mmp 9 plasmid Search Results


plasmid pcmv3 mmmp9 flag  (Sino Biological)
90
Sino Biological plasmid pcmv3 mmmp9 flag
Plasmid Pcmv3 Mmmp9 Flag, supplied by Sino Biological, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pmc07330055-49-1-7?v=Sino+Biological
Average 90 stars, based on 1 article reviews
plasmid pcmv3 mmmp9 flag - by Bioz Stars, 2026-08
90/100 stars
  Buy from Supplier
mmp9  (Addgene inc)
90
Addgene inc mmp9
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Mmp9, supplied by Addgene inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pmc08042603-310-13-15?v=Addgene+inc
Average 90 stars, based on 1 article reviews
mmp9 - by Bioz Stars, 2026-08
90/100 stars
  Buy from Supplier
mmp9 inhibition  (OriGene)
91
OriGene mmp9 inhibition
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Mmp9 Inhibition, supplied by OriGene, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/ppr0301339-74-10-28?v=OriGene
Average 91 stars, based on 1 article reviews
mmp9 inhibition - by Bioz Stars, 2026-08
91/100 stars
  Buy from Supplier
pcdna3 1 mmp 9 ha  (Addgene inc)
93
Addgene inc pcdna3 1 mmp 9 ha
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Pcdna3 1 Mmp 9 Ha, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pmc12459465-231-20-21?v=Addgene+inc
Average 93 stars, based on 1 article reviews
pcdna3 1 mmp 9 ha - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
human mmp9 shrna plasmid kit  (OriGene)
91
OriGene human mmp9 shrna plasmid kit
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Human Mmp9 Shrna Plasmid Kit, supplied by OriGene, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/ppr0301339-74-13-28?v=OriGene
Average 91 stars, based on 1 article reviews
human mmp9 shrna plasmid kit - by Bioz Stars, 2026-08
91/100 stars
  Buy from Supplier
mmp 9 plasmid  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology mmp 9 plasmid
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Mmp 9 Plasmid, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pmc06861869-37-72-76?v=Santa+Cruz+Biotechnology
Average 93 stars, based on 1 article reviews
mmp 9 plasmid - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
mmp 9 shrna plasmid  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology mmp 9 shrna plasmid
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Mmp 9 Shrna Plasmid, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pm31772631-41-36-41?v=Santa+Cruz+Biotechnology
Average 93 stars, based on 1 article reviews
mmp 9 shrna plasmid - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
c terminal his tag  (Sino Biological)
91
Sino Biological c terminal his tag
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
C Terminal His Tag, supplied by Sino Biological, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pm36648434-64-28-37?v=Sino+Biological
Average 91 stars, based on 1 article reviews
c terminal his tag - by Bioz Stars, 2026-08
91/100 stars
  Buy from Supplier
mmp  (Santa Cruz Biotechnology)
93
Santa Cruz Biotechnology mmp
Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), <t>MMP9,</t> MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).
Mmp, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pmc07126203-35-0-8?v=Santa+Cruz+Biotechnology
Average 93 stars, based on 1 article reviews
mmp - by Bioz Stars, 2026-08
93/100 stars
  Buy from Supplier
mmp9 vector  (OriGene)
91
OriGene mmp9 vector
<t>MMP9</t> is an essential downstream effector of RPL23. (A, B) EMT-related markers (MMP9, MMP2, N-cadherin, Vimentin, Smad2, Twist1 and E-cadherin) were measured on RPL23-depleted HCC cells by qRT-PCR. β-actin was used as an internal quantitative control. (***p < 0.001) (C) RPL23 regulated MMP9 protein expression in HLE and MHCC97H cells measured by western blot assay. GAPDH was used as a loading control for western blotting. (D) RIP assays showed that RPL23 directly bound to MMP9 mRNA. (E, F) RNA pull-down results showed that RPL23 was directly associated with the 3`UTR of MMP9 mRNA. (E) shows the biotinylated-MMP9-3’UTR, CDS or 5’UTR transcript in vitro , (F) shows the result of the RNA pull-down assay as analyzed by western blot. Control indicates a control pulldown containing beads only. (G) The half-life of <t>MMP-9</t> mRNA was reduced after RPL23 knockdown in HLE and MHCC97H cells followed by treatment with 5ug/mL actinomycin D at the indicated times. Error bars represent SEM. p-values (HLE): **p = 0.00116 (shCont vs shRPL23#1), **p = 0.00296 (shCont vs shRPL23#2). p-values (MHCC97H): **p = 0.00314 (shCont vs shRPL23#1), **p = 0.00477 (shCont vs shRPL23#2). (H) Scatter plot between RPL23 and MMP9 mRNA level in HCC (n = 371, from TCGA database). Spearman’s correlation coefficients were calculated. Representative data are from at least three independent experiments. *p<0.05, **P < 0.01, ***P < 0.001.
Mmp9 Vector, supplied by OriGene, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/pmc08677661-68-1-6?v=OriGene
Average 91 stars, based on 1 article reviews
mmp9 vector - by Bioz Stars, 2026-08
91/100 stars
  Buy from Supplier
human mmp9/mmp-9/clg4b gene orf cdna clone expression plasmid  (Sino Biological)
91
Sino Biological human mmp9/mmp-9/clg4b gene orf cdna clone expression plasmid
<t>MMP9</t> is an essential downstream effector of RPL23. (A, B) EMT-related markers (MMP9, MMP2, N-cadherin, Vimentin, Smad2, Twist1 and E-cadherin) were measured on RPL23-depleted HCC cells by qRT-PCR. β-actin was used as an internal quantitative control. (***p < 0.001) (C) RPL23 regulated MMP9 protein expression in HLE and MHCC97H cells measured by western blot assay. GAPDH was used as a loading control for western blotting. (D) RIP assays showed that RPL23 directly bound to MMP9 mRNA. (E, F) RNA pull-down results showed that RPL23 was directly associated with the 3`UTR of MMP9 mRNA. (E) shows the biotinylated-MMP9-3’UTR, CDS or 5’UTR transcript in vitro , (F) shows the result of the RNA pull-down assay as analyzed by western blot. Control indicates a control pulldown containing beads only. (G) The half-life of <t>MMP-9</t> mRNA was reduced after RPL23 knockdown in HLE and MHCC97H cells followed by treatment with 5ug/mL actinomycin D at the indicated times. Error bars represent SEM. p-values (HLE): **p = 0.00116 (shCont vs shRPL23#1), **p = 0.00296 (shCont vs shRPL23#2). p-values (MHCC97H): **p = 0.00314 (shCont vs shRPL23#1), **p = 0.00477 (shCont vs shRPL23#2). (H) Scatter plot between RPL23 and MMP9 mRNA level in HCC (n = 371, from TCGA database). Spearman’s correlation coefficients were calculated. Representative data are from at least three independent experiments. *p<0.05, **P < 0.01, ***P < 0.001.
Human Mmp9/Mmp 9/Clg4b Gene Orf Cdna Clone Expression Plasmid, supplied by Sino Biological, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/custom%40hg10327-ut%4039613801?v=Sino+Biological
Average 91 stars, based on 1 article reviews
human mmp9/mmp-9/clg4b gene orf cdna clone expression plasmid - by Bioz Stars, 2026-08
91/100 stars
  Buy from Supplier
human mmp9/mmp-9/clg4b gene orf cdna clone expression plasmid, c-gfpspark tag  (Sino Biological)
92
Sino Biological human mmp9/mmp-9/clg4b gene orf cdna clone expression plasmid, c-gfpspark tag
<t>MMP9</t> is an essential downstream effector of RPL23. (A, B) EMT-related markers (MMP9, MMP2, N-cadherin, Vimentin, Smad2, Twist1 and E-cadherin) were measured on RPL23-depleted HCC cells by qRT-PCR. β-actin was used as an internal quantitative control. (***p < 0.001) (C) RPL23 regulated MMP9 protein expression in HLE and MHCC97H cells measured by western blot assay. GAPDH was used as a loading control for western blotting. (D) RIP assays showed that RPL23 directly bound to MMP9 mRNA. (E, F) RNA pull-down results showed that RPL23 was directly associated with the 3`UTR of MMP9 mRNA. (E) shows the biotinylated-MMP9-3’UTR, CDS or 5’UTR transcript in vitro , (F) shows the result of the RNA pull-down assay as analyzed by western blot. Control indicates a control pulldown containing beads only. (G) The half-life of <t>MMP-9</t> mRNA was reduced after RPL23 knockdown in HLE and MHCC97H cells followed by treatment with 5ug/mL actinomycin D at the indicated times. Error bars represent SEM. p-values (HLE): **p = 0.00116 (shCont vs shRPL23#1), **p = 0.00296 (shCont vs shRPL23#2). p-values (MHCC97H): **p = 0.00314 (shCont vs shRPL23#1), **p = 0.00477 (shCont vs shRPL23#2). (H) Scatter plot between RPL23 and MMP9 mRNA level in HCC (n = 371, from TCGA database). Spearman’s correlation coefficients were calculated. Representative data are from at least three independent experiments. *p<0.05, **P < 0.01, ***P < 0.001.
Human Mmp9/Mmp 9/Clg4b Gene Orf Cdna Clone Expression Plasmid, C Gfpspark Tag, supplied by Sino Biological, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/mmp+9+plasmid/custom%40hg10327-acg%4039613801?v=Sino+Biological
Average 92 stars, based on 1 article reviews
human mmp9/mmp-9/clg4b gene orf cdna clone expression plasmid, c-gfpspark tag - by Bioz Stars, 2026-08
92/100 stars
  Buy from Supplier

