Journal: Oncogenesis

Article Title: Neuron-specific enolase promotes stem cell-like characteristics of small-cell lung cancer by downregulating NBL1 and activating the BMP2/Smad/ID1 pathway.

doi: 10.1038/s41389-022-00396-5

Figure Lengend Snippet: Fig. 5 NSE downregulates and interacts with NBL1 in SCLC cells. A, B Western blot images showed the interaction between NSE and NBL1 by Co-IP experiment. C The double-immunofluorescence labeling experiment detected NSE and NBL1 and displayed co-location in clinical samples (original magnification ×40, scale bar 50 μm). D Western blot images show the interaction of BMP2 and BMPR1A enhanced after overexpression of NSE. E The photo shows a significant and negative correlation between ENO2 (encode NSE) and NBL1 in mRNA level using the CCLE database.

Article Snippet: The membranes were incubated with the following antibodies: NSE (1:2000, Abcam), GFP (1:2000, Proteintech), Flag (1:1000, Proteintech), OCT4 (1:1000, CST), Nanog (1:2000, CST), SOX2 (1:1000, CST), β-actin (1:5000, Proteintech), BMP2 (1:1000, Proteintech), Smad1 (1:2000, Proteintech), pSmad1/5/8 (1:1000, CST), and ID1 (1:500, Proteintech).

Techniques: Western Blot, Co-Immunoprecipitation Assay, Labeling, Over Expression

Journal: Function (Oxford, England)

Article Title: Deletion of Mechanosensory β1-integrin From Bladder Smooth Muscle Results in Voiding Dysfunction and Tissue Remodeling.

doi: 10.1093/function/zqac042

Figure Lengend Snippet: Figure 1. Characterization of the mouse model. (A) RT-PCR of β1-integrin from detrusor extracted mRNA at 6 wk after i.p. injections. Expression of functional Cre in β1-integrin floxed mice is evidenced by a smaller DNA product lacking exon 3 (ex3), which is not present in wild type mice. Myh11β 1-fl/fl mice were induced with TMX or injected with corn oil vehicle (Myh11-TMX and Myh11- Oil, respectively). (B) Western blotting of β1-integrin in detrusor from oil- and TMX-injected mice at 6 wk after injections. β-actin loading controls are shown in the lower panel. (C) Quantitation of densitometry from (B) normalized to the amount of actin in each lane. Data are mean ± SD, ∗P < 0.05, based on t-test with Welch’s correction for unequal variances. (D) Immunofluorescence of β1- integrin in BSM of oil- and TMX-injected Myh11β 1-fl/fl mice (left panels, green). Phalloidin staining shows actin (red) and DAPI highlights nuclei (blue). Scale bar (white) in top left panel = 10 μm.

Article Snippet: Western blotting: Rabbit antibodies to β1-integrin (ab52971), and muscarinic receptor 3 (M3, ab126168) were from Abcam (Waltham, MA, USA); α1-integrin (#71 747), β3-integrin (#13 166), and β-actin (#4697) were from Cell Signaling Technology (Danvers, MA, USA); M2 (G206) was from Assay Biotech (Fremont, CA, USA) and P2X1 (APR001) was from Alomone Laboratories (Jerusalem, Israel).

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Functional Assay, Injection, Western Blot, Quantitation Assay, Immunofluorescence, Staining

Journal: American Journal of Physiology - Endocrinology and Metabolism

Article Title: Skeletal inflammation and attenuation of Wnt signaling, Wnt ligand expression, and bone formation in atherosclerotic ApoE-null mice

doi: 10.1152/ajpendo.00501.2015

Figure Lengend Snippet: TaqMan reagents used for quantification of mRNA by qPCR

Article Snippet: Transcript levels were calculated by normalizing to the housekeeping gene mitochondrial ribosomal protein S2 using the ΔC T method ( 34 ). table ft1 table-wrap mode="anchored" t5 Table 1. caption a7 NCBI Gene Name Gene Name TaqMan # Alox12 Arachidonate 12-lipoxygenase Mm00545833_m1 Alox15 Arachidonate 15-lipoxygenase Mm01250458_m1 Axin2 Axin2 Mm00443610_m1 Mrps2 Mitochondrial ribosomal protein S2 (chob) Mm01962382_s91 Cx43 Gap junction membrane channel protein alpha 1 Mm01962382_s53 Dkk1 Dickkopf homolog 1 Mm00438422_m1 IL-1ß Interleukin-1β Mm00434228_m1 IL-6 Interleukin-6 Mm00446190_m1 Nkd2 Naked cuticle 2 homolog Mm00472240_m1 Sost Sclerostin Mm00470479_m1 Sfrp4 Secreted frizzled-related protein 4 Mm00840104_m1 TNF Tumor necrosis factor-α Mm00443258_m1 Tnfrsf11b Tumor necrosis factor receptor superfamily, member 11b (osteoprotogerin, OPG) Mm00435452_m1 Wisp1 WNT1 inducible signaling pathway protein 1 Mm00457574_m1 Wnt4 Wingless-related MMTV integration site 4 Mm00437341_m1 Wnt5a Wingless-related MMTV integration site 5A Mm00437347_m1 Wnt6 Wingless-related MMTV integration site 6 Mm00437353_m1 Wnt10b Wingless-related MMTV integration site 10B Mm00442104_m1 Wnt16 Wingless-related MMTV integration site 16 Mm00446420_m1 Open in a separate window TaqMan reagents used for quantification of mRNA by qPCR

