Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: MicroRNA-150 relieves vascular remodeling and fibrosis in hypoxia-induced pulmonary hypertension.

doi: 10.1016/j.biopha.2018.11.058

Figure Lengend Snippet: Fig. 4. Effect of miR-150 on pulmonary fibrosis of pulmonary hypertension rats. (A) Pulmonary fibrosis was detected by Masson’s staining (200× magnification). Scale bars, 100 μm. (B) The area of pulmonary fibrosis was calculated and shown. The mRNA expressions of TGF-β1 (C) and collagen I (D) in lung tissues were evaluated by qPCR. (E) The protein levels of TGF-β1 and collagen I in lung tissues were measured by western blot assay. (F)&(G) Relative grey values of the protein bands were shown. (H) The expressions of TGF-β1 and collagen I in lung tissues were detected by immunohistochemical staining (400× magnification). Scale bars, 50 μm. Data were presented as mean ± SD. **P < 0.01, ***P < 0.001 versus the indicated group.

Article Snippet: The membranes were blocked in 5% nonfat milk for 1 h at room temperature, then incubated with primary antibodies against TGF-β1 (1:400, BOSTER, China), Collagen I (1:300, BOSTER, China), p-AKTser473 (1:500, KeyGen, China), AKT (1:500, KeyGen, China), p-mTORser2481 (1:500, Sangon Biotech, China), mTOR (1:1000, Cell signaling Technology, USA), and β-actin (1:500, Bioss, China) at 4°C overnight, followed by incubation with HRP-labeled Goat AntiRabbit or Goat Anti-Mouse IgG (1:5000, Beyotime, China) at 37°C for 45min.

Techniques: Staining, Western Blot, Immunohistochemical staining

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: MicroRNA-150 relieves vascular remodeling and fibrosis in hypoxia-induced pulmonary hypertension.

doi: 10.1016/j.biopha.2018.11.058

Figure Lengend Snippet: Fig. 6. Effect of miR-150 on the expressions of fibrosis-related molecules. The mRNA expressions of TGF-β1 (A) and collagen I (B) in PASMCs were measured by qPCR. (C) The protein levels of TGF-β1 and collagen I in PASMCs were detected by western blot assay. (D)&(E) Relative grey values of the protein bands were shown. Data were presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001 versus the indicated group.

Article Snippet: The membranes were blocked in 5% nonfat milk for 1 h at room temperature, then incubated with primary antibodies against TGF-β1 (1:400, BOSTER, China), Collagen I (1:300, BOSTER, China), p-AKTser473 (1:500, KeyGen, China), AKT (1:500, KeyGen, China), p-mTORser2481 (1:500, Sangon Biotech, China), mTOR (1:1000, Cell signaling Technology, USA), and β-actin (1:500, Bioss, China) at 4°C overnight, followed by incubation with HRP-labeled Goat AntiRabbit or Goat Anti-Mouse IgG (1:5000, Beyotime, China) at 37°C for 45min.

Techniques: Western Blot

Journal: Cell death & disease

Article Title: Cancer-derived exosomal HSPC111 promotes colorectal cancer liver metastasis by reprogramming lipid metabolism in cancer-associated fibroblasts.

doi: 10.1038/s41419-022-04506-4

Figure Lengend Snippet: Fig. 6 An increase in levels of acetyl-CoA and H3K27ac promotes CXCL5 secretion in CAFs. The effects of acetate treatment on acetyl-CoA levels in LX-2 cells incubated with ExoHCT116 (A) and ExoSW480 (B). Western blot showed the effects of acetate treatment on CXCL5 and H3K27ac levels in LX-2 cells incubated with ExoHCT116 (C) and ExoSW480 (D). E The effects of Trichostatin A (TSA) treatment on CXCL5 and H3K27ac levels in LX-2 cells incubated with ExoHCT116 KD and their relative control. F The effects of C646 treatment on CXCL5 and H3K27ac levels in LX-2 cells incubated with ExoSW480 OE and their relative control. G, H Chromatin immunoprecipitation (ChIP) assays using IgG as a control were performed with antibody against H3K23ac. Examination of H3K27 acetylation status in CXCL5 gene promoter region in LX-2 cells incubated with ExoHCT116 KD and ExoHCT116 KD Ctrl or in LX-2 cells incubated with ExoHCT116 KD and ExoHCT116 KD Ctrl and treated with TSA (G). Examination of H3K27 acetylation status in CXCL5 gene promoter region in LX-2 cells incubated with ExoSW480 OE and ExoSW480 OE Ctrl or in LX- 2 cells incubated with ExoSW480 OE and ExoSW480 OE Ctrl and treated with C646 (H). Each experiment was performed in triplicate. Data are shown as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: Cells were cultured in the presence of appropriate exosomes (20μg/mL) for 48 h before supernatants were collected, and the CXCL5 level was detected by CXCL5 ELISA kit (EK0728, BOSTER) according to the manufacturer’s protocols.

