Journal: Endocrinology

Article Title: Msx Homeobox Genes Act Downstream of BMP2 to Regulate Endometrial Decidualization in Mice and in Humans

doi: 10.1210/en.2019-00131

Figure Lengend Snippet: Msx2 is a downstream target of BMP2 signaling in the uterus during decidualization. (A) The primary cultures of mouse endometrial stromal cells (MESCs) were transduced with adenovirus expressing GFP or BMP2. The cells were lysed at different time points, as indicated. Total RNA was isolated, and real-time PCR was performed to analyze the levels of Msx2. The relative levels of gene expression were determined by setting the expression level of the GFP-treated sample at 24 h to 1.0 (n = 3). Rplp0, encoding a ribosomal protein, was used to normalize the level of RNA. *P < 0.05. (B) The nucleotide positions of the SBEs on the Msx2 promoter were analyzed by ChIP. (C) Mouse stromal cells were treated with E + P or E, P, and BMP2 (E + P + BMP2) for 90 min. ChIP, using the Smad4 antibody, was performed, as described in “Materials and Methods.” Chromatin enrichment was quantified by real-time PCR using primers flanking the potential SBE in the Msx2 promoter and also a negative control region in the ORF of Msx2. Enrichments were normalized to 1% of input DNA. The experiment was repeated twice, and representative data are shown.

Article Snippet: The next day, the cells were either treated with E + P or E + P + BMP2 (recombinant human BMP2; R&D Systems; 355-BM-010) for 90 minutes.

Techniques: Transduction, Expressing, Isolation, Real-time Polymerase Chain Reaction, Gene Expression, Negative Control

Journal: Endocrinology

Article Title: Msx Homeobox Genes Act Downstream of BMP2 to Regulate Endometrial Decidualization in Mice and in Humans

doi: 10.1210/en.2019-00131

Figure Lengend Snippet: MSX1 and MSX2 mediate BMP2-induced HESC decidualization. The primary cultures of HESCs were established as described in “Materials and Methods.” The cells were transduced with adenovirus expressing GFP or BMP2. The cells were lysed at different time points as indicated. Total RNA was isolated, and real-time PCR was performed to analyze the levels of MSX1 and MSX2. The relative levels of gene expression were determined by setting the expression level on day 0 of the GFP-treated sample at 1.0. RPLP0, encoding a ribosomal protein, was used to normalize the level of RNA. Data were collected from three independent clinical samples, which were subjected to the same experimental conditions. *P < 0.05; **P < 0.005.

Article Snippet: The next day, the cells were either treated with E + P or E + P + BMP2 (recombinant human BMP2; R&D Systems; 355-BM-010) for 90 minutes.

Techniques: Transduction, Expressing, Isolation, Real-time Polymerase Chain Reaction, Gene Expression

Journal: British Journal of Pharmacology

Article Title: The polyphenol resveratrol promotes skeletal growth in mice through a sirtuin 1‐bone morphogenic protein 2 longevity axis

doi: 10.1111/bph.14477

Figure Lengend Snippet: eNOS and NO‐donors stimulate bone growth. eNOS mRNA expression in MC3T3 (A) and 2T3 (B) cells treated with RSV (1–100 μM) or vehicle for 24 h ( n = 5) determined by real time PCR. Nitrite (NO 2 − ) levels (C) or total eNOS protein (D) from 2T3 or MC3T3 cells treated with RSV (1–100 μM) or vehicle ( n = 5) for 48 h, determined by colorimetric assay. ALP levels from primary osteoblasts treated with NO‐donor (NOC22, 0.1–10 μM) or vehicle for 4 days ( n = 5) normalized to total cell protein (E). Calvariae from newborn mice cultured with NO‐donor (NOC22, 0.1–1 μM) or vehicle control for 4 days ( n = 5) processed for histology and H&E staining (F). mRNA expression of osteoblast marker genes Runx2, osteocalcin (OCN) and BMP2 determined in 2T3 cells treated with NOC22 (1.0 μM) or vehicle for 24 h ( n = 5) by real time PCR (G). BMP2 protein levels assessed in conditioned media from NO‐donor‐treated osteoblasts (NOC22 or SNP, 3–200 μM, 48 h, n = 5), by ELISA (with rhBMP2 as standard) (H). The μCT analysis of dissected tibia from homozygous eNOS knockout ( −/− ) or wild‐type ( +/+ ) control mice (4 month, n = 10) (I) and trabecular bone volume (J) and BMD (K) analysis (* P < 0.05 vs. vehicle; * P < 0.01 vs. wild‐type control animals).

