Journal: Animal Reproduction

Article Title: Evaluation of conditioned medium from Sertoli cells as a potential inducer of in vitro germ cell differentiation of bovine fetal mesenchymal stem cells (MSCs)

doi: 10.1590/1984-3143-AR2024-0094

Figure Lengend Snippet: Phase contrast microphotographs and relative gene expression of specific MSC markers in AT-MSCs/SCCM and control during 21 days of culture. (A) AT-MSC/SCCM exhibit typical fibroblast-like morphology on day 0 (A) and mainly polygonal morphology from days 7, 14 and 21 of culture (B-D). Similarly, control AT-MSCs showed polygonal morphology from days 7, 14 and 21 of culture (F-H), with a slight less contrast compared to AT-MSCs/SCCM; (B) CD73 gene expression decreased (P<0.05) on day 14 in AT-MSCs/SCCM compared to control on the same day. Relative gene expression of CD90 and CD105 were not different (P>0.05) on days 7, 14 and 21 of culture compared to day 0 and controls. OCT4, DAZL and PIWIL2 gene expression were detected from days 0, 21 and 7, respectively; however, no differences (P>0.05) were detected between treatments or compared to day 0. DMC1 gene expression values were detected from day 7 to 21 with no significant differences between treatments. Scale Bar (A, E): 500 µm; (B-D, F-H): 100 µm. Superscripts (a,b) indicate a significant (P<0.05) difference between treatments or day 0.

Article Snippet: Cells were incubated in primaries mouse monoclonal anti-OCT4 antibody (Cat.#sc-5279; Santa Cruz Biotechnology, CA, USA) or rabbit polyclonal anti-DAZL (Cat.#ab34139; Abcam) or anti-PIWIL2 antibodies (Cat.#ab85084; Abcam) diluted (1:50) in 3% BSA in PBS overnight at 4 °C.

Techniques: Gene Expression, Control

Journal: Animal Reproduction

Article Title: Evaluation of conditioned medium from Sertoli cells as a potential inducer of in vitro germ cell differentiation of bovine fetal mesenchymal stem cells (MSCs)

doi: 10.1590/1984-3143-AR2024-0094

Figure Lengend Snippet: Expression of OCT4, DAZL and PIWIL2 markers in AT-MSC/SCCM and control during 21 days of culture. (A) Representative dot plots showing distribution of cells positive for OCT4, DAZL and PIWIL2 in AT-MSC/SCCM and control based on size (FSC) and complexity (SSC) at day 21 days of culture; (B) Representative histograms displaying cells positive for OCT4 (green area), DAZL (red area) and PIWIL2 (blue area) in AT-MSC/SCCM and control at day 21 of culture. Gray area represents cells incubated only with secondary antibody (C) OCT4, DAZL, PIWIL2-positive cells were detected in both AT-MSC/SCCM and control, with no differences (P>0.05) between them. Abbreviations: (MFI) Mean Fluorescence Intensity; (FL1-INT-LOG) signal intensity of protein expression on a logarithmic scale.

Article Snippet: Cells were incubated in primaries mouse monoclonal anti-OCT4 antibody (Cat.#sc-5279; Santa Cruz Biotechnology, CA, USA) or rabbit polyclonal anti-DAZL (Cat.#ab34139; Abcam) or anti-PIWIL2 antibodies (Cat.#ab85084; Abcam) diluted (1:50) in 3% BSA in PBS overnight at 4 °C.

Techniques: Expressing, Control, Incubation, Fluorescence

Journal: Frontiers in Veterinary Science

Article Title: Enhanced immunomodulatory effects of canine adipose tissue-derived mesenchymal stem cells in 3D culture

doi: 10.3389/fvets.2025.1500267

Figure Lengend Snippet: Stemness-related protein expression in cAT-MSCs spheroids. (A) cAT-MSCs spheroids harvested 48 h after incubation and stained with anti-SOX2 antibody (green) and Hoechst 33342 (blue) were examined using a confocal microscope (scale bar = 50 μm). (B) cAT-MSCs spheroids harvested 48 h after incubation and stained with anti-OCT4 antibody (green) and Hoechst 33342 (blue) were examined using a confocal microscope (scale bar = 50 μm). (C) SOX2 and OCT4, and TSG6 were extracted from 2D-cultured cAT-MSCs and 3D-cultured cAT-MSCs spheroids and quantified using western blotting. (D) Relative protein expressions in 2D and 3D-cultured cAT-MSCs were analyzed using ImageJ. SOX2 and OCT4 expression was significantly increased in 3D-cultured cAT-MSCs spheroids compared to 2D-cultured cAT-MSCs. * p < 0.05 and *** p < 0.001, as determined by Student’s t-test.

Article Snippet: Membranes were blocked with 5% skim milk in Tris-buffered saline and incubated overnight at 4°C with primary antibodies against SOX2 (1:1000; ABclonal Technology,), OCT4 (1:1000; ABclonal Technology), and β -actin (1:1000, Santa Cruz Biotechnology, Dallas, TX, United States).