Techniques: Incubation, Western Blot, Control, Chromatin Immunoprecipitation

Journal: Cell death & disease

Article Title: Cancer-derived exosomal HSPC111 promotes colorectal cancer liver metastasis by reprogramming lipid metabolism in cancer-associated fibroblasts.

doi: 10.1038/s41419-022-04506-4

Figure Lengend Snippet: Fig. 8 A schematic diagram of mechanism of exosomal HSPC111 facilitates CRLM via reprogramming lipid metabolism in CAFs. Exosomal HSPC111 derived from CRC cells phosphorylates ACLY in CAFs, leading to the increased levels of acetyl-CoA and histone acetylation to secrete CXCL5, and resulting in CRC cells colonized in liver via the CXCL5-CXCR2 axis.

Article Snippet: Cells were cultured in the presence of appropriate exosomes (20μg/mL) for 48 h before supernatants were collected, and the CXCL5 level was detected by CXCL5 ELISA kit (EK0728, BOSTER) according to the manufacturer’s protocols.

Techniques: Derivative Assay

Journal: Bioactive materials

Article Title: Bioactive extracellular matrix scaffolds engineered with proangiogenic proteoglycan mimetics and loaded with endothelial progenitor cells promote neovascularization and diabetic wound healing.

doi: 10.1016/j.bioactmat.2021.08.017

Figure Lengend Snippet: Fig. 2. Specificity and binding affinity of LXW7 ligand to ZDF-EPCs and expression of αvβ3 integrin on ZDF-EPCs A. On-bead cell binding assay for testing the binding affinity and specificity of ligands to ZDF-EPCs showing high binding of ZDF-EPCs to LXW7 coated beads (right panel) compared to THP-1 monocytes (left panel). B. Quantification of the number of THP-1 monocytes and ZDF-EPCs bound to the LXW7 coated beads. C. Expression of αvβ3 integrin on ZDF-EPCs. D. Flow cytometry analysis of the binding affinity of LXW7 showing the blocking of its binding by anti-αvβ3 antibody. Scale bar = 100 μm. Data are expressed as mean ± SD. ****p < 0.0001, n = 3.

Article Snippet: Before purifying, LXW7 was cyclized by oxidizing cysteine residues using ClearOx resin (Peptides International) according to manufacturer’s protocol.

Techniques: Binding Assay, Expressing, Cell Binding Assay, Flow Cytometry, Blocking Assay

Journal: Bioactive materials

Article Title: Bioactive extracellular matrix scaffolds engineered with proangiogenic proteoglycan mimetics and loaded with endothelial progenitor cells promote neovascularization and diabetic wound healing.

doi: 10.1016/j.bioactmat.2021.08.017

Figure Lengend Snippet: Fig. 3. Effect of LXW7 and LXW7-DS-SILY ligands on the attachment, growth and viability of ZDF-EPCs on ligand modified surfaces. A-B. Representative images of attached ZDF-EPCs on surfaces treated by D-biotin (A; control), LXW7 (B) after 20 min incubation. C. Quantification and the correlative statistical analysis of remaining cells shown in A-B. D-E. Representative images of adhered EPCs on untreated (D; control) or LXW7-DS-SILY modified SIS surface (E) after 20 min incubation. F. Quantification and the correlative statistical analysis of remaining cells shown in D-E. G. Growth and viability of ZDF-EPCs on LXW7 treated tissue culture surfaces and D-biotin treated surfaces (control) was assessed by CCK-8 assay. H. Growth and viability of ZDF-EPCs on LXW7-DS-SILY or DS-SILY treated SIS scaffold surfaces was assessed by CCK-8 assay. Scale bar = 200 μm. Data are expressed as mean ± SD. ****p < 0.0001, **p < 0.01, *p < 0.05, n = 3.

Article Snippet: Before purifying, LXW7 was cyclized by oxidizing cysteine residues using ClearOx resin (Peptides International) according to manufacturer’s protocol.

Techniques: Modification, Control, Incubation, CCK-8 Assay

Journal: Bioactive materials

Article Title: Bioactive extracellular matrix scaffolds engineered with proangiogenic proteoglycan mimetics and loaded with endothelial progenitor cells promote neovascularization and diabetic wound healing.

doi: 10.1016/j.bioactmat.2021.08.017

Figure Lengend Snippet: Fig. 4. SIS/LXW7-DS-SILY scaffolds accelerate wound closure and support ZDF-EPC survival under non-ischemic condition. A. Representative images of healing in wounds treated with different groups at days 0, 3, 7, 11 and 14. B. Quantification of the healing rate of different groups. a) SIS/LXW7-DS-SILY group was significantly better compared to SIS/DS-SILY and SIS groups. b) SIS/LXW7-DS-SILY/EPCs group was significantly better than the SIS/DS-SILY/EPCs and SIS/EPCs groups. c) SIS/LXW7-DS-SILY/EPCs group compared to SIS/LXW7-DS-SILY group showed no significant difference. d) SIS/DS-SILY/EPCs group was signifi cantly better compared to SIS/DS-SILY group. e) There was a trend that the SIS/EPCs group was better than the SIS group but the difference was not statis tically significant. Data are expressed as mean ± SEM. SIS group, SIS/LXW7-DS-SILY group, and SIS/EPCs group: n = 8; SIS/DS-SILY group: n = 7; SIS/DS-SILY/ EPCs group: n = 12; SIS/LXW7-DS-SILY/EPCs group: n = 13. C. Bioluminescence imaging of wounds transplanted with different modified SIS scaffolds loaded with Td-Tomato/luciferin-labeled ZDF-EPCs. D. Quantification of the signal intensity of biolumi nescence (Most of the SIS scaffolds spontaneously fall off the wounds by day 11). Data are expressed as mean ± SEM. n = 5 per group. ***p < 0.001, **p < 0.01, *p < 0.05.

Article Snippet: Before purifying, LXW7 was cyclized by oxidizing cysteine residues using ClearOx resin (Peptides International) according to manufacturer’s protocol.

Techniques: Imaging, Modification, Labeling

Journal: Bioactive materials

Article Title: Bioactive extracellular matrix scaffolds engineered with proangiogenic proteoglycan mimetics and loaded with endothelial progenitor cells promote neovascularization and diabetic wound healing.

doi: 10.1016/j.bioactmat.2021.08.017

Figure Lengend Snippet: Fig. 7. SIS/LXW7-DS-SILY constructs accelerate wound closure and support survival of ZDF-EPCs under ischemic condition. A. Representative images of healing in wounds treated with different groups at days 0, 3, 7, 11 and 14. B. Quantification of the wound closure rates of different groups. a) SIS/LXW7-DS-SILY group was significantly better compared to SIS group. There was a trend that the SIS/DS-SILY group was better than the SIS group but the difference was not statistically significant. b) SIS/LXW7-DS-SILY/EPCs group was significantly better than the SIS/DS-SILY/EPCs and SIS/EPCs groups. c) SIS/LXW7-DS-SILY/EPCs group was significantly better compared to SIS/LXW7-DS- SILY group. d) SIS/DS-SILY/EPCs group compared to SIS/DS-SILY group showed no significant differ ence. e) SIS/EPCs group compared to SIS group showed no significant difference. Data are expressed as mean ± SEM. SIS group, SIS/LXW7-DS-SILY group, and SIS/EPCs group: n = 8; SIS/DS-SILY group: n = 7; SIS/DS-SILY/EPCs group: n = 11; SIS/LXW7-DS- SILY/EPCs group: n = 12. C. Bioluminescence image of wounds transplanted with different modified scaf folds loaded with ZDF-EPCs. D. Quantification of the bioluminescence signals intensity (Most of the SIS scaffolds spontaneously fall off the wounds by day 11). Data are expressed as mean ± SEM. n = 5 per group. **p < 0.01, *p < 0.05.

Article Snippet: Before purifying, LXW7 was cyclized by oxidizing cysteine residues using ClearOx resin (Peptides International) according to manufacturer’s protocol.

Techniques: Construct, Modification

Journal: Molecular therapy : the journal of the American Society of Gene Therapy

Article Title: Self-Replicating RNAs Drive Protective Anti-tumor T Cell Responses to Neoantigen Vaccine Targets in a Combinatorial Approach.

doi: 10.1016/j.ymthe.2020.11.027

Figure Lengend Snippet: Figure 1. Ancer-Designed Polytope Neoantigen Constructs Expressed in the SMARRT Platform Generate Polyfunctional CD4 and CD8 T Cell Responses upon Vaccination (A) Schematic showing polytope design considerations following neoantigen identification. SMARRT replicons expressing neoantigen cassettes designed with Ancer were injected into BALB/c mice at a single dose of 10 mg, and spleens were removed 14 days later and restimulated with a peptide pool containing the top 50 Ancer predicted neoantigens from CT26. T cell function was measured by IFN-g ELISpot (B), and intracellular cytokine staining is shown for selected replicons (C). Graphs show mean with standard deviation, n = 5 mice per group. Statistical testing was carried out with ordinary one-way ANOVA. *p < 0.05; **p < 0.01; ****p < 0.0001.

Article Snippet: The top 20 neoantigen, predicted by Ancer, were used individually at 10 mg/mL for immunodominance studies (New England Peptide).

Techniques: Construct, Expressing, Injection, Immunopeptidomics, Cell Function Assay, Enzyme-linked Immunospot, Staining, Standard Deviation

Journal: Molecular therapy : the journal of the American Society of Gene Therapy

Article Title: Self-Replicating RNAs Drive Protective Anti-tumor T Cell Responses to Neoantigen Vaccine Targets in a Combinatorial Approach.

doi: 10.1016/j.ymthe.2020.11.027

Figure Lengend Snippet: Figure 2. SMARRT.Ancer Primes a Limited Number of Dominant T Cell Epitopes SMARRT.Ancer (containing the top 20 CT26 neoantigens) was used to immunize BALB/c mice with varying prime/ boost interval lengths. Splenocytes were analyzed by intracellular cytokine staining (on the indicated days post- final injection) by restimulating individually with all 20 neo- antigens encoded in the cassette. (A) and (B) show the percentage of IFN-g+ CD8 and CD4 T cells specific for peptides stimulating a response above mock (data not shown). (C) and (D) show the polyfunctionality of CD8 and CD4 T cells in response to the dominant neoantigen 20 and 4, respectively. Graphs show mean with standard deviation, n = 5 mice per group. Statistical analysis was performed using one-way ANOVA with Tukeys multiple comparison test. For (C) and (D), the triple functional sub- sets were compared. *p < 0.05; **p < 0.01; ****p < 0.0001

Article Snippet: The top 20 neoantigen, predicted by Ancer, were used individually at 10 mg/mL for immunodominance studies (New England Peptide).

Techniques: Immunopeptidomics, Staining, Injection, Standard Deviation, Comparison, Functional Assay

Journal: Molecular therapy : the journal of the American Society of Gene Therapy

Article Title: Self-Replicating RNAs Drive Protective Anti-tumor T Cell Responses to Neoantigen Vaccine Targets in a Combinatorial Approach.

doi: 10.1016/j.ymthe.2020.11.027

Figure Lengend Snippet: Figure 3. SMARRT Can Prime Polyfunctional T Cell Responses Specific to the Shared Neoantigen KRAS G12V HLA-A*1101 transgenic mice were vaccinated with SMARRT replicons encoding an epitope from either WT or G12V mutant KRAS and boosted on days 21 and 41. Spleens were analyzed 7 days post final boost by intracellular cytokine staining (A) and IFN-g ELISpot (B) following restimulation ex vivo with the indicated peptides. Graphs show mean with standard deviation, n = 3 mice per group. Statistical analysis was performed using Mann- Whitney U Test, panel B compared double cytokine posi- tive T cells. *p<0.05; ****p<0.0001.

Article Snippet: The top 20 neoantigen, predicted by Ancer, were used individually at 10 mg/mL for immunodominance studies (New England Peptide).

Techniques: Transgenic Assay, Mutagenesis, Staining, Enzyme-linked Immunospot, Ex Vivo, Standard Deviation, MANN-WHITNEY