Journal: Scientific Reports

Article Title: Bladder organoid conditioned media enhances myoblast proliferation under serum free conditions

doi: 10.1038/s41598-026-38603-7

Figure Lengend Snippet: Schematic diagram of the production of mouse normal tissue-derived organoids and comparative analysis of the culture supernatants of tissue-derived organoids from each organ ( A ). At 5 weeks of age, male C57BL/6J mice were euthanized, and the lungs (MLO), gallbladder (MGO), spleen (MSO), kidneys (MKO), and bladder (MBO) were removed. Each tissue was digested, washed, mixed with Matrigel, and cultured as three-dimensional organoids. Each conditioned medium (CM) of the organoid was applied to C2C12 myoblasts to assess their proliferation activity in vitro. Brightfield microscopy (scale bar: 50 µm), hematoxylin and eosin (H&E) staining, and immunohistochemical staining (scale bars: 200 µm) of MLO, MGO, MSO, MKO, and MBO ( B ). Organoids were probed with the pan-epithelial marker AE1/AE3 and the mesenchymal marker vimentin to assess lineage composition. The following markers were confirmed as tissue-specific: thyroid transcription factor (TTF) for MLO; cytokeratin 19 (CK19) and mucin-3 (MUC3) for MGO; vascular endothelial growth factor receptor 2 (VEGFR-2) and CD31 for MSO; paired box protein 8 (PAX8) for MKO; and cytokeratin 5 (CK5) and uroplakin 3A (UPK3A) for MBO.

Article Snippet: The primary antibodies used were as follows: cytokeratin cocktail (AE1/AE3, Novus Biologicals, Centennial, CO, USA); vimentin (R&D Systems, Minneapolis, MN, USA); thyroid transcription factor-1 (TTF-1, Novus Biologicals, Centennial, CO, USA); cytokeratin 19 (CK19, Novus Biologicals); mucin 3 (MUC3, Bioss Inc., Woburn, MA, USA); cluster of differentiation 31 (CD31, Thermo Fisher Scientific); vascular endothelial growth factor receptor 2 (VEGFR-2, Santa Cruz Biotechnology, Dallas, TX, USA); paired box gene 8 (PAX8, GeneTex, Inc., Irvine, CA, USA); cytokeratin 5 (CK5; GeneTex, Inc); uroplakin IIIa (UPK3A, Novus Biologicals), myogenic factor 5 (Myf5, GeneTex, Inc), myoblast determination protein 1 (MyoD, GeneTex, Inc).

Techniques: Derivative Assay, Cell Culture, Activity Assay, In Vitro, Microscopy, Staining, Immunohistochemical staining, Marker