Article Snippet: Recombinant human BMP2 (hBMP2; R&D Systems Inc.) was used as a standard.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Colorimetric Assay, Cell Culture, Control, Staining, Marker, Enzyme-linked Immunosorbent Assay, Knock-Out

Journal: British Journal of Pharmacology

Article Title: The polyphenol resveratrol promotes skeletal growth in mice through a sirtuin 1‐bone morphogenic protein 2 longevity axis

doi: 10.1111/bph.14477

Figure Lengend Snippet: The eNOS‐SIRT1 axis is necessary for the pro‐osteogenic effects of RSV. Different effects on ALP levels (A) BMP2 gene expression (qPCR, B) and promoter activity (C) in primary osteoblasts from eNOS knockout ( −/− ) or wild‐type ( +/+ ) control mice ( n = 5) treated with RSV (5 μM) or vehicle (24 h). RSV‐induced ALP levels (5 μM) in the presence of the BMP inhibitor noggin (500 ng·mL −1 , 48 h, n = 5), normalized to total protein (D). SIRT1 gene expression in 2T3 osteoblasts following RSV (1–100 μM) treatment, quantified by real time PCR (E), along with eNOS (F) and BMP2 (G) mRNA levels transfected with SIRT1 siRNA or scramble control ( n = 5) with and without RSV treatment (5 μM). RSV‐induced ALP levels (5 μM) in the presence of SIRT1 (or scrambled control) siRNA (H) ( n = 5) (* P < 0.05 vs. vehicle treated WT/scrambled ctrl; # P < 0.001 vs. RSV‐treated WT/scrambled ctrl).

Article Snippet: Recombinant human BMP2 (hBMP2; R&D Systems Inc.) was used as a standard.

Techniques: Gene Expression, Activity Assay, Knock-Out, Control, Real-time Polymerase Chain Reaction, Transfection

Journal: British Journal of Pharmacology

Article Title: The polyphenol resveratrol promotes skeletal growth in mice through a sirtuin 1‐bone morphogenic protein 2 longevity axis

doi: 10.1111/bph.14477

Figure Lengend Snippet: Ageing decreases bone volume and eNOS‐BMP2 expression. μCT analysis of tibia of young (3 month) or aged (12 month) mice ( n = 10) (A) determining BV/TV (B) and BMD (C). Gene expression changes (real time PCR) of eNOS (D), BMP2 (E) in ageing long bones ( n = 10). Proposed mechanism of RSV action within osteoblasts (F) (* P < 0.05).

Article Snippet: Recombinant human BMP2 (hBMP2; R&D Systems Inc.) was used as a standard.

Techniques: Expressing, Gene Expression, Real-time Polymerase Chain Reaction

Journal: Orphanet Journal of Rare Diseases

Article Title: Identification and characterization of regulatory elements in the promoter of ACVR1 , the gene mutated in Fibrodysplasia Ossificans Progressiva

doi: 10.1186/1750-1172-8-145

Figure Lengend Snippet: Effect of BMP2 on ACVR 1 promoter activity. A) HeLa and C2C12 cells were transfected with Pr-2.9 and Pr-0.072 reporter constructs and Luciferase expression was determined in the presence or absence of 100 ng/ml BMP2. Results are expressed as fold activation relative to the activity by the same promoter construct in absence of BMP2 (UN, untreated), with p < 0.05 * , p < 0.01 ** , or p < 0.001***. B) ACVR1 mRNA expression levels were detected by RT-qPCR in C2C12 cells upon treatment with BMP2. Values were normalized to the expression level of GAPDH and β - Actin genes.

Article Snippet: Human recombinant BMP2 was purchased from R&D Systems (Minneapolis, MN).

Techniques: Activity Assay, Transfection, Construct, Luciferase, Expressing, Activation Assay, Quantitative RT-PCR

Journal: ACS Applied Materials & Interfaces

Article Title: Nanotopography Influences Host–Pathogen Quorum Sensing and Facilitates Selection of Bioactive Metabolites in Mesenchymal Stromal Cells and Pseudomonas aeruginosa Co-Cultures

doi: 10.1021/acsami.4c09291

Figure Lengend Snippet: Characterization of functional active coatings on Ti nanotopographies. (a) Representative SEM images of flat control, nanospike (NS), and nanonetwork (NN) together with surface height and surface area measurements (table); scale bar 2 μm. (b) Chemical structure of PEA. (c) Titanium, carbon, and oxygen spectra of PEA coated flat, NS, and NN taken by X-ray photoelectron spectroscopy (XPS) surfaces. Each color corresponds to the number on the PEA chemical structure. (d) 1 × 1 μm AFM micrographs from the flat, NS, and NN surfaces before coating (top row) and after coating with PEA for 90 s at 100 W using plasma polymerization, followed by fibronectin (FN) for 1 h (bottom row). (e) Roughness (Rq) and (f) contact angle (3 μL sessile water drop) measurements for flat, NS, and NN with PEA coating and PEA+FN coating. ELISA was used to quantify the amount of (g) FN and (h) BMP2 adsorbed onto flat, NS, and NN samples after one h of coating. (i) The percentage of BMP2 released into solution after PEA+FN coating, quantified at days 1, 3, 5, 7, 9, 12, and 14. (e–h) Average represented as bars with individual values and standard deviation. Comparison of differences was tested using a Kruskal–Wallis test with a p -value <0.05 (*) considered significant, and <0.001 (**) highly significant. Together these results show that the PEA+FN+BMP2 coating could be used on Ti flat, NS, and NN nanotopographies to test with bacteria, hMSCs, and co-cultures.

Article Snippet: FN and BMP2 levels in the supernatants were calculated using enzyme-linked immunosorbent assay (ELISA) duo-set Human Fibronectin (DY1918, R&D systems), and Human BMP2 (DY355, R&D systems), respectively, following instructions from the manufacturer.

Techniques: Functional Assay, Control, Spectroscopy, Clinical Proteomics, Enzyme-linked Immunosorbent Assay, Standard Deviation, Comparison, Bacteria