Haematopoietic TLR4 deletion attenuates perivascular brown adipose tissue inflammation in atherosclerotic mice

Aims: To investigate whether haematopoietic TLR4 deletion attenuates perivascular brown adipose tissue inflammation in atherosclerotic mice. Methods and Results: Experiments were performed using irradiated LDL receptor-deficient (LDLR-/-) mice with marrow from either TLR4-deficient (TLR4(-/-)) or age-matched wild-type (WT) mice. After 12 weeks of being fed a high-cholesterol diet, TLR4(-/-) -> LDLR-/- mice developed fewer atherosclerotic lesions in the aorta compared to WT -> LDLR-/- mice. This effect was associated with an increase in multilocular lipid droplets and mitochondria in perivascular adipose tissue (PVAT). Immunofluorescence analysis confirmed that there was an increase in capillary density and M2 macrophage infiltration, accompanied by a decrease in tumour necrosis factor (TNF)-alpha expression in the localized PVAT of TLR4(-/-) -> LDLR-/- mice. In vitro studies indicated that bone marrow-derived macrophages (BMDMs) from WT mice demonstrated an M1-like phenotype and expression of inflammatory cytokines induced by palmitate. These effects were attenuated in BMDMs isolated from TLR4(-/-) mice. Furthermore, brown adipocytes incubated with conditioned medium (CM) derived from palmitate-treated BMDMs, exhibited larger and more unilocular lipid droplets, and reduced expression of brown adipocyte-specific markers and perilipin-1 compared to those observed in brown adipocytes exposed to CM from palmitate-treated BMDMs of TLR4(-/-) mice. This decreased potency was primarily due to TNF-alpha, as demonstrated by the capacity of the TNF-alpha neutralizing antibody to reverse these effects. Conclusions: These results suggest that haematopoietic-specific deletion of TLR4 promotes PVAT homeostasis, which is involved in reducing macrophage-induced TNF-alpha secretion and increasing mitochondrial biogenesis in brown adipocytes.