tgf-beta 2 antibody Search Results


anti tgf β2  (Bioss)
93
Bioss anti tgf β2
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Anti Tgf β2, supplied by Bioss, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
anti tgf β2 - by Bioz Stars, 2026-03
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tgf beta 2  (R&D Systems)
93
R&D Systems tgf beta 2
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Tgf Beta 2, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
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anti tgf β2  (Proteintech)
95
Proteintech anti tgf β2
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Anti Tgf β2, supplied by Proteintech, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 95 stars, based on 1 article reviews
anti tgf β2 - by Bioz Stars, 2026-03
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anti tgf 2  (R&D Systems)
93
R&D Systems anti tgf 2
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Anti Tgf 2, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
anti tgf 2 - by Bioz Stars, 2026-03
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mouse tgf β ab  (R&D Systems)
93
R&D Systems mouse tgf β ab
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Mouse Tgf β Ab, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
mouse tgf β ab - by Bioz Stars, 2026-03
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biotinylated tgf β2 affinity purified goat igg  (R&D Systems)
92
R&D Systems biotinylated tgf β2 affinity purified goat igg
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Biotinylated Tgf β2 Affinity Purified Goat Igg, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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tgfb2 1 2 antibody  (R&D Systems)
94
R&D Systems tgfb2 1 2 antibody
To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) <t>TGF-β2</t> was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.
Tgfb2 1 2 Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti tgfβ2 neutralizing antibody  (R&D Systems)
92
R&D Systems anti tgfβ2 neutralizing antibody
(A) HUVECs were transfected with either control mimic (con) or miR-30b mimic (30b) and levels of TGFβ1 and <t>TGFβ2</t> mRNA were assessed by qRT-PCR. Expression levels relative to control mimic transfected cells and normalized to β-actin expression are presented as the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 expression. * P < 0.05, ** P < 0.01 as determined by unpaired Student’s t -test. (B) Cells were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of TGFβ2 protein levels by western blot. β-actin was used as endogenous control. (C) ELISAs for TGFβ1 and TGFβ2 were performed with 24 hour conditioned supernates from HUVECs transfected with 20 nM of either control or miR-30b mimic. Data represents the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 secretion into cell culture supernate. * P = 0.044 as determined by unpaired Student’s t -test. (D) HUVECs were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of Smad2 phosphorylation by western blot.
Anti Tgfβ2 Neutralizing Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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mab612  (R&D Systems)
94
R&D Systems mab612
(A) HUVECs were transfected with either control mimic (con) or miR-30b mimic (30b) and levels of TGFβ1 and <t>TGFβ2</t> mRNA were assessed by qRT-PCR. Expression levels relative to control mimic transfected cells and normalized to β-actin expression are presented as the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 expression. * P < 0.05, ** P < 0.01 as determined by unpaired Student’s t -test. (B) Cells were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of TGFβ2 protein levels by western blot. β-actin was used as endogenous control. (C) ELISAs for TGFβ1 and TGFβ2 were performed with 24 hour conditioned supernates from HUVECs transfected with 20 nM of either control or miR-30b mimic. Data represents the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 secretion into cell culture supernate. * P = 0.044 as determined by unpaired Student’s t -test. (D) HUVECs were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of Smad2 phosphorylation by western blot.
Mab612, supplied by R&D Systems, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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tgf β1 polyclonal ab pab  (R&D Systems)
91
R&D Systems tgf β1 polyclonal ab pab
(A) Real-time PCR showing Tgfb1, Tgfb2, and Tgfb3 mRNA level in aortas of WT and P311–/– mice. (B) ELISA showing total levels of all 3 <t>TGF-βs</t> in aortas. (C) ELISA showing levels of total and active TGF-βs in blood serum. (D) Western blots showing expression of pro–TGF-βs and LAP-1 in aortas. Relative densities of Western blot signals in P311–/– aortas relative to WT (n = 15 per group) are also shown. (E) Myographic determination of the contractile response of TGF-β receptor inhibitor–treated (LY364947, 100 μM) resistance arteries to increasing K+ concentration (n = 3 per group). (F) Western blots showing total and phosphorylated Smad2 and Smad3 in aortas (n = 3 per group). Relative levels of Smad2 and Smad3 in P311–/– aortas relative to WT are also shown. (G) GFP immunoreactivity (brown staining) in CAGA12/GFP transgenic P311+/+ and P311–/– mouse aortas. Scale bar: 100 μm. Data represent mean ± SD. *P < 0.05, **P < 0.01, by 1-way ANOVA.
Tgf β1 Polyclonal Ab Pab, supplied by R&D Systems, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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room temperature  (R&D Systems)
92
R&D Systems room temperature
(A) Real-time PCR showing Tgfb1, Tgfb2, and Tgfb3 mRNA level in aortas of WT and P311–/– mice. (B) ELISA showing total levels of all 3 <t>TGF-βs</t> in aortas. (C) ELISA showing levels of total and active TGF-βs in blood serum. (D) Western blots showing expression of pro–TGF-βs and LAP-1 in aortas. Relative densities of Western blot signals in P311–/– aortas relative to WT (n = 15 per group) are also shown. (E) Myographic determination of the contractile response of TGF-β receptor inhibitor–treated (LY364947, 100 μM) resistance arteries to increasing K+ concentration (n = 3 per group). (F) Western blots showing total and phosphorylated Smad2 and Smad3 in aortas (n = 3 per group). Relative levels of Smad2 and Smad3 in P311–/– aortas relative to WT are also shown. (G) GFP immunoreactivity (brown staining) in CAGA12/GFP transgenic P311+/+ and P311–/– mouse aortas. Scale bar: 100 μm. Data represent mean ± SD. *P < 0.05, **P < 0.01, by 1-way ANOVA.
Room Temperature, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti tgf β2 antibody  (R&D Systems)
93
R&D Systems anti tgf β2 antibody
(A) Real-time PCR showing Tgfb1, Tgfb2, and Tgfb3 mRNA level in aortas of WT and P311–/– mice. (B) ELISA showing total levels of all 3 <t>TGF-βs</t> in aortas. (C) ELISA showing levels of total and active TGF-βs in blood serum. (D) Western blots showing expression of pro–TGF-βs and LAP-1 in aortas. Relative densities of Western blot signals in P311–/– aortas relative to WT (n = 15 per group) are also shown. (E) Myographic determination of the contractile response of TGF-β receptor inhibitor–treated (LY364947, 100 μM) resistance arteries to increasing K+ concentration (n = 3 per group). (F) Western blots showing total and phosphorylated Smad2 and Smad3 in aortas (n = 3 per group). Relative levels of Smad2 and Smad3 in P311–/– aortas relative to WT are also shown. (G) GFP immunoreactivity (brown staining) in CAGA12/GFP transgenic P311+/+ and P311–/– mouse aortas. Scale bar: 100 μm. Data represent mean ± SD. *P < 0.05, **P < 0.01, by 1-way ANOVA.
Anti Tgf β2 Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Image Search Results


