Journal: Oncology Letters

Article Title: RNA methyltransferase NSUN2 enhances vasculogenic mimicry and malignant progression of cervical cancer through upregulation of MMP-9

doi: 10.3892/ol.2026.15518

Figure Lengend Snippet: MMP-9 , a factor that promotes Vasculogenic mimicry, is highly expressed in CC and is associated with poor prognosis. (A) CC database of TCGA was used to analyze key factors associated with VM. (B) Association of Sox2 expression with overall survival in CC (log-rank test). (C) Association of MMP-9 expression with overall survival in CC (log-rank test). (D) Panoramic scans after immunohistochemical detection of MMP-9 and H&E staining in samples from cancerous and paracancerous tissues from subjects with CC. Scale bar, 50 µm. Original magnification, ×20. (E) Protein levels of MMP-9 in 20 paired samples, with the MMP-9 level in CC tissue expressed compared with that in the paired normal tissue. (F) Expression levels of MMP-9 mRNA in 44 paired CC and paracancerous tissues, with MMP-9 expression in CC tissue expressed compared with that in the paired normal tissue. (G) Comparison of the average expression levels of MMP-9 mRNA in CC tissues compared with paracancerous tissues. (H) HeLa and SiHa cells were incubated under hypoxia (0.1% O 2 ) and proteins collected at 24, 48 and 72 h for western blotting of ALDH1, EPHA2, MMP-9 and GAPDH. ImageJ was used to semi-quantify western blotting signals from HeLa (I) and SiHa (J) cells. GAPDH served as an internal reference. *P<0.05, **P<0.01 and ***P<0.001. MMP-9, matrix metalloproteinase 9; VM, vasculogenic mimicry; ALDH1, aldehyde dehydrogenase 1; EPHA2, ephrin type-A receptor 2; TCGA, The Cancer Genome Atlas; Sox2, SRY-box transcription factor 2; CC, cervical cancer; CESC, cervical squamous cell carcinoma.

Article Snippet: The membranes were blocked with 5% milk at 20±5°C for 1 h. The blocked membranes were incubated at 4°C overnight with the following antibodies: A rabbit monoclonal anti-NSUN2 antibody (1:1,000; cat. no. AB259941; Abcam), a rabbit monoclonal anti-transfer RNA aspartic acid methyltransferase 1 (TRDMT1) antibody (1:1,000; cat. no. 19221-1-AP; Proteintech Group, Inc.; Wuhan Sanying Biotechnology), a rabbit polyclonal anti-MMP-9 antibody (1:1,000; cat. no. 10375-2-AP; Proteintech Group, Inc.; Wuhan Sanying Biotechnology), a rabbit polyclonal anti-aldehyde dehydrogenase 1 (ALDH1) antibody (1:1,000; cat. no. 15910-1-AP; Proteintech Group, Inc.; Wuhan Sanying Biotechnology), a rabbit polyclonal anti-ephrin type-A receptor 2 (EPHA2) antibody (1:1,000; cat. no. AF5 238; Affinity Biosciences) and a rabbit polyclonal anti-GAPDH antibody (1:1,000; TA309157 OriGene Technologies, Inc.).

Techniques: Expressing, Immunohistochemical staining, Staining, Comparison, Incubation, Western Blot

Journal: Oncology Letters

Article Title: RNA methyltransferase NSUN2 enhances vasculogenic mimicry and malignant progression of cervical cancer through upregulation of MMP-9

doi: 10.3892/ol.2026.15518

Figure Lengend Snippet: m 5 C RNA methyltransferase NUSN2 is significantly upregulated in CC and its gene expression is associated with poor prognosis. (A) The level of m 5 C modification of RNA in CC cells was analyzed using a dot blot assay. Methylene blue staining served as an internal reference. (B) Semi-quantitative analysis of dot blot results in HeLa cells. (C) Semi-quantitative analysis of dot blot results in SiHa cells. (D) Expression levels of genes encoding m 5 C methyltransferases was analyzed within a CC database of TCGA. (E) HeLa and SiHa cells were incubated under hypoxia (0.1% O 2 ) and proteins collected at 24, 48 and 72 h were analyzed for NSUN2, TRDMT1 and GAPDH by western blotting. ImageJ was used to quantify western blotting signals from (F) HeLa and (G) SiHa cells. GAPDH served as an internal reference. (H) mRNA expression level of NSUN2 in 44 pairs of CC and paracancerous tissues were quantified using RT-qPCR. Gene expression in cancer tissues is expressed compared with expression in normal tissues. (I) Comparison of the average expression level of NSUN2 mRNA in CC tissues compared with paracancerous tissues. (J) Panoramic scans after IHC detection of NSUN2 and H&E staining in CC and paracancerous tissues. Scale bar, 50 µm. Magnification, ×20. (K) Protein levels of NSUN2 in 20 paired CC and paracancerous tissues, with the NSUN2 level in CC tissue expressed compared with that in the paired normal tissue. (L) Level of NSUN2 expression in CC was associated with a poor prognosis in the form of biochemical progression-free survival (P=0.018; log-rank test). *P<0.05, **P<0.01 and ***P<0.001. MMP9, matrix metalloproteinase 9; VM, vasculogenic mimicry; CC, cervical cancer; IHC, immunohistochemistry; NSUN2, NOP2/Sun RNA methyltransferase 2; m 5 C, 5-methylcytidine; TRDMT1, transfer RNA aspartic acid methyltransferase 1; TCGA, The Cancer Genome Atlas; RT-qPCR, reverse transcription-quantitative PCR; IHC, immunohistochemistry.

Article Snippet: The membranes were blocked with 5% milk at 20±5°C for 1 h. The blocked membranes were incubated at 4°C overnight with the following antibodies: A rabbit monoclonal anti-NSUN2 antibody (1:1,000; cat. no. AB259941; Abcam), a rabbit monoclonal anti-transfer RNA aspartic acid methyltransferase 1 (TRDMT1) antibody (1:1,000; cat. no. 19221-1-AP; Proteintech Group, Inc.; Wuhan Sanying Biotechnology), a rabbit polyclonal anti-MMP-9 antibody (1:1,000; cat. no. 10375-2-AP; Proteintech Group, Inc.; Wuhan Sanying Biotechnology), a rabbit polyclonal anti-aldehyde dehydrogenase 1 (ALDH1) antibody (1:1,000; cat. no. 15910-1-AP; Proteintech Group, Inc.; Wuhan Sanying Biotechnology), a rabbit polyclonal anti-ephrin type-A receptor 2 (EPHA2) antibody (1:1,000; cat. no. AF5 238; Affinity Biosciences) and a rabbit polyclonal anti-GAPDH antibody (1:1,000; TA309157 OriGene Technologies, Inc.).

Techniques: Gene Expression, Modification, Dot Blot, Staining, Quantitative Dot Blot, Expressing, Incubation, Western Blot, Quantitative RT-PCR, Comparison, Immunohistochemistry, Reverse Transcription, Real-time Polymerase Chain Reaction