Techniques: Expressing, Incubation, Staining, Microscopy, Cell Culture, Western Blot

Journal: Frontiers in Pharmacology

Article Title: The NEDD4/FLRT2 axis regulates NSCLC cell stemness

doi: 10.3389/fphar.2024.1459978

Figure Lengend Snippet: Primers of RT-qPCR.

Article Snippet: The NC membranes were incubated overnight at 4°C with primary antibodies for FLRT2 (Proteintech, China), OCT4 (CST, USA), CD44 (Abcam, United Kingdom), CD133 (Invitrogen, USA), E-cadherin (Bioworld, China), N-cadherin (Invitrogen, USA), Bax (Proteintech, China), Bcl-2 (Proteintech, China), and NEDD4 (Abcam, United Kingdom), respectively.

Techniques:

Journal: Frontiers in Pharmacology

Article Title: The NEDD4/FLRT2 axis regulates NSCLC cell stemness

doi: 10.3389/fphar.2024.1459978

Figure Lengend Snippet: FLRT2 expression in NSCLC and NSCLC CSC . (A) FLRT2 expression in lung cancer stem cells was analyzed in the GEO database. (B) FLRT2 level was involved in NSCLC prognosis. (C) RT-qPCR analysis compared the mRNA level of FLRT2 in NSCLC and normal tissues. (D) mRNA abundance of FLRT2 was examined in normal lung epithelial and lung cancer cells. (E) mRNA level of FLRT2 was examined in NSCLC and NSCLC CSC cells. * represented comparison with the A549 group, # represented comparison with the H1975 group. (F) FLRT2 protein abundance was examined in normal lung epithelial cells, lung cancer cells, and NSCLC CSC . All data are expressed as mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001 assessed via a two-tailed t-test; ns: non-significant.

Article Snippet: The NC membranes were incubated overnight at 4°C with primary antibodies for FLRT2 (Proteintech, China), OCT4 (CST, USA), CD44 (Abcam, United Kingdom), CD133 (Invitrogen, USA), E-cadherin (Bioworld, China), N-cadherin (Invitrogen, USA), Bax (Proteintech, China), Bcl-2 (Proteintech, China), and NEDD4 (Abcam, United Kingdom), respectively.

Techniques: Expressing, Quantitative RT-PCR, Comparison, Quantitative Proteomics, Two Tailed Test

Journal: Frontiers in Pharmacology

Article Title: The NEDD4/FLRT2 axis regulates NSCLC cell stemness

doi: 10.3389/fphar.2024.1459978

Figure Lengend Snippet: FLRT2 overexpression inhibits the stemness of NSCLC cells. (A) The efficacy of FLRT2 overexpression was measured after transfection for 48 h. (B) Evaluation of tumor sphere formation after FLRT2 upregulation. (C) Statistical data revealed the size and number of the spheres in the pcDNA 3.1 and FLRT2 groups. (D) Change in the protein level of stem cell markers after FLRT2 overexpression. (E) Change in mRNA level of stem cell markers after FLRT2 overexpression. All data are expressed as mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001 assessed via a two-tailed t-test; ns: non-significant.

Article Snippet: The NC membranes were incubated overnight at 4°C with primary antibodies for FLRT2 (Proteintech, China), OCT4 (CST, USA), CD44 (Abcam, United Kingdom), CD133 (Invitrogen, USA), E-cadherin (Bioworld, China), N-cadherin (Invitrogen, USA), Bax (Proteintech, China), Bcl-2 (Proteintech, China), and NEDD4 (Abcam, United Kingdom), respectively.

Techniques: Over Expression, Transfection, Two Tailed Test

Journal: Frontiers in Pharmacology

Article Title: The NEDD4/FLRT2 axis regulates NSCLC cell stemness

doi: 10.3389/fphar.2024.1459978

Figure Lengend Snippet: Overexpression of FLRT2 inhibits NSCLCCSC proliferation and drug resistance. (A) Viability of NSCLCCSC detected by CCK-8 assay. (B) Proliferation of NSCLCCSC detected by colony formation assay. (C) Changes in colony number. (D, E) Western blotting detected proliferation markers. (F) Bioinformation analyzed FLRET2 correlation with NSCLC cells cisplatin resistance. (G) CCK-8, flow cytometry (H, I) and western blotting (J, K) detecte NSCLC cisplatin resistance cells (A549/CDDP and H1975/CDDP) IC50, cell death and cell drug resistance-related proteins to cisplatin when knockdown FLRT2, respectively. All data are expressed as mean ± SEM of three independent experiments. *p < 0.05, **p < 0.01, and ***p < 0.001 assessed via a two-tailed t-test; ns: non-significant.

Article Snippet: The NC membranes were incubated overnight at 4°C with primary antibodies for FLRT2 (Proteintech, China), OCT4 (CST, USA), CD44 (Abcam, United Kingdom), CD133 (Invitrogen, USA), E-cadherin (Bioworld, China), N-cadherin (Invitrogen, USA), Bax (Proteintech, China), Bcl-2 (Proteintech, China), and NEDD4 (Abcam, United Kingdom), respectively.