To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) TGF-β2 was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.

Journal: PeerJ

Article Title: CircHIPK3 regulates cardiac fibroblast proliferation, migration and phenotypic switching through the miR-152-3p/TGF-β2 axis under hypoxia

doi: 10.7717/peerj.9796

Figure Lengend Snippet: To determine the role of miR-152-3p in CFs, CFs were transfected with the miR-152-3p mimic or inhibitor. (A–G and J–P) Representative images of EdU-stained CFs from different groups. Quantification of EdU+ cells presented as the % EdU-positive cells and Hoechst-stained nuclei. The orange color is EdU-positive cells, and the blue color is nuclei stained by Hoechst 33342. Scale bar = 20 μm. n = 10. * P < 0.05, vs. MNC group or INC group. (H, I and Q, R) The protein expression levels of Col I and Col III were measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group. (S) Relative expression of miR-152-3p in CFs after transfection with miR-152-3p mimic or inhibitor was examined by RT-qPCR analysis. n = 3. * P < 0.05, vs. hypoxia group. (T) TGF-β2 was predicted as a target gene of miR-152-3p using the TargetScan database. HEK-293T cells were cotransfected with RLuc-TGF-β2-WT or RLuc-TGF-β2-Mut and miR-152-3p mimics or NC mimic. Luciferase activity was detected using the dual luciferase reporter assay at 48 h post-transfection. n = 3. * P < 0.05, vs. TGF-β2-WT + NC group. # P < 0.05, vs. TGF-β2-WT + miR-152-3p group. (U–X) The protein expression level of TGF-β2 was measured by western blot analysis. n = 3. * P < 0.05, vs. MNC group or INC group.

