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Assembly and characterization of human 3D liver spheroids via DNA origami NAC-linkers. (A) Schematic of 3D liver spheroid self-assembly from primary human <t>hepatocytes,</t> liver sinusoidal endothelial cells, and Kupffer cells using NAC-linkers. (B) Atomic force microscopy image of NAC-linkers. Scale bars, 200 nm. (C) 1% agarose gel electrophoresis confirming cholesterol-modified NAC-linkers assembly (lanes: DNA marker, M13mp18 scaffold, and NAC-linkers). (D) Bright-field image of a mature spheroid. (E) Hematoxylin and eosin (H&E) staining of a spheroid section. (F) Immunofluorescence staining of cell type markers in human 3D liver spheroids: albumin (ALB, hepatocytes), CD31 (endothelial cells), and CD68 (Kupffer cells). Scale bars, 200 μm.
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Assembly and characterization of human 3D liver spheroids via DNA origami NAC-linkers. (A) Schematic of 3D liver spheroid self-assembly from primary human <t>hepatocytes,</t> liver sinusoidal endothelial cells, and Kupffer cells using NAC-linkers. (B) Atomic force microscopy image of NAC-linkers. Scale bars, 200 nm. (C) 1% agarose gel electrophoresis confirming cholesterol-modified NAC-linkers assembly (lanes: DNA marker, M13mp18 scaffold, and NAC-linkers). (D) Bright-field image of a mature spheroid. (E) Hematoxylin and eosin (H&E) staining of a spheroid section. (F) Immunofluorescence staining of cell type markers in human 3D liver spheroids: albumin (ALB, hepatocytes), CD31 (endothelial cells), and CD68 (Kupffer cells). Scale bars, 200 μm.
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Assembly and characterization of human 3D liver spheroids via DNA origami NAC-linkers. (A) Schematic of 3D liver spheroid self-assembly from primary human <t>hepatocytes,</t> liver sinusoidal endothelial cells, and Kupffer cells using NAC-linkers. (B) Atomic force microscopy image of NAC-linkers. Scale bars, 200 nm. (C) 1% agarose gel electrophoresis confirming cholesterol-modified NAC-linkers assembly (lanes: DNA marker, M13mp18 scaffold, and NAC-linkers). (D) Bright-field image of a mature spheroid. (E) Hematoxylin and eosin (H&E) staining of a spheroid section. (F) Immunofluorescence staining of cell type markers in human 3D liver spheroids: albumin (ALB, hepatocytes), CD31 (endothelial cells), and CD68 (Kupffer cells). Scale bars, 200 μm.
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ATCC human endothelial ea hy926 cells
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Assembly and characterization of human 3D liver spheroids via DNA origami NAC-linkers. (A) Schematic of 3D liver spheroid self-assembly from primary human hepatocytes, liver sinusoidal endothelial cells, and Kupffer cells using NAC-linkers. (B) Atomic force microscopy image of NAC-linkers. Scale bars, 200 nm. (C) 1% agarose gel electrophoresis confirming cholesterol-modified NAC-linkers assembly (lanes: DNA marker, M13mp18 scaffold, and NAC-linkers). (D) Bright-field image of a mature spheroid. (E) Hematoxylin and eosin (H&E) staining of a spheroid section. (F) Immunofluorescence staining of cell type markers in human 3D liver spheroids: albumin (ALB, hepatocytes), CD31 (endothelial cells), and CD68 (Kupffer cells). Scale bars, 200 μm.

Journal: One Health

Article Title: Human 3D liver spheroids support productive infection of a novel tick-borne phenuivirus

doi: 10.1016/j.onehlt.2026.101321

Figure Lengend Snippet: Assembly and characterization of human 3D liver spheroids via DNA origami NAC-linkers. (A) Schematic of 3D liver spheroid self-assembly from primary human hepatocytes, liver sinusoidal endothelial cells, and Kupffer cells using NAC-linkers. (B) Atomic force microscopy image of NAC-linkers. Scale bars, 200 nm. (C) 1% agarose gel electrophoresis confirming cholesterol-modified NAC-linkers assembly (lanes: DNA marker, M13mp18 scaffold, and NAC-linkers). (D) Bright-field image of a mature spheroid. (E) Hematoxylin and eosin (H&E) staining of a spheroid section. (F) Immunofluorescence staining of cell type markers in human 3D liver spheroids: albumin (ALB, hepatocytes), CD31 (endothelial cells), and CD68 (Kupffer cells). Scale bars, 200 μm.

Article Snippet: Primary human hepatocytes, liver sinusoidal endothelial cells, and Kupffer cells (IxCell Biotechnology) were mixed at specific ratios and co-incubated with NAC-Linker A and B (Puheng Biomedicine, NAC001) to facilitate NAC structure formation on the cell surfaces.

