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Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor <t>(VEGF)</t> (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).
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Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor <t>(VEGF)</t> (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).
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Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor <t>(VEGF)</t> (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).
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Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor <t>(VEGF)</t> (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).
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Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor <t>(VEGF)</t> (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).
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Figure 2. The effect of anti-Ts7TMR scFv on A549 lung cancer and its preliminary mechanism. A. The effect of anti-Ts7TMR scFv on the proliferation of A549 cells in vitro. A549 cells were co-incubated with different doses of anti-Ts7TMR scFv (5, 25, and 45 μg) for 48h to detect cell proliferation by CCK-8 assay. B. The effect of anti-Ts7TMR scFv on A549 lung cancer growth in nude mice. Nude mice were injected subcutaneously with A549 cells and then treated subcutaneously with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg) around the tumour for 10 consecutive days. C. Scoring of PCNA, BCL-2, and <t>VEGF</t> expression in tumour tissues by immunohistochemistry (400 ×). Expression of PCNA, BCL-2, and VEGF in tumours treated with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg). Scale bar = 20 μm. *p < 0.05, **p < 0.01.
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Figure 2. The effect of anti-Ts7TMR scFv on A549 lung cancer and its preliminary mechanism. A. The effect of anti-Ts7TMR scFv on the proliferation of A549 cells in vitro. A549 cells were co-incubated with different doses of anti-Ts7TMR scFv (5, 25, and 45 μg) for 48h to detect cell proliferation by CCK-8 assay. B. The effect of anti-Ts7TMR scFv on A549 lung cancer growth in nude mice. Nude mice were injected subcutaneously with A549 cells and then treated subcutaneously with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg) around the tumour for 10 consecutive days. C. Scoring of PCNA, BCL-2, and <t>VEGF</t> expression in tumour tissues by immunohistochemistry (400 ×). Expression of PCNA, BCL-2, and VEGF in tumours treated with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg). Scale bar = 20 μm. *p < 0.05, **p < 0.01.
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Figure 2. The effect of anti-Ts7TMR scFv on A549 lung cancer and its preliminary mechanism. A. The effect of anti-Ts7TMR scFv on the proliferation of A549 cells in vitro. A549 cells were co-incubated with different doses of anti-Ts7TMR scFv (5, 25, and 45 μg) for 48h to detect cell proliferation by CCK-8 assay. B. The effect of anti-Ts7TMR scFv on A549 lung cancer growth in nude mice. Nude mice were injected subcutaneously with A549 cells and then treated subcutaneously with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg) around the tumour for 10 consecutive days. C. Scoring of PCNA, BCL-2, and <t>VEGF</t> expression in tumour tissues by immunohistochemistry (400 ×). Expression of PCNA, BCL-2, and VEGF in tumours treated with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg). Scale bar = 20 μm. *p < 0.05, **p < 0.01.
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Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor (VEGF) (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).

Journal: International journal of molecular sciences

Article Title: CD40 Ligand-CD40 Interaction Is an Intermediary between Inflammation and Angiogenesis in Proliferative Diabetic Retinopathy.

doi: 10.3390/ijms242115582

Figure Lengend Snippet: Figure 8. Soluble(s) CD40L induces blood-retinal barrier (BRB) breakdown. sCD40L was injected intravitreally at the dose of 5 ng in 5 µL in one eye and the same volume of phosphate-buffered saline (PBS) was injected in the contralateral eye of the normal rats. The BRB breakdown was quantified with the FITC-conjugated dextran technique. Results are expressed as median (interquartile range) of 8 rats (A). Western blot analysis of rat retinas. Intravitreal administration of sCD40L (5 ng in 5 µL) induced a significant upregulation of the expression of phospho-ERK1/2 (B), vascular endothelial growth factor (VEGF) (C), intercellular adhesion molecule-1 (ICAM-1) (D) and vascular cell adhesion molecule- 1 (VCAM-1) (E) compared to intravitreal administration of PBS (C). Results are expressed as median (interquartile range) of 10 rats. (* p < 0.05 compared to the values obtained from PBS-injected eyes; Mann–Whitney test).

Article Snippet: Immunodetection was performed with the use of a rabbit polyclonal anti-CD40 antibody (1:1000, ab13545, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab65854, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab2391, Abcam), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho NF-κB) antibody (1:1000, ab86299, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-TNF-α antibody (1:1000, MAB610, R&D Systems), mouse monoclonal anti-glial fibrillary acidic protein (GFAP) antibody (1:1000, ab4648, Abcam) and rabbit polyclonal anti-caspase-3 antibody (1:1000, sc-7148, Santa Cruz Biotechnology Inc.).

