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

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

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

Journal: bioRxiv

Article Title: Using atorvastatin-induced vascular weakness to model brain haemorrhage in vascularised cerebral organoids

doi: 10.64898/2026.04.20.719465

Figure Lengend Snippet: ( A ) Phase contrast images of HBMECs after 24 hours of treatment with 100 μM atorvastatin (ATV). Scale bars 50 μm. ( B ) Cell survival after ATV treatment for 24 hours in both stationary and rotating culture conditions ( n =4 independent repeats). Scale bars 100 μm. ( C ) qPCR analysis of marker expression after ATV treatment in rotating cells shows an increase in VE-Cadherin and NG2 gene expression (non-significant, One-Way ANOVA). ( D ) Fluorescent images of CD31 (green) and ZO-1 (red) cellular expression after 24 hours with ATV and DMSO control. ( E ) Total expression of ZO-1 after treatment ( n =4 independent repeats, two-way ANOVA *** P =0.0009, **** P <0.0001) ( F ) Analysis of ZO-1 co-localised with CD31 on the cell surface when treated with ATV ( n =4 independent repeats, two-way ANOVA, *** P =0.0002). ( G ) Fluorescent images of CD31 (green) and VE-Cadherin (red) cellular expression after 24 hours of ATV treatment in both stationary (top panels) and rotating culture (bottom panels). Zoom inset shows the internalisation of both CD31 and VE-Cadherin from the cell surface when treated with ATV. Scale bars 20 μm. ( H ) Analysis of VE-Cadherin co-localised with CD31 on the cell surface when treated with ATV ( n =3 independent repeats, two-way ANOVA, * P <0.04). ( I ) Total area expression of CD31 and VE-Cadherin as a percentage area of DAPI in both stationary and rotating cells, ( n =3 independent repeats, no significance from analysis with a two-way ANOVA).

Article Snippet: Primary human brain microvascular endothelial cells (HBMECs) (Innoprot P10361) were maintained on 1% gelatin coated flasks in endothelial cell growth medium MV (PromoCell C-22020).

Techniques: Marker, Expressing, Gene Expression, Control

Journal: bioRxiv

Article Title: Using atorvastatin-induced vascular weakness to model brain haemorrhage in vascularised cerebral organoids

doi: 10.64898/2026.04.20.719465

Figure Lengend Snippet: ( A ) Representative images of filipin-stained HBMECs, after 24hours of drug treatment. Scale bars 50 μm. ( B ) Filipin expression as a percentage of area from 6 ROIs in 2 wells across 2 independent repeats after 24 hours with ATV, ** P =0.0081, t -test. ( C ) qPCR analysis of rotated HBMECs after 24 hours of drug treatment shows a significant increase in HMGCR expression ( n =5 independent repeats, * P =0.0323 One-Way ANOVA).

Article Snippet: Primary human brain microvascular endothelial cells (HBMECs) (Innoprot P10361) were maintained on 1% gelatin coated flasks in endothelial cell growth medium MV (PromoCell C-22020).

Techniques: Staining, Expressing

Journal: bioRxiv

Article Title: Using atorvastatin-induced vascular weakness to model brain haemorrhage in vascularised cerebral organoids

doi: 10.64898/2026.04.20.719465

Figure Lengend Snippet: ( A ) Phase images of tube formation in untreated and ATV-treated HBMECs. Scale bars 20 μm. ( B ) Analysis of the network parameter average vessel length shows a significant reduction with ATV treatment, both pre-tube formation and post (One-way ANOVA with Tukey’s post hoc multiple comparisons test ** P =0.003, *** P =0.0007, **** P <0.0001). ( C ) Cytotoxicity analysis of ATV treated tubes showed no difference from controls ( n =3, t -test).

Article Snippet: Primary human brain microvascular endothelial cells (HBMECs) (Innoprot P10361) were maintained on 1% gelatin coated flasks in endothelial cell growth medium MV (PromoCell C-22020).

Techniques:

Journal: bioRxiv

Article Title: Using atorvastatin-induced vascular weakness to model brain haemorrhage in vascularised cerebral organoids

doi: 10.64898/2026.04.20.719465

Figure Lengend Snippet: ( A ) Representative confocal images of whole organoids treated with ATV for 24 hours and the loss of VE-Cadherin expression from the surface. Scale bars 500 μm. ( B ) VE-Cadherin, not CD31 ( P =0.091), expressed as a percentage of DAPI was significantly reduced with ATV treatment compared to DMSO controls (two-tailed t -test, * P =0.011). ( C ) Size progression for 4 batches of organoids that were treated with ATV at day 40. ( D ) Vascular metrics were unchanged for CD31 ( n =15 organoids from 4 batches, non-significant t- test) with slightly more endpoints, signifying single cells. ( E ) Angiotool analysis of VE-Cadherin staining revealed shorter overall vessel length ( n =15 organoids from 4 batches, t- test, * P =0.0389)). ( F ) qPCR analysis of ATV-treated organoids showed a reduction in VE-Cadherin RNA expression; however, other markers of endothelial function are unchanged (One-Way ANOVA, n =5 organoids from 2 batches). ( G ) ATV treatment increased the expression of some cholesterol biosynthesis markers compared to DMSO controls, opposite to what was observed in HBMECs in 2D (One-Way ANOVA, n =5 organoids from 2 batches).

