Journal: International Journal of Nanomedicine

Article Title: A Three-Dimensional Printed Polycaprolactone Scaffold Combined with Co-Axially Electrospun Vancomycin/Ceftazidime/Bone Morphological Protein-2 Sheath-Core Nanofibers for the Repair of Segmental Bone Defects During the Masquelet Procedure

doi: 10.2147/IJN.S238478

Figure Lengend Snippet: SEM image and fiber diameter distribution of ( A ) pure PLGA nanofibers, ( B ) sheath-core PLGA/BMP-2 nanofibers, and ( C ) PLGA/vancomycin/ceftazidime nanofibers.

Article Snippet: Commercial antibodies were adapted for IHC analysis; these included anti-BMP-2 (polyclonal, 1:200; cat. no. A0231; ABclonal, MA, USA); anti-VEGF (polyclonal, 1:200; cat. no. A0280; ABclonal); anti-vWF (vWF Picoband antibody, 1:200; cat. no. PB9062; Boster Biological Technology, Pleasanton CA, USA); anti-TGF-β (polyclonal, 1:200; cat. no. A2561; ABclonal); and anti-IL-6 (polyclonal, 1:200; cat. no. A0286; ABclonal).

Techniques:

Journal: International Journal of Nanomedicine

Article Title: A Three-Dimensional Printed Polycaprolactone Scaffold Combined with Co-Axially Electrospun Vancomycin/Ceftazidime/Bone Morphological Protein-2 Sheath-Core Nanofibers for the Repair of Segmental Bone Defects During the Masquelet Procedure

doi: 10.2147/IJN.S238478

Figure Lengend Snippet: Water contact angles of ( A ) pure PLGA nanofibers (126.24 ± 10.26°), ( B ) sheath-core PLGA/BMP-2 nanofibers (120.00 ± 8.67°), and ( C ) PLGA/vancomycin/ceftazidime nanofibers (69.2.33 ± 5.14°). Substantially improved hydrophilicity of electrospun nanofibers was observed with the addition of water-soluble antibiotics.

Article Snippet: Commercial antibodies were adapted for IHC analysis; these included anti-BMP-2 (polyclonal, 1:200; cat. no. A0231; ABclonal, MA, USA); anti-VEGF (polyclonal, 1:200; cat. no. A0280; ABclonal); anti-vWF (vWF Picoband antibody, 1:200; cat. no. PB9062; Boster Biological Technology, Pleasanton CA, USA); anti-TGF-β (polyclonal, 1:200; cat. no. A2561; ABclonal); and anti-IL-6 (polyclonal, 1:200; cat. no. A0286; ABclonal).

Techniques:

Journal: International Journal of Nanomedicine

Article Title: A Three-Dimensional Printed Polycaprolactone Scaffold Combined with Co-Axially Electrospun Vancomycin/Ceftazidime/Bone Morphological Protein-2 Sheath-Core Nanofibers for the Repair of Segmental Bone Defects During the Masquelet Procedure

doi: 10.2147/IJN.S238478

Figure Lengend Snippet: In vitro drug release profiles. ( A ) Daily and ( B ) accumulated release of antibiotics from the composite nanofibers. ( C ) Daily and ( D ) accumulated release of BMP-2 from the composite nanofibers. In vivo examination of the daily release of ( E ) antibiotics and ( F ) BMP-2 from the composite scaffold in the muscular tissue.

Article Snippet: Commercial antibodies were adapted for IHC analysis; these included anti-BMP-2 (polyclonal, 1:200; cat. no. A0231; ABclonal, MA, USA); anti-VEGF (polyclonal, 1:200; cat. no. A0280; ABclonal); anti-vWF (vWF Picoband antibody, 1:200; cat. no. PB9062; Boster Biological Technology, Pleasanton CA, USA); anti-TGF-β (polyclonal, 1:200; cat. no. A2561; ABclonal); and anti-IL-6 (polyclonal, 1:200; cat. no. A0286; ABclonal).