Image Search Results


Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), MMP9, MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: Suppression of DMBT1 promotes invasion and metastasis. (A) Lysates from UM-SCC-29 and UM-SCC-47 stably transduced with shEZH2 were immunoblotted with DMBT1, EZH2, and actin (loading control) antibodies ( n = 2). (B) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding. Scatter plots show three independent experiments; each color represents one experiment with three replicates in each (*, P < 0.05; ****, P < 0.0001; one-way ANOVA; error bars represent SD). (C) Lysates were immunoblotted with DMBT1, E-cadherin (E-Cad), MMP9, MMP2, EZH2, and actin antibodies. Quantification represents densitometric unit (DU) normalized to actin and expressed as percent of corresponding control ( n = 2). (D) GFP-labeled UM-SCC-29-shEZH2 cells transfected with siNT or siDMBT1 (1 × 10 6 ) were seeded on the CAM. After 4 d, the upper CAM was photographed (brightfield and GFP) and merged (scale bar = 2 mm). Representative images shown. (E) Tumor area (*, P < 0.05; siNT: n = 6; siDMBT1: n = 4; t test; error bars represent SD). (F) Tumor islands in H&E-stained sections were quantified (top scale bar = 1 mm; bottom scale bar = 500 µm). Sections stained with anti-collagen IV and DAPI to highlight basement membrane and nuclei, respectively (bottom). Tumor cells have GFP label (scale bar = 100 µm). Arrows show invasive islands. (G) Quantification of invasive islands (*, P < 0.05; siNT: n = 5; siDMBT1: n = 4; t test; error bars represent SD). (H) Metastases (Mets) to the lower CAM ( n = 6) and liver ( n = 6) were quantified. y axis = metastatic cells/µg of DNA from lower CAM (*, P < 0.05; t test; error bars represent SD).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Stable Transfection, Transduction, Control, Transfection, Labeling, Staining, Membrane

