Journal: Cancer Research

Article Title: Gene Therapy with Secretory Leukoprotease Inhibitor Promoter-Controlled Replication-Competent Adenovirus for Non-Small Cell Lung Cancer

doi: 10.1158/0008-5472.can-03-2549

Figure Lengend Snippet: Fig. 1. Secretion of secretory leukoprotease inhibitor (SLPI) from various human cell lines, construction of AdSLPI.E1AdB, and E1A protein expression and replication of adenovirus in cancer cells infected with AdSLPI.E1AdB in vitro. A, SLPI secretion from human cell lines. The level of SLPI protein in the medium from nine human cell lines [NHBE (normal human bronchial epithelium), A549 (lung adenocarcinoma), H358 (lung adenocarcinoma), HS24 (lung squamous cell carcinoma), SBC-3 (small cell lung cancer), HepG2 (hepatocellular carcinoma), HT1080 (sarcoma), SAOS-2 (osteosarcoma), and INR-90 (normal fibroblast)] was determined by using an ELISA kit for human SLPI protein. B, a construct of AdSLPI.E1AdB. E1A region controlled by the SLPI promoter and E1B-19K driven by the CMV promoter composed the expression cassette of the Ad5 vector. C, Western blotting of E1A protein. Cell lysates were subjected to Western blot analysis using anti-adenovirus E1A protein. D, Southern blotting of the E4 region. DNA extracted from A549 or HepG2 cells was analyzed by Southern blotting using E4 probe. C and D, A549 or HepG2 cells were infected with either AdSLPI.LacZ (Lanes 1), AdSLPI.E1AdB (Lanes 2), or AxE1AdB (Lanes 3).

Article Snippet: To evaluate the secretion of SLPI protein from the various cell lines, concentrations of SLPI protein in the culture medium after 72 h of incubation (1 106 cells/10-cm dish) were determined by an ELISA kit for human SLPI (Quantikine; R&D Systems, Inc.).

Techniques: Expressing, Infection, In Vitro, Enzyme-linked Immunosorbent Assay, Construct, Plasmid Preparation, Western Blot, Southern Blot

Journal: Cancer gene therapy

Article Title: Type I IFN gene delivery suppresses regulatory T cells within tumors.

doi: 10.1038/cgt.2014.60

Figure Lengend Snippet: Figure 1. Antitumor effect of intratumoral IFN-α gene transfer. (a) Growth of tumors injected with Ad-mIFN. Tumor volumes were measured at indicated days following the intratumoral injection of Ad-mIFN (n = 6) or Ad-AP (n = 8). Relative tumor volumes compared with those at day10 were presented. Data are shown as means ± standard deviation (s.d.). (b) ELISpot assay of IFN-γ-producing cells in response to stimulation of CT26 cells. Twenty-two days after tumor inoculation, splenocytes were isolated from mice injected with Ad-mIFN (n = 4) or Ad-AP (n = 3), and were cultured with CT26 or syngeneic lymphocytes. Data are presented as means ± s.d. (c) Intracellular cytokine staining of IFN-γ-producing cells in response to CT26 cells. The splenocytes from mice injected with Ad-mIFN (n = 4) or Ad-AP (n = 3) at day 22 were incubated with CT26 cells and stained by anti-mouse IFN-γ antibody. The activated cell fractions were analyzed by staining with anti-mouse CD8 antibody. Representative FACS plots (right panel) are shown.

Article Snippet: The amounts of cytokines in cell culture supernatants and tumors were assayed with antibodies for IL-6 and mouse IFN-α (Quantikine; R&D systems, Minneapolis, MN, USA) in accordance with the manufacturer’s recommendations.

