Journal: Frontiers in Genetics

Article Title: BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC

doi: 10.3389/fgene.2022.875617

Figure Lengend Snippet: Association between the clinicopathological variables and BTC expression in 38 OSCC patients.

Article Snippet: The OSCC cell lines SCC4 and CAL27 were obtained from the Center for Molecular Medicine, Xiangya Hospital, Central South University (Changsha, China).

Techniques: Expressing, Virus, Infection

Journal: Frontiers in Genetics

Article Title: BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC

doi: 10.3389/fgene.2022.875617

Figure Lengend Snippet: DEGs in OSCC. (A,B) Volcano plots were constructed using FC values and FDRs. The red points in the plot represent the overexpressed mRNAs, and the blue points indicate the downregulated mRNAs with statistical significance. (A) DEGs between normal and tumor tissues. (B) DEGs between metastasis-positive and metastasis-negative tumor tissues. (C) Hierarchical clustering analysis of mRNAs that were differentially expressed between metastasis-negative and metastasis-positive tissues. The normalized expression levels in the heatmaps are colored from blue to red in ascending order. (D) Multivariate Cox regression analysis according to gene expression. (E) Kaplan–Meier survival curves were performed to show the prognosis of patients with high and low expression of BTC through the analysis of the mRNA expression profile data of 260 OSCC tumor samples from TCGA database.

Article Snippet: The OSCC cell lines SCC4 and CAL27 were obtained from the Center for Molecular Medicine, Xiangya Hospital, Central South University (Changsha, China).

Techniques: Construct, Expressing, Gene Expression

Journal: Frontiers in Genetics

Article Title: BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC

doi: 10.3389/fgene.2022.875617

Figure Lengend Snippet: BTC expression is correlated with clinicopathological parameters in OSCC patients. (A) Representative images of immunohistochemical staining and (B) immunoreactive score of BTC in human normal mucosa samples, OSCC tissue samples, and metastatic LN samples. The experiment was repeated three times independently. Results are shown as mean ± SD. T-test, n = 38. (C) mRNA expression of BTC in normal and tumor tissues was detected by RT-PCR. (D) Kaplan–Meier survival curves of 38 patients from our department. (E–L) Analysis of 330 OSCC samples from TCGA database and 32 pairs of OSCC samples selected from TCGA database showed the comparison of (E,F) BTC expressed in tumor tissues and normal tissues. Correlation analysis with BTC expression and (G) age, (H) histological grade, (I) sex, (J) T category, (K) tumor stage, and (L) LN metastasis status. * p < 0.05. ** p < 0.01. *** p < 0.001. **** p < 0.0001.

Article Snippet: The OSCC cell lines SCC4 and CAL27 were obtained from the Center for Molecular Medicine, Xiangya Hospital, Central South University (Changsha, China).

Techniques: Expressing, Immunohistochemical staining, Staining, Reverse Transcription Polymerase Chain Reaction, Comparison

Journal: Frontiers in Genetics

Article Title: BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC

doi: 10.3389/fgene.2022.875617

Figure Lengend Snippet: Statistical analyses of clinicopathological features associated with survival in 38 OSCC patients with the multivariate Cox proportional hazards models.

Article Snippet: The OSCC cell lines SCC4 and CAL27 were obtained from the Center for Molecular Medicine, Xiangya Hospital, Central South University (Changsha, China).

Techniques: Expressing

Journal: Frontiers in Genetics

Article Title: BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC

doi: 10.3389/fgene.2022.875617

Figure Lengend Snippet: Overexpression of BTC inhibits the proliferation, migration, and invasion of OSCC cell lines. (A) Cancer cell transfectants of the BTC-expressing vector and empty vector control were identified in SCC4 and CAL27 cells by Western blot. (B,C) Overexpression of BTC inhibited cell proliferation, as indicated by the CCK-8 assay, in CAL27 and SCC4 cells. (D) Wound healing assay showed that overexpression of BTC inhibited CAL27 and SCC4 cell migration. (E,F) Transwell assays showed that the migration and invasion abilities of CAL27 and SCC4 cells were impaired after overexpression of BTC.