Techniques: Microscopy, Agarose Gel Electrophoresis, Modification, Marker, Staining, Immunofluorescence

Adaptation and pathogenesis of MKWV in human 3D liver spheroids. (A) Schematic of serial passaging of the HLJ1 strain in spheroids, yielding the adapted NAC-Org5 strain. (B, C) Viral RNA copies (B) and TCID₅₀ titers (C) across passages (P1-P5). (D) Bright-field image of spheroids infected with passage 5 (P5) virus, showing structural disruption. Scale bar, 100 μm. (E) Quantification of spheroid diameter post-infection. (F) Transmission electron micrographs of virions within cytoplasmic vesicles of infected spheroids. Scale bars: 1 μm (left), 200 nm (right). (G) Representative images and quantification of nuclei showing infection-induced cell death. Scale bar, 200 μm. (H) Western blot detecting cleaved caspase-3 in spheroids at 48 and 72 h post-infection (hpi). (I) Multiplex immunofluorescence showing NAC-Org5 tropism for CD31 + endothelial cells and CD68 + Kupffer cells, with weaker detection in ALB + hepatocytes. Scale bar, 200 μm. (J) Functional assessment of infected spheroids: ATP (viability), ALT/AST/LDH (damage), ALB/urea (synthetic function). (K) RT-qPCR analysis of pro-inflammatory cytokine mRNA expression, normalized to β-actin. Data are mean ± SD ( n = 5 biological replicates). * p < 0.05, ** p < 0.01.

Journal: One Health

Article Title: Human 3D liver spheroids support productive infection of a novel tick-borne phenuivirus

doi: 10.1016/j.onehlt.2026.101321

Figure Lengend Snippet: Adaptation and pathogenesis of MKWV in human 3D liver spheroids. (A) Schematic of serial passaging of the HLJ1 strain in spheroids, yielding the adapted NAC-Org5 strain. (B, C) Viral RNA copies (B) and TCID₅₀ titers (C) across passages (P1-P5). (D) Bright-field image of spheroids infected with passage 5 (P5) virus, showing structural disruption. Scale bar, 100 μm. (E) Quantification of spheroid diameter post-infection. (F) Transmission electron micrographs of virions within cytoplasmic vesicles of infected spheroids. Scale bars: 1 μm (left), 200 nm (right). (G) Representative images and quantification of nuclei showing infection-induced cell death. Scale bar, 200 μm. (H) Western blot detecting cleaved caspase-3 in spheroids at 48 and 72 h post-infection (hpi). (I) Multiplex immunofluorescence showing NAC-Org5 tropism for CD31 + endothelial cells and CD68 + Kupffer cells, with weaker detection in ALB + hepatocytes. Scale bar, 200 μm. (J) Functional assessment of infected spheroids: ATP (viability), ALT/AST/LDH (damage), ALB/urea (synthetic function). (K) RT-qPCR analysis of pro-inflammatory cytokine mRNA expression, normalized to β-actin. Data are mean ± SD ( n = 5 biological replicates). * p < 0.05, ** p < 0.01.

Article Snippet: Primary human hepatocytes, liver sinusoidal endothelial cells, and Kupffer cells (IxCell Biotechnology) were mixed at specific ratios and co-incubated with NAC-Linker A and B (Puheng Biomedicine, NAC001) to facilitate NAC structure formation on the cell surfaces.

Techniques: Passaging, Infection, Virus, Disruption, Transmission Assay, Western Blot, Multiplex Assay, Immunofluorescence, Functional Assay, Quantitative RT-PCR, Expressing

Pathogenicity of the NAC-Org5 strain in murine models. (A) Experimental schematic for intracranial (3-day-old) and intraperitoneal (3-week-old) inoculation of BALB/c mice. (B, C) Survival (B) and weight change (C) of suckling mice after NAC-Org5 infection. (D) Viral load in tissues and blood of suckling mice at 7 dpi. (E, F) Survival (E) and weight change (F) of 3-week-old mice. (G) Viral load in tissues and blood of 3-week-old mice at 7 dpi. Data are from 3 independent experiments. (H) Representative H& E -stained liver sections from 3-week-old mice at 7 and 15 dpi, showing inflammatory infiltrates and hepatocyte necrosis that resolves by 15 dpi. Scale bar, 100 μm. *** p < 0.001.

Journal: One Health

Article Title: Human 3D liver spheroids support productive infection of a novel tick-borne phenuivirus

doi: 10.1016/j.onehlt.2026.101321

Figure Lengend Snippet: Pathogenicity of the NAC-Org5 strain in murine models. (A) Experimental schematic for intracranial (3-day-old) and intraperitoneal (3-week-old) inoculation of BALB/c mice. (B, C) Survival (B) and weight change (C) of suckling mice after NAC-Org5 infection. (D) Viral load in tissues and blood of suckling mice at 7 dpi. (E, F) Survival (E) and weight change (F) of 3-week-old mice. (G) Viral load in tissues and blood of 3-week-old mice at 7 dpi. Data are from 3 independent experiments. (H) Representative H& E -stained liver sections from 3-week-old mice at 7 and 15 dpi, showing inflammatory infiltrates and hepatocyte necrosis that resolves by 15 dpi. Scale bar, 100 μm. *** p < 0.001.

