Journal: Annals of Translational Medicine

Article Title: MMP2/9 downregulation is responsible for hepatic function recovery in cirrhotic rats following associating liver partition and portal vein ligation for staged hepatectomy

doi: 10.21037/atm-22-1312

Figure Lengend Snippet: Detection of liver hypertrophy in rats after the operation. ELISA assay showing the changes in expression of HGF (A), TGF-β (B), TNF-α (C), and IL-6 (D) in rats. (E) The PCNA levels in rat livers were assessed by immunohistochemistry (×10 magnification, ×10: 1 cm: 40 µm). (F) Hematoxylin and eosin staining of rat liver tissues (×10 magnification, ×10: 1 cm: 40 µm). All quantitative variables are presented as mean ± standard deviation and compared using two-way ANOVA. P<0.05. #, A or P groups vs. S groups, P<0.05, marked as #NA, #NP, #CP, #CA; Δ, CA group vs. NA group, P<0.05; *, CP groups vs. CA groups. ELISA, enzyme-linked immunosorbent assay; HGF, hepatocyte growth factor; TGF, transforming growth factor; TNF, tumor necrosis factor; IL, interleukin; PCNA, proliferating cell nuclear antigen; ANOVA, analysis of variance; PVL, portal vein ligation; ALPPS, associating liver partition and portal vein ligation for staged hepatectomy; A, ALPPS; P, PVL; S, sham operated; CA, cirrhotic rats that underwent ALPPS; NA, normal rats that underwent ALPPS; CP, cirrhotic rats that underwent PVL.

Article Snippet: Enzyme-linked immunosorbent assay (ELISA) The serum levels of recombinant human hepatocyte growth factor (HGF), transforming growth factor-β (TGF-β), tumor necrosis factor (TNF)-α, and interleukin (IL)-6 in rats were detected by ELISA kits (Elabscience Biotechnology Company, Co., Ltd., China).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Immunohistochemistry, Staining, Standard Deviation, Ligation

Journal: Liver Transplantation

Article Title: Adverse effects of graft congestion and ameliorative effects of hepatocyte growth factor after liver transplantation in rats

doi: 10.1097/LVT.0000000000000449

Figure Lengend Snippet: Experiment summary. (A) Experiment 1 included the liver resection model. The 30% (0%), 30% (7%), and 40% (10%) models were created by liver resection and vein ligation. Gray color within the liver represents the congested area. (B) Experiment 2 included the liver transplantation model. The livers from the 30% (0%), 30% (7%), 40% (10%), 50% (7%), and 60% (10%) models were used as grafts for experiment 2. These grafts were chilled at 4°C for 1 hour in HTK solution and transplanted into syngeneic rats. (C) Experiment 3 included the liver transplantation plus HGF administration model. Rats in the 50% (7%) liver transplantation model received 100 µg/kg recombinant human HGF administered through the penile vein under anesthesia immediately after surgery and on POD1 and POD2. Abbreviations: HTK, histidine-tryptophan-ketoglutarate; i.v., intravenous injection; LHV, left hepatic vein; LML, left median lobe; POD, postoperative day; RHV, right hepatic vein; RML, right median lobe.

Article Snippet: The 50% (7%) model rats received recombinant human HGF (50 μg/mL; catalog number 294-HGN; R&D Systems) administered at 100 μg/kg through the penile vein under anesthesia immediately after LT and on PODs 1 and 2 (Figure C).

Techniques: Ligation, Transplantation Assay, Recombinant, Injection

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Killing efficiency of different physicochemical treatments on recombinant L. lactis strains displaying different binders of proinflammatory cytokines on their surface. The killing efficiency was determined by culturing and counting colonies on agar plates. Complete killing of bacteria i.e., 0 CFU/mL (no bacterial colonies on agar plates) was considered as 100% efficiency. pSD-ZIL6, L. lactis displaying anti-IL6 affibody; pSD-ZTNF, L. lactis displaying anti-TNF affibody; pSD-Fyn17, L. lactis displaying anti-IL17 fynomer; pSD-EVA, L. lactis displaying anti-IL8 evasin; pNZ8148, L. lactis harboring empty plasmid. The results presented are means ± standard deviation (SD) of three technical replicates from a representative experiment.

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Recombinant, Bacteria, Plasmid Preparation, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Cytokine binding ability of recombinant L. lactis bacteria displaying different binders of proinflammatory cytokines on their surface upon exposure to bacteria-killing treatments. ELISA-determined concentrations of recombinant IL6 (A) , TNF (B) , IL17 (C) , and IL8 (D) that remained in the solution following incubation with the corresponding strain of bacteria before and after treatment with heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, UV, and gamma irradiation. pSD-ZIL6, L. lactis displaying ZIL6 affibody; pSD-ZTNF, L. lactis displaying anti-TNF affibody; pSD-Fyn17, L. lactis displaying anti-IL17 fynomer; pSD-EVA, L. lactis displaying anti-IL8 evasin; Ctrl: L. lactis control cells containing empty plasmid pNZ8148. The results are presented as means ± SD of three technical replicates of a representative experiment. ns, p = 0.09; *, p ≤ 0.05; **, p = 0.002; ***, p < 0.001 (unpaired two-tailed t -test).

