Journal: Cell reports

Article Title: PAI-1 uncouples integrin-β1 from restrain by membrane-bound β-catenin to promote collagen fibril remodeling in obesity-related neoplasms

doi: 10.1016/j.celrep.2024.114527

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Article Snippet: Mouse monoclonal anti-COL1 (E3E1X) , Cell Signaling Technology , Cat# 66948S; RRID: AB_2920541.

Techniques: Recombinant, Membrane, Cell Culture, Reverse Transcription, SYBR Green Assay, Clinical Proteomics, Protein Extraction, Extraction, Bicinchoninic Acid Protein Assay, In Situ, Blocking Assay, Migration, shRNA, Control, Software, Pyromark Assay, Western Blot, Simple Western

Journal: International Journal of Molecular Sciences

Article Title: Interleukin-17A Orchestrates Lung Injury and Remodeling Through p53 and uPA System Crosstalk

doi: 10.3390/ijms27041841

Figure Lengend Snippet: Regulation of BLM-LI and TSE-LI by IL-17A. ( A ) Lung sections of WT mice exposed to saline or BLM for 24–72 h were subjected to IHC staining for IL-17A antigen ( i ). AECs isolated from WT and IL-17A-deficient mice exposed to saline or BLM were analyzed for apoptosis and Sirt1 expression by Western blotting (WB) three days later ( ii ). Lung sections of WT and IL-17A −/− mice exposed to saline or BLM were subjected to Masson’s trichrome staining 21 days later ( iii ). Whole lung homogenates of WT and IL-17A −/− mice exposed to saline or BLM as in were analyzed for total HYP content ( iv ) or for Col-1 and FN by WB 21 days after BLM ( v ). ( B ) WT, p53 −/− and PAI-1 −/− mice were kept in ambient air or exposed to passive TS for 20 weeks. Total lung homogenates of control mice kept in ambient air, TSE WT mice, and p53 −/− or PAI-1 −/− mice were analyzed for IL-17A and IL-17RA protein by WB ( i ). Total RNA from mice kept in ambient air, TSE WT mice, and p53 −/− or PAI-1 −/− mice was analyzed for IL-17A mRNA ( ii ). AECs from TSE WT and IL-17A −/− mice were analyzed for p53, ACp53, cleaved (Cl.) caspase-3/caspase-3 and Sirt1 by WB ( iii ). AECs from lungs of IL-17A −/− mice kept in ambient air or TSE, as well as mice treated with or without CSP7 or CP, were analyzed for telomere length by qPCR ( iv ). AECs from lungs of IL-17A −/− mice treated as in ( B(iv )) were immunoblotted for the listed proteins ( v ). Lung sections of IL-17A −/− mice treated as in ( B ( iv )) were subjected to IHC for telomere binding shelterin complex (TRF1 and TRF2) proteins ( vi ).

Article Snippet: 14 , Anti-Col-1 mAb , Southern Biotech , #1340-01 , 1:1000 , .

Techniques: Saline, Immunohistochemistry, Isolation, Expressing, Western Blot, Staining, Control, Binding Assay

Journal:

Article Title:

doi:

Figure Lengend Snippet: Mineralization assay. After three days of osteogenic induction, ALP staining was hardly detected in both p75NTR +/+ and p75NTR ExIII−/− EMSCs (A), but on day 7, more abundant and deeper ALP staining was present in p75NTR +/+ EMSCs compared with that in p75NTR ExIII−/− EMSCs. On day 14, this difference became more prominent in not only ALP but also alizarin red staining. ALP staining was deeper and the mineralized nodules of alizarin red staining were larger in p75NTR +/+ EMSCs. Western blot on day 14 showed increased expression of the mineralization‐related markers ALP, Col1, Runx2, BSP, OCN and OPN in p75NTR +/+ EMSCs (B). Similar results were obtained by RT‐PCR on day 14 (C). The mRNA expression of ALP, Col‐1, Runx2, BSP, OCN and OPN in p75NTR +/+ EMSCs was significantly higher than that in p75NTR ExIII−/− EMSCs. Scale bar represents 100 μm. Abbreviations: K, knockout (p75NTR ExIII−/− ); W, wild type (p75NTR +/+ )

Article Snippet: After blocking with 5% bovine serum albumin for 2 hours at room temperature, the membranes were incubated overnight at 4°C with the following primary antibodies: rabbit polyclonal GAPDH (AP0063, 1:2000; Immunoway), rabbit polyclonal Runx2 (ab23981, 1:1000; Abcam), mouse monoclonal collagen type 1 (Col‐1, ab6308, 1:1500; Abcam), mouse monoclonal ALP (ab95462, 1:1500; Abcam), mouse monoclonal bone sialoprotein (BSP, ab125227, 1:1500; Abcam), mouse monoclonal osteocalcin (OCN, ab93876, 1:1500; Abcam), mouse monoclonal osteopontin (OPN, sc‐21742, 1:1500; Santa Cruz Biotechnology), rabbit polyclonal Mage‐D1 (sc‐28243, 1:500; Santa Cruz Biotechnology), rabbit polyclonal distal‐less homeobox 1 (Dlx1, sc‐81959, 1:500; Santa Cruz Biotechnology) or rabbit polyclonal Msh homeobox 1 (Msx1, ab174207, 1:500; Abcam).

Techniques: Mineralization Assay, Staining, Western Blot, Expressing, Reverse Transcription Polymerase Chain Reaction, Knock-Out