Techniques: Injection, Standard Deviation, Enzyme-linked Immunospot, Isolation, Cell Culture, Staining, Incubation

Journal: Cancer gene therapy

Article Title: Type I IFN gene delivery suppresses regulatory T cells within tumors.

doi: 10.1038/cgt.2014.60

Figure Lengend Snippet: Figure 2. Intratumoral IFN-α gene transfer reduced the frequency of Tregs in tumors. (a) Frequency of CD4+Foxp3+ Tregs per CD4+ T cells in tumors. Tumors injected with viruses were harvested at days 16, 22 and 28, and processed into single-cell suspension. The percentage of CD4+

Article Snippet: The amounts of cytokines in cell culture supernatants and tumors were assayed with antibodies for IL-6 and mouse IFN-α (Quantikine; R&D systems, Minneapolis, MN, USA) in accordance with the manufacturer’s recommendations.

Techniques: Injection, Suspension

Journal: Cancer gene therapy

Article Title: Type I IFN gene delivery suppresses regulatory T cells within tumors.

doi: 10.1038/cgt.2014.60

Figure Lengend Snippet: Figure 3. Intratumoral IL-6 concentration was significantly increased by IFN-α gene transfer. (a) IL-6 concentration in tumors. Tumors injected with Ad-mIFN (n = 5) or Ad-AP (n = 5) were harvested at days 16, 22 and 28, and IL-6 concentration was measured by ELISA. (b) Relationship between IL-6 and IFN-α concentration in tumors. Tumors injected with Ad-mIFN (n = 5) or Ad-AP (n = 5) were harvested at day 16, and the concentrations of IL-6 and IFN-α were compared by ELISA. (c) IL-6 production from tumor CD11c+ cells. The CD11c+ and CD11c −cells were isolated from tumors injected with Ad-mIFN (n = 2) or Ad-AP (n = 2) at day 16, and 5 × 104 cells were plated in 96-well plates. After 48 h, supernatants were assayed for the measurement of IL-6 concentration by ELISA. (d) IL-6 production from splenic CD11c+ cells in response to a recombinant IFN-α protein. The CD11c+ and CD11c−cells isolated from naïve splenocytes, and 5 × 104 of CT26 cells were cultured in 96-well plates with depicted concentration of recombinant mouse IFN-α (Miltenyi Biotech). After 48 h, supernatants were assayed for the measurement of IL-6 concentration by ELISA (n = 2 for each group).

Article Snippet: The amounts of cytokines in cell culture supernatants and tumors were assayed with antibodies for IL-6 and mouse IFN-α (Quantikine; R&D systems, Minneapolis, MN, USA) in accordance with the manufacturer’s recommendations.

Techniques: Concentration Assay, Injection, Enzyme-linked Immunosorbent Assay, Isolation, Recombinant, Cell Culture

Journal: Cancer gene therapy

Article Title: Type I IFN gene delivery suppresses regulatory T cells within tumors.

doi: 10.1038/cgt.2014.60

Figure Lengend Snippet: Figure 4. IL-6 receptor blockade suppressed IFN-α-mediated Treg reduction in tumors. (a) Schema of experiment. The 1000 μg of the monoclonal anti-IL-6 receptor antibody was intraperitoneally injected into the mice at days 7, 14 and 21 after tumor inoculation. Ad-mIFN or Ad-AP was injected once at day 10 after inoculation. (b) Frequency of CD4+Foxp3+ cells per CD4+ T cells in tumors treated with IL-6R ab. Tumors were harvested at day 22, and CD4+ T cells and CD4+Foxp3+ Tregs were analyzed by flow cytometry (n = 5 for the group of Ad-AP i.t. +IL-6R ab i.p., n = 4 for the other groups). (c) Ratio of CD8+ T cells to CD4+Foxp3+ Tregs in tumors. Frequency of CD8+ T cells within whole tumor cells (left panel). Frequency of CD4+Foxp3+ Tregs within whole tumor cells (middle panel). The number of CD8+ T cells was compared with that of CD4+Foxp3+ Tregs in tumors at day 16 (right panel) (n = 4 for the group of Ad-mIFN i.t.+IL-6R ab i.p., n = 6 for the other groups). IL-6R ab, anti-IL-6 receptor antibody; i.t., intratumoral injection; i.p., intraperitoneal administration; TDLNs, tumor-draining lymph nodes.

Article Snippet: The amounts of cytokines in cell culture supernatants and tumors were assayed with antibodies for IL-6 and mouse IFN-α (Quantikine; R&D systems, Minneapolis, MN, USA) in accordance with the manufacturer’s recommendations.