Techniques: Injection, Saline, Western Blot, Expressing, MANN-WHITNEY

Figure 10. Müller cells were left untreated or treated with soluble CD40L for 24 h. Levels of vascular endothelial growth factor (VEGF) (A), matrix metalloproteinase-9 (MMP-9) (B), monocyte chemotactic protein (MCP)-1 (C) and high-mobility group box-1 (HMGB1) (D) were quantified in the culture media by ELISA. Protein expression of CD40 (E), caspase-3 (F) and glial fibrillary acidic protein (GFAP) (G) in the cell lysates was determined by Western blot analysis. sCD40L stimulated proliferation of Müller glial cells that was almost as potent as 10 ng/mL of VEGF compared to untreated cells (H). Results are expressed as median (interquartile range) from three different experiments. (* p < 0.05 compared with the values obtained from untreated cells; Mann–Whitney test).

Journal: International journal of molecular sciences

Article Title: CD40 Ligand-CD40 Interaction Is an Intermediary between Inflammation and Angiogenesis in Proliferative Diabetic Retinopathy.

doi: 10.3390/ijms242115582

Figure Lengend Snippet: Figure 10. Müller cells were left untreated or treated with soluble CD40L for 24 h. Levels of vascular endothelial growth factor (VEGF) (A), matrix metalloproteinase-9 (MMP-9) (B), monocyte chemotactic protein (MCP)-1 (C) and high-mobility group box-1 (HMGB1) (D) were quantified in the culture media by ELISA. Protein expression of CD40 (E), caspase-3 (F) and glial fibrillary acidic protein (GFAP) (G) in the cell lysates was determined by Western blot analysis. sCD40L stimulated proliferation of Müller glial cells that was almost as potent as 10 ng/mL of VEGF compared to untreated cells (H). Results are expressed as median (interquartile range) from three different experiments. (* p < 0.05 compared with the values obtained from untreated cells; Mann–Whitney test).

Article Snippet: Immunodetection was performed with the use of a rabbit polyclonal anti-CD40 antibody (1:1000, ab13545, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab65854, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab2391, Abcam), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho NF-κB) antibody (1:1000, ab86299, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-TNF-α antibody (1:1000, MAB610, R&D Systems), mouse monoclonal anti-glial fibrillary acidic protein (GFAP) antibody (1:1000, ab4648, Abcam) and rabbit polyclonal anti-caspase-3 antibody (1:1000, sc-7148, Santa Cruz Biotechnology Inc.).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Western Blot, MANN-WHITNEY

Figure 13. Human retinal microvascular endothelial cells were left untreated (C) or were stimulated with sCD40L for 24 h. Protein expression of phospho-ERK1/2 (A), the p65 subunit of NF-κß (B), intercellular adhesion molecule-1 (ICAM-1) (C), vascular cell adhesion molecule-1 (VCAM-1) (D), vascular endothelial growth factor (VEGF) (E), CD40 (F) and caspase-3 (G) in the cell lysates was determined by Western blot analysis. Results are expressed as mean ± standard deviation from three experiments (* p < 0.05; independent t-test).

Journal: International journal of molecular sciences

Article Title: CD40 Ligand-CD40 Interaction Is an Intermediary between Inflammation and Angiogenesis in Proliferative Diabetic Retinopathy.

doi: 10.3390/ijms242115582

Figure Lengend Snippet: Figure 13. Human retinal microvascular endothelial cells were left untreated (C) or were stimulated with sCD40L for 24 h. Protein expression of phospho-ERK1/2 (A), the p65 subunit of NF-κß (B), intercellular adhesion molecule-1 (ICAM-1) (C), vascular cell adhesion molecule-1 (VCAM-1) (D), vascular endothelial growth factor (VEGF) (E), CD40 (F) and caspase-3 (G) in the cell lysates was determined by Western blot analysis. Results are expressed as mean ± standard deviation from three experiments (* p < 0.05; independent t-test).

Article Snippet: Immunodetection was performed with the use of a rabbit polyclonal anti-CD40 antibody (1:1000, ab13545, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab65854, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab2391, Abcam), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho NF-κB) antibody (1:1000, ab86299, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-TNF-α antibody (1:1000, MAB610, R&D Systems), mouse monoclonal anti-glial fibrillary acidic protein (GFAP) antibody (1:1000, ab4648, Abcam) and rabbit polyclonal anti-caspase-3 antibody (1:1000, sc-7148, Santa Cruz Biotechnology Inc.).

Techniques: Expressing, Western Blot, Standard Deviation

Figure 15. Summary of the in vitro and in vivo experiments that documented the relevance of the CD40/CD40L pathway in proliferative diabetic retinopathy. TNF-α = tumor necrosis factor- α; VEGF = vascular endothelial growth factor; HRMEC = human retinal microvascular endothe- lial cell; ERK = extracellular-signal regulated kinase; ICAM-1 = intercellular adhesion molecule-1; VCAM-1 = vascular cell adhesion molecule-1; NF-κB = nuclear factor-Kappa B; MCP-1 = monocyte chemotactic protein-1; HMGB1 = high mobility group box-1; MMP-9 = matrix metalloproteinase-9; BRB = blood–retinal barrier; ↑= enhanced.

