Journal: Journal of biological engineering

Article Title: The infectivity of AAV9 is influenced by the specific location and extent of chemically modified capsid residues.

doi: 10.1186/s13036-024-00430-7

Figure Lengend Snippet: Fig. 1 Analysis of AAV9 WT and mutant capsid proteins expression and AAV productions. A Western Blot analysis of VP1, VP2, and VP3 protein expression 72 hours post-transfection of AAV9 wild-type (WT) and mutated PDP9 plasmids together with the pAcBac1-tR4-MbPyl plasmid in the presence or absence of the unnatural amino acid AzK. B Presentation of AAV9 and mutant AAV titers obtained after the initial purification step using an iodixanol gradient, shown in the first column. The second column displays the titers obtained after the second purification step, which involved affinity chromatography in addition to the first purification step

Article Snippet: corresponding antibodies to detect AAV capsid proteins (Origene [BM5051]) and actin (Abcam) and the Lumigen ECL ULTRA substrate A+B (Lumigen).

Techniques: Mutagenesis, Expressing, Western Blot, Transfection, Plasmid Preparation, Purification, Affinity Chromatography

Journal: Cell Biology and Toxicology

Article Title: AP-1 activates KCNN4-mediated Ca2 + signaling to induce the Th1/Th17 differentiation of CD4 + cells in chronic non-bacterial prostatitis

doi: 10.1007/s10565-024-09967-1

Figure Lengend Snippet: KCNN4 is a downstream target of AP-1 in CD4 + T cells. ( a ) Expression of KCNN4 in prostate CD4 + T cells detected by transcriptome sequencing was examined using control and CNP rats. ( b ) DNA binding matrix of the transcription factor AP-1. ( c ) The binding site for AP-1 on the KCNN4 promoter fragment was predicted in Jaspar. ( d ) Effects of T + E treatment and combined use of the AP-1 inhibitor SR11302 on KCNN4 expression in CD4 + T cells were analyzed using RT-qPCR. ( e ) The binding ability of AP-1(FOS, JUN) to the KCNN4 promoter fragment in CD4 + T cells was detected by ChIP-qPCR. ( f ) The effects of T + E treatment and combined use of the AP-1 inhibitor SR11302 on the transcriptional activity of the KCNN4 promoter in CD4 + T cells were examined using reporter assay. Experiments were performed at least in triplicate. Data are shown as mean ± SEM, n = 5, Unpaired t-test or one-way ANOVA

Article Snippet: The overexpression adeno-associated virus (AAV) of KCNN4 (AAV-KCNN4) and the AAV-negative control (NC) were acquired from VectorBuilder (Guangzhou, Guangdong, China), and the viral titer was 10 13 genome copy/mL.

Techniques: Expressing, Sequencing, Control, Binding Assay, Quantitative RT-PCR, Activity Assay, Reporter Assay

Journal: Cell Biology and Toxicology

Article Title: AP-1 activates KCNN4-mediated Ca2 + signaling to induce the Th1/Th17 differentiation of CD4 + cells in chronic non-bacterial prostatitis

doi: 10.1007/s10565-024-09967-1

Figure Lengend Snippet: AP-1/KCNN4 promotes Th1/Th17 differentiation in a Ca2 + signaling-dependent manner. ( a ) The effects of T + E treatment and the combined use of the AP-1 inhibitor SR11302 on Ca2 + concentration in CD4 + T cells were analyzed using Fluo-4-AM fluorescent staining. Overexpression efficiency of Vector-KCNN4 detected by RT-qPCR ( b ) and western blot analysis ( c ). ( d ) Effect of overexpression of KCNN4 on intracellular Ca2 + concentration detected by Fluo-4-AM fluorescence staining. ( e ) The effect of overexpression of KCNN4 on Th1/Th17 differentiation of CD4 + T cells was flow cytometry. ( f ) The effect of overexpression of KCNN4 on intracellular Ca2 + concentration in the presence of the calcium chelator BAPTA-AM was analyzed using Fluo-4-AM fluorescence staining. ( g ) The effect of overexpression of KCNN4 on Th1/Th17 differentiation of CD4 + T cells in the presence of the calcium chelator BAPTA-AM was analyzed using flow cytometry. Experiments were performed at least in triplicate. Data are shown as mean ± SEM, Unpaired t-test, or one-way ANOVA

Article Snippet: The overexpression adeno-associated virus (AAV) of KCNN4 (AAV-KCNN4) and the AAV-negative control (NC) were acquired from VectorBuilder (Guangzhou, Guangdong, China), and the viral titer was 10 13 genome copy/mL.