Techniques: In Vitro, In Vivo

Journal: International Journal of Nanomedicine

Article Title: A Three-Dimensional Printed Polycaprolactone Scaffold Combined with Co-Axially Electrospun Vancomycin/Ceftazidime/Bone Morphological Protein-2 Sheath-Core Nanofibers for the Repair of Segmental Bone Defects During the Masquelet Procedure

doi: 10.2147/IJN.S238478

Figure Lengend Snippet: Analysis of the gross specimen ( A ) revealed a thick induced membrane (IM) formed circumferentially around the applied scaffold. The PLGA nanofibers had been dissolved completely, and only the PCL mesh was preserved. The histological evaluation of the induced membrane was performed (blue arrow) and represents in ( B ). ( B ) Histological evaluation by hematoxylin and eosin staining of the induced membrane. Radiographic examination of fracture healing in ( C ) the PCL group, ( D ) the PCL-PLGA/antibiotic group, and ( E ) the PCL-PLGA/antibiotic/BMP-2 group.

Article Snippet: Commercial antibodies were adapted for IHC analysis; these included anti-BMP-2 (polyclonal, 1:200; cat. no. A0231; ABclonal, MA, USA); anti-VEGF (polyclonal, 1:200; cat. no. A0280; ABclonal); anti-vWF (vWF Picoband antibody, 1:200; cat. no. PB9062; Boster Biological Technology, Pleasanton CA, USA); anti-TGF-β (polyclonal, 1:200; cat. no. A2561; ABclonal); and anti-IL-6 (polyclonal, 1:200; cat. no. A0286; ABclonal).

Techniques: Staining

Journal: International Journal of Nanomedicine

Article Title: A Three-Dimensional Printed Polycaprolactone Scaffold Combined with Co-Axially Electrospun Vancomycin/Ceftazidime/Bone Morphological Protein-2 Sheath-Core Nanofibers for the Repair of Segmental Bone Defects During the Masquelet Procedure

doi: 10.2147/IJN.S238478

Figure Lengend Snippet: Immunohistochemical analysis of the expression of ( A ) BMP-2 (*membrane-lining cells, † spindle cells), ( B ) TGF-β (*membrane-lining cells, † spindle cells), ( C ) vWF, ( D ) VEGF, and ( E ) IL-6.

Article Snippet: Commercial antibodies were adapted for IHC analysis; these included anti-BMP-2 (polyclonal, 1:200; cat. no. A0231; ABclonal, MA, USA); anti-VEGF (polyclonal, 1:200; cat. no. A0280; ABclonal); anti-vWF (vWF Picoband antibody, 1:200; cat. no. PB9062; Boster Biological Technology, Pleasanton CA, USA); anti-TGF-β (polyclonal, 1:200; cat. no. A2561; ABclonal); and anti-IL-6 (polyclonal, 1:200; cat. no. A0286; ABclonal).

Techniques: Immunohistochemical staining, Expressing

Journal: Annals of Clinical and Translational Neurology

Article Title: A novel de novo GABRA2 gene missense variant causing developmental epileptic encephalopathy in a Chinese patient

doi: 10.1002/acn3.52262

Figure Lengend Snippet: Analysis of GABA A R pentameric structure. (A) A pentameric model featuring 2 α2 subunits (GABRA2), 2 β3 subunits (GABRB3), and 1 γ2 subunit (GABRG2). (B) The structure is rotated 90 degrees for a top‐down view. In the wild‐type GABRA2, Ala308 forms hydrogen bonds with Pro209 of GABRB3 and with another GABRA2 protein at position 238 of GABRG2. (C) In the mutated GABRA2 (p.Ala308Val), the longer valine side chain disrupts these hydrogen bonds, potentially altering structural stability.

Article Snippet: The primary antibodies utilized included: DYKDDDDK Tag (9A3) Mouse mAb (Cat#8146, Cell Signaling Technology, MA, USA, diluted 1:200); GABRB3 Antibody (Cat#PB0627, Boster Biological Technology, Wuhan, China, diluted 1:1000); GABRG2 Antibody (Cat#A02361‐1, Boster Biological Technology, Wuhan, China, diluted 1:1000); and β‐Actin (8H10D10) Mouse mAb (Cat#3700, Cell Signaling Technology, MA, USA, diluted 1:1000).

Techniques:

Journal: Annals of Clinical and Translational Neurology

Article Title: A novel de novo GABRA2 gene missense variant causing developmental epileptic encephalopathy in a Chinese patient

doi: 10.1002/acn3.52262

Figure Lengend Snippet: In vitro functional assay. (A) Western blotting analysis indicated that the GABRA2‐MUT group exhibited protein expression, but it was significantly lower than the GABRA2‐WT group, with a reduction of approximately 70.62% (*** P < 0.001). (B) CO‐IP results showed that wild‐type GABRA2 interacted with GABRB3/GABRG2, while the mutant may disrupt this interaction. The right panel presents normalized CO‐IP analysis, *** P < 0.001.

