Journal: Bone Reports

Article Title: Identifying the best reference gene for RT-qPCR analyses of the three-dimensional osteogenic differentiation of human induced pluripotent stem cells

doi: 10.1016/j.bonr.2024.101816

Figure Lengend Snippet: Three-dimensional osteogenic differentiation culture using human induced pluripotent stem cells (hiPSCs). ( a ) Schema showing three-dimensional osteogenic differentiation culture using hiPSCs. ( b ) Sections of the sponge were incubated for 28 days, and then stained with Alizarin Red S. Positive areas can be observed at the edges of the sponge. COL1A1 expression was analyzed using confocal laser microscopy. ( c ) Atelocollagen sponge without seeded hiPSCs. ( d)–(g ) Section of the sponge cultured for 28 days after seeding hiPSCs. Blue fluorescence indicates the nuclei and green fluorescence indicates COL1A1 expression. ( d ), ( e ) Cells stained only with DAPI without staining for COL1A1. ( e ) An enlarged image of a part of ( d ). Both ( d ) and ( e ) show the presence of nuclei at the edges and inside the sponge. ( f ), ( g ) Cells with additional staining for COL1A1. ( g ) An enlarged image of a region from ( f ). COL1A1-positive areas can be observed at the edges and inside the sponge. COL1A1, type I collagen alpha 1; hiPSC, human induced pluripotent stem cell.

Article Snippet: These sections were incubated with a primary antibody against type I collagen alpha 1 (COL1A1; diluted 1:100; 322-COLTt, PhosphoSolutions, Denver, CO, USA) at 37 °C in a humid chamber for 1 h. Subsequently, the sections were incubated at 25 °C for 1 h with the secondary antibody Donkey Anti-Rabbit IgG H&L (Alexa Fluor® 488; diluted 1:1000; Thermo Fisher Scientific).

Techniques: Incubation, Staining, Expressing, Microscopy, Cell Culture, Fluorescence

Journal: Bone Reports

Article Title: Identifying the best reference gene for RT-qPCR analyses of the three-dimensional osteogenic differentiation of human induced pluripotent stem cells

doi: 10.1016/j.bonr.2024.101816

Figure Lengend Snippet: Primers used for amplifying the target genes in the present study.

Article Snippet: These sections were incubated with a primary antibody against type I collagen alpha 1 (COL1A1; diluted 1:100; 322-COLTt, PhosphoSolutions, Denver, CO, USA) at 37 °C in a humid chamber for 1 h. Subsequently, the sections were incubated at 25 °C for 1 h with the secondary antibody Donkey Anti-Rabbit IgG H&L (Alexa Fluor® 488; diluted 1:1000; Thermo Fisher Scientific).

Techniques: Functional Assay, SYBR Green Assay, Marker, Gene Expression

Journal:

Article Title: The matrix component biglycan is proinflammatory and signals through Toll-like receptors 4 and 2 in macrophages

doi: 10.1172/JCI23755

Figure Lengend Snippet: Increased expression of BGN in infiltrating cells of lung parenchyma in sepsis. (A and B) Northern blot of BGN mRNA normalized to α-tubulin (2 hours) (A) and Western blot of BGN core protein normalized to α-tubulin (8 hours) (B) from lungs of Bgn+/0 mice after a lethal dose of LPS versus PBS. (C and D) Immunostaining of BGN (red stain) in lungs from control (C) and septic Bgn+/0 mice (D) 8 hours after a lethal dose of LPS. (E and F) Double staining, marked by arrows, for BGN (blue) and macrophages (F4/80, red) (E) and in situ hybridization for BGN (F) in septic lungs from Bgn+/0 mice (8 hours). The lower right inset shows a magnified view of mononuclear cells expressing BGN (white arrowhead). The upper left inset represents the sense riboprobe. (G and H) Immunostaining of macrophages (F4/80, red), with numerous F4/80–positive macrophages (lower right insets), in septic lungs from Bgn+/0 (G) versus Bgn–/0 mice (H). (I–L) Immunostaining of type I collagen in lungs from Bgn+/0 (brown) (I) versus Bgn–/0 mice (J) and Western blots of fibronectin (K) and laminin (L) normalized to β-tubulin in lungs from Bgn+/0 versus Bgn–/0 mice 8 hours after a lethal dose of LPS. The lower right insets in C, D, F, G, and H represent higher magnifications. Scale bars in D, H, and J apply to panels in C, G, and I, respectively.

Article Snippet: Primary antibodies included LF-113, a rabbit anti–murine decorin antiserum ( 54 ), LF-106 (1:500), rabbit anti–mouse fibromodulin (1:100) ( 33 ), rabbit anti–rat type I collagen (1:50; Quartett GmbH), rabbit anti–human fibronectin (1:200; Sigma-Aldrich), biotinylated goat anti–type VI collagen (1:20; Southern Biotechnology Associates Inc.), and, for populations of monocytes/macrophages, rat anti-mouse F4/80 (1:2,000; BD Biosciences — Pharmingen).

Techniques: Expressing, Northern Blot, Western Blot, Immunostaining, Staining, Double Staining, In Situ Hybridization