Journal: Cell Communication and Signaling : CCS

Article Title: Integrated analysis reveals critical glycolytic regulators in hepatocellular carcinoma

doi: 10.1186/s12964-020-00539-4

Figure Lengend Snippet: OPN promotes the Warburg effect in HCC cells. a The knockdown efficiency of OPN in HCC-LM3 cells was measured by Western blotting and ELISA. b Effects of OPN knockdown on the glucose uptake and lactate production in HCC-LM3 cells ( n = 3). c The extracellular acidification rate (ECAR) in sh-OPN and sh-Ctrl HCC-LM3 cells was measured by Seahorse analyzer ( n = 5). d Effects of OPN blockade on the glucose uptake and lactate production in HCC-LM3 cells ( n = 3). e The overexpression efficiency of OPN in NIH3T3 cells and MEFs was measured by Western blotting. f Effects of OPN overexpression on the glucose uptake and lactate production in NIH3T3 cells and MEFs ( n = 3). g Effects of OPN overexpression on ECAR in NIH3T3 cells and MEFs were measured by Seahorse analyzer ( n = 5). * P < 0.05 and ** P < 0.01

Article Snippet: The human HCC cell lines (Huh7, HCC-LM3, SMCC-7721, MHCC-97H, Hep3B, and MHCC-97H), NIH3T3 cells, and MEFs were obtained from American Type Culture Collection (ATCC, UK).

Techniques: Knockdown, Western Blot, Enzyme-linked Immunosorbent Assay, Over Expression

Journal: Cell Communication and Signaling : CCS

Article Title: Integrated analysis reveals critical glycolytic regulators in hepatocellular carcinoma

doi: 10.1186/s12964-020-00539-4

Figure Lengend Snippet: Certification of the negative regulators of HCC glycolysis. a Western blotting showed the overexpression efficiency of SPP2, LECT2, SLC10A1, CYP3A4, HSD17B13, and IYD in Huh7 cells. b Real-time qPCR analysis showed the overexpression efficiency of SPP2, LECT2, SLC10A1, CYP3A4, HSD17B13, and IYD in Huh7 cells ( n = 3). c-e Measurement of SPP2, LECT2, SLC10A1, CYP3A4, HSD17B13, or IYD overexpression on the glucose utilization ( f , n = 3), lactate production ( g , n = 3) and ECAR ( h , n = 5) in Huh7 cells. * P < 0.05, ** P < 0.01, and *** P < 0.001

Article Snippet: The human HCC cell lines (Huh7, HCC-LM3, SMCC-7721, MHCC-97H, Hep3B, and MHCC-97H), NIH3T3 cells, and MEFs were obtained from American Type Culture Collection (ATCC, UK).

Techniques: Western Blot, Over Expression

Journal: Cell Communication and Signaling : CCS

Article Title: Integrated analysis reveals critical glycolytic regulators in hepatocellular carcinoma

doi: 10.1186/s12964-020-00539-4

Figure Lengend Snippet: Effects of glycolysis-related genes on HCC tumor growth. a Colony formation assay showed that OPN knockdown or blockade inhibits HCC-LM3 cell proliferation ( n = 3). b Colony formation assay for Huh3B cells treated with recombinant OPN protein ( n = 3). c Colony formation assay showed that SPP2, LECT2, SLC10A1, CYP3A4, HSD17B13, or IYD overexpression inhibits Huh7 cell proliferation ( n = 3). d The effects of glycolysis-related genes on HCC tumor growth in the presence or absence of 5 mM 2-DG ( n = 3). e In the culture medium containing 25 mM glucose or galactose, the effects of glycolysis-related genes on HCC tumor growth were analyzed by clonogenic assay. * P < 0.05 and ** P < 0.01

Article Snippet: The human HCC cell lines (Huh7, HCC-LM3, SMCC-7721, MHCC-97H, Hep3B, and MHCC-97H), NIH3T3 cells, and MEFs were obtained from American Type Culture Collection (ATCC, UK).

