Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Combining photodynamic therapy and cascade chemotherapy for enhanced tumor cytotoxicity: the role of CTT 2 P@B nanoparticles

doi: 10.3389/fbioe.2024.1361966

Figure Lengend Snippet: ROS generation detection in vitro . (A) The release of singlet oxygen from each drug was detected in vitro under dark conditions. Data were presented as the mean ± SD ( n = 3). (B) Detection of singlet oxygen release of each drug under 660 nm laser irradiation at a power density of 280 mW cm -2 was performed in vitro . Data were presented as the mean ± SD ( n = 3). (C) ROS generation was observed in 4T1 cells treated with each medication while being exposed to laser irradiation at a wavelength of 660 nm and intensity of 280 mW cm −2 . Bright: Bright field. DCF: Green fluorescence represents intracellular ROS. Merge: Superimpose the image. Scale Bar = 100 μm.

Article Snippet: The murine breast cancer cells (4T1) were procured from Icell Biotech Co., Ltd. (Shanghai, China).

Techniques: In Vitro, Irradiation, Fluorescence

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Combining photodynamic therapy and cascade chemotherapy for enhanced tumor cytotoxicity: the role of CTT 2 P@B nanoparticles

doi: 10.3389/fbioe.2024.1361966

Figure Lengend Snippet: Evaluation of mitochondrial targeting function. (A) Confocal images of the mitochondrial sites of TPPOH 2 , CTT 2 , CTT 2 P, and CTT 2 P@B NPs in 4T1 cells. Mito-Tracker Green was used to stain the mitochondria in the green channel. The red channel was derived from the emission of the photosensitizer fraction (PS) itself. Merge stands for superimposed image. The Green and red curves in the Plot Profile represent the gray value of Mito-Tracker Green and PS, respectively. Scale bar = 20 μm. (B) Flow cytometry JC-1 method was used to analyze the mitochondrial function of cells treated with different drugs. Red fluorescence: normal mitochondria (J-aggregate); Green fluorescence: depolarized mitochondria (J-monomer).

Article Snippet: The murine breast cancer cells (4T1) were procured from Icell Biotech Co., Ltd. (Shanghai, China).

Techniques: Staining, Derivative Assay, Flow Cytometry, Fluorescence

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Combining photodynamic therapy and cascade chemotherapy for enhanced tumor cytotoxicity: the role of CTT 2 P@B nanoparticles

doi: 10.3389/fbioe.2024.1361966

Figure Lengend Snippet: Evaluation of cell death in vitro through cytotoxicity assessment and examination of apoptosis and necrosis. (A) The in vitro cytotoxicity of 4T1 cells treated with CPT, TPPOH 2 , CTT 2 , CTT 2 P, and CTT 2 P@B NPs in the dark was assessed using the CCK-8 assay. Data were presented as the mean ± SD ( n = 5). ** p < 0.01, **** p < 0.0001. (B) The in vitro cytotoxicity of 4T1 cells treated with TPPOH 2 , CTT 2 , CTT 2 P and CTT 2 P@B NPs under laser irradiation (660 nm, 280 mW cm −2 ) was determined using the CCK-8 assay. Data were presented as the mean ± SD ( n = 5). ** p < 0.01, **** p < 0.0001. (C) Cell apoptosis and necrosis were analyzed using flow cytometry with Annexin V-FITC/PI double staining following treatment with various drugs at a concentration of 5 μM.

Article Snippet: The murine breast cancer cells (4T1) were procured from Icell Biotech Co., Ltd. (Shanghai, China).

Techniques: In Vitro, CCK-8 Assay, Irradiation, Flow Cytometry, Double Staining, Concentration Assay

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Combining photodynamic therapy and cascade chemotherapy for enhanced tumor cytotoxicity: the role of CTT 2 P@B nanoparticles

doi: 10.3389/fbioe.2024.1361966

Figure Lengend Snippet: Biodistribution in vivo . (A) Blood compatibility test of CPT, TPPOH 2 , CTT 2 , CTT 2 P, CTT 2 P@B NPs. Data were presented as the mean ± SD ( n = 3). *** p < 0.001, **** p < 0.0001. (B) Time-lapse live fluorescence imaging of mice with 4T1 tumors following the administration of free CTT 2 P and CTT 2 P@B NPs via intravenous injection. (C) Fluorescent images of major organs and tumors were obtained 24 h after injection, using ex vivo methods. (D) The mean fluorescence intensity of each organ and tumor was used to determine the biodistribution of free CTT 2 P and CTT 2 P@B NPs in mice. Data were presented as the mean ± SD ( n = 3). * p < 0.05.

