Figure 5 from Human Bone Marrow Organoids for Disease Modeling, Discovery, and Validation of Therapeutic Targets in Hematologic Malignancies

<p>Bone marrow organoids model TGFβ-induced bone marrow fibrosis and enable inhibitor screening. <b>A,</b> Organoids were treated with 10, 25, or 50 ng/mL recombinant TGFβ and evaluated by qRT-PCR for expression of <i>ACTA2</i> (αSMA) and <i>COL1A1</i>, indicators of fibrosis. <b>B,</b> Soluble IL11 detected in organoid media following treatment of organoids with TGFβ. <b>C,</b> Confocal Z-stack images of whole, untreated, and TGFβ (50 ng/mL)-treated organoids stained for αSMA and COL1A1. <b>D,</b> Reticulin staining of formalin-fixed, paraffin-embedded sections of TGFβ-treated organoids versus control. <b>E,</b> Measurement of total reticulin stained area in untreated and TGFβ (50 ng/mL)-treated organoids. <b>F</b> and <b>G,</b> CD34 immunostaining of organoid vessels (<b>F</b>) and quantification of total vascular area of organoids with/without TGFβ treatment (<b>G</b>). <b>H</b> and <b>I,</b> Effect of two potential inhibitors of TGFβ-induced fibrosis (SB431542 and JQ1) on IL11 secretion (<b>H</b>) and <i>ACTA2</i> and <i>COL1A1</i> expression (<b>I</b>). <b>J,</b> αSMA and COL1A1 expression in TGFβ-treated organoids with/without indicated inhibitors. Representative images are shown. *, <i>P</i> < 0.01; **, <i>P</i> < 0.05; ***, <i>P</i> < 0.001; ****, <i>P</i> < 0.0001 for one-way ANOVA with multiple comparisons (Fisher least significant difference). <i>T</i> tests performed for image analysis of paraffin-embedded sections (reticulin and CD34). <i>n</i> = 3 with each repeat comprising 15 organoids pooled from 3 independent differentiations and treatments.</p>