Journal: International journal of molecular sciences

Article Title: CD40 Ligand-CD40 Interaction Is an Intermediary between Inflammation and Angiogenesis in Proliferative Diabetic Retinopathy.

doi: 10.3390/ijms242115582

Figure Lengend Snippet: Figure 15. Summary of the in vitro and in vivo experiments that documented the relevance of the CD40/CD40L pathway in proliferative diabetic retinopathy. TNF-α = tumor necrosis factor- α; VEGF = vascular endothelial growth factor; HRMEC = human retinal microvascular endothe- lial cell; ERK = extracellular-signal regulated kinase; ICAM-1 = intercellular adhesion molecule-1; VCAM-1 = vascular cell adhesion molecule-1; NF-κB = nuclear factor-Kappa B; MCP-1 = monocyte chemotactic protein-1; HMGB1 = high mobility group box-1; MMP-9 = matrix metalloproteinase-9; BRB = blood–retinal barrier; ↑= enhanced.

Article Snippet: Immunodetection was performed with the use of a rabbit polyclonal anti-CD40 antibody (1:1000, ab13545, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab65854, Abcam), rabbit polyclonal anti-CD40L antibody (1:1000, ab2391, Abcam), rabbit polyclonal anti-p65 subunit of nuclear factor-kappa B (phospho NF-κB) antibody (1:1000, ab86299, Abcam), rabbit monoclonal anti-phospho-extracellular signal-regulated kinase (ERK)1/2 antibody (1:1000, MAB1018, R&D Systems), mouse monoclonal anti-VEGF antibody (1:750, MAB293, R&D Systems), mouse monoclonal anti-intercellular adhesion molecule-1 (ICAM1) antibody (1:100, sc-8439, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-vascular cell adhesion molecule-1 (VCAM-1) antibody (1:100, sc-13160, Santa Cruz Biotechnology Inc.), mouse monoclonal anti-TNF-α antibody (1:1000, MAB610, R&D Systems), mouse monoclonal anti-glial fibrillary acidic protein (GFAP) antibody (1:1000, ab4648, Abcam) and rabbit polyclonal anti-caspase-3 antibody (1:1000, sc-7148, Santa Cruz Biotechnology Inc.).

Techniques: In Vitro, In Vivo

Figure 2. The effect of anti-Ts7TMR scFv on A549 lung cancer and its preliminary mechanism. A. The effect of anti-Ts7TMR scFv on the proliferation of A549 cells in vitro. A549 cells were co-incubated with different doses of anti-Ts7TMR scFv (5, 25, and 45 μg) for 48h to detect cell proliferation by CCK-8 assay. B. The effect of anti-Ts7TMR scFv on A549 lung cancer growth in nude mice. Nude mice were injected subcutaneously with A549 cells and then treated subcutaneously with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg) around the tumour for 10 consecutive days. C. Scoring of PCNA, BCL-2, and VEGF expression in tumour tissues by immunohistochemistry (400 ×). Expression of PCNA, BCL-2, and VEGF in tumours treated with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg). Scale bar = 20 μm. *p < 0.05, **p < 0.01.

Journal: Artificial cells, nanomedicine, and biotechnology

Article Title: The effects of anti-lung cancer in nude mice by a fully human single-chain antibody against associated antigen Ts7TMR between A549 cells and Trichinella spiralis .

doi: 10.1080/21691401.2024.2347377

Figure Lengend Snippet: Figure 2. The effect of anti-Ts7TMR scFv on A549 lung cancer and its preliminary mechanism. A. The effect of anti-Ts7TMR scFv on the proliferation of A549 cells in vitro. A549 cells were co-incubated with different doses of anti-Ts7TMR scFv (5, 25, and 45 μg) for 48h to detect cell proliferation by CCK-8 assay. B. The effect of anti-Ts7TMR scFv on A549 lung cancer growth in nude mice. Nude mice were injected subcutaneously with A549 cells and then treated subcutaneously with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg) around the tumour for 10 consecutive days. C. Scoring of PCNA, BCL-2, and VEGF expression in tumour tissues by immunohistochemistry (400 ×). Expression of PCNA, BCL-2, and VEGF in tumours treated with PBS, negative serum (Ns), anti-T. spiralis serum (Ts), and different doses of anti-Ts7TMR scFv (25, 50, and 100 μg). Scale bar = 20 μm. *p < 0.05, **p < 0.01.

Article Snippet: Then mouse anti-human PCNA monoclonal antibody (60097-1-Ig, Proteintech), mouse anti-human BCL-2 monoclonal antibody (60178-1-Ig, Proteintech), and mouse anti-human VEGF monoclonal antibody (66828-1-Ig, Proteintech) were added separately dropwise and incubated at 37 °C for 30 min. Then the sections were incubated with biotin-conjugated Goat anti-mouse IgG at 37 °C for 30 min. Next, Streptavidin Immunohistochemistry kits (KIT-9710, maxim biotechnologies) and diaminobenzidine (dAB-0031, maxim biotechnologies) were added dropwise in sequence.

Techniques: In Vitro, Incubation, CCK-8 Assay, Injection, Expressing, Immunohistochemistry