Article Snippet: Primary human hepatocytes, liver sinusoidal endothelial cells, and Kupffer cells (IxCell Biotechnology) were mixed at specific ratios and co-incubated with NAC-Linker A and B (Puheng Biomedicine, NAC001) to facilitate NAC structure formation on the cell surfaces.

Techniques: Infection, Staining

Angiogenic activity of endothelial cells in response to CDP scaffolds. (a) Phase-contrast images of EA.hy926 endothelial tube networks formed on Matrigel in indirect co-culture with collagen, CHA-7, CDP-3, CDP-5 or CDP-7 scaffolds. Top row: original images; bottom row: binarized skeletons obtained using the Angiogenesis Analyzer plugin. (b–d) Quantitative analysis of (b) number of junctions, (c) number of tubes and (d) number of loops, showing progressively enhanced network complexity with increasing DDM-p content and maximal values for CDP-7. (e–g) qRT-PCR analysis of angiogenesis-related genes in EA.hy926 cells, including (e) CD31, (f) VEGFA and (g) HIF-1α, demonstrating significant upregulation particularly on CDP-7 scaffolds. Data are mean ± SD; ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001; ns, not significant.

Journal: Materials Today Bio

Article Title: Cryo-printed collagen scaffolds reinforced with dentin-derived bioactive particles promote osteo-angiogenic bone regeneration

doi: 10.1016/j.mtbio.2026.102853

Figure Lengend Snippet: Angiogenic activity of endothelial cells in response to CDP scaffolds. (a) Phase-contrast images of EA.hy926 endothelial tube networks formed on Matrigel in indirect co-culture with collagen, CHA-7, CDP-3, CDP-5 or CDP-7 scaffolds. Top row: original images; bottom row: binarized skeletons obtained using the Angiogenesis Analyzer plugin. (b–d) Quantitative analysis of (b) number of junctions, (c) number of tubes and (d) number of loops, showing progressively enhanced network complexity with increasing DDM-p content and maximal values for CDP-7. (e–g) qRT-PCR analysis of angiogenesis-related genes in EA.hy926 cells, including (e) CD31, (f) VEGFA and (g) HIF-1α, demonstrating significant upregulation particularly on CDP-7 scaffolds. Data are mean ± SD; ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001; ns, not significant.

Article Snippet: For the experiments, pre-osteoblastic MC3T3-E1 cells (ATCC, Manassas, VA, USA), human endothelial EA.hy926 cells (ATCC, Manassas, VA, USA), and human adipose-derived stem cells (hASCs, ATCC, Manassas, VA, USA) were used.

Techniques: Activity Assay, Co-Culture Assay, Quantitative RT-PCR

Pro-angiogenic and pro-migratory effects of hydrogels. Immunofluorescence staining shows blood vessel formation of HUVECs in LPS-macrophage condition medium (A). Scratch assays and Transwell migration assays of HUVECs (B) and L929 cells (C). Quantification of junctions (D), branches (E), wound closure percentage (F–G) and migrated cells (H–I). One-way ANOVA with Tukey's post hoc test and n = 3 for D-I; ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, n.s. ANOVA, analysis of variance; CS, chitosan; IBU, ibuprofen; GP, genipin; MA, methacrylic anhydride; LPS, lipopolysaccharides; HUVEC, human umbilical vein endothelial cell; n.s., not significant.

Journal: Materials Today Bio

Article Title: Injectable chitosan-based hydrogel via in situ gelation modulates the inflammatory microenvironment and facilitates minimally invasive repair of peripheral nerve injury

doi: 10.1016/j.mtbio.2026.102814

Figure Lengend Snippet: Pro-angiogenic and pro-migratory effects of hydrogels. Immunofluorescence staining shows blood vessel formation of HUVECs in LPS-macrophage condition medium (A). Scratch assays and Transwell migration assays of HUVECs (B) and L929 cells (C). Quantification of junctions (D), branches (E), wound closure percentage (F–G) and migrated cells (H–I). One-way ANOVA with Tukey's post hoc test and n = 3 for D-I; ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, n.s. ANOVA, analysis of variance; CS, chitosan; IBU, ibuprofen; GP, genipin; MA, methacrylic anhydride; LPS, lipopolysaccharides; HUVEC, human umbilical vein endothelial cell; n.s., not significant.

Article Snippet: Mouse fibroblasts (L929, ATCC), human umbilical vein endothelial cells (HUVECs, ATCC), and mouse macrophages (RAW 264.7, ATCC) were provided by the Cell Bank of the Chinese Academy of Sciences.

Techniques: Immunofluorescence, Staining, Migration