Article Snippet: The primary anti-bodies: Anti-Collagen I (Cell Signaling Technology, 91144, dilution: 1:1,000), Anti-Collagen III (Abcam, ab7778, dilution: 1:7,000), Anti-α-SMA (Cell Signaling Technology, 19245, dilution: 1:1,000), Anti-p-Smad2 (Cell Signaling Technology, 18338, dilution: 1:1,000), Anti-p-Smad3 (Cell Signaling Technology, 9520, dilution: 1:1,000), Anti-TGF-β2 (Bioss, bs-20412R, dilution: 1:1,000) and Anti-β-actin Antibody (Abcam, ab8227, dilution: 1:1,000), were used.

Techniques: Transfection, Staining, Expressing, Western Blot, Quantitative RT-PCR, Luciferase, Activity Assay, Reporter Assay

(A–E) The cell cycle was investigated by a cell cycle assay in cardiac fibroblasts. The results represent the percentage of cells in G1 and S + G2/M phases for the indicated conditions. n = 3. * P < 0.05, # P < 0.05. (F–J) Wound scratch assay for the different groups. The average sizes of the gaps were measured at 48 h. Scale bar = 200 μm. n = 5. * P < 0.05, # P < 0.05. (K and L) The protein expression levels of Col I, Col III, TGF-β2, p-Smad2 and p-Smad3 were measured by western blot analysis. n = 3. * P < 0.05, # P < 0.05. * P < 0.05, vs. hypoxia group, # P < 0.05, vs. si-circHIPK3 group.

Journal: PeerJ

Article Title: CircHIPK3 regulates cardiac fibroblast proliferation, migration and phenotypic switching through the miR-152-3p/TGF-β2 axis under hypoxia

doi: 10.7717/peerj.9796

Figure Lengend Snippet: (A–E) The cell cycle was investigated by a cell cycle assay in cardiac fibroblasts. The results represent the percentage of cells in G1 and S + G2/M phases for the indicated conditions. n = 3. * P < 0.05, # P < 0.05. (F–J) Wound scratch assay for the different groups. The average sizes of the gaps were measured at 48 h. Scale bar = 200 μm. n = 5. * P < 0.05, # P < 0.05. (K and L) The protein expression levels of Col I, Col III, TGF-β2, p-Smad2 and p-Smad3 were measured by western blot analysis. n = 3. * P < 0.05, # P < 0.05. * P < 0.05, vs. hypoxia group, # P < 0.05, vs. si-circHIPK3 group.

Article Snippet: The primary anti-bodies: Anti-Collagen I (Cell Signaling Technology, 91144, dilution: 1:1,000), Anti-Collagen III (Abcam, ab7778, dilution: 1:7,000), Anti-α-SMA (Cell Signaling Technology, 19245, dilution: 1:1,000), Anti-p-Smad2 (Cell Signaling Technology, 18338, dilution: 1:1,000), Anti-p-Smad3 (Cell Signaling Technology, 9520, dilution: 1:1,000), Anti-TGF-β2 (Bioss, bs-20412R, dilution: 1:1,000) and Anti-β-actin Antibody (Abcam, ab8227, dilution: 1:1,000), were used.

Techniques: Cell Cycle Assay, Wound Healing Assay, Expressing, Western Blot

(A) HUVECs were transfected with either control mimic (con) or miR-30b mimic (30b) and levels of TGFβ1 and TGFβ2 mRNA were assessed by qRT-PCR. Expression levels relative to control mimic transfected cells and normalized to β-actin expression are presented as the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 expression. * P < 0.05, ** P < 0.01 as determined by unpaired Student’s t -test. (B) Cells were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of TGFβ2 protein levels by western blot. β-actin was used as endogenous control. (C) ELISAs for TGFβ1 and TGFβ2 were performed with 24 hour conditioned supernates from HUVECs transfected with 20 nM of either control or miR-30b mimic. Data represents the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 secretion into cell culture supernate. * P = 0.044 as determined by unpaired Student’s t -test. (D) HUVECs were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of Smad2 phosphorylation by western blot.

