Transmembrane channel-like 5 drives hepatocellular carcinoma progression by regulating epithelial-mesenchymal transition

Figure 1 Bioinformatics and prognosis analyses of transmembrane channel-like 5 expression. A and B: The expression profile of transmembrane channel-like 5 (TMC5) in different cancers and normal tissues was determined using GENT2; C: Pan-cancer analysis of TMC5 mRNA expression using the GEPIA database; D: Expression of TMC5 between hepatocellular carcinoma (HCC) and paired normal liver tissues based on the GEPIA database; E: Immunohistochemistry showing TMC5 expression in normal liver tissues from The Human Protein Atlas database; F: Overall survival curve of TMC5 in HCC patients in GEPIA dataset; G: Construction and validation of the nomogram based on TMC5 expression and clinical characteristics (gender, age, stage, T stage, N stage, and M stage); H: Calibration analysis based on the nomogram. ^aP < 0.05; ^bP < 0.01. TMC5: Transmembrane channel-like 5.

Figure 2 Expression of transmembrane channel-like 5 in hepatocellular carcinoma tissue and cell lines. A: Immunohistochemistry analysis of transmembrane channel-like 5 (TMC5) expression in Hepatocellular carcinoma tumor and paracancerous tissues (n = 10). Original magnification, 20 ×, scale bar = 100 μm; B: RT-qPCR was performed to detect the expression of TMC5 in LO2, MHCC-97H and MHCC97-LM3 (n = 3). HCC: Hepatocellular carcinoma; IHC: Immunohistochemistry; TMC5: Transmembrane channel-like 5. ^aP < 0.01.

Figure 3 Transmembrane channel-like 5 promotes cell invasion, migration and proliferation. A: MHCC97-LM3 were transfected with three short hairpin RNAs (shRNAs) lentivirus, and MHCC-97H were transfected with transmembrane channel-like 5 (TMC5) overexpression lentivirus, conformed by qRT-PCR and western blot (n = 3); B: Colony formation formation assay of MHCC97-LM3 was transfected with three shRNAs lentivirus, MHCC-97H were transfected with TMC5 overexpression lentivirus (n = 3), scale bar = 500 μm; C: Transwell invasion assay of MHCC97-LM3 was transfected with three shRNAs lentivirus, MHCC-97H was transfected with TMC5 overexpression lentivirus, scale bar = 500 μm (n = 3); D: Wound healing assay of MHCC97-LM3 was transfected with three shRNAs lentivirus, MHCC-97H was transfected with TMC5 overexpression lentivirus (n = 3), scale bar = 200 μm. ^aP < 0.05; ^bP < 0.01. TMC5: Transmembrane channel-like 5; shNC: Short hairpin negative control; shRNA: Short hairpin RNA; OV: Overexpression lentivirus.

Figure 4 Protein expression of epithelial-mesenchymal transition-related markers was detected by Western blotting. Band intensity analysis was performed MHCC97-LM3 cells were transfected with short hairpin negative control and short hairpin RNAs lentivirus, MHCC-97H transfected with overexpression transmembrane channel-like 5 lentivirus (n = 3). ^aP < 0.05; ^bP < 0.01. shNC: Short hairpin negative control; shRNA: Short hairpin RNA; OV: Overexpression lentivirus.

Figure 5 Tumor growth and metastasis were examined by IVIS imaging system and quantified by bioluminescence intensity, and representative images were shown (n = 6). shNC: Short hairpin negative control; shRNA: Short hairpin RNA; OV: Overexpression lentivirus. ^aP < 0.01.

Figure 6 Mouse liver was collected and immunohistochemistry analysis of snail, E-cadherin, N-cadherin and ZO-1, and representative images were shown (n = 6). Original magnification, 20 ×, scale bar = 100 μm. ^aP < 0.05; ^bP < 0.01. IHC: Immunohistochemistry; shNC: Short hairpin negative control; shRNA: Short hairpin RNA; OV: Overexpression lentivirus.