Techniques: Membrane

Journal: Experimental and Therapeutic Medicine

Article Title: Triptolide inhibits the function of TNF-α in osteoblast differentiation by inhibiting the NF-κB signaling pathway

doi: 10.3892/etm.2017.4749

Figure Lengend Snippet: TNF-α significantly inhibited the BMP-2-induced osteoblast differentiation. (A) Effect of BMP-2 on osteogenic activity. C2C12 cells were treated with 200 ng/ml BMP-2 for 3 days and the ALP activity was assessed. **P<0.01 vs. Ctrl group. (B) Effect of TNF-α on osteoblast activity. C2C12 cells were treated with 5 ng/ml TNF-α for 3 days and the ALP activity was determined. *P<0.05 vs. Ctrl group. (C) TNF-α inhibited the effect of BMP-2 on osteogenic activity. C2C12 cells were treated with 200 ng/ml BMP-2 or 200 ng/ml BMP + 5 ng/ml TNF-α for 3 days and the ALP activity was determined. ##P<0.01 vs. Ctrl group, **P<0.01 vs. BMP-2 group. TNF, tumor necrosis factor; BMP, bone morphogenetic protein; ALP, alkaline phosphatase; Ctrl, control.

Article Snippet: In the present study, C2C12 cells were treated with BMP-2 (10426-H01H-10; Sinobiological, Inc., Beijing, China) in order to produce osteoblast cells.

Techniques: Activity Assay

Journal: Experimental and Therapeutic Medicine

Article Title: Triptolide inhibits the function of TNF-α in osteoblast differentiation by inhibiting the NF-κB signaling pathway

doi: 10.3892/etm.2017.4749

Figure Lengend Snippet: TNF-α inhibited BMP-2-induced osteoblast differentiation in a dose-dependent manner. (A) C2C12 cells were treated with 100, 200 or 300 ng/ml BMP-2 for 3 days and ALP activity was evaluated. **P<0.01 vs. untreated cells. (B) C2C12 cells were treated with 100 ng/ml BMP-2 + 2.5, 5.0 or 10.0 ng/ml TNF-α for 3 days and ALP activity was assessed. **P<0.01 vs. BMP-2 group (0 ng/ml TNF-α). TNF, tumor necrosis factor; BMP, bone morphogenetic protein; ALP, alkaline phosphatase.

Article Snippet: In the present study, C2C12 cells were treated with BMP-2 (10426-H01H-10; Sinobiological, Inc., Beijing, China) in order to produce osteoblast cells.

Techniques: Activity Assay

Journal: Experimental and Therapeutic Medicine

Article Title: Triptolide inhibits the function of TNF-α in osteoblast differentiation by inhibiting the NF-κB signaling pathway

doi: 10.3892/etm.2017.4749

Figure Lengend Snippet: TPL significantly attenuated TNF-α-induced inhibition of osteoblast differentiation. C2C12 cells were treated with 100 ng/ml of BMP-2 and 5 ng/ml of TNF-α with or without triptolide (4, 8, 16 ng/ml) for 3 days and ALP activity was assessed. **P<0.01, vs. BMP-2-TNF-α group and #P<0.05, ##P<0.01, vs. BMP-2 group. TPL, triptolide; TNF, tumor necrosis factor; BMP, bone morphogenetic protein; ALP, alkaline phosphatase.

Article Snippet: In the present study, C2C12 cells were treated with BMP-2 (10426-H01H-10; Sinobiological, Inc., Beijing, China) in order to produce osteoblast cells.

Techniques: Inhibition, Activity Assay

Journal: PLoS ONE

Article Title: Palmitate Promotes the Paracrine Effects of Macrophages on Vascular Smooth Muscle Cells: The Role of Bone Morphogenetic Proteins

doi: 10.1371/journal.pone.0029100

Figure Lengend Snippet: A. RAW 264.7 cells were treated with BSA control or palmitate (250 µM) for 4 hours and conditioned media were collected 20 hours later. BMP 2 and 4 secreted in the media were pulled down on heparin-Sepharose columns and detected by immunoblotting. B. RAW 264.7 cells were treated with BSA control or palmitate for 4 hours, and then incubated with fresh media for 20 hours. BMP2 and BMP4 mRNA levels were analyzed by real-time PCR. Data are illustrated in arbitrary unit relative control and represent the means ± SE from three independent experiments. *p<0.05 compared to control.