Techniques: Concentration Assay, Staining, Over Expression, Plasmid Preparation, Quantitative RT-PCR, Western Blot, Fluorescence, Flow Cytometry

Journal: Cell Biology and Toxicology

Article Title: AP-1 activates KCNN4-mediated Ca2 + signaling to induce the Th1/Th17 differentiation of CD4 + cells in chronic non-bacterial prostatitis

doi: 10.1007/s10565-024-09967-1

Figure Lengend Snippet: Activation of autophagy inhibits Th1/Th17 differentiation. ( a ) Effects of T + E treatment and the combination of the AP-1 inhibitor SR11302 on the activity of mTOR and the expression of autophagy-related proteins in CD4 + T cells were analyzed using western blot. ( b ) Effects of overexpression of KCNN4 and combined use of the calcium chelator BAPTA-AM on the activity of mTOR and the expression of autophagy-related proteins in cells were analyzed using western blot. ( c ) The effect of rapamycin on AP-1 activity was analyzed using an AP-1 luciferase reporter assay. ( d ) Effect of rapamycin on the activity of mTOR and the expression of KCNN4 and autophagy-related proteins in CD4 + T cells detected by western blot. ( e ) Effect of rapamycin on Th1/Th17 differentiation of CD4 + T cells detected using flow cytometry. Experiments were performed at least in triplicate. Data are shown as mean ± SEM, Unpaired t-test, or one-way ANOVA

Article Snippet: The overexpression adeno-associated virus (AAV) of KCNN4 (AAV-KCNN4) and the AAV-negative control (NC) were acquired from VectorBuilder (Guangzhou, Guangdong, China), and the viral titer was 10 13 genome copy/mL.

Techniques: Activation Assay, Activity Assay, Expressing, Western Blot, Over Expression, Luciferase, Reporter Assay, Flow Cytometry

Journal: Cell Biology and Toxicology

Article Title: AP-1 activates KCNN4-mediated Ca2 + signaling to induce the Th1/Th17 differentiation of CD4 + cells in chronic non-bacterial prostatitis

doi: 10.1007/s10565-024-09967-1

Figure Lengend Snippet: CD4 + T cell Th1/Th17 differentiation blocked by autophagy activation slows CNP progression. ( a ) Effect of AAV-KCNN4 as well as rapamycin treatment on body weight, prostate weight, and prostate index in CNP rats. ( b ) Effects of AAV-KCNN4 and rapamycin treatment on the expression of KCNN4, mTOR, and autophagy-related proteins in prostate CD4 + T cells from rats with CNP detected by western blot. ( c ) Effects of AAV-KCNN4 and rapamycin treatment on Th1/Th17 differentiation of prostate CD4 + T cells in CNP rats. ( d ) Effects of AAV-KCNN4 and rapamycin treatment on the reduction of inflammatory infiltrates (red arrowheads), PIN (black arrowheads), and adenocarcinoma (red arrowheads) in the prostate of CNP rats. + Data are shown as mean ± SEM, n = 5

Article Snippet: The overexpression adeno-associated virus (AAV) of KCNN4 (AAV-KCNN4) and the AAV-negative control (NC) were acquired from VectorBuilder (Guangzhou, Guangdong, China), and the viral titer was 10 13 genome copy/mL.

Techniques: Activation Assay, Expressing, Western Blot

Journal: Cell Biology and Toxicology

Article Title: AP-1 activates KCNN4-mediated Ca2 + signaling to induce the Th1/Th17 differentiation of CD4 + cells in chronic non-bacterial prostatitis

doi: 10.1007/s10565-024-09967-1

Figure Lengend Snippet: Schematic diagram. Aberrant AP-1 activity in CD4 + T cells promotes the transcription of KCNN4, which inhibits autophagy by enhancing Ca2 + signaling to activate mTORC1 activity in the cells. This leads to Th1/Th17 differentiation of CD4 + T cells and the progression of CNP to PIN and adenocarcinoma

Article Snippet: The overexpression adeno-associated virus (AAV) of KCNN4 (AAV-KCNN4) and the AAV-negative control (NC) were acquired from VectorBuilder (Guangzhou, Guangdong, China), and the viral titer was 10 13 genome copy/mL.

Techniques: Activity Assay

Journal: Brain Tumor Research and Treatment

Article Title: Trends in Developing Extracellular Vesicle-Based Therapeutics

doi: 10.14791/btrt.2024.0027

Figure Lengend Snippet: Development cases of extracellular vesicle-based therapeutics by global biopharmaceutical companies

Article Snippet: Evox Therapeutics will collaborate with the Icahn School of Medicine at Mount Sinai (USA) to encapsulate adeno-associated virus (AAV) into extracellular vesicles, and to develop extracellular vesicle-encapsulated AAV vectors (exoAAV), a gene therapy technology to treat cardiac diseases [ ].

Techniques: