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. 2019 Aug 9;235(3):1877–1887. doi: 10.1002/jcp.29121

Figure 3.

Figure 3

CytoMatrix for the study of MPE‐derived CSCs. (a) Representative images of a lung stable cell line NCI‐H460 and BBIRE‐T238‐derived primary cell line grown in adherent (2D) and in 3D conditions. The pictures were captured on Day 4. Original magnification = ×100. (b) Gene expression of stemness markers and scd1 performed by qRT‐PCR analyses on 3D and 2D cultures obtained from NCI‐H460 and the primary cell line isolated from MPEs of BBIRE‐T238 patient. The bar chart shows the fold change in the mRNA levels in 3D spheroids compared with 2D parental cells (histone H3 was used as housekeeping gene). Data represent the mean and standard deviation (SD) or standard error of mean (SEM) of at least three independent experiments and are statistically significant if *p < .05 (Student's t test). (c) Example of IHC staining of haematoxylin and eosin, CK7, PAN‐CK, CD45 and TTF‐1 performed on lung adenocarcinoma biopsy (upper panels), and of CytoMatrix‐based ICC staining of primary cell line derived from BBIRE‐T238 patient (middle and bottom panels). The results indicate a similar pattern of expression between biopsy and primary cell cultures. Original magnification = ×400. (d) SCD1 expression obtained from CytoMatrix‐based ICC staining of NCI‐H460 and BBIRE‐T238 primary cell line in 2D and 3D conditions, and from IHC staining of biopsy. Original magnification = ×200, detail ×1,000. (e) YAP/TAZ IHC (upper panel) and YAP/TAZ CytoMatrix‐based ICC (bottom panel) performed on samples obtained from BBIRE‐T238 patient. Original magnification = ×200, detail ×1,000. (f) Comparison between YAP/TAZ CytoMatrix‐based ICC (panel on the left) and YAP/TAZ CytoMatrix‐based IF (panel on the right) analyses obtained from NCI‐H460 grown in 3D conditions. Scale bar = 20 µm. 2D, two‐dimensional; CSC, cancer stem cells; ICC, immunocytochemistry;IF, immunofluorescence; IHC, immunohistochemistry; MPE, malignant pleural effusion; PAN‐CK, Pan‐cytokeratin; TAZ, transcriptional coactivator with PDZ‐binding motif; qRT‐PCR, quantitative reverse‐transcriptase polymerase chain reaction; YAP, Yes‐associated protein [Color figure can be viewed at wileyonlinelibrary.com]