Article Snippet: The OSCC cell lines SCC4 and CAL27 were obtained from the Center for Molecular Medicine, Xiangya Hospital, Central South University (Changsha, China).

Techniques: Over Expression, Migration, Expressing, Plasmid Preparation, Control, Western Blot, CCK-8 Assay, Wound Healing Assay

Journal: Frontiers in Genetics

Article Title: BTC as a Novel Biomarker Contributing to EMT via the PI3K-AKT Pathway in OSCC

doi: 10.3389/fgene.2022.875617

Figure Lengend Snippet: (A) Comparison of EMT scores in normal and tumor (in BTC low- and high-expression groups) tissue. (B) TCGA and GSEA showed highly regulated genes in patients with high-BTC expression versus those with low-BTC expression. (C) Heatmap of EMT marker expression in the BTC high- and low-expression groups. (D–F) Relationship between BTC and EMT markers, such as E-cadherin, N-cadherin, and vimentin. (G) PPI network analysis and Western blot analysis. The PPI network of the DEGs was constructed using STRING. The network nodes represent different proteins. The edges represent protein–protein associations, and the line thickness indicates the strength of the supporting data. (H) Protein expression level of EMT-related markers and the PI3K-AKT signaling pathway after overexpression of BTC in SCC4 and Cal27 cells.

Article Snippet: The OSCC cell lines SCC4 and CAL27 were obtained from the Center for Molecular Medicine, Xiangya Hospital, Central South University (Changsha, China).

Techniques: Comparison, Expressing, Marker, Western Blot, Construct, Over Expression

Journal: Cancers

Article Title: WISP-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the miR-153-3p/Snail Axis

doi: 10.3390/cancers11121903

Figure Lengend Snippet: WISP-1 induces epithelial-to-mesenchymal transition (EMT) functioning via E-cadherin downregulation. ( A , B ) SCC4 cells were treated with different concentrations of WISP-1 (0–30 ng/mL) for 24 h; cell migration and cell-scattering phenotype were analyzed by wound healing and scatter assays, respectively. Images of living cells were captured by microscope and the width of the gap in each scratch was measured by ImageJ software. ( C , D ) Cells were incubated with different concentrations of WISP-1 (0–30 ng/mL) for 24 h and EMT marker expression was evaluated by qPCR and Western blot assay. The α-tubulin content was used to normalize for levels of E-cadherin protein. ( E ) E-cadherin mRNA expression in tumor tissue and adjacent normal tissue was determined in TCGA database records. Mean levels (ranges) of mRNA expression in normal and tumor samples, respectively: 10,660 (2858.6–17,763.1), 9095 (4.929–18,812.1); log 2 (fold-change): –0.229. Results are expressed as the mean ± SEM. * p < 0.05 compared with the control group.

Article Snippet: The human OSCC cell line SCC4 was purchased from the Bioresource Collection and Research Center (BCRC; Hsinchu, Taiwan).

Techniques: Migration, Microscopy, Software, Incubation, Marker, Expressing, Western Blot, Control

Journal: Cancers

Article Title: WISP-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the miR-153-3p/Snail Axis

doi: 10.3390/cancers11121903

Figure Lengend Snippet: WISP-1 stimulates the EMT process via the integrin αvβ3 receptor in oral squamous cell carcinoma (OSCC) cells. ( A , B ) SCC4 cells were pretreated with Arg–Gly–Asp (RGD) (100 nM) or Arg–Ala–Asp (RAD) (100 nM) for 30 min, then stimulated with WISP-1 for 24 h. E-cadherin expression was examined by IF and qPCR assays. ( C ) Cells were pretreated with RGD or RAD for 30 min then stimulated with WISP-1 for 24 h, and cell migration was examined by the wound healing assay. Results are expressed as the mean ± SEM. * p < 0.05 compared with controls; # p < 0.05 compared with the WISP-1-treated group.

