Journal: The Journal of Clinical Investigation

Article Title: TRAIL agonists rescue mice from radiation-induced lung, skin, or esophageal injury

doi: 10.1172/JCI173649

Figure Lengend Snippet: ( A ) Experimental timeline. Female C57BL/6 mice orthotopically injected with e0771 on day 0 and when tumors reached 2–5 mm in size mice were irradiated with a whole-thorax irradiation dose of 20 Gy and treated with either TLY012, ONC201, or the combination ( n = 3–4 per treatment per group). (Created in BioRender. Strandberg J. 2024. https://BioRender.com/c65v064. ) ( B and C ) Tumors were removed after mice were euthanized 18 days after cell injection, and weight and volume were calculated. ( D ) Pulse oximetry readings before radiation and 9 days after radiation showed oxygen saturation more conserved in the TLY012-treated group compared with irradiated controls. Values are mean ± SEM. ( E ) Statistical analysis of cytokine fold change showed significant decrease in levels of MDC/CCL22 ( P = 0.035) in comparison with irradiated controls (1-way ANOVA with Tukey’s post hoc test).

Article Snippet: Seven-week-old Trail –/– female mice received a single thoracic x-ray irradiation dose of 20 Gy with shielding of other organs and were treated with either 20 μg of anti-CCL22 antibody (AF439, R&D Systems Inc.) dissolved in 500 μL of sterile normal saline via i.p. injection every other day for 2 weeks or 20 μg of polyclonal goat IgG (AB-108-C, R&D Systems Inc.) dissolved in 500 μL of normal saline via i.p. injection every other day for 2 weeks ( n = 5 per treatment per group).

Techniques: Injection, Irradiation, Comparison

Journal: The Journal of Clinical Investigation

Article Title: TRAIL agonists rescue mice from radiation-induced lung, skin, or esophageal injury

doi: 10.1172/JCI173649

Figure Lengend Snippet: ( A ) Experimental timeline of Trail –/– female mice treated with 20 μg of anti-CCL22 in 500 μL of saline or 20 μg of goat IgG in 500 μL of saline every other day for 2 weeks ( n = 5 per treatment per group). (Created in BioRender. Strandberg J. 2024. https://BioRender.com/j52q830. ) ( B ) H&E images of each mouse 2 weeks after thoracic irradiation (original magnification, ×20). Scale bar: 100 μm. ( C ) Quantification of inflammatory scores provided by a blinded pathologist showed decrease in inflammation in Trail –/– female mice treated with anti-CCL22 ( n = 5 per treatment per group) but not to a significant extent (1-tailed Mann-Whitney test) ( P = 0.1032). Values are mean ± SEM.

Article Snippet: Seven-week-old Trail –/– female mice received a single thoracic x-ray irradiation dose of 20 Gy with shielding of other organs and were treated with either 20 μg of anti-CCL22 antibody (AF439, R&D Systems Inc.) dissolved in 500 μL of sterile normal saline via i.p. injection every other day for 2 weeks or 20 μg of polyclonal goat IgG (AB-108-C, R&D Systems Inc.) dissolved in 500 μL of normal saline via i.p. injection every other day for 2 weeks ( n = 5 per treatment per group).

Techniques: Saline, Irradiation, MANN-WHITNEY

Journal: Chemical Engineering Journal

Article Title: Microfluidic fabrication of lipid nanoparticles for co-delivery of siRNA and hydroxychloroquine: An engineered theranostic platform for enhanced breast cancer treatment

doi: 10.1016/j.cej.2025.160172

Figure Lengend Snippet: Fig. 3. Effect of various treatments on cell proliferation. Screening of human- (A) and mouse- (B) derived candidate siRNA sequences. (C) Cell counting kit-8 (CCK-8) assay to evaluate anti-proliferative results against 4T1 cells. (D) Crystalline violet staining to visualize agent-treated cell colonies. (E) Ultraviolet absorption of crystalline violet eluents from stained colonies (n = 3). *p<0.05, **p < 0.01, ***p < 0.001.

Article Snippet: The LysoTracker Green and Autophagy Staining Assay Kit with MDC were obtained from Beyotime (Shanghai, China).

Techniques: Derivative Assay, Cell Counting, CCK-8 Assay, Staining

Journal: Cancer science

Article Title: ADAM23 is downregulated in side population and suppresses lung metastasis of lung carcinoma cells.

doi: 10.1111/cas.12895

Figure Lengend Snippet: Fig. 3. ADAM23 expression in A549-derived side population (SP) and main population (MP) cells and other lung adenocarcinoma cell lines. (a) mRNA expression of ADAM23 in A549 parent, SP(1), MP(1), SP(9) and MP(9) cells by RT-PCR (upper panel) and qPCR (lower panel) (n = 4). (b) mRNA expression of ADAM23 in parent, SP and MP cells derived from lung adenocarcinoma cell lines measured by qPCR (n = 3). Bars, mean SD. **P < 0.01; ***P < 0.001.