Techniques: Injection, Cytometry

Journal: Cancer gene therapy

Article Title: Type I IFN gene delivery suppresses regulatory T cells within tumors.

doi: 10.1038/cgt.2014.60

Figure Lengend Snippet: Figure 5. IL-6 receptor blockade partially attenuated IFN-α-mediated tumor growth suppression. (a) Growth of tumors treated with IL-6R ab. Tumor volumes in mice treated with the viruses and/or IL-6R ab were measured at the indicated days (n = 5 for the group of Ad-AP i.t.+IL-6R ab i.p., n = 6 for the other groups). Relative tumor volumes compared with those at day 10 were presented. (b) ELISpot assay of IFN-γ- producing cells in mice treated with IL-6R ab. The splenocytes were isolated from mice as shown in Figure 4a at day 28, and the cells were cultured with CT26 or syngeneic splenocytes (n = 5 for the group of Ad-AP i.t.+IL-6R ab i.p., n = 6 for the other groups).

Article Snippet: The amounts of cytokines in cell culture supernatants and tumors were assayed with antibodies for IL-6 and mouse IFN-α (Quantikine; R&D systems, Minneapolis, MN, USA) in accordance with the manufacturer’s recommendations.

Techniques: Enzyme-linked Immunospot, Isolation, Cell Culture

Journal: Cancer gene therapy

Article Title: Type I IFN gene delivery suppresses regulatory T cells within tumors.

doi: 10.1038/cgt.2014.60

Figure Lengend Snippet: Figure 6. Intratumoral IFN-α expression increased the number of Th17 cells in tumors. (a) Expression of RORγt and Foxp3 genes in tumors. The tumors injected with viruses were harvested at day 16 and were subjected to real-time PCR analysis (Ad-mIFN: n = 4, Ad-AP: n = 3). (b) Expression of IL-17 in tumors. The tumors were harvested at day 28, and subjected to RT-PCR for IL-17 expression (n = 3 for the group of Ad-AP i.t.+PBS i.p., n = 2 for the group of Ad-mIFN i.t.+PBS i.p., n = 3 for the group of Ad-mIFN i.t.+IL-6R ab i.p.). (c) Intracellular cytokine staining of IL-17A in CD4+ T cells. The tumors and tumor-draining lymph nodes were harvested at day 22, and IL-17A expressions were analyzed by flow cytometry (n = 3 for the group of Ad-AP i.t.+PBS i.p., n = 4 for the group of Ad-mIFN i.t.+PBS i.p., n = 4 for the group of Ad-AmIFN i.t.+IL-6R ab i.p.) (upper panel). Representative FACS plots of tumors (lower left panel) and tumor-draining lymph nodes (lower right panel) were shown.

Article Snippet: The amounts of cytokines in cell culture supernatants and tumors were assayed with antibodies for IL-6 and mouse IFN-α (Quantikine; R&D systems, Minneapolis, MN, USA) in accordance with the manufacturer’s recommendations.

Techniques: Expressing, Injection, Real-time Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Staining, Cytometry

Journal: BMC Pediatrics

Article Title: Correlation of biochemical markers and inflammatory cytokines in autism spectrum disorder (ASD)

doi: 10.1186/s12887-024-05182-3

Figure Lengend Snippet: The minimum and maximum values, the median, and the IQR for each marker. The IQR represents the middle 50% of the data, with the 25th percentile and the 75th percentile values

Article Snippet: Human ELISA Kits were used to measure CRP (Quantikine, R&D Systems, cat. DCRP00B, USA, Sensitivity: 0.022 ng/mL), TNF-α (Immunological Sciences, cat. IK4185, Sensitivity: 1 pg/ml), TGF-β1(Diaclone Human TGF-β1 ELISA kit, Sensitivity: 9 pg/mL), IL-1β (Quantikine, R&D Systems, cat. HSLB00D, USA, Sensitivity: 0.063 pg/mL), IL-10 (Quantikine, R&D Systems, cat. HS100C, USA, Sensitivity: 0.17 pg/m), Il-8 (MabTech Human IL-8 ELISA kit, Sensitivity: 2 pg/mL), and IL-6 (Quantikine, R&D Systems, cat.HS600C, USA, sensitivity: 0.09 pg/mL) in the serum samples of ASD patients and control group, according to the Kit manufacturer.

