Neuropeptide Y-Positive Neurons in the Dorsomedial Hypothalamus Are Involved in the Anorexic Effect of Angptl8

FIGURE 1. Effect of peripheral and central Angptl8 on food intake and 24 h body weight. (A) After 12-h food deprivation, the I.V. injection of Angptl8 (0.1 ml) decreased nocturnal cumulative food intake from 1 to 12 h after injection. (B) After 12-h food deprivation, I.C.V. injection of Angptl8 (2 μl) decreased nocturnal cumulative food intake from 2 to 6 h after injection. (C) Peripheral Angptl8 (0.1 ml; 30 μg/ml) decreased 24-h body weight gain (n = 8). (D) Central Angptl8 for (2 μl; 0.3 μg/μl) decreased 24-h body weight gain (n = 6). Data are means ± SEM. ∗P < 0.05 and ∗∗P < 0.05 relative to the NS control.

FIGURE 2. Angptl8 expression in appetite-related nuclei in the hypothalamus. Angptl8 was expressed in the hypothalamic PVN (B), DMH (A), ARC (C), and VMH (D) in normal mice on chow diet. Scale bar = 200 μm.

FIGURE 3. Peripheral Angptl8 decreased c-Fos-positive neurons expression in the DMH of the hypothalamus. (A) c-Fos-positive neurons change in hypothalamus 2 h after tail intravenous injection with NS and Angptl8. (B–D) The numbers of c-Fos-immunopositive cells in the DMH (n = 8), PVN (n = 6), and ARC (n = 8) are expressed as means ± SEM. ∗P < 0.05 relative to the NS control group. Scale bar in A = 100 μm.

FIGURE 4. Central Angptl8 influenced c-Fos-positive neurons expression in the PVN, DMH, and ARC. (A) Changes in c-Fos-positive neurons in the hypothalamus 2 h after I.C.V. injection with NS and Angptl8. (B–D) The numbers of c-Fos immunoreactive cells in the DMH (n = 5), PVN (n = 3), and ARC (n = 6) are expressed as means ± SEM. ∗P < 0.05, ∗∗P < 0.001 relative to the NS control group. Scale bar in A = 100 μm.

FIGURE 5. Double staining of c-Fos and NPY in the hypothalamic PVN, DMH, and ARC after the I.C.V. injection of Angptl8. (A–C) Double-immunofluorescence images showed c-Fos immunoreactive cells (green) and NPY neurons (red) separately, and neurons with colocalization in the PVN, DMH, and ARC are depicted in yellow in the merged photograph. Scale bars = 200 μm. (D–F) Colocalized neurons in the DMH (n = 4), PVN (n = 3), and ARC (n = 3) were counted and are expressed as means ± SEM. ∗P < 0.05 relative to the NS control group. Scale bar = 200 μm → means colocalization of c-Fos and NPY neurons.

FIGURE 6. Effect of peripheral and central Angptl8 on body weight gain, fat mass and plasma FFA levels. Angptl8 (30 μg/ml) via I.V. injection decreased 9-day body weight gain (A) and epididymal fat deposits (B) but had no effect on inguinal fat deposits (C) or plasma free fatty acid levels (D). Angptl8 (0.3 μg/μl) via I.C.V. injection decreased 9-day body weight gain (E) and plasma free fatty acid (H) but increased the epididymal (F) and inguinal (G) fat deposits. Data are means ± SEM. ∗P < 0.05 relative to the NS control group.

FIGURE 7. Effect of peripheral and central Angptl8 on BAT mass, BAT morphology, and Ucp-1 expression. Angptl8 (30 μg/ml) via I.V. injection (n = 6 and 3 in experimental and control group, respectively) decreased BAT deposit (A) but had no effect on BAT cell density (B) and Ucp-1 expression (C–E). (D) Use β-actin as the HKG and (E) use GAPDH as the HKG. Angptl8 (0.3 μg/μl) via I.C.V. injection (n = 7 and 3 in experimental and control group, respectively) increased BAT deposit (F) but had no effect on BAT morphology (G) or Ucp-1 expression (H–J). (I) Use β-actin as the HKG and (J) use GAPDH as the HKG. Data are means ± SEM. ∗P < 0.05 relative to the NS control group.