Figure 4. Mild mitoribosomal stress increases β-END expression in POMC neurons.

(A) Pomc fluorescence in situ hybridization (FISH) showing no alteration in ARH POMC neuron numbers (upper panel) and an increase in the percentage of c-Fos⁺ POMC neurons (lower panel) in 8 week-old Pomc-Cre; Crif1^f/+ males compared to Crif1^f/f controls (n = 6–7). Scale bars, 25 μm. Arrowheads indicate c-Fos⁺ POMC neurons.

(B) Increased numbers of β-END⁺ neurons, but no change in the α-MSH⁺ neuron population, in the hypothalamic ARH of Pomc-Cre; Crif1^f/+ male mice at 8 weeks (n = 6–7). Scale bars, 50 μm. IF, immunofluorescence staining.

(C) Increase in β-END⁺ axonal projections in the hypothalamic paraventricular (PVH) and dorsomedial nucleus (DMH) of Pomc-Cre; Crif1^f/+ males (n = 7). Scale bars, 100 μm.

(D) Quantitation of the hypothalamic β-END and α-MSH protein contents at 8 weeks of age (n =4–6).

(E) Changes in the hypothalamic mRNA expression levels of Pomc and of enzymes involved in the production and degradation of POMC-derived peptides in Pomc-Cre; Crif1^f/+ mice (n = 5).

(F–H) Repeated ICV injection of β-END (0.1 μg twice a day for 4 days) increases EE and induces browning, enhanced sympathetic innervation, thermogenic gene expression, and the UPR^mt in the iWAT of C57 mice (n = 5–6). Scale bars, 200 μm (G, upper) and 50 μm (G, lower).

Results are presented as a mean ± SEM. *p < 0.05, **p < 0.01, and ***p < 0.001 between indicated groups. NS, not significant. See also Figure S4.