Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Vaccination with parasite-specific TcTASV proteins combined with recombinant baculovirus as a delivery platform protects against acute and chronic Trypanosoma cruzi infection

doi: 10.3389/fcimb.2024.1297321

Figure Lengend Snippet: Vaccination with TcTASV and baculovirus elicits robust immune responses. (A) Immunization scheme and experimental design. Animals were immunized with recombinant proteins (rTcTASV-C or rGST) in the first dose and with recombinant proteins plus BV in the second (rTcTASV-C+BV::A CAP or rGST+ wt BV); sham-immunized animals were injected with PBS. Immunological and parasitological parameters (after challenge) were evaluated. (B-D) Evaluation of antibody responses by ELISA in immunized animals. (B) Total IgG, (C) IgG1 and IgG2a isotype profile against TcTASV-C, and (D) total IgG against TcTASV-A were evaluated in sera obtained seven days after the last immunization dose. The OD values obtained for GST were subtracted and the anti-TcTASV specific response was plotted. Two-tailed unpaired Student’s t-test between TcTASV- and control-vaccinated groups (*p ≤ 0.05; ** p≤ 0.005; ***p ≤ 0.0005). Mouse strain: C3H/He; N= 5/group. Similar results were observed in 3 independent experiments. (E) Cytokine and chemokine profiles in vaccinated mice. Commercial arrays displaying immobilized capture antibodies specific for 32 cytokines were used to evaluate the cytokine secretion profile in sera of immunized mice at 7 days after last dose. The membranes were incubated with pooled sera from TcTASV-C-vaccinated or sham-immunized mice, developed by chemiluminescence and quantified by densitometry using the ImageJ software. Each array was first normalized to its internal positive controls, and then the ratio between TcTASV-vaccinated/sham-immunized signals was calculated for each cytokine and expressed as a log2 fold change. (F) Cellular immune responses. Immunized mice were euthanized seven days after the second dose, and splenocytes were cultured with media alone or stimulated with rGST, rTcTASV-C, rTcTASV-A, or both rTcTASV-C and rTcTASV-A (10 μg/mL) for 24 hours. Cells were harvested and surface labeled for CD4, CD8, and intracytoplasmic IFNγ. Fluorescence acquisition and quantification was performed using flow cytometry. Upper panel: IFNγ production by CD4+ cells. Lower panel: IFNγ production by CD8+ cells. Two-way ANOVA plus Bonferroni’s post-test (**p ≤ 0.005; ***p ≤ 0.0005). (G) Complement-mediated lysis of trypomastigotes by sera from TcTASV-vaccinated mice. Sera from TcTASV-, control- vaccinated mice or preimmune sera were decomplemented and incubated with cell-derived trypomastigotes (RA strain) in the presence (or not) of fresh human serum as an external source of complement at 37°C. After 1 h, live parasites were quantified by microscopy. The lysis percentage was calculated as the ratio of live parasites incubated with and without complement (established as 100% survival). One-way ANOVA plus Tukey’s post-test (***p ≤ 0.0005; differences calculated vs. pre-immune sera).

Article Snippet: Commercial arrays RayBio C-Series Mouse Cytokine Antibody Array C2 (AAM-CYT-2, RayBiotech) displaying immobilized capture antibodies specific for 32 cytokines/chemokines (6Ckine (CCL21), TACK (CCL27), Eotaxin-1 (CCL11), GCSF, GM-CSF, IL-2, IL-3, IL-4, IL-5, IL-6, IL-9, IL-10, IL-12 p40/p70, IL-12 p70, IL-13, IL-17A, IFN-gamma, KC (CXCL1), Leptin, MCP-1 (CCL2), MCP-5, MIP-1 alpha (CCL3), MIP-2, MIP-3 beta (CCL19), RANTES (CCL5), SCF, TNF RI (TNFRSF1A), TARC (CCL17), TIMP-1, TNF alpha, Thrombopoietin (TPO) and VEGF-A) were used to evaluate the cytokine secretion profile in immunized mice, following the manufacturer’s protocol.

Techniques: Recombinant, Injection, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Incubation, Software, Cell Culture, Labeling, Fluorescence, Flow Cytometry, Lysis, Derivative Assay, Microscopy

Journal: Oncology reports

Article Title: Peiminine serves as an adriamycin chemosensitizer in gastric cancer by modulating the EGFR/FAK pathway.

doi: 10.3892/or.2018.6184

Figure Lengend Snippet: Figure 5. PMI combined with ADR suppresses the phosphorylation of receptor tyrosine kinases in GC cell lines. (A and B) SGC7901 cells were treated with ADR (1.0 µM) or ADR (1.0 µM) combined with PMI (50.0 µM) and proteins were detected by the RayBio® Human RTK phosphorylation antibody array kit (chemilumi- nescent readout). 1, p-EGFR; 2, ACK1; 3, CsK; 4, EphA4; 5, EphA5; 6, EphA6; 7, EphA7; 8, EphB3; 9, FAK; 10, FER; 11, Fgr; 12, FRK; 13, LTK; 14, PDGFR-α; 15, ROS; 16, Tie-1; 17, Tie-2; 18, TyK2; 19, TYRO10. (C and D) Expression levels of p-EGFR, EGFR, p-FAK, FAK, cyclin D1 and cleaved (cl)-PARP were detected using western blotting. β-actin was used as a reference control. ADR, adriamycin; PMI, peiminine; *P<0.05, **P<0.01 compared with the ADR alone group.

Article Snippet: The RayBio® Human RTK phosphorylation antibody array kit was used to test the differential protein expression.

Techniques: Phospho-proteomics, Ab Array, Expressing, Western Blot, Control