Techniques: Marker

Journal: BMC Pediatrics

Article Title: Correlation of biochemical markers and inflammatory cytokines in autism spectrum disorder (ASD)

doi: 10.1186/s12887-024-05182-3

Figure Lengend Snippet: Comparing the serum concentrations of CRP, TNF-α, TGF-β, IL-1β, and IL-6 between ASD patients and control individuals

Article Snippet: Human ELISA Kits were used to measure CRP (Quantikine, R&D Systems, cat. DCRP00B, USA, Sensitivity: 0.022 ng/mL), TNF-α (Immunological Sciences, cat. IK4185, Sensitivity: 1 pg/ml), TGF-β1(Diaclone Human TGF-β1 ELISA kit, Sensitivity: 9 pg/mL), IL-1β (Quantikine, R&D Systems, cat. HSLB00D, USA, Sensitivity: 0.063 pg/mL), IL-10 (Quantikine, R&D Systems, cat. HS100C, USA, Sensitivity: 0.17 pg/m), Il-8 (MabTech Human IL-8 ELISA kit, Sensitivity: 2 pg/mL), and IL-6 (Quantikine, R&D Systems, cat.HS600C, USA, sensitivity: 0.09 pg/mL) in the serum samples of ASD patients and control group, according to the Kit manufacturer.

Techniques: Control

Journal: BMC Pediatrics

Article Title: Correlation of biochemical markers and inflammatory cytokines in autism spectrum disorder (ASD)

doi: 10.1186/s12887-024-05182-3

Figure Lengend Snippet: Correlation matrix of inflammatory markers in the ASD patients

Article Snippet: Human ELISA Kits were used to measure CRP (Quantikine, R&D Systems, cat. DCRP00B, USA, Sensitivity: 0.022 ng/mL), TNF-α (Immunological Sciences, cat. IK4185, Sensitivity: 1 pg/ml), TGF-β1(Diaclone Human TGF-β1 ELISA kit, Sensitivity: 9 pg/mL), IL-1β (Quantikine, R&D Systems, cat. HSLB00D, USA, Sensitivity: 0.063 pg/mL), IL-10 (Quantikine, R&D Systems, cat. HS100C, USA, Sensitivity: 0.17 pg/m), Il-8 (MabTech Human IL-8 ELISA kit, Sensitivity: 2 pg/mL), and IL-6 (Quantikine, R&D Systems, cat.HS600C, USA, sensitivity: 0.09 pg/mL) in the serum samples of ASD patients and control group, according to the Kit manufacturer.

Techniques:

Journal: BMC Pediatrics

Article Title: Correlation of biochemical markers and inflammatory cytokines in autism spectrum disorder (ASD)

doi: 10.1186/s12887-024-05182-3

Figure Lengend Snippet: Correlation matrix of inflammatory markers in the control group

Article Snippet: Human ELISA Kits were used to measure CRP (Quantikine, R&D Systems, cat. DCRP00B, USA, Sensitivity: 0.022 ng/mL), TNF-α (Immunological Sciences, cat. IK4185, Sensitivity: 1 pg/ml), TGF-β1(Diaclone Human TGF-β1 ELISA kit, Sensitivity: 9 pg/mL), IL-1β (Quantikine, R&D Systems, cat. HSLB00D, USA, Sensitivity: 0.063 pg/mL), IL-10 (Quantikine, R&D Systems, cat. HS100C, USA, Sensitivity: 0.17 pg/m), Il-8 (MabTech Human IL-8 ELISA kit, Sensitivity: 2 pg/mL), and IL-6 (Quantikine, R&D Systems, cat.HS600C, USA, sensitivity: 0.09 pg/mL) in the serum samples of ASD patients and control group, according to the Kit manufacturer.

Techniques: Control