Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: rmIL-12 treatment schedules for mice which received a combined oral vaccine. Four groups of C57BL/6 mice were given TT plus CT as adjuvant on days 0, 7, and 14 by the oral route. Group A were positive controls receiving oral vaccine but not rmIL-12. Group B received 15 doses of rmIL-12 (day 0 to 5, 7 to 11, and 14 to 18) by the intraperitoneal route. This group was subdivided into mice receiving 10, 100, or 1,000 ng of rmIL-12/dose. Groups C and D received three (days 0, 7, 14) or six (days 0, 3, 7, 10, 14, 17) oral doses of rIL-12 complexed to liposomes.

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques:

Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: The effect of parenteral administration of rmIL-12 on polyclonal and TT-specific serum Ab responses in mice orally immunized with TT and CT as mucosal adjuvant. Mice received the oral vaccine alone ( unshaded ) or together with rmIL-12 by the i.p. route ( shaded , 10 ng/dose or striped , 100 ng/dose). ( A ) Polyclonal and TT-specific serum IgE Abs were measured on day 14 by ELISA and PCA assays, respectively. ( B ) The distribution of TT-specific serum IgG subclasses were analyzed on day 21 by ELISA. ( C ) Serum TT-specific IgM, IgA, and IgG Abs were measured by ELISA on day 21. Results are expressed as the mean ± one SD from three different experiments (five mice per group).

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: The effect of oral administration of rmIL-12 on polyclonal and TT-specific serum Ab responses. Mice received the oral combined vaccine alone ( unshaded ) or vaccine together with rmIL-12 complexed to liposomes by the oral route ( striped , 1,000 ng/dose three times and shaded , 1,000 ng/dose six times). ( A ) Polyclonal and TT-specific serum IgE Abs were measured on day 14 by ELISA and PCA assays, respectively. ( B ) The distribution of TT-specific serum IgG subclasses were determined on day 21 by ELISA. ( C ) Serum TT-specific IgM, IgA, and IgG Abs were measured by ELISA on day 21. Results are expressed as the mean ± one SD from four different experiments (five mice per group).

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: The effect of oral and parenteral administration of rmIL-12 on mucosal IgA responses to an oral vaccine consisting of TT and CT as mucosal adjuvant. TT-specific S-IgA Ab titers in fecal extracts were measured on day 21 by ELISA. The frequency of TT-specific IgA AFCs in LPLs was determined on day 21 by ELISPOT. Mice received the oral vaccine alone ( unshaded ) or together with rmIL-12 by i.p. injection ( striped , 100 ng/dose) or six oral administrations ( shaded , 1,000 ng/dose). The results are expressed as the mean ± one SD from four different experiments (five mice per group).

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques: Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot, Injection

Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: The induction of Th1-type (IL-2 and IFN-γ) and Th2-type (IL-4, IL-5, IL-6, and IL-10) cytokine secretion by CD4 + T cells isolated from mucosally immunized mice which had been given oral ( shaded ) or parenteral ( striped ) rmIL-12. SP and PP CD4 + T cells, were purified from mice receiving the combined oral vaccine only ( unshaded ) or combined vaccine together with rmIL-12 orally (six times, 1,000 ng/dose) or parenterally (15 times, 100 ng/dose) and were restimulated in vitro for 6 d in the presence of feeder cells and TT-coated beads. Th1type ( A ) and Th2-type ( B ) cytokine production in culture supernatants were analyzed by ELISA. Cytokine levels are representative of three separate experiments.

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques: Isolation, Purification, In Vitro, Enzyme-linked Immunosorbent Assay

Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: The induction of Th1-type (IFN-γ) and Th2-type (IL-5 and IL-10) cytokines in sera of mice receiving rmIL-12 and orally immunized with TT and CT as adjuvant. Mice received the combined oral vaccine ( unshaded ) together with rmIL-12 by the i.p. route ( striped , 15 times, 100 ng/dose) or by the oral route ( shaded six times, 1,000 ng/dose). Levels of IFN-γ, IL-5, and IL-10 in the serum were determined on day 21 by ELISA. The results are expressed as the mean ± one SD and are representative of three different experiments.

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: The Journal of Experimental Medicine

Article Title: Oral but Not Parenteral Interleukin (IL)-12 Redirects T Helper 2 (Th2)-type Responses to an Oral Vaccine Without Altering Mucosal IgA Responses

doi:

Figure Lengend Snippet: Induction of delayed-type hypersensitivity responses in rmIL-12–treated mice given the combined oral vaccine. Three groups of mice were assessed and included combined oral vaccine only ( unshaded ), those receiving combined oral vaccine and rmIL-12 by the i.p. ( striped ; 15 times 100 ng/dose) route, or mice given rmIL-12 by the oral route ( shaded ; six times 1,000 ng/dose). The results are expressed as the mean ± one SD of the difference in the ear swelling between the TT-injected and OVA-injected ear pinna and are representative of three separate experiments.

