Journal: Scientific Reports

Article Title: Vasoactive intestinal peptide blockade suppresses tumor growth by regulating macrophage polarization and function in CT26 tumor-bearing mice

doi: 10.1038/s41598-023-28073-6

Figure Lengend Snippet: Conditioned medium from CT26 cells induced M2 macrophage polarization and VPAC expression in RAW264.7 cells. Real-time PCR analyses of M1 macrophage markers TNF-α , iNOS , and CXCL10 ( A ), M2 macrophage markers Mrc-1 , IL-1rn , and CCL-22 ( B ), immune checkpoint markers SIRP-α , PD-1 , and PD-L1 ( C ), and VIP receptors Vipr1 and Vipr2 ( D ) in RAW264.7 cells at day 4 after incubation with 20% conditioned medium derived from CT26 with DMEM complete medium (CT26-CM) or 20% RPMI medium with DMEM complete medium (control); n = 5–6/group. Representative western blotting images showing protein expression levels of VPAC1 and VPAC2 in RAW264.7 cells at day 4 after incubation with CT26-CM or control ( E , left). Western blotting analysis of VPAC1 and VPAC2 protein expression in RAW264.7 cells at day 4 after incubation with control medium or CT26-CM (E, right); n = 5/group. The full-length blots of ( E ) are included in Supplemental Fig. . *p < 0.05 vs control, **p < 0.01 vs control.

Article Snippet: In a separate experiment, [Lys15, Arg16, Leu27] VIP (1–7)/GRF (8–27) (Phoenix Pharmaceuticals)—a selective VPAC1 agonist —or BAY 55-9837 (Tocris Bioscience, Minneapolis, MN)—a selective VPAC2 agonist—was added daily at a dose of 100 nM . mRNA extraction was performed on day 3 or day 4 after CT26-CM incubation, depending on the experiment.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Incubation, Derivative Assay, Control, Western Blot

Journal: Scientific Reports

Article Title: Vasoactive intestinal peptide blockade suppresses tumor growth by regulating macrophage polarization and function in CT26 tumor-bearing mice

doi: 10.1038/s41598-023-28073-6

Figure Lengend Snippet: Knockdown or activation of VPAC2 disturbed macrophage polarization in RAW264.7 cells incubated with CT26-CM. Effect of Vipr1 ( A ) or Virp2 ( B ) silencing on gene expression of M1 and M2 macrophage markers in RAW264.7 cells at day 4 after incubation with CT26-CM. Negative control siRNA was used as control; n = 6–9/group. Effect of [Lys15, Arg16, Leu27] VIP (1–7)/GRF (8–27), a VPAC1 agonist, ( C ) or BAY 55–9837, a VPAC2 agonist, ( D ) on the mRNA expression of M1 and M2 macrophage markers in RAW264.7 cells at day 3 after incubation with CT26-CM. PBS was used as vehicle; n = 6–10/group. *p < 0.05 vs Negative control siRNA or vehicle.

Article Snippet: In a separate experiment, [Lys15, Arg16, Leu27] VIP (1–7)/GRF (8–27) (Phoenix Pharmaceuticals)—a selective VPAC1 agonist —or BAY 55-9837 (Tocris Bioscience, Minneapolis, MN)—a selective VPAC2 agonist—was added daily at a dose of 100 nM . mRNA extraction was performed on day 3 or day 4 after CT26-CM incubation, depending on the experiment.

Techniques: Knockdown, Activation Assay, Incubation, Gene Expression, Negative Control, Control, Expressing

Journal: Scientific Reports

Article Title: Vasoactive intestinal peptide blockade suppresses tumor growth by regulating macrophage polarization and function in CT26 tumor-bearing mice

doi: 10.1038/s41598-023-28073-6

Figure Lengend Snippet: CD68 macrophages were highly expressed in tumor stroma and co-expressed with VPAC2, but not VPAC1, in human colorectal cancer specimens. Representative images of CD68 (green) and VPAC1 (red; A , B ), and CD68 (green) and VPAC2 (red; C , D ) staining in the tumor stroma of human colorectal cancer specimens. B and D were enlarged images from white square region indicated in ( A ) and ( C ), respectively. All images represented cancer lesions. Nuclei were stained by DAPI (blue). White arrow indicated double positive cells. Scale bar 100 µm.

