVAT) from C57BL/6 mice on a normal chow diet, compared to calorie restriction or a high fat diet, where lipid storage phenotypes are known. Results indicate that lipid content can be estimated within mouse PVAT in a quantitative and reproducible manner, and shows correlation with previously studied molecular and physiological measures.