Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Promoter activity profiles in cancer‐associated fibroblasts and normal fibroblasts. (A) Average expression levels ( X ‐axis, CPM in log 2 scale) and fold changes ( Y ‐axis, log 2 scale) between CAFs and normal fibroblasts. Differentially expressed promoters with statistical significance (FDR < 1%) are shown in red. The most active promoters among COL1A1 , ITGA11, and COL11A1 are indicated by crossed dots; +, x, and asterisk;* respectively. (B) Heatmap demonstrating the individual promoter activity of the differentially expressed promoters (FDR < 1%); those with a certain level of expression (more than two CPM on average) and difference of expression (more than 2‐fold change) are shown. The hierarchical clustering across promoters and samples is based on Pearson's correlation coefficient with complete linkage method. ( n = 16; pairs of control and corresponding CAF).

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Activity Assay, Expressing, Control

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Immunohistochemical expression of ITGA11 and COL11A1 in cancer stroma (A) Immunohistochemical staining of human NSCLC tissues; representative sections of Adeno, Sq, and normal lung with ITGA11 and COL11A1 expressions are shown. Original magnification: 100×; Scale bars: 200 μm. (B) ITGA11 and (C) COL11A1 expression levels in each subject. (B, C; n = 41, one‐way ANOVA) Association between the recurrence and ITGA11 expression in (D) cancer stroma and (E) cancer epithelium. (D, E; n = 25, Mann–Whitney) Association between the pathological staging and ITGA11 expression in (F) cancer stroma and (G) cancer epithelium. (F, G; n = 38, one‐way ANOVA) Vertical axis: H ‐score values (see Section ). Each symbol represents one patient. The values represent the mean ± SD. * P < 0.05, *** P < 0.001, **** P < 0.0001. Adeno, Adenocarcinoma; Sq, squamous cell carcinoma.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Immunohistochemical staining, Expressing, Staining, MANN-WHITNEY

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Correlation between ITGA11 and COL11A1 expression levels in cancer‐associated fibroblasts. r , correlation coefficient.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Expressing, Immunostaining, Western Blot

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Cancer‐associated fibroblast migration and expression of ITGA11, integrin α5, integrin β1, and COL11A1. Sub‐confluent fibroblasts from 16 pairs of control fibroblasts and CAFs were cultured, and (A) proteins were extracted and analyzed by western blot to detect (B) ITGA11, (C) COL11A1, (D) integrin α5, and (E) integrin β1 expression. (B‐E; n = 16, Wilcoxon) Vertical axis: expression of protein normalized to β‐actin. (F) After sub‐confluent fibroblasts, from 16 pairs of control fibroblasts and CAFs, were cultured, chemotactic activity toward fibronectin (20 µg·mL −1 ) was measured. (F; n = 16, Mann–Whitney) Vertical axis: number of migrated cells per 5HPF. Filled symbols represent lung adenocarcinoma. Open symbols represent squamous cell lung cancer. (G) Comparison of CAFs and control fibroblasts from the same donor. (G; n = 16, Wilcoxon) Vertical axis: number of migrated cells per 5HPF. Filled symbols represent lung adenocarcinoma. Open symbols represent squamous cell lung cancer. (H) Relationship between fold CAF migration toward fibronectin (20 µg·mL −1 ) divided by corresponding control fibroblast migration and fold CAF expression of ITGA11 divided by corresponding control fibroblast. Vertical axis: fold CAF migration compared to control fibroblasts. Horizontal axis: fold ITGA11 expression in CAFs compared to control fibroblasts. (H; n = 16, Spearman) (I) Relationship between fold CAF migration toward fibronectin (20 µg·mL −1 ) divided by corresponding control fibroblast migration, and fold CAF expression of COL11A1 divided by corresponding control fibroblast. Vertical axis: fold CAF migration compared to control fibroblasts. Horizontal axis: fold COL11A1 expression in CAFs compared to control fibroblasts. (I; n = 16, Spearman) (J) After sub‐confluent patients’ control fibroblasts and CAFs were cultured, migration activity toward collagen type I (1 µg·mL −1 ) was measured. (J; n = 9, Mann–Whitney) Vertical axis: number of migrated cells per 5HPF. Filled symbols represent lung adenocarcinoma. Open symbols represent squamous cell lung cancer. Each symbol represents one patient. The values represent the mean ± SD. Pts: patients. ** P < 0.01, *** P < 0.001.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Migration, Expressing, Control, Cell Culture, Western Blot, Activity Assay, MANN-WHITNEY, Comparison

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Response of cancer‐associated fibroblasts to TGF‐β1 and fibronectin. Sub‐confluent HFL‐1 were cultured and treated over different times and with different doses of (A) TGF‐β1 and (B) fibronectin. Proteins were extracted and subjected to western blot analysis to detect the time course of (C, D) ITGA11 and (E, F) COL11A1 expression level. (C‐F; n = 3, unpaired Student's t ‐test) Vertical axis: expression of protein normalized to β‐actin. (G) Effect of TGF‐β1 on CAF fibronectin production compared to control fibroblasts. (G; n = 9, unpaired Student's t ‐test) Vertical axis: fibronectin production expressed as the amount per 10 5 cells mL −1 . (H) Effect of TGF‐β1 in CAF migration toward fibronectin (20 µg·mL −1 ) compared to control fibroblasts. (H; n = 7, unpaired Student's t ‐test) Vertical axis: number of migrated cells per 5HPF. Each symbol represents one patient. The values represent the mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Cell Culture, Western Blot, Expressing, Control, Migration

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Regulation of ERK1/2 signal activation in cancer‐associated fibroblasts. (A) Sub‐confluent fibroblasts from 16 pairs of control fibroblasts and CAFs were cultured, and proteins were extracted and subjected to western blot analysis to detect (B) p‐ERK1/2 expression levels. Vertical axis: expression of protein normalized to t‐ERK1/2. (B; n = 16, Wilcoxon) (C) Effect of various concentrations of ERK inhibitor CAS 1049738‐54‐6, with or without TGF‐β1 (10 p m ), on HFL‐1 cells was assessed with migration assay toward fibronectin (20 µg·mL −1 ). (C; n = 3, one‐way ANOVA) Vertical axis: number of migrated cells per 5HPF. (D) Effect of various concentrations of CAS 1049738‐54‐6, with or without TGF‐β1 (10 p m ), on HFL‐1 cells was assessed with migration assay toward collagen type I (1 µg·mL −1 ). (D; n = 3, one‐way ANOVA) Vertical axis: number of migrated cells per 5HPF. Sub‐confluent HFL‐1 cells were cultured and treated with or without TGF‐β1 (10 p m ), and CAS 1049738‐54‐6 (10 μ m ) for 24 h. Proteins were extracted and subjected to western blot analysis to detect the (E) ITGA11, (F) COL11A1, and (G) fibronectin expression levels. (E, F, G; n ≥ 3, unpaired Student's t ‐test) Vertical axis: expression of protein normalized to β‐actin. Each symbol represents one patient. The values represent the mean ± SD. Pts: patients. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Activation Assay, Control, Cell Culture, Western Blot, Expressing, Migration

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Phenotypic alteration of lung fibroblasts via fibroblast–lung cancer cell interaction. The effect of conditioned media on (A) ITGA11, (B) COL11A1, (C) integrin α5, and (D) integrin β1 expression levels in HFL‐1 cells. The conditioned media were harvested from BEAS‐2B as the normal epithelial cells, A549 as the lung adenocarcinoma cell line, and HFL‐1 as the negative control (see Section ). (A‐D; n = 3, unpaired Student’s t ‐test) Vertical axis: expression of protein normalized to β‐actin. (E) Sub‐confluent control fibroblasts and CAFs were cultured and the supernatant was harvested after being incubated for 48 h. Subsequently, the migration ability of A549 cells toward supernatant from CAFs was compared to that of the control fibroblasts. (E; n = 6, Wilcoxon) Vertical axis: number of migrated cells per 5HPF. Each symbol represents one patient. The values represent the mean ± SD. * P < 0.05, ** P < 0.01.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Expressing, Negative Control, Control, Cell Culture, Incubation, Migration

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Effect of ITGA11 genetic modification on fibroblast features. (A) ITGA11 was knocked down using siRNA in HFL‐1 and proteins were extracted and subjected to western blot analysis to detect the expression levels of each target (see Section ). Vertical axis: (B) ITGA11, (C) COL11A1, (D) integrin α5, (E) integrin β1, (F) fibronectin, and (G) α‐SMA protein expression normalized to β‐actin, (H) p‐ERK1/2 protein expression normalized to t‐ERK1/2 and (I) p‐SMAD3 protein expression normalized to t‐SMAD3. (J) Migration toward fibronectin (20 µg·mL −1 ) was assessed after silencing ITGA11 . Vertical axis: number of migrated cells per 5HPF. (K) Migration toward collagen type I (1 µg·mL −1 ) was assessed after silencing ITGA11 . (B‐K; n ≥ 3, one‐way ANOVA) Vertical axis: number of migrated cells per 5HPF. (L) NIH 3T3 cells were transfected with ITGA11 ‐overexpression vector or GFP and subjected to western blot analysis to detect the ITGA11 and α‐SMA expression levels (see Section ). (M) Migration toward fibronectin (20 µg·mL −1 ) was assessed with ITGA11 ‐overexpressing NIH 3T3 cells. (M; n = 8, Mann–Whitney) Vertical axis: number of migrated cells per 5HPF. The values represent the mean ± SD. * P < 0.05, ** P < 0.01, **** P < 0.0001.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Modification, Western Blot, Expressing, Migration, Transfection, Over Expression, Plasmid Preparation, MANN-WHITNEY

Journal: Molecular Oncology

Article Title: Cancer‐associated fibroblast migration in non‐small cell lung cancers is modulated by increased integrin α11 expression

doi: 10.1002/1878-0261.12937

Figure Lengend Snippet: Schematic illustration of acquired features of the phenotype of cancer‐associated fibroblasts. TGF‐β1‐stimulated fibronectin and collagen type I production induce ITGA11 expression in fibroblasts, resulting in accelerated CAF‐mediated migration toward fibronectin and collagen type I via ERK1/2 signal without changing integrin α5 β1 expressions. The released fibronectin induced COL11A1 production, which was accompanied by ITGA11 expression via ERK1/2 signal in cancer stroma.