Article Snippet: The human OSCC cell line SCC4 was purchased from the Bioresource Collection and Research Center (BCRC; Hsinchu, Taiwan).

Techniques: Expressing, Migration, Wound Healing Assay

Journal: Cancers

Article Title: WISP-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the miR-153-3p/Snail Axis

doi: 10.3390/cancers11121903

Figure Lengend Snippet: Involvement of FAK and ILK in WISP-1-regulated EMT functioning. ( A , B ) SCC4 cells were incubated with WISP-1 (30 ng/mL) for the indicated time intervals; FAK and ILK activation was examined by Western blot assay. ( C , D ) Cells were pretreated with RGD (100 nM) for 30 min then treated with WISP-1 for 10 min; FAK and ILK activation was examined by Western blot assay. FAK and GSK3β content was used to normalize for pFAK and pGSK3βlevels. ( E , F ) Cells were pretreated for 30 min with a FAKi (10 M) and KP392 (10 M), prior to incubation with WISP-1 for 24 h. E-cadherin expression was examined by IF and qPCR assays. ( G ) Cells were pretreated for 30 min with a FAKi (10 M) and KP392 (10 M), then incubated with WISP-1 for 24 h. Cell migration was examined by the wound healing assay. Results are expressed as the mean ± SEM. * p < 0.05 compared with controls; # p < 0.05 compared with the WISP-1-treated group.

Article Snippet: The human OSCC cell line SCC4 was purchased from the Bioresource Collection and Research Center (BCRC; Hsinchu, Taiwan).

Techniques: Incubation, Activation Assay, Western Blot, Expressing, Migration, Wound Healing Assay

Journal: Cancers

Article Title: WISP-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the miR-153-3p/Snail Axis

doi: 10.3390/cancers11121903

Figure Lengend Snippet: WISP-1 induces Akt phosphorylation via the integrin αvβ3/FAK/ILK signaling pathway. ( A ) SCC4 cells were incubated with WISP-1 (30 ng/mL) for the indicated time intervals; levels of Akt phosphorylation were examined by Western blot assay. ( B , C ) Cells were pretreated with RGD (100 nM), a FAKi (10 M), or KP392 (10 M) for 30 min, then incubated with WISP-1 for 30 min. Akt activation was examined by Western blot assay. Akt protein was used to normalize for pAkt levels. ( D – F ) Cells were pretreated for 30 min with Akti (10 M), prior to incubation with WISP-1 for 24 h. E-cadherin expression was examined by IF and qPCR assays. Cell migration was examined by the wound healing assay. Results are expressed as the mean ± SEM. * p < 0.05 compared with the control group; # p < 0.05 compared with the WISP-1-treated group.

Article Snippet: The human OSCC cell line SCC4 was purchased from the Bioresource Collection and Research Center (BCRC; Hsinchu, Taiwan).

Techniques: Phospho-proteomics, Incubation, Western Blot, Activation Assay, Expressing, Migration, Wound Healing Assay, Control

Journal: Cancers

Article Title: WISP-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the miR-153-3p/Snail Axis

doi: 10.3390/cancers11121903

Figure Lengend Snippet: WISP-1 induces EMT function by Snail up-regulation in OSCC. ( A , B ) Cells were incubated with different concentrations of WISP-1 (0–30 ng/mL) for 24 h, then Snail and Twist expression were examined by qPCR and Western blot assays. The α-tubulin protein was used to normalize for levels of Snail and Twist. ( C , D ) Cells were pretreated for 30 min with RGD, FAKi, KP392, or Akti, prior to incubation with WISP-1 for 24 h. Snail translocation into the nucleus was examined by IF assay. Snail mRNA expression was examined by qPCR assay. ( E ) Cells were transfected with Snail siRNA (10 μM) or control siRNA (10 μM) for 24 h then stimulated with WISP-1 for 24 h; E-cadherin mRNA expression was examined using the qPCR assay. ( F ) Snail mRNA expression in tumor tissue and adjacent normal tissue was analyzed using records from the TCGA database. Mean levels (ranges) of Snail mRNA expression in normal and tumor samples, respectively: 6.064 (2.53–8.719), 6.555 (2.594–9.079); log 2 (fold-change): 0.491. Results are expressed as the mean ± SEM. * p < 0.05 compared with controls; # p < 0.05 compared with the WISP-1-treated group.

