Thus, obesity involved only a modest and not significantly higher expression of ChAT mRNA per macrophage, suggesting that the higher ACh and choline levels seen in the SVF of HFD mice are mainly due to the increased number of ChAT-expressing macrophages that infiltrate obese eWAT, as shown in Figure 2, Figure 3G. Moreover, according to the semiquantitative histological scoring method provided by ACDBio for RNAscope® (https://acdbio.com/image-analysis), ChAT staining of macrophages from CD and HFD mice was low to moderate (score 2). Here, CHAT is linked to obesity due to melanocortin 4 receptor deficiency.