Article Snippet: The murine breast cancer cells (4T1) were procured from Icell Biotech Co., Ltd. (Shanghai, China).

Techniques: In Vivo, Fluorescence, Imaging, Injection, Ex Vivo

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Combining photodynamic therapy and cascade chemotherapy for enhanced tumor cytotoxicity: the role of CTT 2 P@B nanoparticles

doi: 10.3389/fbioe.2024.1361966

Figure Lengend Snippet: In vivo anti-tumor study of each drug in 4T1 tumor-bearing mice. (A) Tumor images of different drug administration treatments after the antitumor study. (B) During the administration, the growth of tumors in mice was observed in each group receiving treatment. Data were presented as the mean ± SD ( n = 5), ** p < 0.01. (C) The weight of mice in each treatment group was monitored throughout the administration period. Data were presented as the mean ± SD ( n = 5).

Article Snippet: The murine breast cancer cells (4T1) were procured from Icell Biotech Co., Ltd. (Shanghai, China).

Techniques: In Vivo

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Combining photodynamic therapy and cascade chemotherapy for enhanced tumor cytotoxicity: the role of CTT 2 P@B nanoparticles

doi: 10.3389/fbioe.2024.1361966

Figure Lengend Snippet: Investigation of the effects of each medication on mice with 4T1 tumors through pathological examination. (A) After administering various medications, the major organs (including the heart, liver, spleen, lung, and kidney) were subjected to H&E staining. Scale bar = 50 μm. (B) After administering various medications, tumors were subjected to H&E staining and TUNEL staining. Scale bar = 50 μm.

Article Snippet: The murine breast cancer cells (4T1) were procured from Icell Biotech Co., Ltd. (Shanghai, China).

Techniques: Medications, Staining, TUNEL Assay

Journal: PLoS Genetics

Article Title: Host CLIC4 expression in the tumor microenvironment is essential for breast cancer metastatic competence

doi: 10.1371/journal.pgen.1010271

Figure Lengend Snippet: ( A-C ) 1×10 5 6DT1 cells were implanted into the fourth left mammary fat pad of Clic4 wildtype (WT), n = 15, heterozygous (HET), n = 29, and knockout (KO), n = 22, FVB female mice. Primary tumors were resected and weighed (g = grams) ( A ), and superficial lung metastases quantified ( B ) after 28 days. ( C ) Immunohistochemistry for CLIC4 expression in 6DT1 primary tumors and superficial lung metastases from individual mice at 28 days post-implantation. ( D-F ) Primary tumor weight (g = grams) ( D ), number of lung metastases ( E ), and CLIC4 immunohistochemistry ( F) 28 days after implanting 2.5×10 5 E0771 cells into Clic4 wildtype (WT), n = 22, heterozygous (HET), n = 12, and knockout (KO), n = 25, C57BL/6 female mice. Values represent the mean ± s.d. P values were obtained after ANOVA followed by unpaired t test. Scale bar, 4 mm.

Article Snippet: The murine breast cancer cell line 6DT1 (FVB background) was grown as previously described [ ] and E0771 (C57BL/6 background) cells were purchased from CH3 BioSystems LLC and grown according to the manufacturer’s recommendations (Amherst, NY, No. 940001).