Techniques: Over Expression, CCK-8 Assay, Colony Assay, Western Blot, Flow Cytometry, Knockdown, Two Tailed Test

Journal: Frontiers in Pharmacology

Article Title: The NEDD4/FLRT2 axis regulates NSCLC cell stemness

doi: 10.3389/fphar.2024.1459978

Figure Lengend Snippet: NEDD4 mediates the ubiquitination of FLRT2. (A–C) FLRT2 is regulated by proteasome degradation. Cells were incubated with MG132 or E64, CHX at indicated concentration for protein level detection by Western blot. (D) Co-IP depicted the interaction between NEDD4 and FLRT2. (E) NEDD4 regulated the ubiquitination degradation of FLRT2. All data are expressed as mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001 assessed via a two-tailed t-test; ns: non-significant.

Article Snippet: The NC membranes were incubated overnight at 4°C with primary antibodies for FLRT2 (Proteintech, China), OCT4 (CST, USA), CD44 (Abcam, United Kingdom), CD133 (Invitrogen, USA), E-cadherin (Bioworld, China), N-cadherin (Invitrogen, USA), Bax (Proteintech, China), Bcl-2 (Proteintech, China), and NEDD4 (Abcam, United Kingdom), respectively.

Techniques: Ubiquitin Proteomics, Incubation, Concentration Assay, Western Blot, Co-Immunoprecipitation Assay, Two Tailed Test

Journal: Frontiers in Pharmacology

Article Title: The NEDD4/FLRT2 axis regulates NSCLC cell stemness

doi: 10.3389/fphar.2024.1459978

Figure Lengend Snippet: NEDD4 overexpression counteracts the inhibitory effects of FLRT2 on tumorigenesis of A549 CSC . (A) Tumor volume increased in the NEDD4 overexpression group compared with the FLRT2 knockdown group. (B) Tumor weight increased in the NEDD4 overexpression group compared with the FLRT2 knockdown group. (C) Tumors were excised from A549 CSC -injected mice in the indicated three groups. (D) NEDD4 overexpression neutralized the effects of FLRT2 downregulation on the abundance of proliferation-, EMT-, and apoptosis-associated genes. All data are expressed as mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001 assessed via a two-tailed t-test; ns: non-significant.

Article Snippet: The NC membranes were incubated overnight at 4°C with primary antibodies for FLRT2 (Proteintech, China), OCT4 (CST, USA), CD44 (Abcam, United Kingdom), CD133 (Invitrogen, USA), E-cadherin (Bioworld, China), N-cadherin (Invitrogen, USA), Bax (Proteintech, China), Bcl-2 (Proteintech, China), and NEDD4 (Abcam, United Kingdom), respectively.

Techniques: Over Expression, Knockdown, Injection, Two Tailed Test

Journal: International Journal of Veterinary Science and Medicine

Article Title: Generation of induced pluripotent stem cells from the Asian bats

doi: 10.1080/23144599.2024.2384835

Figure Lengend Snippet: Primers used for relative quantitative PCR, according to the reference of Mo et al. [ 15 ] and Déjosez et al. [1] .

Article Snippet: The cells were subsequently incubated with primary mouse antibodies against Oct4 (P0082-200UL, Sigma-Aldrich), and primary rabbit antibodies against Nanog (RCAB004P-F, ReproCELL, Beltsville, MD, USA) at 4°C overnight.

Techniques: Real-time Polymerase Chain Reaction

Journal: International Journal of Veterinary Science and Medicine

Article Title: Generation of induced pluripotent stem cells from the Asian bats

doi: 10.1080/23144599.2024.2384835

Figure Lengend Snippet: Immunofluorescence and qPCR characterization of BatiPSC. To identify the cell type most closely resembling iPSCs, we conducted OCT4 (scale bar = 250 µm) and nanog (scale bar = 75 µm) immunofluorescence staining (A) and RT-qPCR (B). The findings demonstrated strong expression of pluripotent stem cell markers, particularly in colonies with a dome-shaped structure. Asterisks were used to indicate statistical significance between the groups (* p < 0.05, ** p < 0.01, *** p < 0.001).

Article Snippet: The cells were subsequently incubated with primary mouse antibodies against Oct4 (P0082-200UL, Sigma-Aldrich), and primary rabbit antibodies against Nanog (RCAB004P-F, ReproCELL, Beltsville, MD, USA) at 4°C overnight.

Techniques: Immunofluorescence, Staining, Quantitative RT-PCR, Expressing

Journal: International Journal of Veterinary Science and Medicine

Article Title: Generation of induced pluripotent stem cells from the Asian bats

doi: 10.1080/23144599.2024.2384835

Figure Lengend Snippet: Amino acid identity comparison between bat, human, and mouse reprogramming factors.

Article Snippet: The cells were subsequently incubated with primary mouse antibodies against Oct4 (P0082-200UL, Sigma-Aldrich), and primary rabbit antibodies against Nanog (RCAB004P-F, ReproCELL, Beltsville, MD, USA) at 4°C overnight.

Techniques: Comparison