Journal: Hepatology Communications

Article Title: Arrb2 in hepatocytes promotes M2 macrophage polarization, ameliorates hepatic ischemia–reperfusion injury through upregulating metabolite 6-ketoLCA

doi: 10.1097/HC9.0000000000000916

Figure Lengend Snippet: 6-ketoLCA upregulated by Arrb2 in hepatocytes promotes M2 macrophage polarization, alleviating hepatic IRI. (A–E) Liver and serum samples from liver-specific Arrb2-CKO and WT mice with or without 6-ketoLCA treatment after I/R (n=6). (A) HE and TUNEL staining were analyzed microscopically. Scale bar: 200 μm; Suzuki score and necrosis area were examined. (B) Quantitative analysis of the bile acid metabolite 6-ketoLCA in liver tissue. (C) Liver function levels (ALT, AST, TBA, GGT) were analyzed between groups. (D) qRT-PCR analysis of mouse liver tissue mRNA expression for inflammatory factors (IL-6, TNF-α, IL-10, TGF-β). (E) iNOS, CD86, CD206, and CD163 expression in PMMs was assessed by WB analysis. (F) The CCK8 assay measured the viability of RAW264.7 cells exposed to varying concentrations and durations of 6-ketoLCA (n=3). (G, I) Co-culture of RAW264.7 and AML12 cells. (G) ELISA for culture medium supernatant inflammatory factors (IL-6, TNF-α, IL-10, TGF-β). (I) BCL2, BAX, and GAPDH expression in AML12 cells was assessed by WB analysis. (H, J) Flow cytometry detected the proportions of M1 (CD86, PE) and M2 (CD206, APC) macrophages treated with IL-4 and 6-ketoLCA. The ratio of M1/M2 macrophages was measured. All results are presented as mean ± SD, and statistical significance was assessed. * p <0.05, ** p <0.01, and *** p <0.001. Abbreviations: ALT, alanine transaminase; AST, aspartate transaminase; CKO, conditional knockout; GGT, gamma-glutamyl transferase; HE, hematoxylin and eosin; I/R, ischemia/reperfusion; IL-6, interleukin 6; IL-10, interleukin 10; IRI, ischemia–reperfusion injury; qRT-PCR, quantitative reverse transcription polymerase chain reaction; PMMs, primary mouse macrophages; TBA, total bile acids; TGF-β, transforming growth factor-β; TNF-α, tumor necrosis factor-α; TUNEL, terminal deoxynucleotidyl transferase–mediated dUTP nick-end labeling; WB, western blotting; WT, wild type.

Article Snippet: RAW264.7 were cultured in 10% FBS RPMI-1640 medium with 6-ketoLCA (100 ng/mL) and IL-4 (20 ng/mL) to induce polarization to M2-like macrophage. shRNA targeting TGR5 (GeneCopoeia, Rockville, MD, USA) was transfected into RAW264.7 cells utilizing Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA) in accordance with the manufacturer’s protocol to achieve TGR5 knockdown.

Techniques: TUNEL Assay, Staining, Quantitative RT-PCR, Expressing, CCK-8 Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Knock-Out, Reverse Transcription, Polymerase Chain Reaction, End Labeling, Western Blot

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: Representative fluorescence microscopy images of mesenchymal stromal cells (MSCs) infected by IL-4 and/or PDGF-BB lentiviral vectors. GFP-MSCs, MSCs infected with control GFP-positive lentivirus vector; IL4-MSCs, MSCs infected with rIL-4 secreting GFP-positive lentivirus vector; PDGF-BB-MSCs, MSCs infected with hPDGF-BB secreting GFP-positive lentivirus vector; IL4-PDGF-BB-MSCs, previously established IL4-MSCs infected with hPDGF-BB secreting RFP-positive lentivirus vector.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques: Fluorescence, Microscopy, Infection, Control, Plasmid Preparation

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: The experimental outline of in vitro experiments. GFP-MSCs, IL4-MSCs, PDGF-BB-MSCs, or IL4-PDGF-BB-MSCs were seeded into T75 flasks and cultured in an MSC growth medium for 1 day. For the non-preconditioning MSC groups, the cells were cultured for 3 days with a fresh MSC growth medium. For the preconditioning MSC groups, cells were cultured in the MSC preconditioning medium for 3 days. The cells from all eight groups were washed three times with Dulbecco’s phosphate-buffered saline. The cells were trypsinized and seeded for each experiment, including cell proliferation, IL4 and PDGF-BB expression level, and osteogenic differentiation performed by alkaline phosphatase (ALP) staining and Alizarin red-staining. Abbreviations: MSCs, mesenchymal stromal cells; pMSCs, preconditioning MSCs; TNFα, tumor necrosis factor-alpha; LPS, lipopolysaccharide.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques: In Vitro, Cell Culture, Saline, Expressing, Staining