Article Snippet: The primary antibodies utilized included: DYKDDDDK Tag (9A3) Mouse mAb (Cat#8146, Cell Signaling Technology, MA, USA, diluted 1:200); GABRB3 Antibody (Cat#PB0627, Boster Biological Technology, Wuhan, China, diluted 1:1000); GABRG2 Antibody (Cat#A02361‐1, Boster Biological Technology, Wuhan, China, diluted 1:1000); and β‐Actin (8H10D10) Mouse mAb (Cat#3700, Cell Signaling Technology, MA, USA, diluted 1:1000).

Techniques: In Vitro, Functional Assay, Western Blot, Expressing, Co-Immunoprecipitation Assay, Mutagenesis

Journal: Scientific Reports

Article Title: BMP2 promotes lung adenocarcinoma metastasis through BMP receptor 2-mediated SMAD1/5 activation

doi: 10.1038/s41598-022-20788-2

Figure Lengend Snippet: BMP2 is highly expressed in more invasive lung adenocarcinoma cells. ( A ) Migration ability was determined by seeding 5 × 10 4 cells in a migration chamber without Matrigel coating. Invasion ability was determined by seeding 1 × 10 5 cells in a migration chamber with Matrigel coating. After 24 h, the cells that successfully migrated through the filter were stained with Liu ‘s solution and counted. Slides were observed under 10 × magnification. ( B ) Quantitative data derived from ( A ). Bar graphs represent the average number of migrated cells ± SD. At least three fields were counted for each condition. ( C ) BMP2 and BMPR2 expression levels were determined by western blotting and qPCR. ( D ) The protein levels of epithelial and mesenchymal marker were analyzed by western blotting. The error bars represent SD, and the p -value was determined by Student’s t test (* p < 0.05; ** p < 0.01; *** p < 0.001).

Article Snippet: The antibodies used in this study included those targeting β-actin (NB600-501, Novus), E-cadherin (#3195, Cell Signaling Technology for western blot; 20,874–1-AP, Proteintech for IHC staining), occludin (13,409–1-AP, Proteintech), N-cadherin (ab53519, Abcam), Vimentin (#550,513, BD), Slug (sc-166476, Santa Cruz Biotechnology), BMPR2 (ab78422, Abcam), BMP2 (A0231, ABclonal), SMAD5 (#9517, Cell Signaling Technology), SMAD1 (#6944, Cell Signaling Technology), SMAD4 (sc-7966, Santa Cruz Biotechnology), and phospho-SMAD1/5 (ser463/465) (#9516, Cell Signaling Technology).

Techniques: Migration, Staining, Derivative Assay, Expressing, Western Blot, Marker

Journal: Scientific Reports

Article Title: BMP2 promotes lung adenocarcinoma metastasis through BMP receptor 2-mediated SMAD1/5 activation

doi: 10.1038/s41598-022-20788-2

Figure Lengend Snippet: Inhibition of BMP2 reduces invasiveness and downregulates EMT in lung cancer cells. ( A ) The invasion and migration abilities were reduced in BMP2-depleted CL1-5 and AS2 cells. BMP2 expression was depleted by two specific shRNAs. shLacZ is the non-targeting control. Slides were observed under 10 × magnification. ( B ) Quantitative data derived from ( A ). Bar graphs represent the average number of migrated cells ± SD. At least three fields were counted and averaged for each cell line. ( C ) The protein levels of BMP2 and EMT marker were determined by western blotting in CL1-5 and AS2 cells. ( D ) Cell invasion and migration abilities were reduced in BMP2-depleted CL1-5 and AS2 cells. The expression of BMP2 was depleted by siRNA. A non-targeting siRNA was used as a control. Slides were observed under 10 × magnification. ( E ) Quantitative data derived from ( D ). Bar graphs represent the average number of migrated cells ± SD. At least three fields were measured in each cell line. ( F ) The expression levels of BMP2 and EMT markers were determined by western blotting. The p -value was determined by Student’s t test. (* p < 0.05, ** p < 0.01, *** p < 0.001).