Article Snippet: ADAM23 expression vectors and empty vectors were purchased from Origene (Rockville, MD, USA) and transfected using FuGENE6 (Roche Diagnostics).

Techniques: Expressing, Derivative Assay, Reverse Transcription Polymerase Chain Reaction

Journal: Cancer science

Article Title: ADAM23 is downregulated in side population and suppresses lung metastasis of lung carcinoma cells.

doi: 10.1111/cas.12895

Figure Lengend Snippet: Fig. 4. Effects of ADAM23 expression, anti-ADAM23 antibody and anti-avb3 integrin antibody on colony formation. (a) Expression of ADAM23 in A549 cells (control) or A549 transfectants with empty vec- tors, ADAM23 expression vectors, non-targeting mock vectors or ADAM23-targeting lentiviral vectors (sh1 and sh5) by RT-PCR and by immunoblotting of the immunoprecipitates (IP-IMB). Arrow indicates ADAM23 and the lower band is a non-specific protein band. (b) Col- ony formation by control A549 cells (control) and A549 cells trans- fected with empty vectors, ADAM23 expression vectors, non-targeting vectors (mock) or ADAM23-targeting vectors (sh1 and sh5) was assayed (n = 6). (c) Colony formation by A549 cells transfected with empty vectors or ADAM23 expression vectors, ADAM23 transfectants treated with non-immune IgG (NI, ADAM23) or anti-ADAM23 anti- body (a-ADAM23), and empty vector transfectants treated with non- immune IgG (NI, empty vector) or anti-avb3 integrin antibody (a-avb3) was determined (n = 6). Bars, mean SD. *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: ADAM23 expression vectors and empty vectors were purchased from Origene (Rockville, MD, USA) and transfected using FuGENE6 (Roche Diagnostics).

Techniques: Expressing, Control, Reverse Transcription Polymerase Chain Reaction, Western Blot, Transfection, Plasmid Preparation

Journal: Cancer science

Article Title: ADAM23 is downregulated in side population and suppresses lung metastasis of lung carcinoma cells.

doi: 10.1111/cas.12895

Figure Lengend Snippet: Fig. 5. Involvement of ADAM23 and avb3 integrin in cell adhesion and migration. (a) Cell adhesion of control A549 cells (control) and A549 cells transfected with empty vectors, ADAM23 expression vectors, non-targeting vectors (mock) or ADAM23-targeting vectors (sh1 and sh5) was assayed (n = 4). (b) Cell adhesion of A549 cells transfected with empty vectors or ADAM23 expression vectors, ADAM23 transfectants treated with non-immune IgG (NI, ADAM23) or anti-ADAM23 antibody (a-ADAM23), ADAM23 transfectants treated with scrambled peptide or disinte- grin peptide, ADAM23 transfectants treated with disintegrin peptide and non-immune IgG (NI, disintegrin peptide) or disintegrin peptide and anti-avb3 antibody (a-avb3, disintegrin peptide), and empty vector transfectants treated with non-immune IgG (NI, empty vector) or anti-avb3 integrin antibody (a-avb3, empty vector) was determined (n = 4). (c) Migration of control A549 cells (control) and A549 cells transfected with empty vectors, ADAM23 expression vectors, non-targeting vectors (mock) or ADAM23-targeting vectors (sh1 and sh5) was measured by migration assay (n = 4). (d) Migration of A549 cells treated with non-immune IgG (NI) or anti-avb3 integrin antibody (a-avb3) was measured using the xCEL- Ligence system (n = 4). Bars, mean SD. *P < 0.05; ***P < 0.001.

Article Snippet: ADAM23 expression vectors and empty vectors were purchased from Origene (Rockville, MD, USA) and transfected using FuGENE6 (Roche Diagnostics).

Techniques: Migration, Control, Transfection, Expressing, Plasmid Preparation

Journal: Cancer science

Article Title: ADAM23 is downregulated in side population and suppresses lung metastasis of lung carcinoma cells.

doi: 10.1111/cas.12895

Figure Lengend Snippet: Fig. 7. Lung metastasis in NOD ⁄ SCID mice after i.v. injection of A549ffLuc-cp156 transfectants. (a) Lung metastasis of the transfectants with empty vectors, ADAM23 expression vectors, mock vectors or shRNA vectors (sh5) was monitored at 1 h, 1 day and 2 weeks by bioluminescence imaging (n = 6 per group). Representative mice of each group are shown. (b) Photon counts were analyzed (n = 6). (c) Representative microscopic views of the lung sections. Hematoxylin-positive blue-colored nodules indicate metastatic foci. Bar, 1 mm. (d) Evaluation of lung metastasis by calculat- ing the numbers ⁄ mm2 (n = 3). Bars, mean SD. *P < 0.05; **P < 0.01.

Article Snippet: ADAM23 expression vectors and empty vectors were purchased from Origene (Rockville, MD, USA) and transfected using FuGENE6 (Roche Diagnostics).