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: Fold change in the amount of dsDNA compared to day 1.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques:

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: ( A ) IL4 and PDGF-BB expression levels on day 3 in the IL4-MSC, IL4-pMSC, PDGF-BB-MSC, PDGF-BB-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups. IL4 expression levels on day 3 in the IL4-MSC, IL4-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups were 77.7 ± 4.4 ng/mL, 34.6 ± 5.7 ng/mL, 16.8 ± 9.4 ng/mL, and 41.8 ± 5.0 ng/mL, respectively. PDGF-BB expression levels on day 3 in the PDGF-BB-MSC, PDGF-BB-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups were 121.7 ± 20.1 pg/mL, 50.3 ± 20.4 pg/mL, 8.7 ± 2.0 pg/mL, 71.5 ± 7.8 pg/mL, respectively. ( B ) IL4 and PDGF-BB expression level-dsDNA ratios on day 3 in the IL4-MSC, IL4-pMSC, PDGF-BB-MSC, PDGF-BB-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups. IL4 expression level-dsDNA ratios on day 3 in the IL4-MSC, IL4-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups were 7.3 ± 0.6, 1.8 ± 0.2, 1.8 ± 1.0, and 2.3 ± 0.1, respectively. PDGF-BB expression level-dsDNA ratios on day 3 in the PDGF-BB-MSC, PDGF-BB-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups were 5.6 ± 0.9, 3.7 ± 1.4, 1.0 ± 0.3, and 3.9 ± 0.1, respectively.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques: Expressing

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: Alkaline phosphatase-staining in all eight groups 2 weeks after seeding. The percentages of stained area in the GFP-MSC, GFP-pMSC, IL4-MSC, IL4-pMSC, PDGF-BB-MSC, PDGF-BB-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups were 19.8 ± 5.3%, 28.3 ± 1.4%, 4.3 ± 0.7%, 6.0 ± 2.4%, 44.2 ± 11.7%, 42.5 ± 8.1%, 21.4 ± 1.5%, and 25.7 ± 1.0%, respectively. Abbreviations: ALP, alkaline phosphatase.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques: Staining

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: Alizarin red-staining in all eight groups 4 weeks after seeding. The percentages of stained area in the GFP-MSC, GFP-pMSC, IL4-MSC, IL4-pMSC, PDGF-BB-MSC, PDGF-BB-pMSC, IL4-PDGF-BB-MSC, and IL4-PDGF-BB-pMSC groups were 33.1 ± 4.7%, 45.1 ± 4.9%, 0.64 ± 0.08%, 0.58 ± 0.03%, 51.7 ± 14.5%, 40.7 ± 6.7%, 17.3 ± 2.5%, and 41.2 ± 1.7%, respectively.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques: Staining

Journal: Bioengineering

Article Title: Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

doi: 10.3390/bioengineering8110165

Figure Lengend Snippet: The summary of the results.

Article Snippet: The expression levels of IL4 and PDGF-BB on day 3 were measured using the rabbit IL4 ELISA Kit (R&D Systems, Minneapolis, MN, USA) and the human PDGF-BB ELISA kit (R&D Systems, Minneapolis), respectively, according to the manufacturer’s protocols.

Techniques: Expressing

Journal: Inflammation and Regeneration

Article Title: Novel artificial nerve transplantation of human iPSC-derived neurite bundles enhanced nerve regeneration after peripheral nerve injury

doi: 10.1186/s41232-024-00319-4

Figure Lengend Snippet: Early-stage tissue evaluation after neurite bundle-derived artificial nerve transplantation. a , b Immunohistochemistry images with anti-NFH and anti-CD31 antibodies. Dotted arrows indicate the proximal side of the intact nerve areas, and solid arrows indicate the NFH- and CD31-positive regeneration tip in the reconstructed tissue. Scale bars = 1 mm. c , d Quantitative comparison of axonal extension distance was indicated by NFH-positive areas and that of vascular elongation distance was demonstrated by CD31-positive areas at 1 and 2 weeks after transplantation. Significant differences between groups elongation distances were observed in 2-week samples only. e , f Iba1 immunohistochemistry image of the proximal part of the regenerated tissue at 2 weeks after transplantation (e). Scale bars = 1 mm. Quantitative analysis of Iba1-positive areas demonstrated that a larger number of macrophages aggregated in the Auto, motor TP and sensory TP (f). ( n = 3). * p < 0.05, ** p < 0.01, N.S. = not significant. Data are represented as the mean ± SEM