Article Snippet: The antibodies used in this study included those targeting β-actin (NB600-501, Novus), E-cadherin (#3195, Cell Signaling Technology for western blot; 20,874–1-AP, Proteintech for IHC staining), occludin (13,409–1-AP, Proteintech), N-cadherin (ab53519, Abcam), Vimentin (#550,513, BD), Slug (sc-166476, Santa Cruz Biotechnology), BMPR2 (ab78422, Abcam), BMP2 (A0231, ABclonal), SMAD5 (#9517, Cell Signaling Technology), SMAD1 (#6944, Cell Signaling Technology), SMAD4 (sc-7966, Santa Cruz Biotechnology), and phospho-SMAD1/5 (ser463/465) (#9516, Cell Signaling Technology).

Techniques: Inhibition, Migration, Expressing, Derivative Assay, Marker, Western Blot

Journal: Scientific Reports

Article Title: BMP2 promotes lung adenocarcinoma metastasis through BMP receptor 2-mediated SMAD1/5 activation

doi: 10.1038/s41598-022-20788-2

Figure Lengend Snippet: Inhibition of BMP2 decreased metastasis in a CL1-5 orthotopic mouse model. ( A ) A schematic diagram showing the animal experiment. CL1-5 cells stably expressing shLacZ or shBMP2 Luc/GFP were injected into the right lungs of NOD-SCID mice. IVIS analysis was conducted at 7, 14, 28, and 35 days after injection. ( B ) The body weights of mice were measured three times per week after tumor injection. ( C ) Metastasis in shLacZ and shBMP2-CL1-5-Luc/GFP groups was measured by IVIS until sacrifice. ( D , E ) Metastastic lesions in the lungs ( D ) and nearby organs ( E ) were analyzed by IVIS after sacrifice. ( F ) Hematoxylin and eosin (H&E) and IHC stains in mouse right lung tissue for the indicated proteins. Slides were observed under 40 × magnification. Error bars represent SD, and p -value was determined by Student’s t test (* p < 0.05, ** p < 0.01, *** p < 0.001).

Article Snippet: The antibodies used in this study included those targeting β-actin (NB600-501, Novus), E-cadherin (#3195, Cell Signaling Technology for western blot; 20,874–1-AP, Proteintech for IHC staining), occludin (13,409–1-AP, Proteintech), N-cadherin (ab53519, Abcam), Vimentin (#550,513, BD), Slug (sc-166476, Santa Cruz Biotechnology), BMPR2 (ab78422, Abcam), BMP2 (A0231, ABclonal), SMAD5 (#9517, Cell Signaling Technology), SMAD1 (#6944, Cell Signaling Technology), SMAD4 (sc-7966, Santa Cruz Biotechnology), and phospho-SMAD1/5 (ser463/465) (#9516, Cell Signaling Technology).

Techniques: Inhibition, Stable Transfection, Expressing, Injection

Journal: Scientific Reports

Article Title: BMP2 promotes lung adenocarcinoma metastasis through BMP receptor 2-mediated SMAD1/5 activation

doi: 10.1038/s41598-022-20788-2

Figure Lengend Snippet: Mean survival times among patients with and without high BMP2 expression levels.

Article Snippet: The antibodies used in this study included those targeting β-actin (NB600-501, Novus), E-cadherin (#3195, Cell Signaling Technology for western blot; 20,874–1-AP, Proteintech for IHC staining), occludin (13,409–1-AP, Proteintech), N-cadherin (ab53519, Abcam), Vimentin (#550,513, BD), Slug (sc-166476, Santa Cruz Biotechnology), BMPR2 (ab78422, Abcam), BMP2 (A0231, ABclonal), SMAD5 (#9517, Cell Signaling Technology), SMAD1 (#6944, Cell Signaling Technology), SMAD4 (sc-7966, Santa Cruz Biotechnology), and phospho-SMAD1/5 (ser463/465) (#9516, Cell Signaling Technology).

Techniques: Expressing

Journal: PLoS ONE

Article Title: Krüppel-Like Factor 6 Expression Changes during Trophoblast Syncytialization and Transactivates ßhCG and PSG Placental Genes

doi: 10.1371/journal.pone.0022438

Figure Lengend Snippet: A- Isolated mononuclear villous CTB cells cultured during the indicated hours and stained for KLF6 immunofluorescence detection (middle panels) with the polyclonal R-173 (green) anti-KLF6 antibody. Nuclei were counterstained with Hoechst 33342 dye (blue) and the overlay is shown (right panels). B- Confocal microscopy imaging of KLF6 at the indicated time points of the differentiation process. KLF6 was labelled with the polyclonal R-173 antibody (left panels) and DNA was stained with propidium iodide (IP) (middle panel). Overlay is shown in the right panels. C- Fluorescence intensity profile of KLF6 (green) and IP (red) along the yellow line shown in the confocal microscopy images. D- Morphological and biochemical differentiation of isolated mononuclear CTB cells were confirmed by the disappearance of desmoplakin intercellular staining (red), the appearance of multinucleated structures and the expression of PSG proteins (green). Original magnification, x1000. Scale bar, 10 µm. Immunofluorescence assays were performed with at least three different CTB purifications and representative figures are shown.

