Norepinephrine-dependent selection of brown adipocyte cell lines

Abstract

A transgenic mouse carrying a simian virus-40 T-antigen gene under control of a mouse urinary protein promoter induced the formation of a brown fat tumor. In tissue culture, these tumor cells expressed the brown fat-specific mitochondrial uncoupling protein (Ucp) gene when stimulated by norepinephrine. To develop cell lines for the analysis of Ucp expression, we investigated growth conditions that would maintain expression. We found that the addition of 10(-7)-10(-6) M norepinephrine was critical for establishing cells with high Ucp expression, mitochondrial content, and adipogenesis. Norepinephrine exerts its effects by selectively stimulating the proliferation of brown fat cells. Results with receptor-specific agonists and antagonists indicate that norepinephrine interacts with the beta 1-adrenergic receptor to stimulate cell proliferation. The capacity of these brown fat cells to respond to norepinephrine by proliferating seems to be a manifestation of a normal physiological response of brown fat cells, because exposing an animal to the cold increases cell proliferation. Thus, this system indicates that two independent pathways for cell proliferation coexist in parallel, one based on transformation of the cell by the simian virus-40 T-antigen and the other on the normal response of the cell to stimulation by norepinephrine. Several clonal lines have been isolated that have similar levels of marker gene expression for adipogenesis and mitochondriogenesis, but differ in the level of Ucp expression. The results underscore the extreme sensitivity of mechanisms controlling Ucp expression and the fact that the expression of Ucp in brown fat is not coordinately linked to that of other nuclear genes which encode proteins associated with thermogenesis.

Adrenergic regulation of cellular plasticity in brown, beige/brite and white adipose tissues

“…51,[85][86][87][88] Early in vitro work from the labs of Kozak and Nedergaard suggested that norepinephrine triggers BA proliferation via ADRB1. [88][89][90][91] These data are in agreement with the fact that ADRB1 are present in BA progenitors, whereas ADRB3 is a marker of the differentiated state. 91 Recently, we reported that knockout of ADRB1 abrogates cold-induced iBAT hyperplasia.…”

Expression of the mitochondrial uncoupling protein gene from the aP2 gene promoter prevents genetic obesity.

“…The absence of an accumulation of mitochondria (unpublished observations, X.X. Koza and L.P. Kozak) indicates that expression of Ucp is not sufficient to stimulate mitochondriogenesis, a conclusion reached independently from our analysis of brown adipocyte cell lines (27). Since the aP2 gene is one of the late stage markers of adipocyte differentiation program (28), it is possible that the adipocyte derivatives originate from white preadipocytes.…”

Increased TNFα and CCAAT/enhancer-binding protein homologous protein with aging predispose preadipocytes to resist adipogenesis

“…Markers of macrophages [F4/80 (emr1), scya4, cd11␤, and cd45 (ptprc)], endothelial cells [vegfr (flk1), vegfr2 (kdr), cd46 (mcp), and von Willebrand], and multipotent mesenchymal progenitors (cd34, connexin 43, sox2, and smad4) indicated that age-related shifts in abundance of these cell types in preadipocyte cultures did not account for our results. Brown fat preadipocytes are not present in epididymal cultures, since uncoupling protein 1 is not evident in these cultures after treatment with isoproterenol but is observed in interscapular preadipocyte positive controls (36,41). To induce differentiation, confluent preadipocyte cultures were exposed to a serum-free differentiation-inducing medium (DM) containing 5 g/ml insulin, 10 g/ml transferrin, and 200 pM triiodothyronine in 1:1 DMEM-Ham's F-12 (20).…”