Journal: Archives of Toxicology

Article Title: Establishment of a human 3D in vitro liver-bone model as a potential system for drug toxicity screening

doi: 10.1007/s00204-024-03899-9

Figure Lengend Snippet: The antibodies used for dot blot

Article Snippet: Soluble receptor activator of nuclear factor kappa-B ligand (sRANKL) , 500-m46 , PeproTech, Hamburg, Germany , 1:1000.

Techniques:

Journal: Archives of Toxicology

Article Title: Establishment of a human 3D in vitro liver-bone model as a potential system for drug toxicity screening

doi: 10.1007/s00204-024-03899-9

Figure Lengend Snippet: DCFH-DA assay was used to detect the ROS levels in liver spheroids. Ellman assay was used to detect the GSH and GSSG levels in liver spheroids. The expression of secreted IL-6, sRANKL, and OPG protein levels in the liver-bone system supernatant were measured by dot blot. a After acute exposure to diclofenac, the ROS levels in liver spheroids on day 7, H 2 O 2 stimulation as a positive control. b After acute exposure to diclofenac, the ratio of GSH and GSSG in liver spheroids on day 7. c With the stimulation of 3–6 µM diclofenac, the IL-6 protein levels in supernatant on day 7, and the representative image of dot blot. d With the stimulation of 3–6 µM diclofenac, the sRANKL protein levels in supernatant on day 7, and the representative image of dot blot. e With the stimulation of 3–6 µM diclofenac, the OPG protein levels in supernatant on day 7, and the representative image of dot blot. f The ratio of sRANKL and OPG in the liver-bone co-culture system. The Kruskal–Wallis test followed by Dunn’s multiple comparison test was used to determine statistical differences. Data are presented as means ± SEM, and the significance is shown as * p < 0.05, *** p < 0.001, and **** p < 0.0001 vs. Control group. N ≥ 3, n ≥ 2

Article Snippet: Soluble receptor activator of nuclear factor kappa-B ligand (sRANKL) , 500-m46 , PeproTech, Hamburg, Germany , 1:1000.

Techniques: DCFH-DA Assay, Expressing, Dot Blot, Positive Control, Co-Culture Assay, Comparison, Control

Journal: International Journal of Molecular Sciences

Article Title: Microbiota-Derived Metabolites, Indole-3-aldehyde and Indole-3-acetic Acid, Differentially Modulate Innate Cytokines and Stromal Remodeling Processes Associated with Autoimmune Arthritis

doi: 10.3390/ijms22042017

Figure Lengend Snippet: IAld enhanced the bone remodeling process of osteoclastogenesis. RAW cells were differentiated with receptor activator of NF-κB-ligand (RANKL) alone or with RANKL in the presence of either IAld or I3AA (0.0125–0.2 mM) as described in Methods. ( a , b ) Cells were fixed and then stained for tartrate-acid phosphates (TRAP) expression on d 5 and the number of osteoclast-like cells were enumerated from multiple regions of each well ( n = 5). Data was reported as the percentage of osteoclast-like cells relative to the RANKL-treated controls (Mean ± SEM) from a representative experiment. Statistical difference relative to RANKL-treated control cells was determined by ordinary one-way ANOVA with post-hoc test for multiple comparisons. ( c , d ) The transcription of osteoclast genes TRAP and CatK on d 5 is reported from the combined results (Fold Induction) of 3 independent experiments ( n = 3). Statistical difference was determined by paired Student’s t test. Indicators of statistical difference Two-tailed p -value: NS = Not significant, * p ≤ 0.05, ** p ≤ 0.01, **** p ≤ 0.0001. ( e ) Representative 20× magnification images taken from d 5 TRAP-stained cells: untreated, RANKL-treated control and cells treated with RANKL in the presence of IAld (0.05 mM) or I3AA (0.2 mM). Osteoclast-like cells are indicated by a red arrow. All results depict consistent trends observed in multiple independent experiments.

Article Snippet: Recombinant human vascular endothelial growth factor peptide (VEGF 165 ) and murine soluble receptor activated NF-κB ligand (RANKL) was supplied by Peprotech (Rockhill, NJ, USA).

Techniques: Staining, Expressing, Control, Two Tailed Test

Journal: International Journal of Molecular Sciences

Article Title: Microbiota-Derived Metabolites, Indole-3-aldehyde and Indole-3-acetic Acid, Differentially Modulate Innate Cytokines and Stromal Remodeling Processes Associated with Autoimmune Arthritis

doi: 10.3390/ijms22042017

Figure Lengend Snippet: AhR antagonist CH-223191 inhibited the disruption of angiogenesis by I3AA but failed to alter the effect of IAld on angiogenesis and osteoclastogenesis. ( a , b ) HUVEC were seeded onto Matrigel with VEGF (25 ng/mL). After 1 h, cells were additionally treated with either IAld (0.5 mM) ( a ), I3AA (0.5 mM) ( b ), CH-223191 (3 μM) or a combination as indicated. The percentage of loops relative to VEGF-treated control cells (Veh) is reported ( n = 8). ( c , d ) RAW cells were treated RANKL alone or RANKL in addition with IAld (0.05 mM) ( c ), I3AA (d), CH-223191 (10 μM) or a combination as indicated. The percentage of osteoclast-like cells relative to RANKL-treated control cells (Veh) was reported ( n = 5). Statistical difference was determined on unnormalized data of either the number of loops formed ( a , b ) or from the number of osteoclast-like cells ( c , d ) by Ordinary one-way ANOVA with post-hoc test for multiple comparisons. Indicators of statistical different are between the respective controls (Veh) of the experimental type and the treatment group for which the mark is above, unless otherwise indicated by a bar. NS = not significant, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001. Data are from one experiment representative of several separate experiments.

Article Snippet: Recombinant human vascular endothelial growth factor peptide (VEGF 165 ) and murine soluble receptor activated NF-κB ligand (RANKL) was supplied by Peprotech (Rockhill, NJ, USA).

Techniques: Disruption, Control