Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: The primer sequences used in this study.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques:

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: Smad signaling pathway was related to inflammatory microenvironment. (a) Western blot analysis and relevant quantitative analysis of P-Smad1/5, Smad1/5, and ID1 in TSCs. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. mRNA expression analysis of Smad5 and ID1. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques: Western Blot, Expressing

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: Smad and NF- κ B signaling pathways play a synergistic role in HO formation, and palovarotene inhibits HO formation by blocking the Smad and NF- κ B signaling pathways. (a) The mRNA expression of OCN, SOX9, RUNX2, Smad1, Smad5, and ID1. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. (b) Immunofluorescence against ID1 ( n = 3). Scale bar = 25 μ m. (c) Western blot analysis and relevant quantitative analysis of p-p65 and ID1. Data are means ± SD ( n = 3) ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. (d) Immunofluorescence against p65 ( n = 3). Scale bar = 25 μ m. (e) Western blot analysis and relevant quantitative analysis of p65 and immunofluorescence against P65 in the previous coculture model series experiments ( n = 3). Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. Scale bar = 25 μ m.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques: Protein-Protein interactions, Blocking Assay, Expressing, Immunofluorescence, Western Blot

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: In vitro experimental further verifies the effects of palovarotene and Smad and NF- κ B signaling pathways in HO. In immunohistochemistry, the positive rate of inflammation such as TNF- α , TGF- β , IFN- γ , IL-1 β , and MMP-9 was increased (a), as well as p65, ID1, p-SMAD1/5, and osteogenic genes, concluding OCN and SOX9 (b). Scale bar = 25 μ m. (c) Immunofluorescence and quantitative analysis against Smad5. Scale bar = 25 μ m. Data are means ± SD ( n = 3) ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. (d) The mRNA expression, Western blot analysis of tendon tissues, and relevant quantitative analysis of p65, ID1, OCN, and SOX9. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques: In Vitro, Protein-Protein interactions, Immunohistochemistry, Immunofluorescence, Expressing, Western Blot

Journal: Scientific Reports

Article Title: TPPP3 promote epithelial-mesenchymal transition via Snail1 in glioblastoma

doi: 10.1038/s41598-023-45233-w

Figure Lengend Snippet: TPPP3 induced epithelial-mesenchymal transition by regulating the expression of Snail1 protein. ( A ) Western Blot analysis the effect of knocking down the expression of TPPP3 on the expression of common key proteins in the EMT process. ( B ) The effect of overexpression of Snail1 in cells with knockdown of TPPP3 on the expression of E-cadherin, N-cadherin and Vimentin, the key markers of EMT process. ( C , D ) Snail1 was overexpressed in TPPP3 knockdown cells, and the invasion ability of glioma cells was quantitatively analyzed by Transwell experiment. n = 5 each group. ( E , F ) The wound healing experiment tested the migration ability of glioblastoma cells after overexpression of Snail1 in TPPP3 knockdown cells. n = 5 each group. ( G ) The CCK8 experiment analyzed the effect of overexpression of Snail1 in TPPP3 knockdown cells on the proliferation of glioblastoma cells. ( H ) Flow cytometry analysis detected the apoptosis of glioblastoma cells after overexpression of Snail1 in TPPP3 knockdown cells. n = 4 each group. Data from at least three independent experiments were quantified. ** P < 0.01, *** P < 0.001.

Article Snippet: The involved primary antibodies were as follow and concentration was diluted according to product instructions: TPPP3 (NBP2-13469, NBP2-95209, Novus Biologicals), and N-cadherin (ab76011, Abcam), E-cadherin (ab40772, Abcam), Vimentin (ab8978, Abcam), Snail1 (13099-1-AP, Proteintech), Slug (ab302780, Abcam), Twist1 (ab175430, Abcam), ZEB1 (ab203829, Abcam), β-catenin(ab203829, Abcam).

