Journal: Journal of nanobiotechnology

Article Title: Evaluating the pro-survival potential of apoptotic bodies derived from 2D- and 3D- cultured adipose stem cells in ischaemic flaps.

doi: 10.1186/s12951-024-02533-1

Figure Lengend Snippet: Fig. 6 3D-ABs suppress oxidative stress and cell death, and stimulated angiogenesis in ischaemic flaps. (A) F-CHP staining for damaged collagen detec tion in the skin on POD7. Scale bars, 100 μm. (B) The three groups’ quantified F-CHP intensity (n = 6). (C) CD31 and α-SMA IF staining in the flap on POD7. Scale bars, 50 μm. (D) Quantified CD31/α-SMA-positive blood vessel density among the three groups (n = 6). (E) Dead cells in flap tissue sections on POD7 under the detection of TUNEL staining. Scale bar, 20 μm. (F) TUNEL-positive cell percentage in the dermal layer, quantified for each of the three groups (n = 6). (G) DHE staining of the skin tissues on POD7 among the three groups. Scale bars, 50 μm. (H) Quantified DHE among the three groups (n = 6). SEM error bars are used. Significance (*): p value < 0.05; equal variances ANOVA with LSD post hoc analysis or unequal variances Dunnett’s T3 technique

Article Snippet: The primary antibodies utilized were specific to CD31 (1:200), CD68 (1:200), α-SMA (1:200; Proteintech, 67735-1-Ig), iNOS (1:200), and Arg1 (1:200).

Techniques: Staining, TUNEL Assay

Journal: eLife

Article Title: Nanchangmycin regulates FYN, PTK2, and MAPK1/3 to control the fibrotic activity of human hepatic stellate cells

doi: 10.7554/eLife.74513

Figure Lengend Snippet: ( A ) Overview of the small molecule screen. The number of candidate compounds (# of hits) is indicated at each step. The number in parenthesis represents the number of compounds tested in the final dose response curve (DRC) analysis due to availability. ( B ) Results of the primary lipid accumulation screen. Each dot indicates the mean scaled value of two replicates for each condition at 48 hr. Red dots represent negative control wells (DMSO), green dots represent positive control wells (nortriptyline, 27 µM), and blue dots represent experimental wells. ( C ) Results of the secondary mRNA screen. Each dot indicates the mean fold change of ACTA2 and COL1A1 after treatment with compounds (normalized to DMSO controls). PSMB2 was used as the reference gene (n=4). Green dots represent positive hits (<0.5), and blue dots represent non-hits (negative). ( D ) Dose-response curves were plotted for 39 compounds and were scored by three researchers independently. The mean score for each compound was plotted. The dotted line indicates the score of the positive control nortriptyline. Green dots represent positive hits, and blue dots represent non-hits (negative). This figure has two supplements.

Article Snippet: Mouse Acta2 , Mm00725412_s1.

Techniques: Negative Control, Positive Control

Journal: eLife

Article Title: Nanchangmycin regulates FYN, PTK2, and MAPK1/3 to control the fibrotic activity of human hepatic stellate cells

doi: 10.7554/eLife.74513

Figure Lengend Snippet: ( A-B ) Effect of 48 hr NCMC treatment on ACTA2 and COL1A1 in HSCs from human donors 1 ( A ) and 3 ( B ). Error bars represent mean ± SEM (n=3). Data are representative of three independent experiments. ns indicates not significant, *** indicates p<0.001, and **** indicates p<0.0001 (one-way ANOVA test). ( C ) Effect of 48 hr NCMC treatment on Acta2 and Col1a1 in primary mouse HSCs. Error bars represent mean ± SEM (n=4). Data are representative of three independent experiments. **** indicates p<0.0001 (one-way ANOVA test). ( D–E ) Effect of 48 hr NCMC treatment (1 µM) on collagen deposition in ECM. ( D ): representative images. Scale bar represents 100 µm. Collagen protein is indicated in green and nuclei for the same field are indicated in blue. ( E ): quantified results. Error bars represent mean ± SEM (n=4). Data are representative of three independent experiments. **** indicates p<0.0001 (Student’s t-test). ( F ) qPCR analysis of COL1A1 in HSC-hepatocyte spheroids treated with NCMC with and without TGF-β (Tβ). Error bars represent mean ± SEM (n=3). One experiment was performed independently for each donor shown. * indicates p<0.05 (Student’s t-test) and ** indicates p<0.01 (Student’s t-test). Analysis was performed on day 3 (3D). ( G–H ) RNA sequencing analysis of HSCs (donor 1) treated with DMSO or 1 µM NCMC for 48 hr. ( G ) Representative gene sets from the gene set enrichment analysis (GSEA). NES refers to normalized enrichment score. Nom P refers to Nominal P value. Vertical black lines refer to affected genes in the indicated signatures. Red color indicates positive correlation, and blue color indicates negative correlation. ( H ): Heatmap showing RNA-seq expression for the canonical HSC gene signature . All genes from the signature that are expressed in HSCs (with a minimum of five reads) were shown regardless of their expression patterns. Z-score values are also provided in . This figure has three supplements.