Journal: PLoS ONE

Article Title: MicroRNA-30b controls endothelial cell capillary morphogenesis through regulation of transforming growth factor beta 2

doi: 10.1371/journal.pone.0185619

Figure Lengend Snippet: (A) HUVECs were transfected with either control mimic (con) or miR-30b mimic (30b) and levels of TGFβ1 and TGFβ2 mRNA were assessed by qRT-PCR. Expression levels relative to control mimic transfected cells and normalized to β-actin expression are presented as the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 expression. * P < 0.05, ** P < 0.01 as determined by unpaired Student’s t -test. (B) Cells were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of TGFβ2 protein levels by western blot. β-actin was used as endogenous control. (C) ELISAs for TGFβ1 and TGFβ2 were performed with 24 hour conditioned supernates from HUVECs transfected with 20 nM of either control or miR-30b mimic. Data represents the mean ± SEM (n = 2). Overexpression of miR-30b significantly increases TGFβ2 secretion into cell culture supernate. * P = 0.044 as determined by unpaired Student’s t -test. (D) HUVECs were transfected with 20 nM of either control mimic (control) or miR-30b mimic (miR-30b) and protein lysates were collected after 48 hours for assessment of Smad2 phosphorylation by western blot.

Article Snippet: Primary antibodies used were: TGFβ2 (V, SC-90), ATF-2 (C-19, SC-187), and phospho-ATF-2 (F-1, SC-8398) from Santa Cruz Biotechnology (Santa Cruz, CA), phospho-Smad2 (S465/467) from Cell Signaling Technology (3101; Danvers, MA), Smad2 from Invitrogen (511300; Carlsbad, CA), β-Actin (clone AC-74) from Sigma-Aldrich (A5316; St. Louis, MO), anti-TGFβ2 neutralizing antibody (AB-12-NA) and Normal Rabbit IgG (AB-105-C) from R&D Systems (Minneapolis, MN).

Techniques: Transfection, Control, Quantitative RT-PCR, Expressing, Over Expression, Western Blot, Cell Culture, Phospho-proteomics

(A) JDP2 mRNA expression was assessed in HUVECs transfected with miR-30b mimic (20 nM) as compared to control by qRT-PCR. Data represents the mean ± SEM (n = 3) normalized to β-actin as endogenous control. * P = 0.016 as determined by unpaired Student’s t -test. (B) HUVEC were transfected with 50 nM of either control siRNA or ATF2 siRNA 1 or 2 and RNA was isolated at 48 hours post transfection. Levels of ATF2 and TGFβ2 mRNA were assessed by qRT-PCR with β-actin as endogenous control. Data presented is mean ± SEM (n = 2). Statistically significant decreases in ATF2 and TGFβ2 expression were seen in ATF2 siRNA treated cells as compared to control siRNA treated cells. * P < 0.05, ** P < 0.01, *** P < 0.001 as determined by unpaired Student’s t -tests for each ATF2 siRNA compared to control siRNA. (C) Cells transfected with 5 nM of either control siRNA or ATF2 siRNA 1 were seeded onto growth factor reduced BME and the formation of capillary-like cord structures and number of loops was assessed after 24 hours. (D) A statistically significant increase in cord formation was observed in cells depleted of ATF2 through siRNA. Data represents the mean ± SEM (n = 2). * P = 0.041 as determined by unpaired Student’s t -test. (E) HUVECs were co-transfected with miRNA mimic (20 nM) and ATF2 siRNA 1 or 2 (50 nM) in the combinations displayed and cell lysates were collected at 48 hours post transfection and assessed for TGFβ2 mRNA expression. Data presented is mean ± SEM (n = 2). * P < 0.05, ** P < 0.01, *** P < 0.001 as determined by ANOVA with post hoc analysis. (F) Cells were transfected as in (E) using miRNA mimic (20 nM) and ATF2 siRNA 1 (5 nM) and serum starved overnight in MCDB 131 with 0.5% FBS prior to protein expression analysis by western blot. Data is representative of expression levels observed in two independently performed experiments.

Journal: PLoS ONE

Article Title: MicroRNA-30b controls endothelial cell capillary morphogenesis through regulation of transforming growth factor beta 2

