Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: Sequences of shRNAs targeting STMN1.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques: Sequencing, shRNA

Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: Primer sequences of genes analyzed for reverse transcription-quantitative PCR.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques:

Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: Clinicopathological characteristics of the included cases.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques: Expressing

Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: STMN1 expression differences and immunohistochemical staining for STMN1 and LVI. (A and B) STMN1 expression level was increased in HSCC tissues compared with normal hypopharyngeal tissues, and was further increased in HSCC tissues of patients with metastases. (C and D) STMN1 expression status in different HSCC samples (magnification, x200). (E and F) LVI statuses in different HSCC samples along with STMN1 expression statuses (magnification, x400). STMN1, stathmin1; LVI, lymphatic vessel invasion; HSCC, hypopharyngeal squamous cell carcinoma.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: The transfection efficacy evaluation of STMN1 knockdown and the results of cell functional experiments. (A and B) The relative mRNA and protein expression levels of STMN1 in different groups. All of them were evidently decreased after STMN1 knockdown with different shRNA sequences, compared with the NC shRNA group. (C-E) The OD 450 , the relative migratory ratio at 48 h, and the relative number of invasive cells were all significantly decreased after STMN1 knockdown with different shRNA sequences; all were compared with the NC shRNA group. The magnification of picture D and picture E were 100 times and 200 times, respectively. * P<0.05 and ** P<0.01. STMN1, stathmin1; shRNA, short hairpin RNA; NC, negative control.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques: Transfection, Knockdown, Functional Assay, Expressing, shRNA, Negative Control

Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: Bioinformatics analyses for target genes and pathways of STMN1. (A) STMN1 was evidently enriched in the HIF-1α pathway in head and neck squamous cell carcinoma. (B) Gene expression levels of HIF-1α pathway were different between cases with high expression of STMN1 and those with low expression of STMN1. (C and D) Target gene prediction analysis from website https://www.aclbi.com , which demonstrated that the expression level of MTA1 was significantly correlated with that of STMN1. STMN1, stathmin1; HIF-1α, hypoxia inducible factor-1alpha; MTA1, tumor metastasis-associated protein 1.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques: Gene Expression, Expressing

Journal: Molecular and Clinical Oncology

Article Title: Stathmin1 promotes lymph node metastasis in hypopharyngeal squamous cell carcinoma via regulation of HIF‑1α/VEGF‑A axis and MTA1 expression

doi: 10.3892/mco.2023.2617

Figure Lengend Snippet: The mRNA and protein expression levels of the target genes of STMN1 in FaDu cells. (A-C) The relative mRNA and protein expression levels of HIF-1α were signifiicantly decreased after STMN1 knockdown with different shRNA sequences. (D) The relative expression level of VEGF-A, the downstream protein of HIF-1α, was significantly decreased after STMN1 knockdown. (E and F) Both relative mRNA and protein expression levels of MTA1 significantly decreased after STMN1 knockdown. All were compared with the NC shRNA group. * P<0.05, ** P<0.01 and *** P<0.001. STMN1, stathmin1; HIF-1α, hypoxia inducible factor-1alpha; shRNA, short hairpin RNA; VEGF, vascular endothelial growth factor; MTA1, tumor metastasis-associated protein 1; NC, negative control.

Article Snippet: Equal amounts of proteins (30 μg) were added for 10% SDS-polyacrylamide gel electrophoresis and then transferred to polyvinylidene difluoride membrane (Pall Life Sciences), then blocked with 5% skimmed milk at room temperature for 1.5 h. Subsequently, membranes were incubated with the following primary antibodies against: STMN1 (1:1,000; cat. no. PB9560; Wuhan Boster Biological Technology, Ltd.), hypoxia inducible factor-1alpha (HIF-1α; 1:1,000; cat. no. A6265; ABclonal Biotech Co., Ltd.), vascular endothelial growth factor (VEGF)-A (1:1,000; cat. no. ab214424; Abcam), metastasis-associated protein 1 (MTA1; 1:1,000; cat. no. ab288765; Abcam) and β-actin (1:2,000; cat. no. CPA9066; Cohesion Biosciences) overnight at 4 ̊C, and then washed with Tris-buffered saline and Tween-20 (TBST, 0.01 M Tris, 0.15 M NaCl, 0.1% Tween-20, pH=7.4) for 30 min.

