Hypothalamic CDK4 regulates thermogenesis by modulating sympathetic innervation of adipose tissues

Abstract This study investigated the role of CDK4 in the oxidative metabolism of brown adipose tissue (BAT). BAT from Cdk4 −/− mice exhibited fewer lipids and increased mitochondrial volume and expression of canonical thermogenic genes, rendering these mice more resistant to cold exposure. Interestingly, these effects were not BAT cell‐autonomous but rather driven by increased sympathetic innervation. In particular, the ventromedial hypothalamus (VMH) is known to modulate BAT activation via the sympathetic nervous system. We thus examined the effects of VMH neuron‐specific Cdk4 deletion. These mice display increased sympathetic innervation and enhanced cold tolerance, similar to Cdk4 −/− mice, in addition to browning of scWAT. Overall, we provide evidence showing that CDK4 modulates thermogenesis by regulating sympathetic innervation of adipose tissue depots through hypothalamic nuclei, including the VMH. This demonstrates that CDK4 not only negatively regulates oxidative pathways, but also modulates the central regulation of metabolism through its action in the brain.

Data information: All data are shown as the mean AE SEM. Student's t-test was used for statistical analysis *P < 0.05; ***P < 0.001. Source data are available online for this figure. A B C D Figure EV2. Cdk4 BAT-specific knockout mice have normal body weight, fat mass, and lean mass.
A Western blot for CDK4 expression in iBAT, scWAT, and pgWAT from Cdk4 flox/flox (n = 3 biological replicates) and Cdk4 flox/flox Ucp1-Cre (n = 3) mice. Tubulin was used as loading control. Please notice that upon amplification, the western blot images are abnormally pixelated. This is because the saturation of the images is shown in FigEV2A_Exposure_3.0sec.scn, which is a direct acquisition file from the Imager, and is found in the data source folder. A JPEG image is also available in this folder. B Whole-body image of Cdk4 flox/flox and Cdk4 flox/flox Ucp1-Cre mice. C, D Body weight (C) and body composition (fat and lean mass) (D) of Cdk4 flox/flox (n = 8) and Cdk4 flox/flox Ucp1-Cre (n = 8) mice.
Data information: All data are shown as the mean AE SEM. Source data are available online for this figure. A B C D Figure EV3. b3-receptor inhibition does not blunt the increased thermogenic response of Cdk4 À/À animals.
Data information: All data are shown as the mean AE SEM. Student's t-test was used for statistical analyses. **P < 0.01, ***P < 0.001. Source data are available online for this figure.
▸ Figure EV4. Cdk4 deficiency in Sf1 neurons does not affect body weight, body composition, or indirect calorimetry measures.
A, B Body weight (A) and body composition (fat mass and lean mass) (B) of Cdk4 flox/flox (n = 17) and Cdk4 flox/flox Sf1-Cre (n = 11) mice. C-H Indirect calorimetry was performed using the Oxymax apparatus (Columbus Instruments) in Cdk4 flox/flox (n = 6) and Cdk4 flox/flox Sf1-Cre (n = 5) mice. Whole-body oxygen consumption rate (VO 2 ) (C, D), respiratory exchange ratio (RER) (E, F), and energy expenditure (G, H) were measured at 24°C during the light phase (white rectangle) and dark phase (black rectangle). I, J CalR was used to implement GLM-regression plot for each group corresponding to the association between energy balance and total body mass at 24°C (I). The CalR interface displays each mouse as a dot, and the standard error of mean for each group in gray. The "mass effect" and "group effect" were analyzed using a generalized linear model (GLM) using body weight as a covariate. The results of this analysis are shown in table (J) for Cdk4 flox/flox (n = 6) and Cdk4 flox/flox Sf1-Cre (n = 5) mice.
Data information: All data are shown as the mean AE SEM. . D-F Western blot analysis (D) and quantification of TH (E) and UCP1 protein expression (n = 6 biological replicates) (F). HSP90 was used as loading control.
Data information: All data are shown as the mean AE SEM. Student's t-test (C) and Mann-Whitney U-test (E, F) were used for statistical analyses. **P < 0.01, ***P < 0.001. Source data are available online for this figure.