doi: 10.1371/journal.pone.0185619

Figure Lengend Snippet: (A) JDP2 mRNA expression was assessed in HUVECs transfected with miR-30b mimic (20 nM) as compared to control by qRT-PCR. Data represents the mean ± SEM (n = 3) normalized to β-actin as endogenous control. * P = 0.016 as determined by unpaired Student’s t -test. (B) HUVEC were transfected with 50 nM of either control siRNA or ATF2 siRNA 1 or 2 and RNA was isolated at 48 hours post transfection. Levels of ATF2 and TGFβ2 mRNA were assessed by qRT-PCR with β-actin as endogenous control. Data presented is mean ± SEM (n = 2). Statistically significant decreases in ATF2 and TGFβ2 expression were seen in ATF2 siRNA treated cells as compared to control siRNA treated cells. * P < 0.05, ** P < 0.01, *** P < 0.001 as determined by unpaired Student’s t -tests for each ATF2 siRNA compared to control siRNA. (C) Cells transfected with 5 nM of either control siRNA or ATF2 siRNA 1 were seeded onto growth factor reduced BME and the formation of capillary-like cord structures and number of loops was assessed after 24 hours. (D) A statistically significant increase in cord formation was observed in cells depleted of ATF2 through siRNA. Data represents the mean ± SEM (n = 2). * P = 0.041 as determined by unpaired Student’s t -test. (E) HUVECs were co-transfected with miRNA mimic (20 nM) and ATF2 siRNA 1 or 2 (50 nM) in the combinations displayed and cell lysates were collected at 48 hours post transfection and assessed for TGFβ2 mRNA expression. Data presented is mean ± SEM (n = 2). * P < 0.05, ** P < 0.01, *** P < 0.001 as determined by ANOVA with post hoc analysis. (F) Cells were transfected as in (E) using miRNA mimic (20 nM) and ATF2 siRNA 1 (5 nM) and serum starved overnight in MCDB 131 with 0.5% FBS prior to protein expression analysis by western blot. Data is representative of expression levels observed in two independently performed experiments.

Article Snippet: Primary antibodies used were: TGFβ2 (V, SC-90), ATF-2 (C-19, SC-187), and phospho-ATF-2 (F-1, SC-8398) from Santa Cruz Biotechnology (Santa Cruz, CA), phospho-Smad2 (S465/467) from Cell Signaling Technology (3101; Danvers, MA), Smad2 from Invitrogen (511300; Carlsbad, CA), β-Actin (clone AC-74) from Sigma-Aldrich (A5316; St. Louis, MO), anti-TGFβ2 neutralizing antibody (AB-12-NA) and Normal Rabbit IgG (AB-105-C) from R&D Systems (Minneapolis, MN).

Techniques: Expressing, Transfection, Control, Quantitative RT-PCR, Isolation, Western Blot

(A) HUVECs were serum starved overnight in MCDB 131 with 0.5% FBS and stimulated with VEGF (50 ng/ml) in the presence or absence of Avastin (1 μg/ml) for 24 hours. Data represents the mean ± SEM (n = 2) for expression of TGFβ1 and TGFβ2 assessed by qRT-PCR relative to β-actin endogenous control. * P < 0.05, ** P < 0.01, *** P < 0.001 as determined by ANOVA. (B) HUVECs were treated with 5 ng/ml of TGFβ2 for 3 days prior to seeding onto growth factor reduced BME for assessment of capillary-like cord formation after 24 hours. (C) A significant decrease in cord formation is observed in the TGFβ2 treated group. Data represents the mean ± SEM (n = 2). ** P = 0.0072 as determined by unpaired Student’s t -test. (D) HUVECs transfected with 1 nM control or miR-30b mimic were treated 4 hours post transfection with 0.8 μg/ml anti-TGFβ2 neutralizing antibody or rabbit IgG. Media was refreshed after 24 hours, again with rabbit IgG or anti-TGFβ2 antibody and cells were seeded onto growth factor reduced BME 24 hours later (ie. 48 hours post transfection) in media containing rabbit IgG or anti-TGFβ2 antibody. (E) Data represents the mean ± SEM (n = 3) of the number of capillary-like cord structures or number of loops formed after 24 hours on BME. * P < 0.05, ns denotes not significant as determined by ANOVA with post hoc analysis.

Journal: PLoS ONE

Article Title: MicroRNA-30b controls endothelial cell capillary morphogenesis through regulation of transforming growth factor beta 2

doi: 10.1371/journal.pone.0185619

Figure Lengend Snippet: (A) HUVECs were serum starved overnight in MCDB 131 with 0.5% FBS and stimulated with VEGF (50 ng/ml) in the presence or absence of Avastin (1 μg/ml) for 24 hours. Data represents the mean ± SEM (n = 2) for expression of TGFβ1 and TGFβ2 assessed by qRT-PCR relative to β-actin endogenous control. * P < 0.05, ** P < 0.01, *** P < 0.001 as determined by ANOVA. (B) HUVECs were treated with 5 ng/ml of TGFβ2 for 3 days prior to seeding onto growth factor reduced BME for assessment of capillary-like cord formation after 24 hours. (C) A significant decrease in cord formation is observed in the TGFβ2 treated group. Data represents the mean ± SEM (n = 2). ** P = 0.0072 as determined by unpaired Student’s t -test. (D) HUVECs transfected with 1 nM control or miR-30b mimic were treated 4 hours post transfection with 0.8 μg/ml anti-TGFβ2 neutralizing antibody or rabbit IgG. Media was refreshed after 24 hours, again with rabbit IgG or anti-TGFβ2 antibody and cells were seeded onto growth factor reduced BME 24 hours later (ie. 48 hours post transfection) in media containing rabbit IgG or anti-TGFβ2 antibody. (E) Data represents the mean ± SEM (n = 3) of the number of capillary-like cord structures or number of loops formed after 24 hours on BME. * P < 0.05, ns denotes not significant as determined by ANOVA with post hoc analysis.