Article Snippet: Primary human brain microvascular endothelial cells (HBMECs) (Innoprot P10361) were maintained on 1% gelatin coated flasks in endothelial cell growth medium MV (PromoCell C-22020).

Techniques: Expressing, Two Tailed Test, Staining, RNA Expression

Journal: Cell Reports Medicine

Article Title: Enalaprilat reverses neutrophil polarization imbalance via targeting taurine-STING axis for treatment of diabetic wounds

doi: 10.1016/j.xcrm.2026.102714

Figure Lengend Snippet: Phenotype repolarization of neutrophils induced by Ena affected the pro-angiogenic capacities of HUVECs and inflammatory state of macrophages (A) Flow cytometry was adopted to reveal the difference of CD206+ population ratio among the three groups. n = 3 independent experiments. (B) Effects of AGE stimulation and sequential Ena supplement on the expression of ICAM1, CXCR2, CXCR4, and Fas as evaluated by western blot. (C) Effects of AGE stimulation and sequential Ena supplement on the expression of ICAM1, CXCR2, CXCR4, and Fas as evaluated by immunofluorescence staining. Scale bar, 2 μm; n = 3 independent experiments. (D) Quantitative reverse-transcription PCR (RT-qPCR) was employed to quantify the level of il1b , ccl3 , arg1 , and ccl17 . n = 3 independent experiments. (E) Schematic diagram for the incubation of HUVECs with pretreated neutrophils. (F) Proliferative ability of HUVECs under the stimulation of neutrophils with indicated preconditioning as detected via EdU staining. Scale bar, 200 μm; n = 3 independent experiments. (G) Migration property of HUVECs as measured by transwell assay. Scale bar, 200 μm; n = 3 independent experiments. (H) Migration property of HUVECs as measured by wound scratch test. Scale bar, 500 μm. (I) Tube formation assay was performed to visualize the neovascularization function of HUVECs with different treatments. Scale bar, 200 μm; n = 3 independent experiments. (J) Phenotype characters of macrophages irritated by neutrophils as determined by immunofluorescence staining. Scale bar, 5 μm. (K) Phenotype characters of macrophages irritated by neutrophils as determined by flow cytometry. n = 3 independent experiments. Data were shown as mean ± standard deviation (SD) from biological replicates, and statistical analyses were performed using one-way ANOVA test followed by Tukey’s multiple comparisons test in (A–D and F–K). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: Neutrophil cell line (HL-60) and human umbilical vein endothelial cell (HUVEC) were gained from the American Type Culture Collection (ATCC) and grown at 37°C with 5% CO 2 .

Techniques: Flow Cytometry, Expressing, Western Blot, Immunofluorescence, Staining, Reverse Transcription, Quantitative RT-PCR, Incubation, Migration, Transwell Assay, Tube Formation Assay, Standard Deviation

Journal: Cell Reports Medicine

Article Title: Enalaprilat reverses neutrophil polarization imbalance via targeting taurine-STING axis for treatment of diabetic wounds

doi: 10.1016/j.xcrm.2026.102714

Figure Lengend Snippet: Inhibitory effects of Ena on HUVEC ferroptosis activated by AGE-elicited neutrophils (A) CCK-8 assay was performed to measure the viability of HUVECs. n = 3 independent experiments. (B) Death rate of HUVECs incubated with neutrophils pretreated by different strategies as detected using flow cytometry. n = 3 independent experiments. (C) Levels of MDA and GSH were quantified to assess the ferroptosis activity. n = 3 independent experiments. (D) Intracellular lipid peroxidation was visualized by fluorescence staining. Scale bar, 10 μm; n = 3 independent experiments. (E) Fe 2+ ion content was visualized by fluorescence staining. Scale bar, 10 μm; n = 3 independent experiments. (F) JC-1 kit was used to evaluate the mitochondrial membrane potential of HUVECs. Scale bar, 10 μm; n = 3 independent experiments. (G) DCFH-DA fluorescence probe was applied to determine ROS abundance in HUVECs. Scale bar, 200 μm; n = 3 independent experiments. (H) Mitochondrial morphology in HUVECs as analyzed by transmission electron microscopy. Scale bar, 500 nm; n = 3 independent experiments. (I) Western blot detection of PGC1α and KLF9 in HUVECs with different treatments. n = 3 independent experiments. (J) Expression of KLF9 in HUVECs incubated with neutrophils pretreated by different approaches as detected using immunofluorescence staining. Scale bar, 10 μm; n = 3 independent experiments. (K) Expression of PGC1α in HUVECs incubated with neutrophils pretreated by different approaches as detected using immunofluorescence staining. Scale bar, 10 μm; n = 3 independent experiments. (L) Schematic diagram of Ena-elicited alleviation on endothelial cell ferroptosis induced by pro-inflammatory neutrophils. Data were shown as mean ± standard deviation (SD) from biological replicates, and statistical comparisons were performed using one-way ANOVA followed by Tukey’s multiple comparisons test in (A–G and I–K). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: Neutrophil cell line (HL-60) and human umbilical vein endothelial cell (HUVEC) were gained from the American Type Culture Collection (ATCC) and grown at 37°C with 5% CO 2 .

Techniques: CCK-8 Assay, Incubation, Flow Cytometry, Activity Assay, Fluorescence, Staining, Membrane, Transmission Assay, Electron Microscopy, Western Blot, Expressing, Immunofluorescence, Standard Deviation