Techniques: Colony Assay, Knockdown, Recombinant, Over Expression, Clonogenic Assay

Journal: Cell Communication and Signaling : CCS

Article Title: Integrated analysis reveals critical glycolytic regulators in hepatocellular carcinoma

doi: 10.1186/s12964-020-00539-4

Figure Lengend Snippet: OPN promotes HCC glycolysis by modulating αvβ3-NF-κB signaling. a Blockade of integrin αvβ3 with Cilengitide inhibits glucose utilization ( n = 3), lactate production ( n = 3) and ECAR ( n = 5) in HCC-LM3 cells. b Western blotting analysis the signaling pathway influenced by OPN. c Glucose utilization and lactate production in Hep3B cells upon treatment with OPN recombinant protein and indicated pathway inhibitors ( n = 3). d Effect of OPN on the NF-κB activity in HCC cells ( n = 3). e Effect of CA-IKKβ on the glucose uptake and lactate production in OPN-silenced HCC-LM3 cells ( n = 3). * P < 0.05 and ** P < 0.01; ns: not significant

Article Snippet: The human HCC cell lines (Huh7, HCC-LM3, SMCC-7721, MHCC-97H, Hep3B, and MHCC-97H), NIH3T3 cells, and MEFs were obtained from American Type Culture Collection (ATCC, UK).

Techniques: Western Blot, Recombinant, Activity Assay

Journal: Cell Communication and Signaling : CCS

Article Title: Integrated analysis reveals critical glycolytic regulators in hepatocellular carcinoma

doi: 10.1186/s12964-020-00539-4

Figure Lengend Snippet: Inhibition of OPN-αvβ3 axis suppresses HCC tumor growth and glycolyis. a Tumor volume in sh-Ctrl and sh-OPN HCC-LM3 xenografts as indicated time point was measured ( n = 5). b Effect of Cilengitide treatment on the tumor growth of HCC-LM3 xenografts ( n = 5). c The lactate level in the tumor tissues from ( a ) and ( b ) was detected ( n = 5). d The expression of glycolytic genes in the tumor tissues from ( a ) and ( b ) was analyzed by real-time qPCR ( n = 5). e Hematoxylin and eosin staining in liver tissue samples from tumor-bearing WT and OPN-KO mice. f The expression of glycolytic genes in liver tissue samples from tumor-bearing WT and OPN-KO mice was analyzed by real-time qPCR ( n = 5). * P < 0.05 and ** P < 0.01

Article Snippet: The human HCC cell lines (Huh7, HCC-LM3, SMCC-7721, MHCC-97H, Hep3B, and MHCC-97H), NIH3T3 cells, and MEFs were obtained from American Type Culture Collection (ATCC, UK).

Techniques: Inhibition, Expressing, Staining

Journal: Cell Communication and Signaling : CCS

Article Title: Integrated analysis reveals critical glycolytic regulators in hepatocellular carcinoma

doi: 10.1186/s12964-020-00539-4

Figure Lengend Snippet: Expression pattern of OPN in clinical samples. a The expression of glycolytic genes in human HCC tissue samples with high OPN ( n = 10) and low OPN ( n = 20) expression was analyzed by real-time qPCR. b Representative photographs of OPN expression in HCC tumor tissues; scale bar: 50 μm. The correlation between OPN expression and the SUVmax value was analyzed. * P < 0.05 and ** P < 0.01

Article Snippet: The human HCC cell lines (Huh7, HCC-LM3, SMCC-7721, MHCC-97H, Hep3B, and MHCC-97H), NIH3T3 cells, and MEFs were obtained from American Type Culture Collection (ATCC, UK).

Techniques: Expressing

Journal: BMC Cancer

Article Title: NSUN5 accelerates the progression of liver hepatocellular carcinoma by m5C-EFNA3-mediated glycolysis

doi: 10.1186/s12885-025-14714-8

Figure Lengend Snippet: NSUN5 is highly expressed in LIHC. ( A ) The volcano plot showed differentially expressed genes in LIHC tumor tissues and adjacent normal tissues from TCGA database. The red dots indicated upregulated genes, while the blue dots indicated downregulated genes. ( B ) The heatmap showed the expression patterns of m5C methylation-related enzymes from differentially expressed genes in tumor and normal tissues. Blue: low expression; red: high expression. ( C ) The box plot showed the expression of NSUN5 in the primary tumor and normal samples. ( D ) The box plot showed the expression of NSUN5 at different cancer stages (normal, stages 1–4). ( E ) The box plot showed the expression of NSUN5 in different tumor grades (normal, grades 1–4). ( F ) The survival curve showed the survival rate of patients with LIHC in NSUN5 high and low/medium groups. ( G ) The expression of NSUN5 in normal liver epithelial cells (THLE3) and LIHC cell lines (Hep3B, SNU449, Huh7, HCCLM3, Focus, and HA22T). n = 3. * P < 0.05. ** P < 0.01. ns: no significance

