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. 2020 Jan 4;9(1):121. doi: 10.3390/cells9010121

Figure 2.

Figure 2

Silencing of OPA1 or MFN1 inhibits mitochondrial fusion and the growth of CCA organoids. (A) Representative confocal images of mitochondria with TMRM staining in in vitro cultured organoids from two tumor tissue, one matched adjacent liver tissue and one donor liver tissue. The white boxed regions were further magnified in the expanded images (right row). Scale bar = 50 µm (left row). Scale bar = 10 µm (right row). (B) Images of tumor and non-tumor “normal” liver organoids were quantified (Normal vs. Tumor: 37.93 ± 10.62 vs. 83.5 ± 7.34; n = 6 images/sample). (C) CCA organoids were transduced with mock lentivirus (Ctr) or shOPA1/shMFN1 lentivirus (KD1 and KD2) respectively. Gene knockdown efficiency of OPA1 was quantified by Real-time PCR (n = 6). (D) Representative live cell confocal images of mitochondria with TMRM staining in CCA organoids with OPA1/MFN1 knockdown. The white boxed regions were further magnified in the expanded images (lower row). Scale bar = 20 µm (upper row). Scale bar = 5 µm (lower row). (E) Quantified measurements of mitochondrial length in control and KD cells of CCA organoids (n = 3 images/sample). (F) The Alamar Blue fluorescence level of CCA organoids was measured at Day 0/1/2/3 for cell viability and data at day 0 was set as control of each group (n = 3). Histograms show means ± SEM with p value derived by the Mann Whitney test.