Fig. 5

Sebaceous gland organoids can model the initial stages of acne vulgaris. a–d Blimp1-YFP⁺-derived sebaceous gland organoids cultured for 5 days and treated for 3 days with a combination of DHT, BRL, LIN (DBL), or pure ethanol as control, followed by treatments for 3 days with retinoic acid (RA), 13-cis-retinoic acid (13cis RA) or Ret-Avit (Tretinoin). a DBL-treated organoids display increased organoid size in compared to control organoids, which is reduced upon treatments with retinoic acid derivatives. b Organoids stained for Oil Red O (ORO) show increased lipid production upon DBL treatment and retinoic acid derivatives. c Confocal analysis of treated and control organoids stained with Ki67, showing increased proliferation upon androgen induction and decreased proliferation in retinoic acid derivatives-treated organoids. d Quantitative real time (RT)-PCR analysis of treated organoids showing changes in mRNA levels of cMyc, Cyclin D1, androgen receptor (AR), fatty acid synthase (FASN), and peroxisome proliferator-activated receptors beta and gamma (PPAR-β,ɣ). e–h Blimp1-YFP⁺-derived sebaceous gland organoids cultured for 5 days and treated for 3 days with DBL combination as before, following treatments for 3 days with c-Myc inhibitor 10058-F4. e c-Myc inhibitor-treated organoids display decreased organoid size in compared to DBL-treated organoids. f Organoids stained for ORO, showing decreased lipid production upon c-Myc inhibition. g Confocal analysis of treated and control organoids stained with Ki67, showing decreased proliferation in c-Myc-inhibited organoids. h Quantitative RT-PCR analysis of treated organoids showing changes in mRNA levels of c-Myc, Cyclin D1, androgen receptor (AR), fatty acid synthase (FASN), and peroxisome proliferator-activated receptors beta and gamma (PPAR-β,ɣ). In all RT-PCR experiments, changes in cycle threshold values were normalized to Rplp0. Data are represented as mean relative to control organoids [n = 3 independent wells analyzed in triplicates]. Error bars show ± s.e.m. Significance was determined using two-tailed unpaired Student’s t-test where *p < 0.05, **p < 0.005, ***p < 0.001. All experiments were repeated at least twice in triplicates with similar results and images are representative of organoids grown from a mix of at least three mice and cultured in three independent biological repeats. Scale bars: 50 μm (a–c, e, f, h)