Suppression of DMBT1 down-regulates E-cadherin (E-cad), up-regulates MMPs, and promotes invasion. (A) Lysates from UM-SCC-1 were immunoblotted with DMBT1, EZH2, E-cad, MMP9, MMP2, and actin (loading control) antibodies. Quantification represents DUs normalized to actin and expressed as percent of control ( n = 2). (B and C) UM-SCC-1-shEZH2-DOX (B) and UM-SCC-29-shEZH2 (C) were transfected with siDMBT1 or control siRNA (siNT). MMP9 and MMP2 in CM were detected by gelatin zymography. Data represent three independent experiments. (D) UM-SCC1-shEZH2-Dox was transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding (*, P < 0.05; t test; error bars represent SD). (E) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1-2 or siNT. Invasion was quantified at 24 h after seeding (****, P < 0.0001; one-way ANOVA; error bars represent SD).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: Suppression of DMBT1 down-regulates E-cadherin (E-cad), up-regulates MMPs, and promotes invasion. (A) Lysates from UM-SCC-1 were immunoblotted with DMBT1, EZH2, E-cad, MMP9, MMP2, and actin (loading control) antibodies. Quantification represents DUs normalized to actin and expressed as percent of control ( n = 2). (B and C) UM-SCC-1-shEZH2-DOX (B) and UM-SCC-29-shEZH2 (C) were transfected with siDMBT1 or control siRNA (siNT). MMP9 and MMP2 in CM were detected by gelatin zymography. Data represent three independent experiments. (D) UM-SCC1-shEZH2-Dox was transfected with siDMBT1 or siNT. Invasion was quantified at 48 h after seeding (*, P < 0.05; t test; error bars represent SD). (E) UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 were transfected with siDMBT1-2 or siNT. Invasion was quantified at 24 h after seeding (****, P < 0.0001; one-way ANOVA; error bars represent SD).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Control, Transfection, Zymography

DMBT1 regulates mRNA stability and transcription of MMP9 and E-cadherin via ZEB1. (A) Immunoblot for EMT markers in UM-SCC-29-pCMV6 and UM-SCC-29-DMBT1 cells ( n = 2). (B–E) DMBT1 inhibits invasion via up-regulation of E-cadherin (E-cad). UM-SCC-29-DMBT1 cells were transiently transfected with siNT or siE-cad-1 (B and C) or siE-cad-2 (D and E). (B and D) Immunoblots verify suppression of E-cadherin. (C and E) Invasion assays. Scatter plots show three independent experiments; each color represents one experiment with at least duplicates in each (**, P < 0.001; ***, P < 0.0001; one-way ANOVA; error bars represent SD). (F and G) Overexpression of MMP9 and ZEB1 rescue invasion. UM-SCC-29-DMBT1 cells were transduced with empty vector (pLX304) or MMP9 or ZEB1. UM-SCC-29-pCMV was transduced with pLX304 as a control for UM-SCC-29-DMBT1-pLX304. Lysates were immunoblotted with anti-MMP9, anti-ZEB1, and anti-GAPDH (F). Invasion was quantified at 48 h after seeding (G). Scatter plots show two independent experiments; each color represents an independent experiment with three replicates in each (****, P < 0.0001; one-way ANOVA; error bars represent SD). (H and I) UM-SCC-29-pCMV6 and UM-SCC-29-DMBT1 were incubated with actinomycin D (1 µg/ml), and RNA was isolated at the indicated time. MMP-9 (H) and E-cadherin (I) transcripts, quantified by Q-RT-PCR, were normalized to GAPDH. Results are presented as percent of corresponding transcript at time 0. Values are mean ± SEM ( n = 3). (J and K) Normalized luciferase activity of MMP9 (J) and CDH1 (K) promoters in UM-SCC-29-pCMV6 and UM-SCC-29-DMBT1. Scatter plots show three independent experiments; each color represents an independent experiment (***, P < 0.001; t test; error bars represent SD).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: DMBT1 regulates mRNA stability and transcription of MMP9 and E-cadherin via ZEB1. (A) Immunoblot for EMT markers in UM-SCC-29-pCMV6 and UM-SCC-29-DMBT1 cells ( n = 2). (B–E) DMBT1 inhibits invasion via up-regulation of E-cadherin (E-cad). UM-SCC-29-DMBT1 cells were transiently transfected with siNT or siE-cad-1 (B and C) or siE-cad-2 (D and E). (B and D) Immunoblots verify suppression of E-cadherin. (C and E) Invasion assays. Scatter plots show three independent experiments; each color represents one experiment with at least duplicates in each (**, P < 0.001; ***, P < 0.0001; one-way ANOVA; error bars represent SD). (F and G) Overexpression of MMP9 and ZEB1 rescue invasion. UM-SCC-29-DMBT1 cells were transduced with empty vector (pLX304) or MMP9 or ZEB1. UM-SCC-29-pCMV was transduced with pLX304 as a control for UM-SCC-29-DMBT1-pLX304. Lysates were immunoblotted with anti-MMP9, anti-ZEB1, and anti-GAPDH (F). Invasion was quantified at 48 h after seeding (G). Scatter plots show two independent experiments; each color represents an independent experiment with three replicates in each (****, P < 0.0001; one-way ANOVA; error bars represent SD). (H and I) UM-SCC-29-pCMV6 and UM-SCC-29-DMBT1 were incubated with actinomycin D (1 µg/ml), and RNA was isolated at the indicated time. MMP-9 (H) and E-cadherin (I) transcripts, quantified by Q-RT-PCR, were normalized to GAPDH. Results are presented as percent of corresponding transcript at time 0. Values are mean ± SEM ( n = 3). (J and K) Normalized luciferase activity of MMP9 (J) and CDH1 (K) promoters in UM-SCC-29-pCMV6 and UM-SCC-29-DMBT1. Scatter plots show three independent experiments; each color represents an independent experiment (***, P < 0.001; t test; error bars represent SD).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Western Blot, Transfection, Over Expression, Transduction, Plasmid Preparation, Control, Incubation, Isolation, Reverse Transcription Polymerase Chain Reaction, Luciferase, Activity Assay