Techniques: Knock-Out, Immunohistochemistry, Expressing

Journal: PLoS Genetics

Article Title: Host CLIC4 expression in the tumor microenvironment is essential for breast cancer metastatic competence

doi: 10.1371/journal.pgen.1010271

Figure Lengend Snippet: Fourteen days after implantation of 1×10 5 6DT1 cells into the fourth left mammary fat pad of Clic4 wildtype (WT), n = 8, and knockout (KO), n = 8, FVB females, 1×10 5 GFP-labeled 6DT1 cells were injected via tail vein. ( A ) GFP immunostaining detects individual tail vein-injected GFP-labeled 6DT1 cells (red arrows) in lungs after 48 hours. Scale bar, 50 μm. ( B-C ) Primary tumor weight (g = grams) ( B ) and number of superficial lung metastases ( C ) 14 days post-tail vein injection. Values are means ± s.d., P values were determined by unpaired t test. ( D ) GFP-positive metastases, mixed GFP-positive and -negative metastases (black arrows), or residual single cells (red arrows) in lungs 14 days after tail vein injection of GFP-labeled 6DT1 cells. Scale bars, 5 mm (top) and 50 μm (bottom). Insert in bottom right panel, micro metastasis.

Article Snippet: The murine breast cancer cell line 6DT1 (FVB background) was grown as previously described [ ] and E0771 (C57BL/6 background) cells were purchased from CH3 BioSystems LLC and grown according to the manufacturer’s recommendations (Amherst, NY, No. 940001).

Techniques: Knock-Out, Labeling, Injection, Immunostaining

Journal: PLoS Genetics

Article Title: Host CLIC4 expression in the tumor microenvironment is essential for breast cancer metastatic competence

doi: 10.1371/journal.pgen.1010271

Figure Lengend Snippet: ( A ) Deletion of Clic4 in 6DT1 tumor cells (Parental) by CRISPR/Cas9 was confirmed by immunoblot. Two independent Clic4 wildtype (sgNT) and Clic4 knockout (sg Clic4 ) clones were selected. Densitometry-based quantification is shown below each protein band, normalized to the loading control (HSP90). NT = non-targeting. ( B ) 1×10 5 sgNT or sg Clic4 6DT1 cells were implanted into the fourth left mammary fat pad of Clic4 wildtype FVB females. Primary tumor weight (g = grams) and number of superficial lung metastases were evaluated 28 days post-implantation of the clonal sublines sgNT-1 (n = 15), sgNT-2 (n = 20), sg Clic4 -1 (n = 15), and sg Clic4 -2 (n = 20). Parental cell-derived tumors (n = 18; gray background) are shown for reference. P values were obtained using one-way ANOVA with comparisons between all clonal lines and the Sidak correction for multiple comparisons. ( C-D ) Representative immunohistochemistry showing the tissue distribution of CLIC4 expression in primary tumor and lung metastases from tumor-bearing mice at 28 days post-implantation of 1×10 5 sgNT or sg Clic4 6DT1 cells. Scale bar, 2 mm ( C ) and 30 μm ( D ). Yellow arrows, tumor stromal compartment. White arrows, tumor epithelial compartment.

Article Snippet: The murine breast cancer cell line 6DT1 (FVB background) was grown as previously described [ ] and E0771 (C57BL/6 background) cells were purchased from CH3 BioSystems LLC and grown according to the manufacturer’s recommendations (Amherst, NY, No. 940001).

Techniques: CRISPR, Western Blot, Knock-Out, Clone Assay, Control, Derivative Assay, Immunohistochemistry, Expressing

Journal: PLoS Genetics

Article Title: Host CLIC4 expression in the tumor microenvironment is essential for breast cancer metastatic competence

doi: 10.1371/journal.pgen.1010271

Figure Lengend Snippet: ( A ) Proteome Profiler Array heat map of 19 significantly differentially expressed cytokines and chemokines, from 111 tested, present in plasma from healthy control (0 days) (n = 3) or 6DT1 tumor-bearing Clic4 wildtype (WT) or knockout (KO) mice at 14 days (n = 3) or 28 days (n = 5) after mammary fat pad implantation. Raw values were obtained by determining the pixel intensity for each analyte followed by normalization to the corresponding internal control for each animal. These values were log2-transformed, and the heat map represents the mean log2 fold-change of each group from the WT healthy control group. P values were determined by unpaired t test comparing the raw values of WT and KO at each time point; *P value < 0.05. ( B-C ) Luminex Array heat map of 35 significantly differentially expressed proteins, from 51 tested, present in the primary tumors and lungs of Clic4 wildtype (WT) or knockout (KO) mice at 0 days (lung only, n = 5), 14 days (n = 8), or 28 days (n = 5) after 6DT1 tumor cell implantation in the mammary fat pad. ( B ) Tumor protein expression values shown are log2 fold-change of KO from WT at 14 or 28 days. P values were determined by unpaired t test comparing the raw values of WT and KO at each time point; * P value < 0.05. ( C ) Lung protein expression values shown are log2 fold-change of WT or KO at 14 or 28 days post-implantation from their respective genotype-matched baseline (0 days) values. P values were determined by unpaired t test comparing the raw values of WT or KO at 14 or 28 days to their respective baseline values; *P value < 0.05. ( D-E ) Quantification of selectively stained infiltrating inflammatory cells in primary tumor and lung tissues from 6DT1 tumor-bearing Clic4 wildtype (WT) and knockout (KO) mice by IHC. Values represent the mean ± s.d. of the total number of positive cells found in tissue sections from 3 animals analyzed by Image Scope. F4/80 was used to detect macrophages, Ly6G/GR1 for neutrophils, and CD4 and CD8 for mature T cell subsets in primary tumors ( D ) and lung tissues ( E ). P values were obtained after unpaired t test between genotypes at each time point.