Article Snippet: Normal liver epithelial cells (THLE3) and LIHC cell lines (Hep3B, SNU449, Focus, and HA22T) were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Expressing, Methylation

Journal: BMC Cancer

Article Title: NSUN5 accelerates the progression of liver hepatocellular carcinoma by m5C-EFNA3-mediated glycolysis

doi: 10.1186/s12885-025-14714-8

Figure Lengend Snippet: Knocking down NSUN5 inhibits glycolysis in LIHC cells. ( A ) Following shNSUN5 and shNC transfection, qPCR was performed to determine NSUN5 expression. SNU449 and Huh7 cells were transfected with shNC and shNSUN5, and cell phenotype was evaluated. ( B ) Cell viability was measured using CCK-8. ( C ) Glucose uptake was detected using a glucose uptake assay kit. ( D ) Lactate production was measured using a lactate assay kit. ( E ) ECAR was measured using Seahorse analysis. ( F ) HK2 and ( G ) PKM2 mRNA expression was detected using qPCR. n = 3. ** P < 0.01

Article Snippet: Normal liver epithelial cells (THLE3) and LIHC cell lines (Hep3B, SNU449, Focus, and HA22T) were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Transfection, Expressing, CCK-8 Assay, Lactate Assay

Journal: BMC Cancer

Article Title: NSUN5 accelerates the progression of liver hepatocellular carcinoma by m5C-EFNA3-mediated glycolysis

doi: 10.1186/s12885-025-14714-8

Figure Lengend Snippet: NSUN5 promotes m5C methylation of EFNA3. ( A ) The scatter plot showed the correlated genes with NSUN5. ( B ) The bubble plot showed the KEGG pathway enrichment analysis results of NSUN5-related genes. ( C ) The matrix heatmap showed the correlation between NSUN5 and genes that enriched in the glycolysis/gluconeogenesis pathway. ( D ) The box plot showed the expression of EFNA3 in LIHC primary tumor samples and normal samples. ( E ) The box plot showed the expression of EFNA3 at different cancer stages (normal, stages 1–4). ( F ) The box plot showed the expression of EFNA3 in different tumor grades (normal, grades 1–4). ( G ) The survival curve showed the survival rate of patients with LIHC in EFNA3 high and low/medium groups. ( H ) The expression of NSUN5 in normal liver epithelial cells (THLE3) and LIHC cell lines (SNU449 and Huh7). ( I ) After NSUN5 knockdown, EFNA3 expression was detected using qPCR. ( J ) EFNA3 expression was measured using MeRIP after NSUN5 knockdown. ( K ) Possible m5C modified sites in EFNA3. ( L ) The m5C sites in EFNA3 were verified using dual-luciferase reporter assay. ( M ) The effect of NSUN5 knockdown on EFNA3 mRNA stability when EFNA3 was wild type or mutated at site 92. n = 3. * P < 0.05. ** P < 0.01. ns: no significance

Article Snippet: Normal liver epithelial cells (THLE3) and LIHC cell lines (Hep3B, SNU449, Focus, and HA22T) were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Methylation, Expressing, Knockdown, Modification, Luciferase, Reporter Assay

Journal: BMC Cancer

Article Title: NSUN5 accelerates the progression of liver hepatocellular carcinoma by m5C-EFNA3-mediated glycolysis

doi: 10.1186/s12885-025-14714-8

Figure Lengend Snippet: Knocking down NSUN5 inhibits glycolysis in LIHC cells by reducing EFNA3 expression. ( A ) Following oeEFNA3 and oeNC transfection, qPCR was performed to determine EFNA3 expression. SNU449 and Huh7 cells were transfected with shNC, shNSUN5, shNSUN5 + oeNC, and shNSUN5 + oeEFNA3, and cell phenotype was evaluated. ( B ) Cell viability was measured using CCK-8. ( C ) Glucose uptake was detected using a glucose uptake assay kit. ( D ) Lactate production was measured using a lactate assay kit. ( E , F ) ECAR was measured using Seahorse analysis. ( G ) HK2 and ( H ) PKM2 mRNA expression was detected using qPCR. n = 3. ** P < 0.01

Article Snippet: Normal liver epithelial cells (THLE3) and LIHC cell lines (Hep3B, SNU449, Focus, and HA22T) were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Expressing, Transfection, CCK-8 Assay, Lactate Assay