Fig 2: Immunofluorescence assays for CD34 (green) and c‐kit (Red) in histological sections of Mongolian gerbil prostate of early postnatal development. (A–D) In the perinatal period (P1), labelling of CD34 and of c‐Kit is dispersed in the VMP. (E‐H) Labelling of CD34 progresses in the region surrounding prostatic budding on P7, while c‐Kit labelling remains disperse along the VMP. Original magnification was 200×, scale bar represents 50 μm.VMP, ventral mesenchymal pad; PB, prostatic branching; St, prostate stroma; PA, developing prostate alveoli.

Fig 3: Characterization of isolated aortic TCs. (A) Different shapes of aortic TCs shown by light microscopy eight days after isolation. Piriform spindle‐shaped TCs (1, 3); triangular‐shaped TCs (2, 4, 5); stellate‐shaped TC (6). Scale bar: 20 µm. (B) CD34/c‐kit (left) and vimentin/c‐kit (right) immunofluorescence staining. TCs were identified as double‐positive cells with CD34 and vimentin or c‐kit, respectively. Nuclei were counterstained with DAPI (blue). Scale bar: 20 µm. (C) Indication of cell‐cell communication. Direct cell‐cell contacts between TCs/Tps and co‐cultivated ECs (1, 3 and 4) or SMCs (2). The areas of cell connections are shown in enlarged sections for 1–3 and further pointed out in 2 (asterisk). An immunofluorescence stain against vimentin (green) is presented in (4). Scale bar: 1, 20 µm; 2–4, 50 µm. EC, endothelial cell; SMC, smooth muscle cell; TC, telocyte body; Tp, telopode

Fig 4: Effect of CGB5 on VM formation by H&E and CD34-PAS staining. (A) After tumor growth to 1–2 cm3, the tumors were collected and cut into serial 6-µm sections. (a-e) H&E staining was used to detect VM formation. (f-j) PAS staining was used to identify matrix-associated vascular channels, while CD34 staining was used to identify the endothelium in the cancer tissue sections. PAS-positive and CD34-negative channels were considered as VM. (B) VM quantification was performed as follows: (a) VM channels and endothelium-dependent vessels in the H&E-stained sections were counted using ×400 magnification, and (b) CD34-PAS dual staining sections were viewed at ×400 magnification. *P<0.05 vs. normal cells. (C) Transmission electron microscopy examination. CGB5, chorionic gonadotropin, beta polypeptide 5; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; RNAi, RNA interference; HCG, human chorionic gonadotropin; VM, vasculogenic mimicry; H&E, hematoxylin and eosin; PAS, periodic acid-Schiff.

Fig 5: Immunocytochemistry for the detection of myogenic markers. The expression of muscle-related proteins in the tonsil-derived mesenchymal stem cells (T-MSCs) during the process of myogenic induction (T-MSCs; T-MSC-derived myoblasts; T-MSC-derived myocytes) was evaluated by immunostaining using antibodies against myogenic satellite cells (panel a, CD34; red), precursors [panel b, paired box 7 (Pax7); green], differentiated myogenic cells [panel c, desmin (red); panel d, dystrophin (green); panel e, myosin heavy chain (MHC; red)], skeletal myogenic cells [panel f, α-actinin (green); panel g, troponin I type 1 (TNNI1; green); panel h, myogenin (red)]. The cells were counterstained with DAPI (blue). The samples were analyzed under a fluorescence microscope using appropriate filters. (A) Undifferentiated T-MSCs expressed (panel a) CD34 and (panel b) Pax7, but no other markers of myogenic cells. (B) Following differentiation into myoblasts, 80–90% of cells expressed (panel c) desmin, (panel d) dystrophin, and (panel e) MHC and 20–50% of cells expressed skeletal muscle (SKM) markers including (panel f) α-actinin, (panel g) TNNI1, and (panel h) myogenin. (Panel a) CD34 and (panel b) Pax7 were not expressed at any stage of the differentiation process from T-MSC-derived myoblasts into T-MSC-derived myocytes. (C) T-MSC-derived myocytes exhibited an increased expression of (panels c to h) myogenic and SKM cell markers compared with T-MSC-derived myoblasts and formed multinucleated myotubes. FITC, fluorescein isothiocyanate; TRITC, tetramethylrhodamine isothiocyanate. Original magnification, ×200.