Techniques: Expressing, Knockdown, shRNA, Negative Control

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: SARS-COV-2 N protein interacts with UBC9. ( A ) pCDNA3.1 (3XFlag Tag) with N protein-coding genes and pEGFP-N1 (GFP Tag) with UBC9-coding genes were co-expressed in HEK293T cells. After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays. ( B ) pDsRED-mono-N1 (RFP Tag) with SARS-COV-2 N protein-coding genes and pEGFP-N1 (GFP Tag) with UBC9 or UBC9 C93A coding genes were expressed or co-expressed in HEK293T cells. After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cells were stained with DAPI before confocal microscopy imaging. Endogenous UBC9 in HEK293T cells was also detected by FITC-conjugated Goat Anti-Mouse IgG(H + L), which recognizes anti-UBC9 mouse antibodies. Scale bars: 10 μm. ( C ) pET28a (HIS 6 tag, Kan + ) with SARS-COV-2 N protein coding genes and pGEX-6P-1 (GST tag, Amp + ) with UBC9 or UBC9 C93A coding genes were co-expressed in BL21 (DE3) E. coli . cell lysates were subjected to Co-IP after IPTG (1 mM) induction for 6 h. ( D ) pCDNA3.1 (3XFlag Tag) with SARS-COV-2 N protein-coding genes was over-expressed in HEK293T cells. After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Transfection, Co-Immunoprecipitation Assay, Staining, Confocal Microscopy, Imaging

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: Human beta coronavirus N protein interacts with UBC9. ( A ) pET28a-N (HIS 6 tag, Kan + ) containing five human beta coronavirus N protein-coding genes and pGEX-UBC9 (GST tag, Amp + ) harboring the UBC9 coding gene were co-expressed in BL21 (DE3) E. coli Cell lysates were prepared and subjected to Co-IP following IPTG induction (1 mM) for 6 h. ( B ) pEGFP-N1 (GFP Tag) with three human beta coronavirus N protein-coding genes and pCDNA3.1 (3XFlag Tag) with the UBC9 coding gene were co-expressed in HEK293T cells. After a 24- h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Co-Immunoprecipitation Assay, Transfection

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: SARS-COV-2 N protein enhanced the molecular interaction between UBC9 and MAVS. ( A ) pEGFP-N1 (GFP Tag) with SARS-COV-2 N protein or UBC9 coding genes and pCDNA3.1 (3XFlag Tag) with MAVS coding gene were co-expressed in HEK293T cells. After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays. ( B ) Relative Ubc9 band intensity before or after IP: Flag in ( A ) were calculated by using ImageJ in triplet replicates. ns: no significant difference, *** p < 0.001. ( C ) Schematic diagram of wild-type SARS-CoV-2 N protein and truncated mutants. ( D ) pEGFP-N1 (GFP Tag) with UBC9 coding genes and pCDNA3.1 (3XFlag Tag) with SARS-COV-2 N protein or truncation coding gene were co-expressed in HEK293T cells. After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Transfection, Co-Immunoprecipitation Assay

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: The expression of UBC9 in HEK293T, Vero, Vero E6, Huh7, and HRT18 cells was detected by Western blotting.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Expressing, Western Blot

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: The interaction between the SARS-CoV-2 N protein and UBC9 inhibits MAVS ubiquitination by enhancing its SUMOylation. ( A , B ) pEGFP-N1 (GFP Tag) with SARS-COV-2 N coding genes or pCDNA3.1 with MAVS-Flag, SUMO3-HA, or K63-His6 coding genes were co-expressed in HEK293T cells (endogenous UBC9 highly expressed) or Vero E6 cells (endogenous UBC9 underexpressed). After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Ubiquitin Proteomics, Transfection, Co-Immunoprecipitation Assay

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: The interaction of N protein and UBC9 plays a crucial role in regulating the SUMOylation and ubiquitination modifications of MAVS. ( A ) pEGFP-N1 (GFP Tag) with SARS-COV-2 N coding genes or pCDNA3.1 with MAVS-Flag, SUMO3-HA or SUMO3 KR -HA, K63-His 6 coding genes were co-expressed in HEK293T cells (endogenous UBC9 was highly expressed). After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays. ( B ) pEGFP-N1 (GFP Tag) with SARS-COV-2 N coding genes or pCDNA3.1 with MAVS-Flag, SUMO3-HA, UBC9 or UBC9 C93A , K63-His 6 coding genes were co-expressed in Vero E6 cells (endogenous UBC9 underexpressed). After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays. ( C ) pEGFP-N1 (GFP Tag) with SARS-COV-2 N, N 44–419 or N 174–419 coding genes or pCDNA3.1 with MAVS-Flag, SUMO3-HA, and K63-His 6 coding genes were co-expressed in HEK293T cells (endogenous UBC9 highly expressed). After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Co-IP assays.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Ubiquitin Proteomics, Transfection, Co-Immunoprecipitation Assay

