A mesodermal fate map for adipose tissue

Abstract

The embryonic origin of distinct fat depots and the role for ontogeny in specifying the functional differences among adipocyte lineages between and within depots is unclear. Using a Cre/Lox-based strategy to track the fate of major mesodermal subcompartments in mice we present evidence that <50% of interscapular brown adipocytes are derived from progenitors of the central dermomyotome. Furthermore, we demonstrate that depot-specific adipocyte lineages spatially diverge as early as gastrulation, and that perigonadal adipocytes arise from separate mesodermal subcompartments in males and females. Last, we show adipocyte precursors (APs) of distinct lineages within the same depot exhibit indistinguishable responses to a high fat diet, indicating that ontogenetic differences between APs do not necessarily correspond to functional differences in this context. Altogether, these findings shed light on adipose tissue patterning and suggest that the behavior of adipocyte lineage cells is not strictly determined by developmental history.

Keywords: Adipocyte; Adipose; Fate map; Lineage tracing; Mesoderm.

Fig. 1.

Fate-mapping mesodermal subcompartments using the Cre/Lox system. (A) Cre drivers used in this study and the unique mesodermal subcompartments in which they are active. (B,D,F) Tracing of somite-derived tissues (triceps and tibialis anterior) in the Meox1-Cre:mTmG system (B), in the Pax7-Cre:mTmG system (D) and in the HoxB6-Cre:mTmG system (F). (C,E,G) Tracing of anterior lateral plate (heart) and posterior lateral plate (spleen) derived tissue in the Meox1-Cre:mTmG system (C), in the Pax7-Cre:mTmG system (E) and in the HoxB6-Cre:mTmG system (G). Data are from males and females aged 4-10 weeks of age. Scale bar: 100 µm. Green, traced cells; red, untraced cells.

Fig. 2.

Dorso-axial adipocytes and male perigonadal adipocytes predominately arise from somitic mesoderm. (A-G) Images and quantification of Meox1-Cre:mTmG reporter in interscapular brown adipose tissue (inscBAT) (A), interscapular white adipose tissue (inscWAT) (B), retroperitoneal white adipose tissue (rWAT) (C), triceps white adipose tissue (triWAT) (D), inguinal white adipose tissue (ingWAT) (E), mesenteric white adipose tissue (mWAT) (F), perigonadal white adipose tissue (pgWAT) (G). n=6-8, depending on the depot. Data are from males aged 4-6 weeks and from females aged 5-6 weeks. For each image, the boxed area is shown in black and white for the red (top) and green (bottom) channels. Error bars represent s.e.m. Scale bar: 100 µm.

Fig. 3.

A minority of interscapular white and brown adipocytes originate in the central dermomyotome. (A-G) Images and quantification of the Pax7-Cre:mTmG reporter in interscapular brown adipose tissue (inscBAT) (A), interscapular white adipose tissue (inscWAT) (B), retroperitoneal white adipose tissue (rWAT) (C), triceps white adipose tissue (triWAT) (D), inguinal white adipose tissue (ingWAT) (E), mesenteric white adipose tissue (mWAT) (F), perigonadal white adipose tissue (pgWAT) (G). n=3-7, depending on the depot. Data are from males aged 4-10 weeks and from females aged 4-9 weeks. For each image, the boxed area is shown in black and white for the red (top) and green (bottom) channels. Error bars represent s.e.m. Scale bar: 100 µm.

Fig. 4.

Adipocytes populating major ventro-lateral adipose depots share a common lineage. (A-G) Images and quantification of HoxB6-Cre:mTmG reporter in interscapular brown adipose tissue (inscBAT) (A), interscapular white adipose tissue (inscWAT) (B), retroperitoneal white adipose tissue (rWAT) (C), triceps white adipose tissue (triWAT) (D), inguinal white adipose tissue (ingWAT) (E), mesenteric white adipose tissue (mWAT) (F), perigonadal white adipose tissue (pgWAT) (G). n=4-6, depending on the depot. Data are from males aged 5 weeks±3 days and from females aged 5-7 weeks. For each image, the boxed area is shown in black and white for the red (top) and green (bottom) channels. Error bars represent s.e.m. Scale bar: 100 µm.

Fig. 5.