Article Snippet: The primary antibodies used in this study were human polyclonal anti-pan-ELAVL (ELAV-like protein 2/3/4) antibody (1:2000, kindly provided from Prof. Robert Darnell, Rockefeller University), rabbit polyclonal anti-TrkA antibody (1:500, kindly provided from Prof. Louis F. Reichardt, UCSF), chicken polyclonal anti-TrkB antibody (1:500, kindly provided from Prof. Louis F. Reichardt, UCSF), goat polyclonal anti-TrkC antibody (AF373, 1:200, R&D systems), rabbit polyclonal anti-parvalbumin antibody (PV-28, 1:1000, Swant), rabbit polyclonal anti-CGRP antibody (BML-CA1134, 1:500, Enzo Life Sciences), Goat polyclonal anti-Choline acetyltransferase antibody (AB144P, 1:200, Chemicon), Mouse monoclonal anti-Islet1 and Islet2 antibody (39.4D5, 1:200, DSHB), rabbit polyclonal anti-Neurofilament heavy polypeptide antibody (ab8135, 1:500, Abcam), goat polyclonal anti-mouse and anti-rat CD31 (AF3628, 1:100, R&D), rabbit polyclonal anti-Iba1 antibody (GtX100042, 1:500, GeneTex), goat anti-GFP antibody (600–101-215, 1:1000, Rockland).

Techniques: Derivative Assay, Transplantation Assay, Immunohistochemistry, Comparison

Journal: Acta Medica Lituanica

Article Title: Regeneration of Rabbit Auricular Cartilage After the Intravenous Stem Cell Injection

doi: 10.15388/Amed.2023.30.2.15

Figure Lengend Snippet: The concentration of IL-6 (A) and TGF-β1 (B) in the blood plasma of rabbits after 1, 2 and 3 months after the artificial auricular cartilage damage and the injection of stem cells. Control – animals without stem cells injection, MSC – intravenous injection of 5•10 6 of homologous mesenchymal stem cells in saline (M±SD, *p<0.05).

Article Snippet: RabbitIL-6 (Interleukin 6) ELISAKit and RabbitTGF-β1 (Transforming Growth Factor Beta 1) ELISAKit (Elabscience Biotechnology Inc., USA) were used.

Techniques: Concentration Assay, Clinical Proteomics, Injection, Control, Saline

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Natterin bridges IFN-φ1 and non-canonical inflammasome pathways via CRFB1 /Gbp4 to license Caspy2-mediated antibacterial immunity

doi: 10.3389/fcimb.2025.1686758

Figure Lengend Snippet: IFN-φ neutralization abolishes ST-induced interferon production in zebrafish embryos. (A) Representative immunohistochemical staining of IFN-φ (brown DAB precipitate) in 1 dpf wild-type (WT) embryos under four conditions: (a.1) unstimulated control, (a.2) ST stimulation (10 6 cells/ml for 2h), (a.3) anti-IFN-β antibody pretreatment alone, and (a.4) anti-IFN-β pretreatment followed by ST stimulation. Scale bar: 50 μm. (B) Quantification of IFN-φ-positive cells from two slides per group (eight sections/slide, three embryos/section) using Fiji-ImageJ. Data represent mean ± SEM ( n = 3 independent experiments); * p < 0.05 versus unstimulated control WT, # p < 0.05 versus ST-stimulated WT.

Article Snippet: They were then incubated overnight at 4°C with the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IL-1β antibody that recognizes the rainbow trout mature IL-1β peptide with 20 kDa according to (#CLF016HP, Cedarlane, 1/3,000) or the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IFN-β antibody (#CLF005HP, Cedarlane, 1/3,000) according to .

Techniques: Neutralization, Immunohistochemical staining, Staining, Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Natterin bridges IFN-φ1 and non-canonical inflammasome pathways via CRFB1 /Gbp4 to license Caspy2-mediated antibacterial immunity

doi: 10.3389/fcimb.2025.1686758

Figure Lengend Snippet: IFN-I signaling is essential for IL-1β–mediated protection against ST stimulation. (A) Locomotor activity (total distance traveled) assessed 72h after stimulation at 28°C. Mortality count (C) and Caspy2 expression in WT embryos after ST stimulation (10 6 cells/ml, 2h) with or without anti-IFN-β antibody pretreatment. The corresponding bar graph (D) represents the protein expression levels as a percentage of the control. (E) Quantification and (F) representative IHC images of mature IL-1β (mIL-1β)–positive cells (brown DAB precipitate) in: (f.1) ST-stimulated embryos, (f.2) pretreatment with anti-IFN-β alone, and (f.3) pretreatment with anti-IFN-β followed by ST stimulation. Scale bar: 200 μm. Data represent mean ± SEM ( n = 3 independent experiments); * p < 0.05 versus unstimulated control WT, # p < 0.05 versus ST-stimulated WT.

