Fig 2: Mycoplasma infection activated nuclear factor-?B (NF-?B) signaling to promote the migration and invasion of hepatocellular carcinoma (HCC) cells. (A) Western blot analysis for the expression of annexin A2 (ANXA2), p65, phosphorylated p65 subunit of NF-?B (p-p65), and p37 in HCC cells with mitochondrial transcription factor A (TFAM) knockdown. Tubulin was used as internal loading. M. Hyo, Mycoplasma hyorhinis. (B) Western blot analysis for the expression of TFAM, ANXA2, p65, p-p65, and p37 in HCC cells with mycoplasma infection, and ANXA2 was transiently knocked-down by siRNA against ANXA2. (C, D) Representative images and statistical analysis of migration and invasion assays for SNU-368 cells treated with the NF-?B inhibitor BAY 11-7082 (12.5 µM) for 12 h. (E, F) Representative images and statistical analysis of migration and invasion assays were carried out using SNU-368 cells 48 h after p65 knockdown (n = 3 independent experiments). **p < 0.01. Ctrl, control

Fig 3: Mitochondrial transcription factor A (TFAM) inhibited mycoplasma infection of hepatocellular carcinoma (HCC) cells by downregulating annexin A2 (ANXA2) expression. (A) Expression heatmap of RNA sequencing in TFAM knockdown and control SNU-368 cells (n = 3 per group). (B) Western blot analysis for the expression of TFAM, ANXA2, specificity protein 1 (Sp1), and p37 in HCC cells with TFAM knockdown. Tubulin was used as internal loading. M. Hyo, Mycoplasma hyorhinis. (C) Western blot analysis for the expression of ANXA2 and p37 in HCC cells 48 h after ANXA2 knockdown. Tubulin was used as internal loading. (D) Quantitative PCR (qPCR) analysis for the mRNA expression of ANXA2 and p37 in HCC cells with ANXA2 knockdown. Statistical difference was determined by t-test. *p < 0.05; **p < 0.01. (E) Representative immunohistochemical staining of TFAM and ANXA2 in human HCC tissues. Scale bar, 50 µm. (F) Spearman's correlation analysis between the TFAM and ANXA2 expression level based on the immunohistochemical (IHC) score. (G) qPCR analysis for the mRNA expression of TFAM and Sp1 in HCC cells with TFAM knockdown. Statistical difference was determined by t-test. *p < 0.05; **p < 0.01. (H) Western blot analysis for the expression of Sp1 in HCC cells with TFAM knockdown. Tubulin was used as internal loading. (I) Western blot analysis for the expression of ANXA2 and Sp1 in HCC cells with Sp1 knockdown. Tubulin was used as internal loading. (J, K) Representative confocal microscopy images and statistical analysis of staining with MitoSOX (5 µM) in HCC cells. Scale bar, 40 µm. (L) Western blot analysis for the expression of Sp1 and ANXA2 in HCC cells treated with mitochondrial ROS scavenger XJB-5-131 (10 µM). Ctrl, control; EPCAM, epithelial cellular adhesion molecule; LRP6, low-density lipoprotein receptor-related protein 6; TLR4, Toll-like receptor 4