Posterior lateral plate mesoderm establishes ventro-lateral adipocytes. (A-G) Images and quantification of HoxB6-CreER^T:mTmG reporter in interscapular brown adipose tissue (inscBAT) (A), interscapular white adipose tissue (inscWAT) (B), retroperitoneal white adipose tissue (rWAT) (C), triceps white adipose tissue (triWAT) (D), inguinal white adipose tissue (ingWAT) (E), mesenteric white adipose tissue (mWAT) (F), perigonadal white adipose tissue (pgWAT) (G). n=3-5, depending on the depot. Data are from males aged 5 weeks±2 days and from females aged 4-5 weeks. For each image, the boxed area is shown in black and white for the red (top) and green (bottom) channels. Error bars represent s.e.m. Scale bar: 100 µm.

Fig. 6.

Lineage tracing patterns for depot-resident APs. (A-D) AP tracing for seven major adipose depots, according to the gating strategy shown in Fig. S2A,B: Meox1-Cre:mTmG, in males (n=7-8) and females (n=6-7) (A), Pax7-Cre:mTmG, in males (n=5-7) and females (n=3-5) (B), HoxB6-Cre:mTmG, in males (n=4) and females (n=5-6) (C), HoxB6-CreER^T:mTmG, in males (n=4) and females (n=3-5) (D). Green, mGFP⁺ APs; red, mTomato⁺ APs; blue, unlabeled APs.

Fig. 7.

The tissue microenvironment, not developmental lineage, controls obesogenic AP proliferation in male perigonadal fat. (A) Anatomy of male perigonadal white adipose tissue (pgWAT) with tip, middle and base indicated. (B) Representative confocal images of tip, middle and base of pgWAT from male Meox1-Cre:mTmG animals. (C) Quantification of mGFP⁺/mTomato⁺ adipocytes in noted regions of male Meox1-Cre:mTmG pgWAT (n=4). (D) Quantification of mGFP⁺/mTomato⁺ APs in noted regions of male Meox1-Cre:mTmG pgWAT (n=3-4). (E) Scatterplot indicating concordant tracing of adipocytes and APs in each region of male Meox1-Cre:mTmG pgWAT. Each dot represents the percentage of APs and adipocytes that are mGFP⁺ for the indicated region of the depot in one animal (n=3-4). As the pgWAT depot exists in a pair, one depot was used for quantifying adipocyte labeling and the other for AP labeling. Significance was determined using Pearson's two-tailed correlation analysis. (F) BrdU incorporation into APs of tip, middle and base of male pgWAT after 1 week of HFD (n=5). (G) BrdU incorporation into somite-derived (mGFP⁺) and non-somite-derived (mTomato⁺) APs of male Meox1-Cre:mTmG pgWAT (n=6). Error bars represent s.e.m. *P<0.05, **P<0.01, ***P<0.001 (unpaired two-tailed Student's t-test on indicated groups in C,D,F,G, significance determined using mGFP⁺ cells in C and D). B, base; HFD, high fat diet; M, mid; NS, not significant; SD, standard diet; SM, somitic mesoderm; T, tip. Scale bar: 100 µm.

Fig. 8.

A mesodermal fate map for adipose tissue. (A) Schematic showing the mesodermal fate map for adipose tissue based on tracing with Meox1-Cre:mTmG, Pax7-Cre:mTmG and HoxB6-Cre:mTmG systems. Red, somitic mesoderm and derivative adipocyte lineages; yellow, central dermomyotome and derivative adipocyte lineages; blue, posterior lateral plate mesoderm and derivative adipocyte lineages. (B) Schematic showing a model fate map for interscapular adipocyte lineages based on Pax7-Cre:mTmG tracing in this study and Pax3/Myf5-Cre:mTmG tracing by Sanchez-Gurmaches and Guertin (2014). It is proposed that a subset of Pax3⁺ progenitors in the epaxial dermomyotome and Pax7/Pax3⁺ progenitors in the central dermomyotome are fated to the interscapular adipocyte lineage. These Pax3⁺ or Pax7/Pax3⁺ cells acquire Myf5 expression in a Pax3-dependent manner (Sato et al., 2010) and go on to establish interscapular white and brown adipocytes. (C) Schematic of somite subcompartments. (D) Model lineage hierarchy for interscapular adipocytes based on Pax7-Cre:mTmG tracing in this study and Pax3/Myf5-Cre:mTmG tracing by Sanchez-Gurmaches and Guertin (2014).