Article Snippet: They were then incubated overnight at 4°C with the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IL-1β antibody that recognizes the rainbow trout mature IL-1β peptide with 20 kDa according to (#CLF016HP, Cedarlane, 1/3,000) or the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IFN-β antibody (#CLF005HP, Cedarlane, 1/3,000) according to .

Techniques: Activity Assay, Expressing, Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Natterin bridges IFN-φ1 and non-canonical inflammasome pathways via CRFB1 /Gbp4 to license Caspy2-mediated antibacterial immunity

doi: 10.3389/fcimb.2025.1686758

Figure Lengend Snippet: Effectiveness of CRISPR/Cas9 depletion of loc795232. Basal expression of loc795232 mRNA transcripts in unstimulated WT or KO embryos ( n = 100/group) at 1 dpf by RT-qPCR (A) or by in situ hybridization (B) . All qPCR data normalized to β-actin and expressed as fold change relative to 0h WT unstimulated control. Data represent mean ± SEM of three biological replicates. Independent groups of 1 dpf embryos previously treated or not with Pam3CSK4 or MCC950 and subjected to ST stimulation for 2h, as well as KO larvae stimulated with ST were processed for Natterin detection by WB (C) using anti-Natterin serum (62 kDa dimeric form) and anti-IgG TrueBlot HRP. The corresponding bar graph represents the protein expression levels as a percentage of the control (D) , and Ponceau S staining is visualized in the first horizontal line.

Article Snippet: They were then incubated overnight at 4°C with the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IL-1β antibody that recognizes the rainbow trout mature IL-1β peptide with 20 kDa according to (#CLF016HP, Cedarlane, 1/3,000) or the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IFN-β antibody (#CLF005HP, Cedarlane, 1/3,000) according to .

Techniques: CRISPR, Expressing, Quantitative RT-PCR, In Situ Hybridization, Control, Staining

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Natterin bridges IFN-φ1 and non-canonical inflammasome pathways via CRFB1 /Gbp4 to license Caspy2-mediated antibacterial immunity

doi: 10.3389/fcimb.2025.1686758

Figure Lengend Snippet: Natterin is required for Gbp4 induction. Constitutive expression of gbp4 (A) and gbp1 (C) , was analyzed in unstimulated WT embryos ( n = 100/group) at 24h intervals by RT-qPCR. 1 dpf ST-responsive expression of gbp4 (B) and gbp1 (D) was assessed in WT and KO groups. All qPCR data normalized to β-actin and expressed as fold change relative to 0h WT unstimulated control. * p < 0.05 versus unstimulated control WT; # p < 0.05 versus ST-stimulated WT.

Article Snippet: They were then incubated overnight at 4°C with the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IL-1β antibody that recognizes the rainbow trout mature IL-1β peptide with 20 kDa according to (#CLF016HP, Cedarlane, 1/3,000) or the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IFN-β antibody (#CLF005HP, Cedarlane, 1/3,000) according to .

Techniques: Expressing, Quantitative RT-PCR, Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Natterin bridges IFN-φ1 and non-canonical inflammasome pathways via CRFB1 /Gbp4 to license Caspy2-mediated antibacterial immunity

doi: 10.3389/fcimb.2025.1686758

Figure Lengend Snippet: Natterin is essential for proteolytic activation of Caspy and Caspy2 during ST stimulation. (A) Developmental expression profile of caspy2 mRNA in unstimulated WT embryos ( n = 100/group) from 24 to 120 hpf. (B) One day post-fertilization ST-induced caspy2 expression in WT versus natterin knockout (KO) embryos 2h post-stimulation. All qPCR data normalized to β-actin and expressed as fold change relative to 0h WT unstimulated control. Data represent mean ± SEM; * p < 0.05 versus unstimulated control WT, # p < 0.05 versus ST-stimulated WT. Western blot analysis of mature Caspy ( Supplementary Figure S3 ) and Caspy2 ( Supplementary Figure S4 ) expression in whole embryo lysates at indicated times post-ST stimulation (15 and 30 min, 1 and 2h). Data are normalized to the unstimulated control and presented as percentage values. Quantitative densitometry is shown in bar graphs (C, D) .

Article Snippet: They were then incubated overnight at 4°C with the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IL-1β antibody that recognizes the rainbow trout mature IL-1β peptide with 20 kDa according to (#CLF016HP, Cedarlane, 1/3,000) or the polyclonal rabbit HRPO-labeled IgG anti-rainbow trout IFN-β antibody (#CLF005HP, Cedarlane, 1/3,000) according to .

Techniques: Activation Assay, Expressing, Knock-Out, Control, Western Blot