Journal: bioRxiv

Article Title: RetroCHMP3 Blocks Budding of Enveloped Viruses Without Blocking Cytokinesis

doi: 10.1101/2020.08.30.273656

Figure Lengend Snippet: (A) Schematic of putative regulatory elements upstream of retroCHMP3 ORF in Mus musculus identified by transcription factor binding site prediction tools. Numbers indicate nucleotides relative to retroCHMP3 start codon. LTR – long terminal repeat, MuRRS – murine retrovirus-related sequence, ERV – endogenous retrovirus. (B) RT-PCR detection of retroCHMP3 RNA in mouse cardiac endothelial cells (MCECs) with and without interferon stimulation. RetroCHMP3 bands were excised, cloned, and sequenced to verify a match with the retroCHMP3 sequence. Representative gel from three independent biological repeats. RT – reverse transcriptase. (C) Droplet digital PCR (ddPCR) detection of retroCHMP3 (red) and RNAse L (grey, positive control) RNA in mouse cardiac endothelial cells (MCECs) with and without interferon stimulation. The housekeeping gene succinate dehydrogenase complex, subunit A (SDHA) was used for normalization. Each line represents one independent biological repeat.

Article Snippet: MCEC cells (mouse cardiac endothelial cells, CLU510, sex: unknown) were obtained from Cedarlane.

Techniques: Binding Assay, Sequencing, Reverse Transcription Polymerase Chain Reaction, Clone Assay, Reverse Transcription, Digital PCR, Positive Control

Journal: Journal of Investigative Dermatology

Article Title: Loss of EPC-1/PEDF Expression During Skin Aging In Vivo

doi: 10.1111/j.0022-202x.2004.22510.x

Figure Lengend Snippet: Figure 8 Microvessel density. Skin sections from young and old donors were pro- cessed by immunofluorescence to de- termine CD31 to identify endothelial cells in the dermis and stained with DAPI to identify nuclei of all cells in the tissue sections. Microvessels were enumerated in five to eight sections per donor and the average number per section is shown in A (po0.005, Stu- dent’s t test, Excel). Representative images of the CD31 staining (green fluorescence) in three random young (B–D) and old (F–H) donors are indi- cated. Donors: 24 y old (B), 24 y old (C), 27 y old (D), 68 y old (E) and 74 y old (F), and 78 y old (G). indicates microves- sels stained with CD31. Scale bar ¼ 100 mm.

Article Snippet: The following primary antibodies were used: a rabbit polyclonal antibody to PEDF (BioProducts Maryland, Middletown, Maryland), a mouse monoclonal anti-human CD31 to detect endothelial cells (R&D Systems; Minneapolis, Minnesota), a rat monoclonal antibody to heparan sulfate proteoglycans to detect basement membranes (Biomeda; Foster City, California), a rabbit polyclonal antibody to Ki67 to detect proliferating cells (Novocastra Laboratories, Newcastle-upon-Tyne, UK, Distributed by US Vector Labs, Burlingame, CA), a chicken anti-human TSP-1 (thrombospondin, kindly provided by Robert Swerlick, Emory University), a rabbit polyclonal antibody to TSP-2 (kindly provided by Michael Detmar, Harvard Medical School/Massachusetts General Hospital), and an affinity purified rabbit polyclonal antibody to VEGF (A-20, Santa Cruz Biotechnology; Santa Cruz, California).

Techniques: Staining

Journal: Bone research

Article Title: Metformin accelerates bone fracture healing by promoting type H vessel formation through inhibition of YAP1/TAZ expression.

doi: 10.1038/s41413-023-00279-4

Figure Lengend Snippet: Fig. 1 Metformin promotes angiogenesis under hypoxic conditions in vitro. Representative images (a) of the transwell migration assay with quantification of crystal violet-stained migrated HMECs (b) treated with metformin under different oxygen concentrations (21% and 1% O2). Met: metformin. Scale bar: 100 μm. n = 3 per group. Representative images (c) of tube formation of HMECs on Matrigel with quantification of the total branching points (d), total tube length (e), and total loop numbers (f). Scale bar: 200 μm. n = 3 per group. g CCK-8 analysis of the proliferation of HMECs. n = 4 per group. h qRT‒PCR analysis of the mRNA levels of the HIF-1α target genes Vegfa and Lrg1 in HMECs with or without metformin treatment under different oxygen concentrations. n = 3 per group. Data are presented as the mean ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001

Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The concentrations of OCN and VEGFA were determined using the Mouse OCN (Elabscience, E-EL-M0864c, Wuhan, China) or VEGFA (Elabscience, E-EL-M1292c) ELISA kit according to the manufacturers’ instructions.

Techniques: In Vitro, Transwell Migration Assay, Staining, CCK-8 Assay

Journal: Bone research

Article Title: Metformin accelerates bone fracture healing by promoting type H vessel formation through inhibition of YAP1/TAZ expression.

doi: 10.1038/s41413-023-00279-4

Figure Lengend Snippet: Fig. 3 Metformin promotes type H vessel formation in osteoporotic fracture mice. a Representative CD31 and Emcn coimmunostaining images (left) with quantification of the type H vessel ratio in calluses (right) from the osteoporotic mice treated with PBS, Met, PTH, and ALN at 3, 6, and 9 weeks post-fracture. ca: callus. The dotted line represents the boundary of the callus. Met metformin, ALN alendronate, PTH parathyroid hormone. Scale bar: 100 μm. n = 5 per group. b Representative Ki67 and Emcn coimmunostaining images (left) with quantification of the number of Ki67-positive endothelial cells in calluses (right) from the osteoporotic mice treated with PBS, Met, PTH, and ALN at 3, 6, and 9 weeks post-fracture. Scale bar: 100 μm. n = 5 per group. ELISAs for the serum (c) and bone marrow (d) concentrations of VEGFA at 9 weeks post-osteoporotic fracture. n = 8 per group; Data are presented as the mean ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001

Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The concentrations of OCN and VEGFA were determined using the Mouse OCN (Elabscience, E-EL-M0864c, Wuhan, China) or VEGFA (Elabscience, E-EL-M1292c) ELISA kit according to the manufacturers’ instructions.

Techniques:

Journal: Bone research

Article Title: Metformin accelerates bone fracture healing by promoting type H vessel formation through inhibition of YAP1/TAZ expression.

doi: 10.1038/s41413-023-00279-4

Figure Lengend Snippet: Fig. 4 Metformin promotes the expression of HIF-1α by inhibiting the expression of YAP1/TAZ in HMECs under hypoxic conditions. a qRT‒PCR analysis of the inhibitory efficiency of siRNAs targeting HIF-1α. n = 3 per group. b qRT‒PCR analysis of the expression of HIF-1α and its target genes Vegfa and Lrg1 in the si-HIF-1α-transfected HMECs with or without metformin treatment under hypoxic conditions (1% O2). Met: metformin. n = 3 per group. Immunofluorescence staining images and quantification showing the protein levels of HIF-1α (c), YAP1 (d), and TAZ (e) in the HMECs treated with PBS (control) or metformin under hypoxic conditions. Scale bar: 20 μm. n = 9 per group. f qRT‒PCR analysis of the inhibitory efficiency of siRNAs targeting YAP1 or TAZ. n = 3 per group. g Immunofluorescence staining images and quantification showing the protein level of HIF-1α in the hypoxia-cultured HMECs from the si-Con, si-YAP1, si-TAZ, si-Y/T, and si-Y/T + Met groups. Y/T: YAP1 and TAZ. Scale bar: 20 μm. n = 3 per group. Data are presented as the mean ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001

Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The concentrations of OCN and VEGFA were determined using the Mouse OCN (Elabscience, E-EL-M0864c, Wuhan, China) or VEGFA (Elabscience, E-EL-M1292c) ELISA kit according to the manufacturers’ instructions.

Techniques: Expressing, Transfection, Staining, Control, Cell Culture