Overexpression of DMBT1 regulates mRNA stability and transcription of MMP9 and E-cadherin (E-cad). (A) Expression of E-cad and MMP9 in UM-SCC-1-pCMV6 and UM-SCC-1-DMBT1 cells ( n = 2). (B and C) UM-SCC-1-pCMV6 and UM-SCC-1-DMBT1 were incubated with actinomycin D (1 µg/ml), and RNA was isolated at different time points. MMP9 (B) and E-cad (C) transcripts were quantified by Q-RT-PCR. MMP9 and E-cad mRNA expression was normalized to GAPDH. Results are presented as percent of corresponding transcript at time 0. Values are mean ± SEM. (D and E) Normalized luciferase activity of MMP9 promoter (D) is reduced whereas activity of CDH1 promoter (E) is significantly higher in UM-SCC-1-DMBT1 compared with control UM-SCC-1-pCMV6 cells (*, P < 0.001; n = 2; t test; error bars represent SD). (F) Q-RT-PCR of UM-SCC-1-pCMV6, UM-SCC-1-DMBT1, UM-SCC-29-pCMV6, and UM-SCC-29-DMBT1. Primers for ZEB1, NRF2 (NFE2), β-catenin, VDR, YY1, c-Jun (AP1), c-Fos (AP1), SP1, EGR1, KLF4, NFATC1 (NFAT), HNF1 (HNF1A), STAT1, STAT3, and TP53 are listed in . Data are normalized to GAPDH. Relative expression of each transcription factor in cells overexpressing DMBT1 is shown as fold change with respect to control (pCMV6) cells (*, P < 0.05; t test; error bars represent SD).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: Overexpression of DMBT1 regulates mRNA stability and transcription of MMP9 and E-cadherin (E-cad). (A) Expression of E-cad and MMP9 in UM-SCC-1-pCMV6 and UM-SCC-1-DMBT1 cells ( n = 2). (B and C) UM-SCC-1-pCMV6 and UM-SCC-1-DMBT1 were incubated with actinomycin D (1 µg/ml), and RNA was isolated at different time points. MMP9 (B) and E-cad (C) transcripts were quantified by Q-RT-PCR. MMP9 and E-cad mRNA expression was normalized to GAPDH. Results are presented as percent of corresponding transcript at time 0. Values are mean ± SEM. (D and E) Normalized luciferase activity of MMP9 promoter (D) is reduced whereas activity of CDH1 promoter (E) is significantly higher in UM-SCC-1-DMBT1 compared with control UM-SCC-1-pCMV6 cells (*, P < 0.001; n = 2; t test; error bars represent SD). (F) Q-RT-PCR of UM-SCC-1-pCMV6, UM-SCC-1-DMBT1, UM-SCC-29-pCMV6, and UM-SCC-29-DMBT1. Primers for ZEB1, NRF2 (NFE2), β-catenin, VDR, YY1, c-Jun (AP1), c-Fos (AP1), SP1, EGR1, KLF4, NFATC1 (NFAT), HNF1 (HNF1A), STAT1, STAT3, and TP53 are listed in . Data are normalized to GAPDH. Relative expression of each transcription factor in cells overexpressing DMBT1 is shown as fold change with respect to control (pCMV6) cells (*, P < 0.05; t test; error bars represent SD).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Over Expression, Expressing, Incubation, Isolation, Reverse Transcription Polymerase Chain Reaction, Luciferase, Activity Assay, Control

ZEB1 down-regulates DMBT1 by binding promoter regions of MMP9 and CDH1 . (A) Venn diagram showing transcription factors with binding sites in the promoter regions of MMP9 and CDH1 , including ZEB1, NRF2 (NFE2), VDR, YY1, c-Jun (AP1), SP1, EGR1, KLF4, NFATC1 (NFAT), HNF1 (HNF1A), STAT1, STAT3, and TP53. (B and C) Q-RT-PCR of UM-SCC-1-pCMV6, UM-SCC-1-DMBT1, UM-SCC-29-pCMV6, and UM-SCC-29-DMBT1. Data are normalized to GAPDH. Relative expression of ZEB1 transcription factor in cells overexpressing DMBT1 is shown as fold change with respect to control (pCMV6) cells (***, P < 0.001; ****, P < 0.0001; t test; n = 2; error bars represent SD). (D and E) Q-RT-PCR was performed with ChIP-eluted DNA from UM-SCC-1-pCMV6, UM-SCC-1-DMBT1, UM-SCC-29-pCMV6, and UM-SCC-29-DMBT1 with validated primers specific to the ZEB1-binding site in the promoter region of MMP9 (D) and CDH1 (E; *, P < 0.05; t test; n = 2; error bars represent SD). (F) Immunoblot validation of DMBT1 down-regulation in lysates from UM-SCC-29-shVSVG and -shEZH2 transfected with nontarget siRNA (NT) or siDMBT1 (-1 and -2), as indicated. (G and H) Q-RT-PCR with ChIP-eluted DNA from UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 (NT, siDMBT1-1, and siDMBT1-2) for standardized primers targeting the ZEB1-binding site in the promoter region of MMP9 (G) and CDH1 (H). Scatter plots show two independent experiments; each color represents an independent experiment (**, P < 0.01; ****, P < 0.0001; one-way ANOVA; error bars represent SD).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: ZEB1 down-regulates DMBT1 by binding promoter regions of MMP9 and CDH1 . (A) Venn diagram showing transcription factors with binding sites in the promoter regions of MMP9 and CDH1 , including ZEB1, NRF2 (NFE2), VDR, YY1, c-Jun (AP1), SP1, EGR1, KLF4, NFATC1 (NFAT), HNF1 (HNF1A), STAT1, STAT3, and TP53. (B and C) Q-RT-PCR of UM-SCC-1-pCMV6, UM-SCC-1-DMBT1, UM-SCC-29-pCMV6, and UM-SCC-29-DMBT1. Data are normalized to GAPDH. Relative expression of ZEB1 transcription factor in cells overexpressing DMBT1 is shown as fold change with respect to control (pCMV6) cells (***, P < 0.001; ****, P < 0.0001; t test; n = 2; error bars represent SD). (D and E) Q-RT-PCR was performed with ChIP-eluted DNA from UM-SCC-1-pCMV6, UM-SCC-1-DMBT1, UM-SCC-29-pCMV6, and UM-SCC-29-DMBT1 with validated primers specific to the ZEB1-binding site in the promoter region of MMP9 (D) and CDH1 (E; *, P < 0.05; t test; n = 2; error bars represent SD). (F) Immunoblot validation of DMBT1 down-regulation in lysates from UM-SCC-29-shVSVG and -shEZH2 transfected with nontarget siRNA (NT) or siDMBT1 (-1 and -2), as indicated. (G and H) Q-RT-PCR with ChIP-eluted DNA from UM-SCC-29-shVSVG and UM-SCC-29-shEZH2 (NT, siDMBT1-1, and siDMBT1-2) for standardized primers targeting the ZEB1-binding site in the promoter region of MMP9 (G) and CDH1 (H). Scatter plots show two independent experiments; each color represents an independent experiment (**, P < 0.01; ****, P < 0.0001; one-way ANOVA; error bars represent SD).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Binding Assay, Reverse Transcription Polymerase Chain Reaction, Expressing, Control, Western Blot, Biomarker Discovery, Transfection

HNSCC reduces DMBT1 expression in adjacent histologically normal epithelium to enhance invasion of HNSCC. (A) LCM to collect normal epithelium (NE; n = 3) and salivary gland (NSG; n = 3) from tissue without cancer. Tumor islands (TI; n = 10), adjacent epithelium (AE; n = 9), and adjacent salivary gland (i.e., no tumor in salivary gland; ASG; n = 5) were isolated from HNSCC tissue sections. Arrows represent tumor islands. Arrowheads show NE, AE, and SGs in corresponding panels. Scale bar = 100 µm. (B) cDNAs were synthesized from RNA collected from NE, NSG, TI, AE, and ASG. Q-RT-PCR was performed (bar graph represents mean ± SD; *, P < 0.001; one-way ANOVA). (C) Immunohistochemistry with anti-DMBT1 on the same HNSCC sections shown in A ( n = 6–9/group). (D) DMBT1 immunostaining is scored based on agreement of two independent analyses. (E) HOK16B were treated with complete, blank, or CM from UM-SCC-29 ( n = 2). Lysates were collected after 24, 48, and 72 h, and immunoblotted with DMBT1 or actin antibodies. Signal intensity is quantified by densitometric analysis with normalization to actin and then expressed as percent of corresponding control at the same time point (DU). (F) HOK16B were treated with blank or CM from UM-SCC-29, and lysates were collected after 48 h and immunoblotted with DMBT1, E-cadherin, MMP9, and ZEB1 ( n = 2). (G) Schematic of the experimental design for H and I. HOK16B were incubated for 96 h with blank medium or CM from UM-SCC-1 or UM-SCC-29. These primed keratinocytes were incubated with fresh blank medium to generate CM. This CM from primed keratinocytes was used for invasion of UM-SCC-1 (H) or UM-SCC-29 on growth factor reduced (GFR) matrigel (I). Scatter plots show the percentage of invasion of HNSCC cells. Each color represents an independent experiment with three replicates in each ( n = 3). Data represent mean ± SD (**, P < 0.01; ***, P < 0.001; t test).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: HNSCC reduces DMBT1 expression in adjacent histologically normal epithelium to enhance invasion of HNSCC. (A) LCM to collect normal epithelium (NE; n = 3) and salivary gland (NSG; n = 3) from tissue without cancer. Tumor islands (TI; n = 10), adjacent epithelium (AE; n = 9), and adjacent salivary gland (i.e., no tumor in salivary gland; ASG; n = 5) were isolated from HNSCC tissue sections. Arrows represent tumor islands. Arrowheads show NE, AE, and SGs in corresponding panels. Scale bar = 100 µm. (B) cDNAs were synthesized from RNA collected from NE, NSG, TI, AE, and ASG. Q-RT-PCR was performed (bar graph represents mean ± SD; *, P < 0.001; one-way ANOVA). (C) Immunohistochemistry with anti-DMBT1 on the same HNSCC sections shown in A ( n = 6–9/group). (D) DMBT1 immunostaining is scored based on agreement of two independent analyses. (E) HOK16B were treated with complete, blank, or CM from UM-SCC-29 ( n = 2). Lysates were collected after 24, 48, and 72 h, and immunoblotted with DMBT1 or actin antibodies. Signal intensity is quantified by densitometric analysis with normalization to actin and then expressed as percent of corresponding control at the same time point (DU). (F) HOK16B were treated with blank or CM from UM-SCC-29, and lysates were collected after 48 h and immunoblotted with DMBT1, E-cadherin, MMP9, and ZEB1 ( n = 2). (G) Schematic of the experimental design for H and I. HOK16B were incubated for 96 h with blank medium or CM from UM-SCC-1 or UM-SCC-29. These primed keratinocytes were incubated with fresh blank medium to generate CM. This CM from primed keratinocytes was used for invasion of UM-SCC-1 (H) or UM-SCC-29 on growth factor reduced (GFR) matrigel (I). Scatter plots show the percentage of invasion of HNSCC cells. Each color represents an independent experiment with three replicates in each ( n = 3). Data represent mean ± SD (**, P < 0.01; ***, P < 0.001; t test).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Expressing, Isolation, Synthesized, Reverse Transcription Polymerase Chain Reaction, Immunohistochemistry, Immunostaining, Control, Incubation

TNFα and TGFβ1 down-regulate DMBT1 in adjacent histologically normal epithelium. (A) Schematic for experimental design for B and C. CM from siNT or siDMBT1 HOK16B were used for the invasion assay. (B) siRNA-mediated down-regulation of DMBT1 in HOK16B. Cell lysates were immunoblotted with antibodies against ZEB1, E-cadherin (E-cad), MMP-9, and actin ( n = 2). Signal intensity was quantified by densitometric analysis with normalization to actin and then expressed as percent of corresponding control (DU). (C) Scatter plots show percent invasion of UM-SCC-1 (top) and UM-SCC-29 (bottom) with CM from keratinocytes transfected with siNT or siDMBT1. Each color denotes an independent experiment with three replicates in each experiment ( n = 2 for UM-SCC-1; n = 3 for UM-SCC-29). Data represent mean ± SD (*, P < 0.05; **, P < 0.01; t test). (D) Heat-map for the expression of IL10, GMCSF, TNFα, and TGFβ1 in normal (N) and cancer (T) tissues from 22 patients from GEO accession no. GSE6631 . (E) Dose response of impact of recombinant TNFα and TGFβ1 on DMBT1 expression in HOK16B ( n = 3). Cells were incubated with increasing concentrations of TNFα (lanes 2–5, lane 1 as vehicle control) and TGFβ1 (lanes 7–10, lane 6 as vehicle control) for 48 h. Cell lysates were immunoblotted with DMBT1 and actin antibodies. (F) Time course of recombinant TNFα and TGFβ1 treatment in HOK16B ( n = 3). Cells were incubated with TNFα (10 ng/ml) or TGFβ1 (1 ng/ml). Lysates were harvested at the indicated time and immunoblotted with DMBT1 and actin antibodies. (G) HOK16B were incubated with TNFα (10 ng/ml), TGFβ1 (1 ng/ml), or vehicle. Lysates were immunoblotted with DMBT1, E-cad, MMP9, ZEB1, and actin antibodies ( n = 2). (H) HOK16B transfected with siRNA (NT) or siDMBT1 were incubated with TNFα (10 ng/ml), TGFβ1 (1 ng/ml), or vehicle. Lysates were immunoblotted with DMBT1, E-cad, MMP9, ZEB1, and actin antibodies ( n = 2). GFR, growth factor reduced Matrigel.

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: TNFα and TGFβ1 down-regulate DMBT1 in adjacent histologically normal epithelium. (A) Schematic for experimental design for B and C. CM from siNT or siDMBT1 HOK16B were used for the invasion assay. (B) siRNA-mediated down-regulation of DMBT1 in HOK16B. Cell lysates were immunoblotted with antibodies against ZEB1, E-cadherin (E-cad), MMP-9, and actin ( n = 2). Signal intensity was quantified by densitometric analysis with normalization to actin and then expressed as percent of corresponding control (DU). (C) Scatter plots show percent invasion of UM-SCC-1 (top) and UM-SCC-29 (bottom) with CM from keratinocytes transfected with siNT or siDMBT1. Each color denotes an independent experiment with three replicates in each experiment ( n = 2 for UM-SCC-1; n = 3 for UM-SCC-29). Data represent mean ± SD (*, P < 0.05; **, P < 0.01; t test). (D) Heat-map for the expression of IL10, GMCSF, TNFα, and TGFβ1 in normal (N) and cancer (T) tissues from 22 patients from GEO accession no. GSE6631 . (E) Dose response of impact of recombinant TNFα and TGFβ1 on DMBT1 expression in HOK16B ( n = 3). Cells were incubated with increasing concentrations of TNFα (lanes 2–5, lane 1 as vehicle control) and TGFβ1 (lanes 7–10, lane 6 as vehicle control) for 48 h. Cell lysates were immunoblotted with DMBT1 and actin antibodies. (F) Time course of recombinant TNFα and TGFβ1 treatment in HOK16B ( n = 3). Cells were incubated with TNFα (10 ng/ml) or TGFβ1 (1 ng/ml). Lysates were harvested at the indicated time and immunoblotted with DMBT1 and actin antibodies. (G) HOK16B were incubated with TNFα (10 ng/ml), TGFβ1 (1 ng/ml), or vehicle. Lysates were immunoblotted with DMBT1, E-cad, MMP9, ZEB1, and actin antibodies ( n = 2). (H) HOK16B transfected with siRNA (NT) or siDMBT1 were incubated with TNFα (10 ng/ml), TGFβ1 (1 ng/ml), or vehicle. Lysates were immunoblotted with DMBT1, E-cad, MMP9, ZEB1, and actin antibodies ( n = 2). GFR, growth factor reduced Matrigel.

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Invasion Assay, Control, Transfection, Expressing, Recombinant, Incubation

TNFα and/or TGFβ1 from HNSCC suppress DMBT1 in adjacent histologically normal epithelium via E-cadherin (E-cad), MMP9, and ZEB1 modulation. (A) Schematic of procedure for depletion of TNFα and TGFβ1 in CM from HNSCC cells. (B) Immunoblot verification of antibody (ab) depletion of TNFα and TGFβ1 in CM from HNSCC cells ( n = 2). (C) CM was collected from UM-SCC-29 ( n = 2). CM with or without depletion, or with depletion and add-back of TNFα or TGFβ1, was incubated with HOK16B for 72 h. Cell lysates were immunoblotted with DMBT1 and actin antibodies. DUs were calculated by normalizing to corresponding controls. (D) Schematic of siRNA-mediated depletion of TNFα or TGFβ1 in CM from HNSCC cells. (E–H) Validation of down-regulation of TNFα (E) and TGFβ1 (G) in UM-SCC-29 lysate (top) and CM (bottom), respectively. HOK16B were incubated with TNFα-depleted (F) and TGFβ1-depleted (H) CM, and lysates were immunoblotted for DMBT1, E-cad, MMP9, ZEB1, and actin. DUs were calculated by normalizing to corresponding controls ( n = 2).

Journal: The Journal of Experimental Medicine

Article Title: Squamous cell carcinoma subverts adjacent histologically normal epithelium to promote lateral invasion

doi: 10.1084/jem.20200944

Figure Lengend Snippet: TNFα and/or TGFβ1 from HNSCC suppress DMBT1 in adjacent histologically normal epithelium via E-cadherin (E-cad), MMP9, and ZEB1 modulation. (A) Schematic of procedure for depletion of TNFα and TGFβ1 in CM from HNSCC cells. (B) Immunoblot verification of antibody (ab) depletion of TNFα and TGFβ1 in CM from HNSCC cells ( n = 2). (C) CM was collected from UM-SCC-29 ( n = 2). CM with or without depletion, or with depletion and add-back of TNFα or TGFβ1, was incubated with HOK16B for 72 h. Cell lysates were immunoblotted with DMBT1 and actin antibodies. DUs were calculated by normalizing to corresponding controls. (D) Schematic of siRNA-mediated depletion of TNFα or TGFβ1 in CM from HNSCC cells. (E–H) Validation of down-regulation of TNFα (E) and TGFβ1 (G) in UM-SCC-29 lysate (top) and CM (bottom), respectively. HOK16B were incubated with TNFα-depleted (F) and TGFβ1-depleted (H) CM, and lysates were immunoblotted for DMBT1, E-cad, MMP9, ZEB1, and actin. DUs were calculated by normalizing to corresponding controls ( n = 2).

Article Snippet: UM-SCC-29-pCMV6, UM-SCC-29-DMBT1, UM-SCC-1-pCMV6, and UM-SCC-1-DMBT1 cells were cotransfected with E-cadherin (#42081; Addgene) and MMP9 (#53434; Addgene) promoter-firefly luciferase reporter and Renilla luciferase constructs (normalization control) with Lipofectamine 2000.

Techniques: Western Blot, Incubation, Biomarker Discovery

MMP9 is an essential downstream effector of RPL23. (A, B) EMT-related markers (MMP9, MMP2, N-cadherin, Vimentin, Smad2, Twist1 and E-cadherin) were measured on RPL23-depleted HCC cells by qRT-PCR. β-actin was used as an internal quantitative control. (***p < 0.001) (C) RPL23 regulated MMP9 protein expression in HLE and MHCC97H cells measured by western blot assay. GAPDH was used as a loading control for western blotting. (D) RIP assays showed that RPL23 directly bound to MMP9 mRNA. (E, F) RNA pull-down results showed that RPL23 was directly associated with the 3`UTR of MMP9 mRNA. (E) shows the biotinylated-MMP9-3’UTR, CDS or 5’UTR transcript in vitro , (F) shows the result of the RNA pull-down assay as analyzed by western blot. Control indicates a control pulldown containing beads only. (G) The half-life of MMP-9 mRNA was reduced after RPL23 knockdown in HLE and MHCC97H cells followed by treatment with 5ug/mL actinomycin D at the indicated times. Error bars represent SEM. p-values (HLE): **p = 0.00116 (shCont vs shRPL23#1), **p = 0.00296 (shCont vs shRPL23#2). p-values (MHCC97H): **p = 0.00314 (shCont vs shRPL23#1), **p = 0.00477 (shCont vs shRPL23#2). (H) Scatter plot between RPL23 and MMP9 mRNA level in HCC (n = 371, from TCGA database). Spearman’s correlation coefficients were calculated. Representative data are from at least three independent experiments. *p<0.05, **P < 0.01, ***P < 0.001.