Article Snippet: Mouse Acta2 , Mm00725412_s1.

Techniques: RNA Sequencing, Expressing

Journal: eLife

Article Title: Nanchangmycin regulates FYN, PTK2, and MAPK1/3 to control the fibrotic activity of human hepatic stellate cells

doi: 10.7554/eLife.74513

Figure Lengend Snippet:

Article Snippet: Mouse Acta2 , Mm00725412_s1.

Techniques: Dominant Negative Mutation, Mutagenesis, Amplification, Plasmid Preparation, Isolation, Recombinant, Generated, Expressing, Sequencing, Cloning, Control, Real-time Polymerase Chain Reaction, Software

Journal: eLife

Article Title: Nanchangmycin regulates FYN, PTK2, and MAPK1/3 to control the fibrotic activity of human hepatic stellate cells

doi: 10.7554/eLife.74513

Figure Lengend Snippet:

Article Snippet: Mouse Acta2 , Mm00725412_s1.

Techniques:

Journal: Cells

Article Title: Stage-Dependent Fibrotic Gene Profiling of WISP1-Mediated Fibrogenesis in Human Fibroblasts

doi: 10.3390/cells13232005

Figure Lengend Snippet: TGFβ induces WISP1 expression in primary lung and dermal fibroblasts. Stimulation with TGFβ (1 ng/mL) for 24 h significantly increases COL1A1, COL3A1, IL6, ACTA2, FN1, and WISP1 mRNA as compared to control in ( A ) NHLF, ( B ) DHLF, and ( C ) NHDF for 3 respective donors ( n = 3/each) analyzed by RT-qPCR. Notably, TGFβ decreased CCL2 in NHLF and DHLF, except in NHDF. Each experiment was independently performed in triplicates with three biological replicates ( n = 3) for each donor and represented as mean ± SD. Statistical analysis was performed using one-way ANOVA with Tukey’s post hoc analysis vs. vehicle control for respective donors. * denotes p < 0.5, ** denotes p < 0.01, *** denotes p < 0.001, **** denotes p < 0.0001 versus the unstimulated control, as shown.

Article Snippet: The following commercially available TaqMan probes and primers were utilized: human Col1A1 (Hs00164004_m1), Col3A1 (Hs00943809_m1), CCL2 (Hs00234140_m1), IL6 (Hs00985639_m1), ACTA2 (Hs05005341_m1), FN1 (Hs01549976_m1), CCN4/WISP1 (Hs00987448_m1), and 18S (4310893E) from Applied Biosystems. cDNAs (1:20 dilution ratio) were added along with TaqMan Fast-advanced Mastermix in a total 10 μL reaction volume in a 384-well plate as per the manufacturer’s instructions and the plate was read using Quant Studio 7 Flex (Applied Biosystems, Waltham, MA, USA).

Techniques: Expressing, Control, Quantitative RT-PCR

Journal: Cells

Article Title: Stage-Dependent Fibrotic Gene Profiling of WISP1-Mediated Fibrogenesis in Human Fibroblasts

doi: 10.3390/cells13232005

Figure Lengend Snippet: WISP1 increases CCL2 and IL6 in primary fibroblasts. Stimulation with WISP1 (1000 nM) for 24 h significantly increases CCL2 and IL6 gene expression in ( A ) NHLF, ( B ) DHLF, and ( C ) NHDF as compared to control. TGFβ (1 ng/mL) for 24 h served as a positive control and significantly increases COL1A1, COL3A1, IL6, ACTA2, FN1, and WISP1 versus vehicle control. ( D ) Stimulation with increasing WISP1 concentrations for 24 h initiates concentration-dependent increase in CCL2 and IL6 protein levels, detected via ELISA in the cell-supernatant of NHDF. Each experiment performed in duplicates with three biological replicates ( n = 3) and represented as mean ± SD. Statistical analysis was performed using one-way ANOVA with Tukey’s post hoc analysis vs. vehicle control for respective donors. * denotes p < 0.5, ** denotes p < 0.01, *** denotes p < 0.001, **** denotes p < 0.0001 versus the vehicle control, as shown.

Article Snippet: The following commercially available TaqMan probes and primers were utilized: human Col1A1 (Hs00164004_m1), Col3A1 (Hs00943809_m1), CCL2 (Hs00234140_m1), IL6 (Hs00985639_m1), ACTA2 (Hs05005341_m1), FN1 (Hs01549976_m1), CCN4/WISP1 (Hs00987448_m1), and 18S (4310893E) from Applied Biosystems. cDNAs (1:20 dilution ratio) were added along with TaqMan Fast-advanced Mastermix in a total 10 μL reaction volume in a 384-well plate as per the manufacturer’s instructions and the plate was read using Quant Studio 7 Flex (Applied Biosystems, Waltham, MA, USA).

Techniques: Gene Expression, Control, Positive Control, Concentration Assay, Enzyme-linked Immunosorbent Assay