(A) Schematic of differentiation protocol for iHTNs.
(B) Immunocytochemical staining of iHTNs showing various neuronal, hypothalamic, and neuropeptidergic markers.
(C) Quantification of the various cell types contained in iHTN cultures (n = 8 lines).
(D) Representative cell traces of iHTNs expressing relevant markers show distinctive morphologies.
(E) Quantification of primary neurite length (n = 8 lines).
(F) qRT-PCR of hypothalamic and arcuate nucleus-specific genes showing significantly increased expression ofthe genes at day 40 of differentiation compared to day 0 (n = 3 lines).
(G) Representative immunoblots and quantified histograms showing increases in neuron numbers (TUBB3) and arcuate nucleus markers (CART and AgRP) in day 40 cultures compared to day 0 (n = 3 lines). *p < 0.05, **p < 0.01, ***p < 0.001. All statistical analysis was performed using unpaired t test.
Error bars represent SEM.