Techniques: Injection, Expressing, shRNA, Imaging

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Distinct roles for CCR4 and CXCR3 in the recruitment and positioning of regulatory T cells in the inflamed human liver.

doi: 10.4049/jimmunol.0901216

Figure Lengend Snippet: FIGURE 6. CCR4 ligands are detected in diseased liver tissue and secreted by LIDCs. A, LITregs migrate toward CCR4 ligands in vitro. Migration of LITregs to CCR4 ligands CCL22 and CCL17 was assessed using fibronectin-coated 5-mm-pore transwell migration chambers. The lower chambers were filled with 600 ml assay media (RPMI 1640 medium with 0.1% BSA) alone or supplemented with chemokines (100 ng/ml recombinant human CCL22 and CCL17). BSA was used as control. A total of 5 3 105 Tregs isolated from inflamed livers (n = 4) were loaded per well in the upper chamber; Tregs were collected from the top and bottom chambers after 4 h of incubation at 37˚C in 5% CO2. The proportion of transmigrated cells was calculated by fixed- volume counting and phenotyped for Treg markers and chemokine receptor expression by flow cytometry. To inhibit chemokine-mediated signaling, Tregs were incubated with PTx (100 ng/ml) prior to the assay. Results are expressed as mean 6 SEM. LITregs showed significant migration responses to CCL22 and CCL17 compared with migration to BSA, and these were inhibited by PTx. pp , 0.05, Student t test. B, To determine whether liver DCs secrete CCL17 and CCL22, we isolated and cultured myeloid DCs from human liver tissue and stimulated them with LPS (1 mg/ml) for 24 h. CCL17 and CCL22, measured by ELISA, were detected in cell culture supernatants of LPS-stimulated liver-derived DCs isolated from patients with AIH (n = 2) or PBC (n = 1) or from NL (n = 3). All DCs secreted detectable CCL17 and CCL22, with a .2-fold increase in secretion by DCs derived from inflamed liver tissue compared with NL. Data represent mean 6 SEM of three experiments using supernatants from different DCs. Values represent the chemokine concentration (pg/ml) of three replicate wells relevant to a standard curve of known concentration. pp # 0.05; ppp # 0.01. C, Expression of CCL22 in human liver tissue samples by immunohistochemistry. CCL22 staining (i) was detected on LIDCs within inflammatory infiltrates (positive staining cells brown) in a patient with AIH. No detectable CCL22 was found in NL or when control Abs were used (ii). (Original magnification 3200.) D, Dual staining of FoxP3-expressing cells (brown) and CCL22 (red) in liver tissue from a patient with AIH reveals CCL22+ cells in close association with FoxP3+ Tregs within an inflammatory infiltrate (original magnification 3200). E, Confocal image of dual staining of FoxP3-expressing cells (red/TRITC) and CD11c (green/FITC) in liver tissue from a patient with AIH reveals CD11c+ cells in close association with FoxP3+ Tregs within an inflammatory infiltrate (original magnification 340). F, Colocalization of DCs and CCL22 in inflamed liver tissue. Serial liver sections from a patient with PBC were stained with CD11c (top panels) and CCL22 (bottom panels). Cells that coexpress CD11c and CCL22 can be determined by comparing staining of CD11c (top right panel) and CCL22 (bottom right panel) at higher power (original magnification 3200). Staining with isotype-matched control Abs was negative (left panels, original magnification 3100.) G, Colocalization of DCs and CCL22 in inflamed liver tissue. Staining of FITC-labeled CCL22 (green) with Texas Red-labeled CD11c (red). Areas of coexpression (top left panel) appear yellow. Nuclei were stained with DAPI (blue). Control Abs demonstrated minimal background tissue fluorescence. The findings are representative of the pattern of immunofluorescence staining seen in tissue sections from four donors each for PBC, ALD, AIH, and HCV

Article Snippet: The following primary Abs were used for immunohistochemistry, immunofluorescence, and confocal microscopy: mouse anti-human CCL22 mAb (MAB 336, R&D Systems, Minneapolis, MN), goat anti-human CCL17 mAb (AF 364, R&D Systems), mouse anti-human FoxP3 (Abcam, 236A/ E7, Cambridge, U.K.), rabbit anti-human polyclonal to CCL22 amino terminal end (Ab 53002-100; Abcam), rabbit anti-human monoclonal to CD11c (Abcam, EP13474), rabbit monoclonal to phosphorylated STAT5 (phosphoSTAT5)(Abcam/E208), mouse anti-human CD4 IgG 2b (eBioscience, Hatfield, U.K.), anti-human CD3 IgG2a (eBioscience), and mouse anti-human CD8 IgG2a (eBioscience).

Techniques: In Vitro, Migration, Recombinant, Control, Isolation, Incubation, Expressing, Cytometry, Cell Culture, Enzyme-linked Immunosorbent Assay, Derivative Assay, Concentration Assay, Immunohistochemistry, Staining, Labeling