Article Snippet: Recombinant human COL11A1 (catalog no.MBS718449_COA) was from MyBioSource (San Diego, CA, USA).

Techniques: Expressing, Migration

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: The amino acid sequence of the COL11A1 Fusion Protein (residues 1545 to 1806 of the P12107-1 A isoform), recombinantly expressed in Escherichia coli , after the removal of an N-terminal GST tag (provided by Proteintech). The first 19 N-terminal (1545) PLPILSSKKTRRHTEGMQA (1563) amino acid residues are part of the putative C-telopeptide. The next 243 amino acid residues (1564 to 1806) constitute the C-propeptide.

Article Snippet: The recombinant COL11A1 Fusion Protein from Proteintech was also assayed in an ELISA and Western blot with finally purified preparations of the anti-colXIα1 clone 9 and PLY-7 mAbs. shows their ELISA immunoreactivity characteristics.

Techniques: Sequencing

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: SDS-PAGE gel staining and Western blot of recombinant antigens with finally purified preparations of anti-colXIα1 clone 9 and PLY-7 mAbs. Lane 1: PageRuler™ Plus Prestained Protein Ladder. Lane 2: COL11A1 Fusion Protein (Proteintech). Lane 3: Collagen XIα1 (GenScript). Lane 4: C-propeptide (GenScript). Western blot color development was monitored following the manufacturer’s instructions. Full-length blots/gels are presented in .

Article Snippet: The recombinant COL11A1 Fusion Protein from Proteintech was also assayed in an ELISA and Western blot with finally purified preparations of the anti-colXIα1 clone 9 and PLY-7 mAbs. shows their ELISA immunoreactivity characteristics.

Techniques: SDS Page, Staining, Western Blot, Recombinant, Purification

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: The PEP-FOLD4-derived structural predictions of peptides related to the putative C-telopeptide. ( A ): The 50 N-terminal amino acid sequence of the COL11A1 Fusion Protein from Proteintech, with the first 19 N-terminal PLPILSSKKTRRHTEGMQA amino acid residues of the putative C-telopeptide. ( B ): A free RRHTEGMQA peptide. ( C ): The 50 C-terminal amino acid sequence of GenScript’s recombinant collagen XIα1 form, whose last 21 C-terminal IQPLPILSSKKTRRHTEGMQA amino acid residues correspond to the putative C-telopeptide. The peptide’s N-terminus is on the left in ( A , B ) and on the right in ( C ).

Article Snippet: The recombinant COL11A1 Fusion Protein from Proteintech was also assayed in an ELISA and Western blot with finally purified preparations of the anti-colXIα1 clone 9 and PLY-7 mAbs. shows their ELISA immunoreactivity characteristics.

Techniques: Derivative Assay, Sequencing, Recombinant

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: The amino acid sequence of the COL11A1 Fusion Protein (residues 1545 to 1806 of the P12107-1 A isoform), recombinantly expressed in Escherichia coli , after the removal of an N-terminal GST tag (provided by Proteintech). The first 19 N-terminal (1545) PLPILSSKKTRRHTEGMQA (1563) amino acid residues are part of the putative C-telopeptide. The next 243 amino acid residues (1564 to 1806) constitute the C-propeptide.

Article Snippet: The COL11A1 Fusion Protein, with an N-terminal GST tag, from ChromoTek GmbH and Proteintech, Germany (Cat. No. Ag16509), was recombinantly expressed in Escherichia coli .

Techniques: Sequencing

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: The PEP-FOLD4-derived structural predictions of peptides related to the putative C-telopeptide. ( A ): The 50 N-terminal amino acid sequence of the COL11A1 Fusion Protein from Proteintech, with the first 19 N-terminal PLPILSSKKTRRHTEGMQA amino acid residues of the putative C-telopeptide. ( B ): A free RRHTEGMQA peptide. ( C ): The 50 C-terminal amino acid sequence of GenScript’s recombinant collagen XIα1 form, whose last 21 C-terminal IQPLPILSSKKTRRHTEGMQA amino acid residues correspond to the putative C-telopeptide. The peptide’s N-terminus is on the left in ( A , B ) and on the right in ( C ).

Article Snippet: The COL11A1 Fusion Protein, with an N-terminal GST tag, from ChromoTek GmbH and Proteintech, Germany (Cat. No. Ag16509), was recombinantly expressed in Escherichia coli .

Techniques: Derivative Assay, Sequencing, Recombinant