Article Snippet: BMP2 siRNA, BMP4 siRNA and scrambled control siRNA were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Control, Western Blot, Incubation, Real-time Polymerase Chain Reaction

Journal: PLoS ONE

Article Title: Palmitate Promotes the Paracrine Effects of Macrophages on Vascular Smooth Muscle Cells: The Role of Bone Morphogenetic Proteins

doi: 10.1371/journal.pone.0029100

Figure Lengend Snippet: A. THP-1 cells differentiated with PMA were stimulated with BSA control or palmitate for 4 hours and conditioned media were collected 20 hours later. Palmitate-conditioned media were treated without or with 2 µg/mL of anti-BMP2, anti-BMP4, or both for 1 hour. Each conditioned media was added to serum starved SMCs for 72 hours. Cell proliferation was analyzed by BrdU incorporation assay. Promotion of SMC proliferation by palmitate-conditioned media was abrogated by combined treatment of anti-BMP2/anti-BMP4. B. Each conditioned media described above was added to SMCs for 24 hours. Cell migration assay was conducted using Boyden chamber assay. Promotion of SMC migration by palmitate-conditioned media was inhibited by treatment of anti-BMP2 or anti-BMP4 or combination of both. C. Each conditioned media described above was added to SMCs for 24 hours. Immunoblots showed that anti-BMP4 or anti-BMP2/anti-BMP4 combination inhibited phenotypic change of SMCs. D. The band intensities were determined by quantitative densitometry. E. THP-1 cell-derived macrophages were transfected with BMP2 or BMP4 or both siRNA for 24 hours. Immunoblots demonstrated knockdown of BMP2 or BMP4 or both BMPs in the macrophages. F. SMCs were treated with palmitate-conditioned media from macrophages without or with knockdown of BMPs for 24 hours. Equal amount of protein were separated by SDS-PAGE, and immunoblots showed that knockdown of BMP4 or both BMPs abrogated phenotypic change of SMCs. Data are illustrated in arbitrary integrator unit relative to β-actin and represent the means ± SE from three independent experiments. *p<0.05 compared to control. **p<0.05 compared to palmitate-conditioned media treatment.

Article Snippet: BMP2 siRNA, BMP4 siRNA and scrambled control siRNA were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Control, BrdU Incorporation Assay, Cell Migration Assay, Boyden Chamber Assay, Migration, Western Blot, Derivative Assay, Transfection, Knockdown, SDS Page

Journal: PLoS ONE

Article Title: Palmitate Promotes the Paracrine Effects of Macrophages on Vascular Smooth Muscle Cells: The Role of Bone Morphogenetic Proteins

doi: 10.1371/journal.pone.0029100

Figure Lengend Snippet: A. Quiescent SMCs were treated with various concentration of BMP2 or BMP4 for 24 hours. Cell proliferation was analyzed by BrdU incorporation assay. Recombinant BMP2 and BMP4 did not have obvious effects on SMC proliferation. B. SMC migration assessment was conducted using Boyden chamber assay and BMP2 and BMP4 significantly promoted SMC migration. C. SMCs were treated with various concentration of recombinant BMP2 or BMP4 for 24 hours. Equal amount of protein was separated by SDS-PAGE and immunoblots showed both recombinant BMPs induced phenotypic change of SMCs with lowering expression of SM α-actin and SM22α.

Article Snippet: BMP2 siRNA, BMP4 siRNA and scrambled control siRNA were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Concentration Assay, BrdU Incorporation Assay, Recombinant, Migration, Boyden Chamber Assay, SDS Page, Western Blot, Expressing

Journal: PLoS ONE

Article Title: Imatinib Treatment Induces CD5+ B Lymphocytes and IgM Natural Antibodies with Anti-Leukemic Reactivity in Patients with Chronic Myelogenous Leukemia

doi: 10.1371/journal.pone.0018925

Figure Lengend Snippet: BM samples were drawn at diagnosis (t0) or 3 months (3mo) after therapy with imatinib. Panel A: SDF-1 (left histograms) or BAFF (right histograms) content was measured in BM plasma by ELISA, referred to a standard curve and results expressed as pg/mL. Mean±SD from 32 R and 8 NR patients. * p<0.001 vs t0; **p<0.001 vs NR. Panels B–D: total RNA was extracted from cells isolated from BM samples before (t0) and after (3 mo) imatinib treatment and reverse-transcribed with random primers. Q-RT-PCR was performed with primers and probes for SDF-1 (B, left histograms), BAFF (B, right histograms), BMP2 (C) and BMP7 (D) on the 7900HT FastRT-PCR system with the fluorescent Taqman method. mRNAs were normalized to 18 s as a control gene, and referred to a standard curve. Results are expressed as the fold relative increase in mRNA expression compared to the control gene. Mean±SD from 32 R and 8 NR patients. * p<0.001 vs t0; **p<0.001 vs NR.

Article Snippet: Primers for BAFF, SDF-1, BMP2 and BMP7 amplifications and probes were purchased from Applied Biosystem (userid: Hs00198106_m1, Hs00171022_m1, Hs01055564_m1 and Hs00233477_m1, respectively).

Techniques: Biomarker Discovery, Clinical Proteomics, Enzyme-linked Immunosorbent Assay, Isolation, Reverse Transcription, Reverse Transcription Polymerase Chain Reaction, Control, Expressing