Article Snippet: For i.p. delivery, rmIL-12 (10, 100, or 1000 ng/dose) was diluted in sterile PBS containing 1% normal mouse serum and given daily (5 times/wk) as indicated in Fig. . For oral delivery, rmIL-12 (1,000 ng/dose) was complexed with preformed cationic liposomes (DOTAP; Boehringer Mannheim Corp. , Indianapolis, IN).

Techniques: Injection

Journal: Cancer Cell

Article Title: Cancer-associated fibroblast phenotypes are associated with patient outcome in non-small cell lung cancer

doi: 10.1016/j.ccell.2023.12.021

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-p75 (CD271); Clone (polyclonal_ANT-007); Lot(ANT007AN0702) , alomone labs , Cat#ANT-007; RRID: AB_2039968.

Techniques: Recombinant, Labeling, Software

Journal: The Journal of Biological Chemistry

Article Title: A Role for Stefin B (Cystatin B) in Inflammation and Endotoxemia *

doi: 10.1074/jbc.M114.609396

Figure Lengend Snippet: Enhanced processing and secretion of IL-1β and IL-18 in stefin B-deficient BMDMs is caspase-1-dependent. BMDMs from WT and StB KO mice were primed for 4 h with LPS (100 ng/ml), washed, and stimulated with ATP for 20 min (5 mm) to activate the Nlrp3 inflammasome. Cell lysates and culture supernatants were immunoblotted with the indicated antibodies. Anti-caspase-1 and anti-caspase-11 antibodies recognized pro-caspase (inactive zymogen) and subunits generated by proteolytic cleavages. Anti-IL-1β and anti-IL-18 antibodies recognized inactive pro-form in cell extracts and mature secreted cytokines in cell supernatants. Lysates were immunoblotted with anti-β-actin as a loading control. Data shown are representative of three independent experiments.

Article Snippet: Antibodies used in Western blotting were purchased from Abcam, and anti-caspase-1 (ab108362), anti-caspase-11 (ab10454), anti-IL-18 (ab71495), anti-stefin B (ab53725), anti-STAT1 (ab3987), anti-STAT1 Y 701 (9171S) were from Cell Signaling.

Techniques: Generated

Journal: The Journal of Biological Chemistry

Article Title: A Role for Stefin B (Cystatin B) in Inflammation and Endotoxemia *

doi: 10.1074/jbc.M114.609396

Figure Lengend Snippet: Increased inflammasome activation in StB KO BMDMs is independent of lysosomal cysteine cathepsins. A, biosynthesis and integrity of lysosomes were studied by LysoTracker Green probe, as described under “Experimental Procedures.” The lysosomal fluorescence intensity was quantified upon LPS and ATP stimulations. Lysosomal destabilization was increased by ATP addition and was comparable between genotypes. Data were obtained from three independent biological experiments performed in triplicate, and the results are presented as means of geometric mean fluorescence intensity ± S.D. B, release of cathepsins into the cytosol and their activity were measured after indicated stimulations using fluorogenic substrates specific for cathepsins. Incubation of BMDMs with E-64d completely prevented cathepsin activity. C, BMDMs from WT and StB KO mice were primed for 4 h with LPS (100 ng/ml), washed, and stimulated with ATP for 20 min (5 mm) or primed with broad spectrum cysteine cathepsin inhibitor E-64d for 1 h (15 μm), followed by LPS and ATP stimulation. Cell lysates were immunoblotted with indicated antibodies. Lysates were immunoblotted with anti-β-actin as a loading control. Incubation of BMDMs with E-64d did not affect the processing of caspase-1 and pro-inflammatory cytokine release. Data shown are representative of three independent experiments. D, BMDMs were seeded into 96-well plates and stimulated as above. IL-1β release was quantified in the media by FlowCytomixTM. StB KO BMDMs secreted higher amounts of IL-1β into the media compared with the WT; however, priming with E-64d did not affect IL-1β release (n.s., nonsignificant difference). Data were obtained from four independent experiments performed in triplicate, and the results are presented as means ± S.D. *, p < 0.05; **, p < 0.01. E, viability of BMDMs was assessed by LDH release into the cell culture media upon inflammasome activation. The cytotoxicity was expressed as the percent of the total LDH release. Data were obtained from three independent experiments performed in triplicate, and the results are presented as means ± S.D. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Antibodies used in Western blotting were purchased from Abcam, and anti-caspase-1 (ab108362), anti-caspase-11 (ab10454), anti-IL-18 (ab71495), anti-stefin B (ab53725), anti-STAT1 (ab3987), anti-STAT1 Y 701 (9171S) were from Cell Signaling.

Techniques: Activation Assay, Fluorescence, Activity Assay, Incubation, Cell Culture