Figure 1.

3D Co-culture with Prostate Stroma Supports Organoid Growth and Increases Organoid Branching

(A) Gene expression of PrS mix 1 from microarray of stroma-specific genes TIMP3, von Willebrand factor (VWF), α-SMA (ACT2 shown as SMA), and EpCAM. Quadruplicate measures reported; error bars represent one standard deviation of the mean. Stromal cells are a heterogeneous mixture of fibroblasts and myofibroblasts by immunofluorescent staining of α-SMA and vimentin in PrS mix 3. Scale bar, 100 μm.

(B) Schematic of mono-culture and co-culture workflow and conditions. PrE and PrS cells seeded into 33% Matrigel on top of 50% Matrigel base layer.

(C) Fluorescent images of PrS mix 1 incorporation of an EdU analog cultured in 3D conditions. Scale bar, 200 μm.

(D) Bright-field images of organoids in mono- and co-culture derived from dissociated epithelial cells isolated from benign prostate tissue specimens of two patients (PrE-1, top; PrE-2, bottom). Hematoxylin and eosin staining showing histopathology of patient tissue. Scale bar, 200 μm.

(E) Percentage of branched organoids in mono- and co-cultures on days 7 and 14 according to a branching classification scheme used to categorize each organoid by extent of branching (right). Quantification of organoid branching in mono- and co-cultures according to the branching classification scheme in organoids derived from two patients (PrE-1, top; PrE-2, bottom). Triplicate wells per condition, n reported as sum of branched organoids from triplicate wells per condition.

(F) Bright-field images of time course showing PrE-1 organoid branching from days 6–10 in co-culture with PrS mix 4. Scale bar, 200 μm.