Article Snippet: The human OSCC cell line SCC4 was purchased from the Bioresource Collection and Research Center (BCRC; Hsinchu, Taiwan).

Techniques: Incubation, Expressing, Western Blot, Translocation Assay, Transfection, Control

Journal: Cancers

Article Title: WISP-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the miR-153-3p/Snail Axis

doi: 10.3390/cancers11121903

Figure Lengend Snippet: A schematic model depicting how WISP-1 regulates EMT functioning in OSCC cells. The modeling shows two potential signaling pathways underlying OSCC-derived WISP-1 regulation of EMT activation: (1) WISP-1 induces Snail expression via the integrin αvβ3/FAK/ILK/Akt signaling pathway, which in turn activates EMT functioning; (2) WISP-1 also inhibits miR-153-3p-induced downregulation of Snail mRNA translation and thus stimulates Snail protein expression and EMT activation. (RGD peptide: a target epitope of integrin αvβ3)

Article Snippet: The human OSCC cell line SCC4 was purchased from the Bioresource Collection and Research Center (BCRC; Hsinchu, Taiwan).

Techniques: Protein-Protein interactions, Derivative Assay, Activation Assay, Expressing

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: The expression of circ_0000140 in OSCC tissues and cells. a The expression of circ_0000140 in OSCC tissues (OSCC) and adjacent normal mucosal tissues (Normal) was detected by qRT-PCR. b The circ_0000140 expression in OSCC tissues with or without lymph node metastasis (Yes or No) was measured by qRT-PCR. c qRT-PCR was used to test the circ_0000140 expression in OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) and HOK cells. d , e The relative expression levels of circ_0000140 and KIAA0907 in CAL-27 and SCC-4 cells were assessed by qRT-PCR after treatment with RNase R. f , g The relative expression levels of circ_0000140, U6 and 18 s rRNA in the nuclear and cytoplasmic of CAL-27 and SCC-4 cells were detected by qRT-PCR. * P < 0.05, ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Expressing, Quantitative RT-PCR

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: Effects of circ_0000140 overexpression on the proliferation, migration and invasion of OSCC cells. CAL-27 and SCC-4 cells were transfected with circ_0000140 overexpression vector or vector. a The expression of circ_0000140 in CAL-27 and SCC-4 cells was detected by qRT-PCR to evaluate transfection efficiency. b Colony formation assay was performed to measure the number of colonies in CAL-27 and SCC-4 cells. c , d The number of migrated and invaded CAL-27 and SCC-4 cells was determined by transwell assay. e , f WB analysis was performed to detect the protein levels of ki67, MMP-2 and MMP-9 in CAL-27 and SCC-4 cells. ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Over Expression, Migration, Transfection, Plasmid Preparation, Expressing, Quantitative RT-PCR, Colony Assay, Transwell Assay

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: Effects of circ_0000140 overexpression on the glycolysis metabolism of OSCC cells. CAL-27 and SCC-4 cells were transfected with circ_0000140 overexpression vector or vector. a , b The ECAR of CAL-27 and SCC-4 cells was measured by Seahorse XF Extracellular Flux Analyzer. c The lactate acid level of CAL-27 and SCC-4 cells was tested by Lactate Assay Kit. d , e The protein levels of GLUT1 and LDHA in CAL-27 and SCC-4 cells were detected by WB analysis. ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Over Expression, Transfection, Plasmid Preparation, Lactate Assay