Journal: Viruses

Article Title: The Interaction between SARS-CoV-2 Nucleocapsid Protein and UBC9 Inhibits MAVS Ubiquitination by Enhancing Its SUMOylation

doi: 10.3390/v15122304

Figure Lengend Snippet: UBC9 plays a critical role in the process of impaired IFN I response caused by the in-teraction between the N protein and MAVS during virus infection. ( A , B ) pCDNA3.1 with SARS-CoV-2 N protein-coding genes was gradually increased in HEK293T cells or VERO E6 cells. After a 24-h lipofectamine transfection followed by 8 h of SeV stimulation, cell lysates were collected for Western blotting. ( C , D ) The relative band intensity of Western blotting results (a relative band quantification of phosphorylation protein to total protein) in ( A , B ) were calculated by using ImageJ in triplet replicates. ns: no significant difference, p > 0.05; *: p < 0.05; **: p < 0.01; ***: p < 0.001; ****: p < 0.0001.

Article Snippet: Subsequently, the transfected cells were exposed to Sev for an additional 8 h, followed by PBS washing and fixation with 4% paraformaldehyde in PBS for a duration of 20 min. Endogenous UBC9 in HEK293T cells was also detected by FITC–conjugated Goat Anti-Mouse IgG(H + L) (Proteintech, Cat No. SA00003-1), which recognizes anti-UBC9 mouse antibody (Proteintech, Cat No. 60201-1-Ig).

Techniques: Virus, Infection, Transfection, Western Blot, Phospho-proteomics

Journal: Scientific Reports

Article Title: The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema

doi: 10.1038/s41598-018-21790-3

Figure Lengend Snippet: MMP9, CD147, cathepsin B, p18 NFE2, p45 NFE2 and ADAMTSL-4 expression in ATII cells and lung tissue obtained from non-smokers (NS), smokers (SM) and patients with emphysema (E). MMP9, CD147 and cathepsin B expression in freshly isolated ATII cells ( A ) and lung tissue ( B ) as detected by Western blotting analysis. ( C ) MMP9, CD147, cathepsin B, p45 NFE2 and ADAMTSL-4 mRNA levels in lung tissue by RT-PCR. ( D,E ) p18 NFE2, p45 NFE2 and ADAMTSL-4 expression in freshly isolated ATII cells ( D ) and lung tissue ( E ). Protein levels were analyzed by Western blotting. Densitometric analysis is also shown. *Statistically significant difference ( p < 0.05) is shown for comparison between non-smokers and smokers, between smokers and emphysema patients and between non-smokers and emphysema patients. Data are shown as the mean (±s.e.m.).

Article Snippet: The following antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA): MMP9, CD147, ADAMTSL-4, p45 NFE2, p18 NFE2, DJ-1, phosphoserine, phosphothreonine and phosphotyrosine.

Techniques: Expressing, Isolation, Western Blot, Reverse Transcription Polymerase Chain Reaction

Journal: Scientific Reports

Article Title: The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema

doi: 10.1038/s41598-018-21790-3

Figure Lengend Snippet: p45 NFE2 and p18 NFE2 tyrosine phosphorylation in human ATII cells and lung tissue obtained from control non-smokers (NS) and smokers (SM) and patients with emphysema (E). Immunoprecipitation was performed in freshly isolated ATII cells ( A ) and lung tissue ( B ) followed by Western blotting analysis. Densitometric quantification is also shown. *Statistically significant difference ( p < 0.05). Data are shown as the mean (±s.e.m.).

Article Snippet: The following antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA): MMP9, CD147, ADAMTSL-4, p45 NFE2, p18 NFE2, DJ-1, phosphoserine, phosphothreonine and phosphotyrosine.