Article Snippet: The murine breast cancer cell line 6DT1 (FVB background) was grown as previously described [ ] and E0771 (C57BL/6 background) cells were purchased from CH3 BioSystems LLC and grown according to the manufacturer’s recommendations (Amherst, NY, No. 940001).

Techniques: Clinical Proteomics, Control, Knock-Out, Transformation Assay, Luminex, Expressing, Staining

Journal: PLoS Genetics

Article Title: Host CLIC4 expression in the tumor microenvironment is essential for breast cancer metastatic competence

doi: 10.1371/journal.pgen.1010271

Figure Lengend Snippet: ( A ) Significant (FDR <0.25) Hallmark pathways identify transcriptomic changes that occur in the lungs of 6DT1 tumor-bearing Clic4 wildtype (WT) or knockout (KO) mice relative to the lungs of genotype-matched healthy control mice 14 days after implantation. Gene set enrichment analysis (GSEA) normalized enrichment scores (NES) are shown. Group 1, similar regulation between WT and KO; Group 2, significant for WT only; Group 3, significant for KO only; Group 4, opposite regulation between WT and KO. ( B ) Gene set enrichment analysis (GSEA) to identify significant enrichment (FDR <0.25) in Hallmark pathways based on genes differentially expressed in KO vs. WT lungs at 14 days after primary tumor implantation. ( C ) Enrichment plots for the “TNFA signaling via NFKB” Hallmark pathway for the three comparisons shown in panels A and B. The ten most significant leading edge genes are listed to the right of each plot.

Article Snippet: The murine breast cancer cell line 6DT1 (FVB background) was grown as previously described [ ] and E0771 (C57BL/6 background) cells were purchased from CH3 BioSystems LLC and grown according to the manufacturer’s recommendations (Amherst, NY, No. 940001).

Techniques: Knock-Out, Control, Tumor Implantation

Journal: Rsc Advances

Article Title: Detecting hypoxia in vitro using 18 F-pretargeted IEDDA “click” chemistry in live cells †

doi: 10.1039/d1ra02482e

Figure Lengend Snippet: The in vitro uptake of [ 18 F]FB-Tz in (A) EMT6 and (B) HCT116 cells. Cells were treated with 8 (0.01–10 μM) and cultured under normoxia or hypoxia (0.1% O 2 ) conditions for 24 h prior to radioactive uptake experiment: 0.74 MBq per well of [ 18 F]FB-Tz was added to cells and incubated for 60 min. (C) Time course of [ 18 F]FB-Tz uptake in EMT6 cells in vitro . Cells were treated with 8 (10 μM) for 24 h under hypoxia before incubation with 0.74 MBq per well of [ 18 F]FB-Tz for 5, 10, 15, 30, 45 and 60 min. (D) Radioactive uptake of [ 18 F]FMISO (1.48 MBq per well) for 0 min (pulse) or 3 h under normoxia or hypoxia conditions in EMT6 and HCT116. Data were expressed as a percentage of total radioactivity incorporated into cells, normalized for total cellular protein. Mean values ± SD ( n = 6; * P < 0.05, ** P < 0.01 and **** P < 0.0001).

Article Snippet: EMT6 murine breast cancer cells were a kind gift from Prof. Sofia Pascu (University of Bath, UK) and were grown in Waymouth medium (Thermo Fisher Scientific); HCT116 human colorectal cancer cells (ATCC) were grown in Dulbecco Modified Eagle Medium (DMEM, Sigma-Aldrich).

Techniques: In Vitro, Cell Culture, Incubation, Radioactivity

Journal: Nature Communications

Article Title: A vaccine-based nanosystem for initiating innate immunity and improving tumor immunotherapy

doi: 10.1038/s41467-020-15927-0

Figure Lengend Snippet: a Expressions of tumor immune regulation-associated genes in 1097 human breast tumors and 114 para-carcinoma tissues from UALCAN ( http://ualcan.path.uab.edu/index.html ). Statistical significance was calculated with two-tailed Student’s t test. b The correlation between the CD3E / IRF7 expressions and the survival of breast tumor patients from TCGA. Statistical significance was calculated with two-tailed Student’s t test. c Schematic diagram for the design and function process of the siRNA@HPVP system. Before the encapsulation of siRNA, the HPV16 L1 capsid was disassembled by DTT. Disassembled HPV16 L1 protein mixed with siRNA and PEG was reassembled into nanoparticles through the removal of DTT via dialysis. The HPVP was expected to initiate an immune response in an Irf7 -dependent manner (mainly in APCs), while the knockdown of the Cd274 gene (mainly in cancer cells) promoted the lymphocyte infiltration. d PCR analysis of Cd274 transcription in 4T1 cells after silenced by three siRNA sequences (100 nmol mL −1 ). The gene silencing efficiency of siRNA2 on 4T1 cells and RAW 267.4 cells. Three biological replicates are shown. Statistical significance was calculated with one-way ANOVA with Tukey post-hoc (left) and two-tailed Student’s t test (middle and right). e Gel electrophoresis of siRNA@HPVP at different mass ratios between HPVP and siRNA, naked siRNA was set as control. f Cryo-TEM images of siRNA@HPVP. A representative image of two biological replicates is shown. g Flow cytometry data of the uptake of siRNA@HPVP in 4T1 cells or 4T1 Itga6 - cells. The intracellular fluorescence intensity was measured at 6 h, 12 h, and 24 h of the co-incubation. Three biological replicates are shown. Statistical significance was calculated with two-tailed Student’s t test. Data are presented as mean values ± SD.

Article Snippet: 3T3 mouse embryonal fibroblasts, B16 murine melanoma cells, 4T1 murine breast tumor cells, CT26 murine colon tumor cells, Raw 267.4 murine macrophages, and HEK-293 cells were obtained from China Center for Type Culture Collection (CCTCC).

Techniques: Two Tailed Test, Encapsulation, Knockdown, Nucleic Acid Electrophoresis, Control, Flow Cytometry, Fluorescence, Incubation

Journal: Nature Communications

Article Title: A vaccine-based nanosystem for initiating innate immunity and improving tumor immunotherapy

doi: 10.1038/s41467-020-15927-0

Figure Lengend Snippet: a In vitro cytotoxicity of splenocytes towards 4T1 cells (with the effector/target ratio of 10:1). Splenocytes were pretreated with different samples (PBS, HPVP, siRNA1, siRNA2, siRNA3, siRNA1@HPVP, siRNA2@HPVP, or siRNA3@HPVP) for 24 h before the cytotoxicity assay. Five biological replicates are shown. b FACS data of the mature DCs (CD80 + CD86 + ) gating on CD11c + cells after co-incubated with different siRNA@HPVP formats (siRNA1@HPVP, siRNA2@HPVP, or siRNA3@HPVP), HPVP or fresh culture medium for 24 h. A representative image of three biological replicates is shown. c Transcript abundance of IFN-α and IFN-γ in DCs after different treatments. Three biological replicates are shown. d , e FACS data to exhibit the phenotype and activation proportion of macrophages (CD11b + CD80 + ) 6 h after different treatments. M1 macrophages (CD11b + CD206 − ) were induced by lipopolysaccharide (100 ng mL −1 ). M2 macrophages (CD11b + CD206 + ) were induced by IL4 (20 ng mL −1 ). A representative image of three biological replicates is shown. f Cytokine levels in the culture medium of RAW 264.7 macrophages after different treatments. The concentration of HPVP for all in vitro immune stimulate assays was 20 mg L −1 . Three biological replicates are shown. Statistical significance was calculated via one-way ANOVA with Tukey post-hoc analysis ( a , c , f ). Data are presented as mean values ± SD.

Article Snippet: 3T3 mouse embryonal fibroblasts, B16 murine melanoma cells, 4T1 murine breast tumor cells, CT26 murine colon tumor cells, Raw 267.4 murine macrophages, and HEK-293 cells were obtained from China Center for Type Culture Collection (CCTCC).

Techniques: In Vitro, Cytotoxicity Assay, Incubation, Activation Assay, Concentration Assay

Journal: Nature Communications

Article Title: A vaccine-based nanosystem for initiating innate immunity and improving tumor immunotherapy

doi: 10.1038/s41467-020-15927-0

Figure Lengend Snippet: a In vivo tumor-targeting capacity of HPVP on 4T1 tumor-bearing mice after i.v . injection. The blockade of α6 integrin with antibody reduced the tumor accumulation of HPVP. A representative image of three biological replicates is shown. b Ex vivo immunofluorescence images for the evaluation of PDL1 inhibition effect of siRNA1@HPVP, siRNA2@HPVP, and siRNA3@HPVP (Scale bar: 1 mm). A representative image of three biological replicates is shown. c Bioluminescence assay for the in situ measurement of innate immunity in 4T1 tumors after treated with siRNA2@HPVP. Tumor cells were transfected with a pTNF-α-promoter-luc plasmid for reporting the intratumoral TNF-α level. A representative image of three biological replicates is shown. d In vivo anticancer effect evaluation of siRNA2@HPVP and HPVP + aPDL1 on the subcutaneous murine breast tumor model. The tumor size was measured every other day. Five biological replicates are shown. e FACS analysis for measuring the expression of PDL1 protein in PBS- or siRNA2@HPVP-treated mice tumors. Three biological replicates are shown. f Quantitative analysis of Ki67 + tumor cells after different treatments. Four images per group were taken. g FACS data of the mature DCs (CD11c + CD80 + CD86 + ) within lymph nodes (left) and tumor-infiltrating cytotoxic T cells (CD3 + CD8 + T cells, right) after different treatments. Three biological replicates (right) and four biological replicates (left) are shown, respectively. h FACS analysis for measuring intratumoral M1/M2 macrophages ratio after different treatments. CD11b + CD206 + cells were defined as M2 macrophages, while CD11b + CD206 − cells were defined as M1 macrophages. Four biological replicates are shown. i IFN levels in the serum from the mice 12 h after different treatments. Three biological replicates are shown. Statistical significance was calculated via one-way ANOVA with a Tukey post-hoc test ( d – i ). Data are presented as mean values ± SD.

Article Snippet: 3T3 mouse embryonal fibroblasts, B16 murine melanoma cells, 4T1 murine breast tumor cells, CT26 murine colon tumor cells, Raw 267.4 murine macrophages, and HEK-293 cells were obtained from China Center for Type Culture Collection (CCTCC).

Techniques: In Vivo, Injection, Ex Vivo, Immunofluorescence, Inhibition, ATP Bioluminescent Assay, In Situ, Transfection, Plasmid Preparation, Expressing

Journal: Nature Communications

Article Title: A vaccine-based nanosystem for initiating innate immunity and improving tumor immunotherapy

doi: 10.1038/s41467-020-15927-0

Figure Lengend Snippet: a Identification of differentially expressed genes in 4T1 tumors 3 days after intratumoral injection with siRNA2@HPVP or aPDL1. Differential genes in the GO term of the immune process were labeled. Three biological replicates are shown. b GeneMANIA for predicting gene interactions between differential genes in the GO term of the immune process. c KEGG enrichment analysis of the pathways involved in the biological effect induced by siRNA2@HPVP treatment. d PCR analysis of the transcript abundance of Tlr7 , Cxcl9 , Infb1 , Ifngr1 , and Ifng genes in 4T1 tumors after different treatments. Three biological replicates are shown. e Schematic diagram of the possible mechanism for siRNA2@HPVP to activate the innate immunity. siRNA2@HPVP activates APCs via the TLR7-mediate pathway, and then recruits T H 1 cells for anticancer effect via their inherent immunogenicity and IFNs pathway. f HEK-293 cells transiently expressing TLR homodimers and ELAM-Luc reporter plasmid proved that the immune response mediated by siRNA2@HPVP is TLR7-dependent. Three biological replicates are shown. g Blockade of type I IFN pathway with anti-IFNAR1 antibody (aifnar, 200 μg per mice, i.p .) suppressed the therapeutic effect of siRNA2@HPVP (7.5 mg kg −1 ) treatment. Tumor size was recorded every other day (left, four biological replicates are shown). The proportion of tumor-infiltrating CD8 + T lymphocytes reduced when the IFN pathway was blocked (right, four biological replicates are shown). h Quantitative analysis of T-bet (left) and CD4 (right) expressions in 4T1 tumors through the immunofluorescence staining after different treatments. Tumor tissues were obtained 15 days after different treatments. Six images per group were taken. Statistical significance was calculated with two-tailed Student’s t test ( a ) one-way ANOVA with Tukey post-hoc analysis ( d , f , g , h ). Data are presented as mean values ± SD.

Article Snippet: 3T3 mouse embryonal fibroblasts, B16 murine melanoma cells, 4T1 murine breast tumor cells, CT26 murine colon tumor cells, Raw 267.4 murine macrophages, and HEK-293 cells were obtained from China Center for Type Culture Collection (CCTCC).

Techniques: Injection, Labeling, Immunopeptidomics, Expressing, Plasmid Preparation, Immunofluorescence, Staining, Two Tailed Test

Journal: Nature Communications

Article Title: A vaccine-based nanosystem for initiating innate immunity and improving tumor immunotherapy

doi: 10.1038/s41467-020-15927-0

Figure Lengend Snippet: a Different tumor models established in this study for evaluating the anticancer effect of siRNA2@HPVP treatment. b In vivo bioluminescence images to track the recurrence and metastasis of 4T1 tumors after the resection of the primary tumor. The resection operation was conducted 15 days after different treatments. Three representative images of five biological replicates are shown. c The data of the lesion condition in the resectable murine breast tumor model after the resection of primary tumor. s@H represents siRNA2@HPVP. d The proportion of T EM cells (CD8 + CD44 + CD62L − ) in spleen tissues 40 days after different treatments. The primary tumor was removed by surgery at day 15. Three biological replicates are shown. e In vivo bioluminescence images to exhibit the anti-metastasis effect of various treatments. The metastatic breast tumor model was constructed by intravenously injecting BABL/C mice with 4T1 luc cells. The treatments were carried out 24 h after the tumor cells injection. Three representative images of five biological replicates are shown. f , g H&E staining and quantitative counting of metastatic nodes in lung tissues 21 days after the i.v . injection of 4T1 cells (Scale bar: 5 mm). Five images per group were taken. h In vivo bioluminescence images of orthotopic breast tumor-bearing mice after different treatments. In this assay, aPDL1 treatment as a clinical treatment for the unresectable tumor was also conducted. Three representative images of six biological replicates are shown. i Representative μ-CT images for the visualization of bone metastasis of orthotopic breast tumor after various treatments. Sites of osteolysis are circled. j Survival curve of orthotopic breast tumor-bearing mice with various treatments. Six biological replicates are shown. Statistical significance was calculated via one-way ANOVA with Tukey post-hoc test ( d , f ) and Logrank test (two-sided) for trend ( j ). Data are presented as mean values ± SD.

Article Snippet: 3T3 mouse embryonal fibroblasts, B16 murine melanoma cells, 4T1 murine breast tumor cells, CT26 murine colon tumor cells, Raw 267.4 murine macrophages, and HEK-293 cells were obtained from China Center for Type Culture Collection (CCTCC).