Article Snippet: Primary antibodies used were: TGFβ2 (V, SC-90), ATF-2 (C-19, SC-187), and phospho-ATF-2 (F-1, SC-8398) from Santa Cruz Biotechnology (Santa Cruz, CA), phospho-Smad2 (S465/467) from Cell Signaling Technology (3101; Danvers, MA), Smad2 from Invitrogen (511300; Carlsbad, CA), β-Actin (clone AC-74) from Sigma-Aldrich (A5316; St. Louis, MO), anti-TGFβ2 neutralizing antibody (AB-12-NA) and Normal Rabbit IgG (AB-105-C) from R&D Systems (Minneapolis, MN).

Techniques: Expressing, Quantitative RT-PCR, Control, Transfection

(A) Real-time PCR showing Tgfb1, Tgfb2, and Tgfb3 mRNA level in aortas of WT and P311–/– mice. (B) ELISA showing total levels of all 3 TGF-βs in aortas. (C) ELISA showing levels of total and active TGF-βs in blood serum. (D) Western blots showing expression of pro–TGF-βs and LAP-1 in aortas. Relative densities of Western blot signals in P311–/– aortas relative to WT (n = 15 per group) are also shown. (E) Myographic determination of the contractile response of TGF-β receptor inhibitor–treated (LY364947, 100 μM) resistance arteries to increasing K+ concentration (n = 3 per group). (F) Western blots showing total and phosphorylated Smad2 and Smad3 in aortas (n = 3 per group). Relative levels of Smad2 and Smad3 in P311–/– aortas relative to WT are also shown. (G) GFP immunoreactivity (brown staining) in CAGA12/GFP transgenic P311+/+ and P311–/– mouse aortas. Scale bar: 100 μm. Data represent mean ± SD. *P < 0.05, **P < 0.01, by 1-way ANOVA.

Journal: The Journal of Clinical Investigation

Article Title: Blood pressure homeostasis is maintained by a P311-TGF-? axis

doi: 10.1172/JCI69884

Figure Lengend Snippet: (A) Real-time PCR showing Tgfb1, Tgfb2, and Tgfb3 mRNA level in aortas of WT and P311–/– mice. (B) ELISA showing total levels of all 3 TGF-βs in aortas. (C) ELISA showing levels of total and active TGF-βs in blood serum. (D) Western blots showing expression of pro–TGF-βs and LAP-1 in aortas. Relative densities of Western blot signals in P311–/– aortas relative to WT (n = 15 per group) are also shown. (E) Myographic determination of the contractile response of TGF-β receptor inhibitor–treated (LY364947, 100 μM) resistance arteries to increasing K+ concentration (n = 3 per group). (F) Western blots showing total and phosphorylated Smad2 and Smad3 in aortas (n = 3 per group). Relative levels of Smad2 and Smad3 in P311–/– aortas relative to WT are also shown. (G) GFP immunoreactivity (brown staining) in CAGA12/GFP transgenic P311+/+ and P311–/– mouse aortas. Scale bar: 100 μm. Data represent mean ± SD. *P < 0.05, **P < 0.01, by 1-way ANOVA.

Article Snippet: Secondary Abs — TGF-β1 polyclonal Ab (pAb) (supplied in the kit), TGF-β2 pAb (BAF302; R&D Systems), and TGF-β3 pAb (BAF243; R&D Systems) — were added to complete the sandwich, followed by the chromogenic substrate 3,3′,5,5′-tetramethyl benzidine (TMB), stop solution (1 N HCl), and recording of absorbance at 450 nm.

Techniques: Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Western Blot, Expressing, Concentration Assay, Staining, Transgenic Assay

(A) pGL3 UTR luciferase reporter constructs. (B) Luciferase reporter assay for TGF-β 5′- and 3′-UTRs in P311–/– mouse VSMCs transfected with P311 or empty vector. (C) P311-mRNA immunoprecipitation showing enrichment of 5′-UTR Tgfb1, Tgfb2, and Tgfb3 mRNAs as well as controls Inhba (TGF-β superfamily member), B9d2 (TGF-β neighboring gene on chromosome 7), Actb, and Tuba1a. Data represent mean ± SD. *P < 0.05, **P < 0.01, 1-way ANOVA. Unless otherwise specified, experiments were performed at least 3 times.

Journal: The Journal of Clinical Investigation

Article Title: Blood pressure homeostasis is maintained by a P311-TGF-? axis

doi: 10.1172/JCI69884

Figure Lengend Snippet: (A) pGL3 UTR luciferase reporter constructs. (B) Luciferase reporter assay for TGF-β 5′- and 3′-UTRs in P311–/– mouse VSMCs transfected with P311 or empty vector. (C) P311-mRNA immunoprecipitation showing enrichment of 5′-UTR Tgfb1, Tgfb2, and Tgfb3 mRNAs as well as controls Inhba (TGF-β superfamily member), B9d2 (TGF-β neighboring gene on chromosome 7), Actb, and Tuba1a. Data represent mean ± SD. *P < 0.05, **P < 0.01, 1-way ANOVA. Unless otherwise specified, experiments were performed at least 3 times.

Article Snippet: Secondary Abs — TGF-β1 polyclonal Ab (pAb) (supplied in the kit), TGF-β2 pAb (BAF302; R&D Systems), and TGF-β3 pAb (BAF243; R&D Systems) — were added to complete the sandwich, followed by the chromogenic substrate 3,3′,5,5′-tetramethyl benzidine (TMB), stop solution (1 N HCl), and recording of absorbance at 450 nm.

Techniques: Luciferase, Construct, Reporter Assay, Transfection, Plasmid Preparation, Immunoprecipitation

(A) ELISA showing total and active TGF-βs in blood serum of WT and P311–/– mice (n = 4 per group), treated or not with rTGF-β1-3. (B) Plethysmographic blood pressure determination (n = 5 per group), with and without rTGF-β1-3 treatment. (C) Echocardiographic aortic diameter in conscious mice (n = 5 per group) at basal levels and after treatment with rTGF-β1-3 or vehicle. (D) Contractile response of mouse aortas (n = 5 per group) to increasing K+ concentration, with and without rTGF-β1-3 treatment. (E) RhoA activity assay showing total and active RhoA in mouse aortas (n = 3 per group), with and without rTGF-β1-3 treatment. Relative density of total and active RhoA is also shown. (F) Collagen gel contraction by mouse aorta–derived VSMCs, with and without rTGF-β1-3 treatment. Relative contraction is also shown, calculated as percent decrease in collagen gel diameter from the original diameter. Gel contraction data were collected from triplicate wells. Data represent mean ± SD. *P < 0.05, **P < 0.01, 1-way ANOVA.

Journal: The Journal of Clinical Investigation

Article Title: Blood pressure homeostasis is maintained by a P311-TGF-? axis

doi: 10.1172/JCI69884

Figure Lengend Snippet: (A) ELISA showing total and active TGF-βs in blood serum of WT and P311–/– mice (n = 4 per group), treated or not with rTGF-β1-3. (B) Plethysmographic blood pressure determination (n = 5 per group), with and without rTGF-β1-3 treatment. (C) Echocardiographic aortic diameter in conscious mice (n = 5 per group) at basal levels and after treatment with rTGF-β1-3 or vehicle. (D) Contractile response of mouse aortas (n = 5 per group) to increasing K+ concentration, with and without rTGF-β1-3 treatment. (E) RhoA activity assay showing total and active RhoA in mouse aortas (n = 3 per group), with and without rTGF-β1-3 treatment. Relative density of total and active RhoA is also shown. (F) Collagen gel contraction by mouse aorta–derived VSMCs, with and without rTGF-β1-3 treatment. Relative contraction is also shown, calculated as percent decrease in collagen gel diameter from the original diameter. Gel contraction data were collected from triplicate wells. Data represent mean ± SD. *P < 0.05, **P < 0.01, 1-way ANOVA.

Article Snippet: Secondary Abs — TGF-β1 polyclonal Ab (pAb) (supplied in the kit), TGF-β2 pAb (BAF302; R&D Systems), and TGF-β3 pAb (BAF243; R&D Systems) — were added to complete the sandwich, followed by the chromogenic substrate 3,3′,5,5′-tetramethyl benzidine (TMB), stop solution (1 N HCl), and recording of absorbance at 450 nm.

Techniques: Enzyme-linked Immunosorbent Assay, Concentration Assay, Activity Assay, Derivative Assay

(A) Western blots showing pro–TGF-βs in WT and P311TG mouse aortas. Relative density of TGF-βs in aortas is also shown (n = 6 per group). (B) ELISA showing levels of total and active TGF-βs in blood serum. (C) GFP immunoreactivity (brown staining) in CAGA12/GFP-transgenic P311+/+ and P311TG mouse aortas. Original magnification, ×10. (D) Plethysmographic blood pressure determination (n = 8 per group). Experiments in which the number of animals is not specified were conducted 3 times. Data represent mean ± SD. *P < 0.05, **P < 0.01, 1-way ANOVA.

Journal: The Journal of Clinical Investigation

Article Title: Blood pressure homeostasis is maintained by a P311-TGF-? axis

doi: 10.1172/JCI69884

Figure Lengend Snippet: (A) Western blots showing pro–TGF-βs in WT and P311TG mouse aortas. Relative density of TGF-βs in aortas is also shown (n = 6 per group). (B) ELISA showing levels of total and active TGF-βs in blood serum. (C) GFP immunoreactivity (brown staining) in CAGA12/GFP-transgenic P311+/+ and P311TG mouse aortas. Original magnification, ×10. (D) Plethysmographic blood pressure determination (n = 8 per group). Experiments in which the number of animals is not specified were conducted 3 times. Data represent mean ± SD. *P < 0.05, **P < 0.01, 1-way ANOVA.

Article Snippet: Secondary Abs — TGF-β1 polyclonal Ab (pAb) (supplied in the kit), TGF-β2 pAb (BAF302; R&D Systems), and TGF-β3 pAb (BAF243; R&D Systems) — were added to complete the sandwich, followed by the chromogenic substrate 3,3′,5,5′-tetramethyl benzidine (TMB), stop solution (1 N HCl), and recording of absorbance at 450 nm.

Techniques: Western Blot, Enzyme-linked Immunosorbent Assay, Staining, Transgenic Assay

(A) Real-time PCR showing mRNA levels of P311 in aortas from normotensive (NT) and hypertensive (HT) patients (n = 5 per group). (B) P311 immunoreactivity (brown staining) in hernia sac arteries from 18 normotensive and 21 hypertensive patients. Scale bar: 50 μm. Quantified P311 signal intensity is also shown. (C) TGF-β immunoreactivity (brown staining) in hernia sac arteries from 18 normotensive and 21 hypertensive patients. Quantified TGF-β signal intensity is also shown. Original magnification, ×25. (D) Smad2/3 immunoreactivity (brown staining) in hernia sac arteries from normotensive and hypertensive patients. Original magnification, ×100. The number of positive Smad2/3 nuclei is also shown. Data represent mean ± SD. **P < 0.01, 1-way ANOVA.

Journal: The Journal of Clinical Investigation

Article Title: Blood pressure homeostasis is maintained by a P311-TGF-? axis

doi: 10.1172/JCI69884

Figure Lengend Snippet: (A) Real-time PCR showing mRNA levels of P311 in aortas from normotensive (NT) and hypertensive (HT) patients (n = 5 per group). (B) P311 immunoreactivity (brown staining) in hernia sac arteries from 18 normotensive and 21 hypertensive patients. Scale bar: 50 μm. Quantified P311 signal intensity is also shown. (C) TGF-β immunoreactivity (brown staining) in hernia sac arteries from 18 normotensive and 21 hypertensive patients. Quantified TGF-β signal intensity is also shown. Original magnification, ×25. (D) Smad2/3 immunoreactivity (brown staining) in hernia sac arteries from normotensive and hypertensive patients. Original magnification, ×100. The number of positive Smad2/3 nuclei is also shown. Data represent mean ± SD. **P < 0.01, 1-way ANOVA.

Article Snippet: Secondary Abs — TGF-β1 polyclonal Ab (pAb) (supplied in the kit), TGF-β2 pAb (BAF302; R&D Systems), and TGF-β3 pAb (BAF243; R&D Systems) — were added to complete the sandwich, followed by the chromogenic substrate 3,3′,5,5′-tetramethyl benzidine (TMB), stop solution (1 N HCl), and recording of absorbance at 450 nm.

Techniques: Real-time Polymerase Chain Reaction, Staining