Techniques: Immunoprecipitation, Isolation, Western Blot

Journal: Scientific Reports

Article Title: The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema

doi: 10.1038/s41598-018-21790-3

Figure Lengend Snippet: NFE2 knockdown increases cell death induced by CSE in A549 cells in vitro . ( A ) A549 cells were transfected with 100 nM NFE2 siRNA or non-targeting (NT) siRNA followed by exposure to CSE for 24 h. Representative flow cytometry images using Annexin V and PI staining are shown. ( B ) NFE2 knockdown significantly increased cell death after treatment with CSE compared to control. *Statistically significant difference ( p < 0.05). Data are shown as the mean (±s.e.m.).

Article Snippet: The following antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA): MMP9, CD147, ADAMTSL-4, p45 NFE2, p18 NFE2, DJ-1, phosphoserine, phosphothreonine and phosphotyrosine.

Techniques: In Vitro, Transfection, Flow Cytometry, Staining

Journal: Scientific Reports

Article Title: The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema

doi: 10.1038/s41598-018-21790-3

Figure Lengend Snippet: p45 NFE2 interaction with DJ-1 in ATII cells and lung tissue obtained from non-smokers (NS), smokers (SM) and patients with emphysema (E). DJ-1 was co-immunoprecipitated in freshly isolated ATII cells ( A ) or lung tissue ( B ) followed by Western blotting analysis to determine its interaction with p45 NFE2 or p18 NFE2 as described in Materials and Methods. Relative expression is also shown. ( C ) p45 NFE2 (green) and DJ-1 (red) expression in ATII cells identified using SP-A staining (violet) in lung tissue by immunohistofluorescence. Cell nuclei were stained with DAPI (blue). The strongest co-localization of p45 NFE2 and DJ-1 is indicated by white arrows. *Statistically significant difference ( p < 0.05). Data are shown as the mean (±s.e.m.).

Article Snippet: The following antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA): MMP9, CD147, ADAMTSL-4, p45 NFE2, p18 NFE2, DJ-1, phosphoserine, phosphothreonine and phosphotyrosine.

Techniques: Immunoprecipitation, Isolation, Western Blot, Expressing, Staining, Immunohistofluorescence

Journal: Scientific Reports

Article Title: The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema

doi: 10.1038/s41598-018-21790-3

Figure Lengend Snippet: MMP9, CD147, cathepsin B, p18 NFE2, p45 NFE2 and ADAMTSL-4 expression in wild-type and DJ-1 KO mice. Wild-type (WT) ( A ) and DJ-1 KO mice ( B ) were exposed to 150 mg/m 3 cigarette smoke (CS) for 2 h/day for 3 weeks as described in Materials and Methods section. Protein levels were analyzed in lung tissue by Western blotting. Lane 1 – WT mice, Lane 2 – WT + CS, Lane 3 – DJ-1 KO mice, Lane 4 – DJ-1 KO mice + CS. Relative expression is also shown. ( C ) p45 NFE2 expression (green) in ATII cells identified using SP-A antibody (violet) in lung tissue obtained from wild-type and DJ-1 KO mice by immunohistofluorescence. Cell nuclei were stained with DAPI (blue). ( D , E ) MMP9, CD147, cathepsin B, p45 NFE2 and ADAMTSL-4 mRNA expression in lung tissue from wild-type ( D ) and DJ-1 KO ( E ) mice by RT-PCR. The strongest p45 NFE2 expression is indicated by white arrows. *Statistically significant difference in comparison with control ( p < 0.05). Data are shown as the mean (±s.e.m.).

Article Snippet: The following antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA): MMP9, CD147, ADAMTSL-4, p45 NFE2, p18 NFE2, DJ-1, phosphoserine, phosphothreonine and phosphotyrosine.

Techniques: Expressing, Western Blot, Immunohistofluorescence, Staining, Reverse Transcription Polymerase Chain Reaction

Journal: Scientific Reports

Article Title: The cytoprotective role of DJ-1 and p45 NFE2 against human primary alveolar type II cell injury and emphysema

doi: 10.1038/s41598-018-21790-3

Figure Lengend Snippet: The role of p45 NFE in ATII cells in non-smokers, smokers and emphysema patients.

Article Snippet: The following antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA): MMP9, CD147, ADAMTSL-4, p45 NFE2, p18 NFE2, DJ-1, phosphoserine, phosphothreonine and phosphotyrosine.

Techniques: