Adipose Tissue Mast Cells Promote Human Adipose Beiging in Response to Cold

Abstract

In a recent study, repeated cold application induced beiging in subcutaneous white adipose tissue (SC WAT) of humans independent of body mass index. To identify factors that promote or inhibit beiging, we performed multiplex analysis of gene expression with the Nanostring nCounter system (the probe set contained genes for specific immune cell markers, cytokines, and chemokines) on the SC WAT from lean subjects. Multiple correlations analysis identified mast cell tryptase and CCL26, a chemokine for mast cells, as genes whose change correlated positively with the change in UCP1 in SC WAT, leading to the hypothesis that mast cells promote SC WAT beiging in response to cold. We quantified mast cell recruitment into SC WAT and degranulation. Mast cells increased in number in SC WAT in lean subjects, and there was an increase in the number of degranulated mast cells in both lean subjects and subjects with obesity. We determined that norepinephrine stimulated mast cell degranulation and histamine release in vitro. In conclusion, cold stimulated adipose tissue mast cell recruitment in lean subjects and mast cell degranulation in SC WAT of all research participants independent of baseline body mass index, suggesting that mast cells promote adipose beiging through the release of histamine or other products.

Figure 1

Correlations of changes in gene expression with changes in UCP1. The change in UCP1 protein expression in SC WAT by acute cold treatment (post – pre) was calculated for the cold treated and contralateral legs using our previously published results. The change in UCP1 protein is plotted versus the change in gene expression of genes identified by multiple correlations analysis. (A,B) The change of tryptase or CCL26 versus the change in UCP1 protein the cold leg are shown. The data were analyzed by linear regression analysis, and Pearson correlation coefficients (r) and P values are shown (n = 12). (C) The quantification of tryptase gene expression at baseline and after cold in SC WAT of the cold treated leg is shown. Data represent means ± SEM (n = 12). *P < 0.05 (Wilcoxon matched-pairs signed rank test).

Figure 2

Mast cell density and degranulation in SC WAT of research participants in response to acute cold treatment. (A,B) Tryptase staining of SC WAT at baseline and after cold (scale bar for C: 50 μm; scale bar for D: 100 μm). White arrows point to intact mast cells and yellow arrows point to mast cells with diffuse, punctate staining. (C) Quantification of mast cell density in lean subjects and subjects with obesity at baseline and in SC WAT from the cold and contralateral legs after 10 days of acute cold exposure. (D,E) Higher magnification images demonstrating degranulated and intact mast cells (scale bar: 20 μm). (F) Quantification of degranulated mast cells in lean subjects and subjects with obesity at baseline and in SC WAT from the cold and contralateral legs after 10 days of acute cold exposure. Data represent means ± SEM. Data were analyzed by RM MANOVA as described in research design and methods. **P < 0.01; ****P < 0.0001 (lean n = 17; obese n = 8).

Figure 3

Mast cells express βARs and degranulate in response to norepinephrine. (A,B) βAR2 and 3 mRNA expression was determined in 3T3L1 and TIB64 mast cells by real-time RT PCR as described in methods. (C) TIB64 cells were changed into media at 32 C or 37 C with 0 or 100 nM norepinephrine (NE) for 0, 120 or 240 minutes as indicated. The media was removed from the cells and histamine determined as described in research design and methods. Data are presented as means ± SEM. The data were analyzed by ANOVA as described in research design and methods. **P < 0.01; ***P < 0.001; ****P < 0.0001. (D) Mast cell density and degranulated mast cells were quantified in SC WAT from human research participants with obesity treated with 50 mg mirabegron per day for 10 weeks. Data are presented as means ± SEM and were analyzed by a paired, two-tailed student’s t test (n = 6).