Article Snippet: Cells were then rinsed with PBS three times, blocked with 2.5% normal goat serum in 0.2% Tween-20 in PBS (PBST) and with 0.5% fish skin gelatin in PBST, and then incubated with the following primary antibodies: rabbit polyclonal anti-KLF6 (anti-Zf9, R-173, Santa Cruz Biotech; 1∶50), mouse monoclonal anti-KLF6 (clone 2c11, 1∶150) whose specificity was previously determined , mouse anti-desmosomal protein (0.045 mg/mL, ZK-31, Sigma Chemical Co.; 1∶400), rabbit polyclonal anti-human chorionic gonadotropin (hCG, A0231, Dako; 1∶500), mouse monoclonal anti-cytokeratin 7 (Dako, Clone OV-TL 12/30) and rabbit polyclonal anti-PSG (A0131, Dako; 1∶100).

Techniques: Isolation, Cell Culture, Staining, Immunofluorescence, Confocal Microscopy, Imaging, Fluorescence, Expressing

Journal: PLoS ONE

Article Title: Krüppel-Like Factor 6 Expression Changes during Trophoblast Syncytialization and Transactivates ßhCG and PSG Placental Genes

doi: 10.1371/journal.pone.0022438

Figure Lengend Snippet: A- JEG-3 cells were cultured in the presence of 1 µM methotrexate to induce cell differentiation during the indicted times and KLF6 mRNA expression was quantified by qRT-PCR (ABI 7500, Applied Biosystems). B- JEG-3 cells transfected with the empty (white bars) or the KLF6 expression vector (black bars) were harvested 24 h after transfection and PSG , βhCG and GCM1 gene expression was quantified by qRT- PCR (ABI 7500, Applied Biosystems). For A and B, results were normalized to cyclophilin A and expressed according to the 2 −ΔΔCt method using as calibrator the mRNA level obtained from the control condition. Data are presented as mean ±SEM of three independent experiments performed in triplicates and a one-sample t-test was used to determine whether experimental values were significantly different from the control value set as 1 (*p<0.05). C- Western blot detection of KLF6, PSG, βhCG and GCM1 in protein extracts of JEG-3 cells transfected with the empty (left lane) or KLF6 expression vector (right lane). α-tubulin was used as a loading control in each assay. Representative western blots are shown and the bar graph shows the densitometric analysis of three different experiments. (*p<0.05) D - JEG-3 cells were transiently transfected with the KLF6 expression vector and 24 h later they were immunostained for the detection of KLF6 (red) and βhCG (green) with a monoclonal anti-KLF6 and a polyclonal anti-βhCG antibodies, respectively. Nuclei were counterstained with Hoechst 33342 dye (blue), and the merge of the three channels is shown on the right side. Bar = 10 µm. Original magnification: ×1000. Representative images from three independent transfections are shown. Arrowheads, JEG-3 cells overexpressing KLF6; arrows, cells positive for βhCG.

Article Snippet: Cells were then rinsed with PBS three times, blocked with 2.5% normal goat serum in 0.2% Tween-20 in PBS (PBST) and with 0.5% fish skin gelatin in PBST, and then incubated with the following primary antibodies: rabbit polyclonal anti-KLF6 (anti-Zf9, R-173, Santa Cruz Biotech; 1∶50), mouse monoclonal anti-KLF6 (clone 2c11, 1∶150) whose specificity was previously determined , mouse anti-desmosomal protein (0.045 mg/mL, ZK-31, Sigma Chemical Co.; 1∶400), rabbit polyclonal anti-human chorionic gonadotropin (hCG, A0231, Dako; 1∶500), mouse monoclonal anti-cytokeratin 7 (Dako, Clone OV-TL 12/30) and rabbit polyclonal anti-PSG (A0131, Dako; 1∶100).

Techniques: Cell Culture, Cell Differentiation, Expressing, Quantitative RT-PCR, Transfection, Plasmid Preparation, Western Blot