Techniques: Expressing, Western Blot, Over Expression, Knockdown, Migration, Flow Cytometry

Journal: Scientific Reports

Article Title: TPPP3 promote epithelial-mesenchymal transition via Snail1 in glioblastoma

doi: 10.1038/s41598-023-45233-w

Figure Lengend Snippet: Data analysis of clinical significance of TPPP3 in glioblastoma. ( A ) Immunohistochemical images of representative glioblastoma tissues with high and low TPPP3 expression. ( B ) Correlation analysis between TPPP3 and Snail1 in glioblastoma tissues. ( C , D ) The expression level of TPPP3 in glioblastoma affected the survival of patients. HR, Hazard Ratio. **** P < 0.0001.

Article Snippet: The involved primary antibodies were as follow and concentration was diluted according to product instructions: TPPP3 (NBP2-13469, NBP2-95209, Novus Biologicals), and N-cadherin (ab76011, Abcam), E-cadherin (ab40772, Abcam), Vimentin (ab8978, Abcam), Snail1 (13099-1-AP, Proteintech), Slug (ab302780, Abcam), Twist1 (ab175430, Abcam), ZEB1 (ab203829, Abcam), β-catenin(ab203829, Abcam).

Techniques: Immunohistochemical staining, Expressing

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: The primer sequences used in this study.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques:

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: Smad signaling pathway was related to inflammatory microenvironment. (a) Western blot analysis and relevant quantitative analysis of P-Smad1/5, Smad1/5, and ID1 in TSCs. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. mRNA expression analysis of Smad5 and ID1. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques: Western Blot, Expressing

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: Smad and NF- κ B signaling pathways play a synergistic role in HO formation, and palovarotene inhibits HO formation by blocking the Smad and NF- κ B signaling pathways. (a) The mRNA expression of OCN, SOX9, RUNX2, Smad1, Smad5, and ID1. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. (b) Immunofluorescence against ID1 ( n = 3). Scale bar = 25 μ m. (c) Western blot analysis and relevant quantitative analysis of p-p65 and ID1. Data are means ± SD ( n = 3) ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. (d) Immunofluorescence against p65 ( n = 3). Scale bar = 25 μ m. (e) Western blot analysis and relevant quantitative analysis of p65 and immunofluorescence against P65 in the previous coculture model series experiments ( n = 3). Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. Scale bar = 25 μ m.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques: Protein-Protein interactions, Blocking Assay, Expressing, Immunofluorescence, Western Blot

Journal: Stem Cells International

Article Title: Palovarotene Can Attenuate Heterotopic Ossification Induced by Tendon Stem Cells by Downregulating the Synergistic Effects of Smad and NF- κ B Signaling Pathway following Stimulation of the Inflammatory Microenvironment

doi: 10.1155/2022/1560943

Figure Lengend Snippet: In vitro experimental further verifies the effects of palovarotene and Smad and NF- κ B signaling pathways in HO. In immunohistochemistry, the positive rate of inflammation such as TNF- α , TGF- β , IFN- γ , IL-1 β , and MMP-9 was increased (a), as well as p65, ID1, p-SMAD1/5, and osteogenic genes, concluding OCN and SOX9 (b). Scale bar = 25 μ m. (c) Immunofluorescence and quantitative analysis against Smad5. Scale bar = 25 μ m. Data are means ± SD ( n = 3) ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01. (d) The mRNA expression, Western blot analysis of tendon tissues, and relevant quantitative analysis of p65, ID1, OCN, and SOX9. Data are means ± SD ( n = 3). ∗ p < 0.05 and ∗∗ p < 0.01. # p < 0.05 and ## p < 0.01.

Article Snippet: After 15 min at room temperature, the liquid turned transparent, and 5% BSA/PBS was added, and the cells were incubated at 37°C for 1 h. The TPPP3 (Biorbyt, #5-F10) or CCR7 (Abcam, # ab32527) or p65 (Abcam, # ab16502) or Smad5 (Affinity, #AF5119) or Id1 (Proteintech, #18475-1--AP) antibody working fluid was added, and the cells were incubated at 4°C overnight.

Techniques: In Vitro, Protein-Protein interactions, Immunohistochemistry, Immunofluorescence, Expressing, Western Blot