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: Circ_0000140 could absorb miR-182-5p. a The sequences of circ_0000140 containing the miR-182-5p binding sites or mutant binding sites were presented. b , c Dual-luciferase reporter assay was used to detect the interaction between circ_0000140 and miR-182-5p in CAL-27 and SCC-4 cell. d , e The enrichment of circ_0000140 and miR-182-5p in anti-Ago2 or anti-IgG was measured by the RIP assay. f qRT-PCR was performed to measure the expression of miR-182-5p in OSCC cells (CAL-27 and SCC-4) and NOK cells. g The expression of miR-182-5p in CAL-27 and SCC-43 cells was assessed by qRT-PCR to assess the effect of circ_0000140 overexpression on miR-182-5p expression. * P < 0.05, ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Binding Assay, Mutagenesis, Luciferase, Reporter Assay, Quantitative RT-PCR, Expressing, Over Expression

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: Effects of miR-182-5p mimic on the progression of OSCC cells. a The expression of miR-182-5p in CAL-27 and SCC-4 cells was measured by qRT-PCR to evaluate the transfection efficiency of miR-182-5p mimic. CAL-27 and SCC-4 cells were co-transfected with circ_0000140 overexpression vector and miR-182-5p mimic. b The number of colonies in CAL-27 and SCC-4 cells was detected by colony formation assay. c , d The number of migrated and invaded CAL-27 and SCC-4 cells was tested by transwell assay. e , f The protein levels of ki67, MMP-2 and MMP-9 in CAL-27 and SCC-4 cells were assessed by WB analysis. g Seahorse XF Extracellular Flux Analyzer was used to evaluate the ECAR of CAL-27 and SCC-4 cells. h The lactate acid level of CAL-27 and SCC-4 cells was determined by Lactate Assay Kit. i , j WB analysis was performed to measure the protein levels of GLUT1 and LDHA in CAL-27 and SCC-4 cells. ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Expressing, Quantitative RT-PCR, Transfection, Over Expression, Plasmid Preparation, Colony Assay, Transwell Assay, Lactate Assay

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: MiR-182-5p could target CDC73. a The sequences of CDC73 3′UTR containing the miR-182-5p binding sites or mutant binding sites were shown. b , c The interaction between miR-182-5p and CDC73 in CAL-27 and SCC-4 cells was assessed by dual-luciferase reporter assay. d The protein level of CDC73 in OSCC cells (CAL-27 and SCC-4) and HOK cells was determined by WB analysis. e WB analysis was used to measure the CDC73 protein level in CAL-27 and SCC-4 cells to evaluate the miR-182-5p expression on CRC73 expression. f , g The CDC73 protein level in CAL-27 and SCC-4 cells was detected by WB analysis to evaluate the circ_0000140 and miR-182-5p expression on CDC73 expression. ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Binding Assay, Mutagenesis, Luciferase, Reporter Assay, Expressing

Journal: Cancer Cell International

Article Title: Circ_0000140 restrains the proliferation, metastasis and glycolysis metabolism of oral squamous cell carcinoma through upregulating CDC73 via sponging miR-182-5p

doi: 10.1186/s12935-020-01501-7

Figure Lengend Snippet: Effects of CDC73 silencing on the progression of OSCC cells. a The protein level of CDC73 in CAL-27 and SCC-4 cells was measured by WB analysis to evaluate the transfection efficiency of si-CDC73. CAL-27 and SCC-4 cells were co-transfected with anti-miR-182-5p and si-CDC73. b Colony formation assay was performed to assess the number of colonies in CAL-27 and SCC-4 cells. c The number of migrated and invaded CAL-27 and SCC-4 cells was detected by transwell assay. d WB analysis was used to measure the protein levels of ki67, MMP-2 and MMP-9 in CAL-27 and SCC-4 cells. e , f Seahorse XF Extracellular Flux Analyzer was employed to assess the ECAR of CAL-27 and SCC-4 cells. g The lactate acid level of CAL-27 and SCC-4 cells was detected by Lactate Assay Kit. h The protein levels of GLUT1 and LDHA in CAL-27 and SCC-4 cells were tested by WB analysis. ** P < 0.01

Article Snippet: OSCC cell lines (CAL-27, SCC-4, SCC-9 and SCC-25) were bought from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Transfection, Colony Assay, Transwell Assay, Lactate Assay