Techniques: In Vivo, Construct, Injection, Staining

Journal: Nature Communications

Article Title: A vaccine-based nanosystem for initiating innate immunity and improving tumor immunotherapy

doi: 10.1038/s41467-020-15927-0

Figure Lengend Snippet: a Schematic illustration of the CRISPR-Cas9-mediated Mlh1 knockout for large-scale gene mutations. Seven monoclonal 4T1 cell lines with different gene mutations were purified. Five cell lines were respectively inoculated on the mice to evaluate the immune response rate of siRNA2@HPVP treatment. b SNP (left) and Indel (right) analyses of the obtained seven mutant cell lines. The union graph of total mutation among five mutant cell lines was constructed to evaluate the connection between the five cell lines. c AS analysis of the mutation of the cell lines after Mlh1 knockout. d – h Tumor growth curves (left, five biological replicates are shown), tumor-infiltrating T cells proportions (middle, three biological replicates are shown) and survival curves (right, five biological replicates are shown) of five distinct breast tumor models with aPDL1 plus PTX (1 mg kg −1 aPDL1 and 10 mg kg −1 PTX) or siRNA2@HPVP plus PTX (7.5 mg kg −1 siRNA2@HPVP and 10 mg kg −1 PTX) treatment by i.v . injection ( n = 5 per group). Three tumors per group were collected for flow cytometry analysis. i Arc plot and heat map for illustrating the intratumoral immune indexes (CD3, CD4, CD8, Foxp3, IL2, T-bet, and IFN-γ) in each individual. Two locations were taken for each tumor. Statistical significance was calculated via two-tailed Student’s t test ( b , d – g , i ) and Logrank test (two-sided) for trend ( d – g ). Data are presented as mean values ± SD.

Article Snippet: 3T3 mouse embryonal fibroblasts, B16 murine melanoma cells, 4T1 murine breast tumor cells, CT26 murine colon tumor cells, Raw 267.4 murine macrophages, and HEK-293 cells were obtained from China Center for Type Culture Collection (CCTCC).

Techniques: CRISPR, Knock-Out, Purification, Mutagenesis, Construct, Injection, Flow Cytometry, Two Tailed Test

Journal: Journal of Advanced Research

Article Title: Innate tumor-targeted nanozyme overcoming tumor hypoxia for cancer theranostic use

doi: 10.1016/j.jare.2021.02.004

Figure Lengend Snippet: In vitro study. (A) T1-weighted MRI in vitro with various MnO 2 concentrations under pH 6.0 and pH 7.4. (B) Concentration-dependent longitudinal relaxation time T1, based on readings from A. (C) Cytotoxicity of HFn carrier toward HeLa, 4 T1, and SKOV3 cancer cells. (D-F) Cell viability of three cell lines under hypoxic conditions determined by MTT assay exposed to various concentrations of nanomaterials for 48 h. The Dox concentration axis represents the quantity of Dox in each group for Dox contained formulations as well as the MnO 2 concentration axis for MnO 2 contained nanomaterials. (G) IC 50 values of various cancer cells based on Dox concentration (*p < 0.05, **p < 0.01 vs SKOV3). Data are presented as means ± SD (n = 3).

Article Snippet: The human cervical cancer cells (HeLa), human ovarian cancer cells (SKOV3) and 4 T1 murine breast cancer cells (4 T1) were originally obtained from the American Type Culture Collection (ATCC) and cultured with the recommended medium at 37 °C within 5% CO 2 atmosphere.

Techniques: In Vitro, Concentration Assay, MTT Assay

Journal: Journal of Advanced Research

Article Title: Innate tumor-targeted nanozyme overcoming tumor hypoxia for cancer theranostic use

doi: 10.1016/j.jare.2021.02.004

Figure Lengend Snippet: T1-MR images of 4 T1 tumor-bearing mice through (A) local and (C) intravenous injection of MnO 2 -Dox@HFn. (B) and (D) Quantification of T1-MRI signals via intratumorally and intravenously injection according to A and C, respectively (n = 3; **p < 0.01, ***p < 0.001 vs before). (E) Tumor growth in various treated groups (n = 3). (F) Tumor weight at the endpoint of the study (**p < 0.01). (G) Representative tumor images were harvested from mice after the treatment. (H) Tumor growth inhibition rate.

Article Snippet: The human cervical cancer cells (HeLa), human ovarian cancer cells (SKOV3) and 4 T1 murine breast cancer cells (4 T1) were originally obtained from the American Type Culture Collection (ATCC) and cultured with the recommended medium at 37 °C within 5% CO 2 atmosphere.

Techniques: Injection, Inhibition