Journal: Frontiers in Oncology

Article Title: Ribosomal Protein L23 Drives the Metastasis of Hepatocellular Carcinoma via Upregulating MMP9

doi: 10.3389/fonc.2021.779748

Figure Lengend Snippet: MMP9 is an essential downstream effector of RPL23. (A, B) EMT-related markers (MMP9, MMP2, N-cadherin, Vimentin, Smad2, Twist1 and E-cadherin) were measured on RPL23-depleted HCC cells by qRT-PCR. β-actin was used as an internal quantitative control. (***p < 0.001) (C) RPL23 regulated MMP9 protein expression in HLE and MHCC97H cells measured by western blot assay. GAPDH was used as a loading control for western blotting. (D) RIP assays showed that RPL23 directly bound to MMP9 mRNA. (E, F) RNA pull-down results showed that RPL23 was directly associated with the 3`UTR of MMP9 mRNA. (E) shows the biotinylated-MMP9-3’UTR, CDS or 5’UTR transcript in vitro , (F) shows the result of the RNA pull-down assay as analyzed by western blot. Control indicates a control pulldown containing beads only. (G) The half-life of MMP-9 mRNA was reduced after RPL23 knockdown in HLE and MHCC97H cells followed by treatment with 5ug/mL actinomycin D at the indicated times. Error bars represent SEM. p-values (HLE): **p = 0.00116 (shCont vs shRPL23#1), **p = 0.00296 (shCont vs shRPL23#2). p-values (MHCC97H): **p = 0.00314 (shCont vs shRPL23#1), **p = 0.00477 (shCont vs shRPL23#2). (H) Scatter plot between RPL23 and MMP9 mRNA level in HCC (n = 371, from TCGA database). Spearman’s correlation coefficients were calculated. Representative data are from at least three independent experiments. *p<0.05, **P < 0.01, ***P < 0.001.

Article Snippet: The MMP9 vector was purchased from OriGene Technologies (Rockville, MD).

Techniques: Quantitative RT-PCR, Expressing, Western Blot, In Vitro, Pull Down Assay

MMP9 overexpression rescues the RPL23 knockdown-induced malignant phenotypes. (A, B) Overexpression of MMP9 rescued the inhibition effect of decreased RPL23 on HCC cell proliferation. (C) overexpression of MMP-9 rescued the repression effect of knockdown RPL23 on HCC cell migration ability by wound-healing assay. The cells were counted from 6 images. (D) Upregulation of MMP-9 could significantly rescued the effects of decreased RPL23 in HLE and MHCC97H cells for both migration and invasion by transwell assays. The cells were counted from 5 images. Representative data are from at least three independent experiments. Data are shown as mean ± SD. **P < 0.01, ***P < 0.001.

Journal: Frontiers in Oncology

Article Title: Ribosomal Protein L23 Drives the Metastasis of Hepatocellular Carcinoma via Upregulating MMP9

doi: 10.3389/fonc.2021.779748

Figure Lengend Snippet: MMP9 overexpression rescues the RPL23 knockdown-induced malignant phenotypes. (A, B) Overexpression of MMP9 rescued the inhibition effect of decreased RPL23 on HCC cell proliferation. (C) overexpression of MMP-9 rescued the repression effect of knockdown RPL23 on HCC cell migration ability by wound-healing assay. The cells were counted from 6 images. (D) Upregulation of MMP-9 could significantly rescued the effects of decreased RPL23 in HLE and MHCC97H cells for both migration and invasion by transwell assays. The cells were counted from 5 images. Representative data are from at least three independent experiments. Data are shown as mean ± SD. **P < 0.01, ***P < 0.001.

Article Snippet: The MMP9 vector was purchased from OriGene Technologies (Rockville, MD).

Techniques: Over Expression, Inhibition, Migration, Wound Healing Assay

RPL23 knockdown suppressed HCC cell lung metastasis in vivo . The nude mice were orthotopically injected with MHCC97H cells stably depleted RPL23. (A) Representative images (left), volume and number (right) of xenograft liver tumor in nude mice. (B) The presence or absence of metastatic nodules in the liver was evaluated by Hematoxylin-Eosin staining. (C) Representative images of lung metastasis nodules in different groups (up), and evaluated by Hematoxylin-Eosin staining(down). (D) IHC assay showed MMP9 expression in metastatic xenograft model tissue in different groups. For (A, C) , the data were presented as mean ± SD, n = 8 in each group. **P < 0.01.

Journal: Frontiers in Oncology

Article Title: Ribosomal Protein L23 Drives the Metastasis of Hepatocellular Carcinoma via Upregulating MMP9

doi: 10.3389/fonc.2021.779748

Figure Lengend Snippet: RPL23 knockdown suppressed HCC cell lung metastasis in vivo . The nude mice were orthotopically injected with MHCC97H cells stably depleted RPL23. (A) Representative images (left), volume and number (right) of xenograft liver tumor in nude mice. (B) The presence or absence of metastatic nodules in the liver was evaluated by Hematoxylin-Eosin staining. (C) Representative images of lung metastasis nodules in different groups (up), and evaluated by Hematoxylin-Eosin staining(down). (D) IHC assay showed MMP9 expression in metastatic xenograft model tissue in different groups. For (A, C) , the data were presented as mean ± SD, n = 8 in each group. **P < 0.01.

Article Snippet: The MMP9 vector was purchased from OriGene Technologies (Rockville, MD).

Techniques: In Vivo, Injection, Stable Transfection, Staining, Expressing