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Binding Assay, Recombinant, Bacteria, Enzyme-linked Immunosorbent Assay, Incubation, Sonication, Irradiation, Control, Plasmid Preparation, Two Tailed Test

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Determination of binding affinity of ZIL6-displaying L. lactis for human IL6. A constant number of 8 × 10 6 CFU bacteria was incubated with increasing concentrations of human IL6 for 2 h. MFI values were measured by flow cytometry and binding curves were fitted to the data using the Hill equation in the GraphPad Prism v.10.3.1. The dissociation constants (Kd) values were calculated from the binding curves for non-treated, live ZIL6-displaying L. lactis bacteria and bacteria treated by heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, and UV irradiation. Data are means ± SD of three technical replicates from a representative experiment.

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Binding Assay, Bacteria, Incubation, Flow Cytometry, Sonication, Irradiation

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: Quantification of the maximum binding capacity of ZIL6-displaying L. lactis . Increasing concentrations of recombinant human IL6 (0.45, 4.5, 45 and 450 ng) were incubated with a constant number of bacteria (4.5 × 10 7 CFU equivalent to 0.1 mg dry cell weight) in 450 μl. Residual IL6 in the supernatants was quantified by ELISA for live non-treated bacteria and the cells treated by heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, and UV irradiation. The percentage of bound IL6 was calculated from the difference measured in the presence of ZIL6-displaying L. lactis and control bacteria (carrying empty plasmid pNZ8148). The data are means ± SD of two biological replicates. The asterisks denote statistically significant differences between treated and non-treated bacteria for each concentration. ***, p < 0.001 (one-way ANOVA with Dunnett multiple comparison test).

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Binding Assay, Recombinant, Incubation, Bacteria, Enzyme-linked Immunosorbent Assay, Sonication, Irradiation, Control, Plasmid Preparation, Concentration Assay, Comparison

Journal: Frontiers in Microbiology

Article Title: Postbiotics derived from recombinant lactic acid bacteria exhibit high IL6-binding capacity and suppress IL6-induced STAT3 signaling

doi: 10.3389/fmicb.2025.1657810

Figure Lengend Snippet: (A) Inhibition of STAT3 signaling in HEKblue-IL6R cells by non-viable ZIL6-displaying L. lactis bacteria in comparison to live non-treated strain. ZIL6-displaying L. lactis cells (1 × 10 8 CFU/mL or 1 × 10 9 CFU/mL) were preincubated with IL6 (1 ng/mL) for 2 h. After removal of bacterial cells, the cell-free supernatant containing remainder of IL6 was added to HEKblue-IL6R cells to induce the reporter system. The percentage of STAT3 inhibition was calculated relative to the STAT3 signaling induced by IL6 in the absence of bacteria. The inhibition of STAT3 signaling was determined for live non-treated bacteria and the cells treated by heat (70 °C, 40 min or 100 °C, 30 min), sonication, ethanol, UV, and gamma irradiation. pSD-ZIL6, L. lactis displaying ZIL6 affibody; Ctrl: L. lactis control cells containing empty plasmid pNZ8148. An anti-IL6 monoclonal antibody (Ab) was used as a positive control. The data are means ± SD of four biological replicates. The asterisks denote statistically significant differences between treated and non-treated bacteria for each concentration. ***, p < 0.001 (one-way ANOVA with Dunnett multiple comparison test). (B) The effect of lactic acid production by L. lactis on viability of HEKblue-IL6R cells. HEKblue-IL6R cells (100 000 cells/well) were incubated with ZIL6-displaying L. lactis (2 × 10 7 bacteria/well) for 6, 12, and 24 h, and the viability of HEKblue-IL6R cells was determined with trypan blue. The data are means ± SD of three technical replicates of a representative experiment.

Article Snippet: Briefly, serial dilutions of IL6 (0.32, 0.65, 1.31, 2.62, 5.25, 10.5, 21, 42 nM; HZ-1019, Proteintech) were incubated with a fixed number of recombinant bacteria (8 × 10 6 CFU) for 2 h in a final volume of 200 μl.