Article Snippet: In a separate experiment, [Lys15, Arg16, Leu27] VIP (1–7)/GRF (8–27) (Phoenix Pharmaceuticals)—a selective VPAC1 agonist —or BAY 55-9837 (Tocris Bioscience, Minneapolis, MN)—a selective VPAC2 agonist—was added daily at a dose of 100 nM . mRNA extraction was performed on day 3 or day 4 after CT26-CM incubation, depending on the experiment.

Techniques: Staining

Journal: Scientific Reports

Article Title: Vasoactive intestinal peptide blockade suppresses tumor growth by regulating macrophage polarization and function in CT26 tumor-bearing mice

doi: 10.1038/s41598-023-28073-6

Figure Lengend Snippet: List of antibodies and their application in the present study.

Article Snippet: In a separate experiment, [Lys15, Arg16, Leu27] VIP (1–7)/GRF (8–27) (Phoenix Pharmaceuticals)—a selective VPAC1 agonist —or BAY 55-9837 (Tocris Bioscience, Minneapolis, MN)—a selective VPAC2 agonist—was added daily at a dose of 100 nM . mRNA extraction was performed on day 3 or day 4 after CT26-CM incubation, depending on the experiment.

Techniques: Western Blot

Journal: Frontiers in Microbiology

Article Title: Transcription factors Asg1p and Hal9p regulate pH homeostasis in Candida glabrata

doi: 10.3389/fmicb.2015.00843

Figure Lengend Snippet: Strains and plasmids used in this study .

Article Snippet: C. glabrata ATCC 55/ GFP , C. glabrata ATCC 55 (pY13- GFP ) , This study.

Techniques: Mutagenesis

Journal: Frontiers in Microbiology

Article Title: Transcription factors Asg1p and Hal9p regulate pH homeostasis in Candida glabrata

doi: 10.3389/fmicb.2015.00843

Figure Lengend Snippet: Growth assays in different YNB media. (A) Deletion of CgASG1 has no effect on the utilization of non-fermentative carbon sources in C. glabrata . (B) CgAsg1p plays a role under acid-stress conditions. (C) CgHAL9 serves an important role in cell growth under hypersaline conditions in C. glabrata . (D) CgHal9p plays a role under acid-stress conditions. Logarithmic-phase cells of each C. glabrata strain were adjusted to 2 × 10 7 cells/mL, and then 4 μL of serial tenfold dilutions were spotted onto the corresponding YNB media, as indicated. Pictures were taken after 4 days of growth at 30°C.

Article Snippet: C. glabrata ATCC 55/ GFP , C. glabrata ATCC 55 (pY13- GFP ) , This study.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Transcription factors Asg1p and Hal9p regulate pH homeostasis in Candida glabrata

doi: 10.3389/fmicb.2015.00843

Figure Lengend Snippet: Intracellular pH of wild-type, mutant and complemented strains of C. glabrata as below ^a .

Article Snippet: C. glabrata ATCC 55/ GFP , C. glabrata ATCC 55 (pY13- GFP ) , This study.

Techniques: Mutagenesis

Journal: Frontiers in Microbiology

Article Title: Transcription factors Asg1p and Hal9p regulate pH homeostasis in Candida glabrata

doi: 10.3389/fmicb.2015.00843

Figure Lengend Snippet: Quantitative reverse-transcribed PCR (qRT-PCR) validation of transcriptional profiles in an acidic environment . Logarithmic-phase C. glabrata cells were incubated in YNB or YNB-pH 2.0 media for 2 h. qRT-PCR analyses of the indicated genes was performed as described in the Materials and Methods Section. The means and standard deviations for three independent experiments are shown. C. glabrata strains: wild-type, Cgasg1 Δ and Cghal9 Δ strains. ( * P < 0.05 compared to the corresponding wild-type control, as determined by t -test).

Article Snippet: C. glabrata ATCC 55/ GFP , C. glabrata ATCC 55 (pY13- GFP ) , This study.

Techniques: Reverse Transcription, Quantitative RT-PCR, Biomarker Discovery, Incubation, Control