Journal: ACS Applied Bio Materials

Article Title: Small Intestine-Targeted Long-Acting Oral Insulin Formulation Based on Engineered Milk Protein Nanoparticles

doi: 10.1021/acsabm.5c02114

Figure Lengend Snippet: Insulin bioavailability in nondiabetic mice and in diabetic mice under fed and fasted conditions after oral casNP/insulin/C 10 administration. ( A ) Blood insulin levels in nondiabetic mice before (0 h) and at 15, 30 min, 1, 2, 4, and 8 h after administration of casNP/insulin/C 10 by oral gavage (50 IU/kg) or IAsp by s.c. injection (5 IU/kg). ( B ) Plasma insulin levels of diabetic mice with normal feeding conditions were measured from 1 to 24 h after oral gavage of casNP/insulin/C 10 at 20 and 50 IU/kg body weight and s.c. injection of IAsp at 5 IU/kg body weight. The insulin levels in mice receiving no treatment were measured as the baseline. ( C ) Changes of insulin levels in the mouse livers 1 to 24 h after oral gavage of casNP/insulin/C 10 at 50 IU/kg body weight. ( D ) Western blotting showing the levels of Akt phosphorylation in the livers of mice after receiving casNP/insulin/C 10 and IAsp for 1, 2, 4, 6, 8, and 24 h. ( E ) Plasma insulin levels of fasted mice 1 to 8 h after the administration of casNP/insulin/C 10 (50 IU/kg body weight, oral) and IAsp (5 IU/kg body weight, s.c.).

Article Snippet: Insulin aspart (IAsp, Y0000349) was purchased from EDQM (Strasbourg, France).

Techniques: Injection, Clinical Proteomics, Western Blot, Phospho-proteomics

Journal: ACS Applied Bio Materials

Article Title: Small Intestine-Targeted Long-Acting Oral Insulin Formulation Based on Engineered Milk Protein Nanoparticles

doi: 10.1021/acsabm.5c02114

Figure Lengend Snippet: Efficacy of casNP/insulin/C 10 in controlling the glycemia of diabetic mice. ( A ) Illustration of the study design. ( B ) Changes of BGLs in diabetic mice on Days 1, 2, and 5 after receiving casNP/insulin/C 10 orally at 20 and 50 IU/kg body weight, IAsp s.c. at 5 IU/kg body weight, and PBS as a placebo, daily at 8:30 am. ( C ) BGLs of diabetic mice under fasting conditions were monitored for 8 h after administration of treatments and placebo.

Article Snippet: Insulin aspart (IAsp, Y0000349) was purchased from EDQM (Strasbourg, France).

Techniques:

Journal: iScience

Article Title: Chemical and biological characterization of vaccine adjuvant QS-21 produced via plant cell culture

doi: 10.1016/j.isci.2024.109006

Figure Lengend Snippet: In vitro biological characterization; ccQS-21 and beQS-21 equivalently induce hemolysis, trigger the innate immune cell stress response, and enhance antigen processing and presentation to enhance the potency of the vaccinal immune synapse (A) Upon incubation with human red blood cells both beQS-21 and ccQS-21 retain equivalent saponin-specific hemolytic properties at pH-7.4 with matching half maximal effective concentration (EC50). (B) Upon pre-conditioning of THP1 human myeloid cells with a TLR4 agonist (LPS), both beQS-21 and ccQS-21 induce inflammasome-dependent secretion of IL-1β and the production of alarmins associated with pyroptosis such as HMGB1. Both IL-1β secretion and pyroptosis can be regulated by the canonical inflammasome pathway and can be suppressed by the disruption of the genes encoding for NLRP3, ASC and Caspase-1 or by the use of Caspase-1 inhibitor, ZVAD. (C) Both beQS-21 and ccQS-21 can comparably induce a broader inflammatory response in vitro . The breadth of the inflammatory response induced by the stimulation of human granulocyte macrophage colony stimulating factor (GMCSF) and IL-4 differentiated Monocyte derived Dendritic Cells (Mo-DCs) using both QS-21 was assessed with multiplex cytokine quantification. Both beQS-21 (BE) and ccQS-21 (CC) induce the same inflammatory signature which is further enhanced by formulating with TLR4 agonist. (D) Pseudo cross-presentation assay using cell viability to demonstrate that beQS-21 and ccQS-21 equivalently induce cytotoxicity through the regulation of the cytosolic translocation from the phagolysosome (e.g., as of the indicated toxin: saporin), a critical process for antigen cross presentation. (E) Flow cytometry analysis demonstrating that both QS-21 equivalently enhance large antigen processing and cross-presentation on MHC-I using a T cell receptor (TCR)-like antibody against the H-2K b -SIINFEKL complex. (F) In vitro antigen cross-presentation assay demonstrating that ccQS-21 and beQS-21 equivalently enhance antigen processing and minimal epitope cross-presentation to activate transgenic T cells with matched OT1 TCR to express NFAT luciferase reporter. (G) OT1 T cell proliferation shows ccQS-21 and beQS-21 equivalently enhance antigen cross-presentation to induce the proliferation of primary T cells with matched OT1 TCR. OVA: Ovalbumin, OT1 long peptide: 29mer peptide including SIINFEKL, BFM: Bafilomycin A1, inhibiting the vacuolar type H + -ATPase (v-ATPase) to transfer protons into the lysosome.

Article Snippet: Cells were then plated in flat-bottom culture 96-well plates (0.5 × 10 6 cells per well). gE overlapping peptide mix (0.25 μg/mL, JPT Peptide Technologies, Germany), or OT1 and OT-II peptide mix (5 μg/mL, GenScript, China) were added for restimulation.

Techniques: In Vitro, Incubation, Concentration Assay, Disruption, Derivative Assay, Multiplex Assay, Translocation Assay, Flow Cytometry, Transgenic Assay, Luciferase

Journal: iScience

Article Title: Chemical and biological characterization of vaccine adjuvant QS-21 produced via plant cell culture

doi: 10.1016/j.isci.2024.109006

Figure Lengend Snippet:

Article Snippet: Cells were then plated in flat-bottom culture 96-well plates (0.5 × 10 6 cells per well). gE overlapping peptide mix (0.25 μg/mL, JPT Peptide Technologies, Germany), or OT1 and OT-II peptide mix (5 μg/mL, GenScript, China) were added for restimulation.

Techniques: Recombinant, Cell Isolation, Enzyme-linked Immunospot, Cytotoxicity Assay, Luciferase, Expressing

Journal: Cancer gene therapy

Article Title: Optimization of vaccine responses with an E1, E2b, E3 deleted Ad5 vector circumvents pre-existing anti-vector immunity

doi: 10.1038/cgt.2009.17

Figure Lengend Snippet: (a) The capacity of Ad-infected human DC to process and present antigen was measured by using a human T cell clone specific for HLA-A*0201/CMVpp65(495–503) peptide-MHC complexes. In vitro cultured human DC were infected with Ad5[E1+], Ad5[E1−], or Ad5[E1−, E2b−] vectors encoding CMVpp65 or CEA at MOI 20,000, or mock infected and incubated for 2 days. 1×10 4 DC-transduced with the vectors were used as stimulators and 1×10 4 or 5×10 4 of CMVpp65 CTL line (3H1) was used as responder cells. DC pulsed with CMVpp65 overlapping polypeptide mix and DC pulsed with a HIV gag overlapping polypeptide mix were used as positive and negative controls respectively. Interferon-gamma ELISPOT results are shown. Error bars represent standard deviation. Representative data are shown from 3 individual experiments. (b) To compare the stimulatory capacity of Ad5CMV-transduced DC, i n vitro cultured human DC were infected with Ad5[E1+], Ad5[E1−], or Ad5[E1−, E2b−] vectors encoding CMVpp65 at MOI 20,000, mock infected, or pulsed with CMVpp65 or HIV overlapping polypeptide mixes as positive control negative controls respectively. DC were cultured with autologous HLA-A*0201+ peripheral blood lymphocytes (PBL) at 10:1 (PBL:DC) ratio. After 10 days of culture the percentage of CD8+ HLA-A*0201/CMVpp65(495–503) peptide-MHC tetramer positive T cells was measured by flow cytometry. Percentages of CD8+ tetramer+ cells in CD3+CD4−CD14−CD19−lymphocytes are shown in dot plots, which were calculated from duplicate samples. Representative data are shown from 3 individual experiments.

Article Snippet: Microtiter plates (Immulon 4BX) were coated with CEA peptide mix or CMVpp65 peptide mix (BD Bioscience) at concentration of 2.6 μg/ml.

Techniques: Infection, In Vitro, Cell Culture, Incubation, Transduction, Enzyme-linked Immunospot, Standard Deviation, Positive Control, Flow Cytometry