Techniques: Inhibition, Bacteria, Comparison, Sonication, Irradiation, Control, Plasmid Preparation, Positive Control, Concentration Assay, Incubation

Journal: Frontiers in Veterinary Science

Article Title: Interleukin-1β in tendon injury enhances reparative gene and protein expression in mesenchymal stem cells

doi: 10.3389/fvets.2022.963759

Figure Lengend Snippet: IL-1β and IL-6 are the predominant inflammatory cytokines present following surgically induced equine SDFT injury. Descriptive report of (A) inflammatory cytokines and (B) growth factors FGF-2 and TGF-β1,−2, and−3 in tendon ultrafiltrate were measured by ELISA following bilateral surgically induced SDFT injury of the forelimb over 21 days in 6 horses. Due to insufficient volume, samples from days 8 onward were grouped for TGF-β isoform analysis. When present, the dashed line at the y-axis represents the lower limit of quantification (LLoQ) for that cytokine or growth factor as determined by the assay manufacturer (MilliporeSigma, MA, USA). Concentrations measured below the LLoQ were assigned a value of 0.1 of the LLoQ.

Article Snippet: Protein expression of IL-1β licensed MSCs were evaluated with ELISA assays for equine IL-6 (R&D Systems, Minneapolis, MN, USA), human PGE2 (Enzo Biochem, Inc., Farmingdale, NY, USA), and equine VEGF (Kingfisher Biotech, Saint Paul, MN, USA) and performed per manufacturer's instructions.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Veterinary Science

Article Title: Interleukin-1β in tendon injury enhances reparative gene and protein expression in mesenchymal stem cells

doi: 10.3389/fvets.2022.963759

Figure Lengend Snippet: IL-1β licensed equine MSCs produce significantly more IL-6, VEGF, and PGE2 than naïve MSCs. From RNA sequencing data, three proteins known to have tendon-relevant effects were selected to confirm downstream protein expression. Analysis of MSC cell culture supernatants by ELISA assay confirmed increased protein expression of IL-6, VEGF, and PGE2 following IL-1β licensing compared to naïve MSC controls. A one-tailed paired t -test was used to examine difference in protein expression between groups.

Article Snippet: Protein expression of IL-1β licensed MSCs were evaluated with ELISA assays for equine IL-6 (R&D Systems, Minneapolis, MN, USA), human PGE2 (Enzo Biochem, Inc., Farmingdale, NY, USA), and equine VEGF (Kingfisher Biotech, Saint Paul, MN, USA) and performed per manufacturer's instructions.

Techniques: RNA Sequencing, Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay, One-tailed Test

Journal: Biochimica et biophysica acta

Article Title: PAX2 promoted prostate cancer cell invasion through transcriptional regulation of HGF in an in vitro model.

doi: 10.1016/j.bbadis.2015.08.008

Figure Lengend Snippet: Fig. 3. PAX2 interacted upstream of the HGF gene. (A) DNA sequence alignment of the upstream region of the HGF genes (upper sequences). Gray boxes are consensus sequences of PAX2 and/or PAX6 binding sites. A schematic alignment of the oligonucleotides used in DNA pull-down assay (bottom sequences). Six nucleotides in the PAX2 and/or PAX6 binding site were replaced with other nucleotides (red characters in mt sequences). (B) DNA pull-down assay using recombinant PAX2 protein and oligonucleotides derived from upstream of the HGF gene (shown in Fig. 4A). (C) Luciferase assay using PC3 cells cotransfected with HGF-luc reporter plasmids (wt or mt, −904–+81 of HGF gene) and FLAG-PAX2 expression plasmids. The same results were reproduced for each experiments three times.

Article Snippet: Cells were incubated with or without 100 mM recombinant HGF (TP315593, OriGene, Rockville, MD, USA) at 37 °C in a humidified 5% CO2 atmosphere for 24 h. Cells that had invaded through the inserts were stained using a Diff-Quik staining kit (Sysmex, Hyogo, Japan), and the total number of stained cells was counted.

Techniques: Sequencing, Binding Assay, Pull Down Assay, Recombinant, Derivative Assay, Luciferase, Expressing

Journal: Biochimica et biophysica acta

Article Title: PAX2 promoted prostate cancer cell invasion through transcriptional regulation of HGF in an in vitro model.

doi: 10.1016/j.bbadis.2015.08.008

Figure Lengend Snippet: Fig. 5. PAX2 enhanced invasive activity of androgen-independent prostate cancer cells upstream of the HGF pathway. (A) Invasion assay using 22Rv1 cells. 22Rv1 cells were transfected with siRNAs for HGF or PAX2 for 2 days. Then, these cells were cultured in Matrigel invasion chambers with or without 100 pM recombinant HGF for 24 h. (B) The number of invading cells was counted. Invasion index shows the average values of 3 independent measurements, which was normalized to cell number of control inserts. *P b 0.05. The same results were reproduced in independent three experiments.

Article Snippet: Cells were incubated with or without 100 mM recombinant HGF (TP315593, OriGene, Rockville, MD, USA) at 37 °C in a humidified 5% CO2 atmosphere for 24 h. Cells that had invaded through the inserts were stained using a Diff-Quik staining kit (Sysmex, Hyogo, Japan), and the total number of stained cells was counted.

Techniques: Activity Assay, Invasion Assay, Transfection, Cell Culture, Recombinant, Control