Journal: International Journal of Molecular Medicine

Article Title: Osthole attenuates ovalbumin-induced lung inflammation via the inhibition of IL-33/ST2 signaling in asthmatic mice

doi: 10.3892/ijmm.2020.4682

Figure Lengend Snippet: Primer sequences of the genes detected by quantitative polymerase chain reaction.

Article Snippet: Subsequently, they were washed with PBS, and incubated with rabbit polyclonal anti-ST2 antibody (1:100; cat. no. 11920-1-AP, ProteinTech Group, Inc.) for 12 h at 4°C in a humid chamber.

Techniques:

Journal: International Journal of Molecular Medicine

Article Title: Osthole attenuates ovalbumin-induced lung inflammation via the inhibition of IL-33/ST2 signaling in asthmatic mice

doi: 10.3892/ijmm.2020.4682

Figure Lengend Snippet: (A-E) Expression of ST2 is increased in lung tissue of asthmatic mice by immunohistochemistry. The ST2 immunostaining (brown stain) at ×200 magnification is shown. (A) normal control group, (B) asthma mice model, (C) DEX treatment group, (D) osthole treatment group (25 mg/kg), (E) osthole treatment group (50 mg/kg). (F) The secretion of IL-33 in BALF was assessed using the Luminex multiplex cytokine assat kit. (G) Relative expression of IL-33 mRNA in lung. (H) Relative expression of ST2 mRNA in lung. Values were expressed as the means ± SD. ## P<0.01 vs. the normal control group; * P<0.05, ** P<0.01 vs. the asthma model group. OVA, ovalbumin; DEX, dexamethasone.

Article Snippet: Subsequently, they were washed with PBS, and incubated with rabbit polyclonal anti-ST2 antibody (1:100; cat. no. 11920-1-AP, ProteinTech Group, Inc.) for 12 h at 4°C in a humid chamber.

Techniques: Expressing, Immunohistochemistry, Immunostaining, Staining, Control, Luminex, Multiplex Assay

Journal: Mucosal immunology

Article Title: IL-33/ST2/IL-9/IL-9R Signaling Disrupts Ocular Surface Barrier in Allergic Inflammation

doi: 10.1038/s41385-020-0288-4

Figure Lengend Snippet: In conjunctiva (Conj), cornea and/or CLN of PBS-treated mice and EAC mice, IL-33 (A) and ST2 (D) mRNA expressions were evaluated by RT-qPCR; IL-33 production was detected by ELISA (B). In PBS-treated mice and EAC mice, IL-33 expression in cornea and Conj was shown by IHC staining (C); ST2 and CD4 expression in Conj and CLN (E) were shown by IF double staining. Direct effects of rmIL-33 on IL-9 expression in isolated mice CD4 + T cells was detected at mRNA (F, up) and protein (F, down) levels. IL-9 signal pathway related proteins (including PU.1, NFATc1 and NFATc2) were evaluated in mice CD4 + T cells induced by rmIL-33 (G). Results shown are the Mean ± SD (n=5), *P<0.05, **P<0.01, compared with PBS controls. Bar: 20μm.

Article Snippet: IHC or IF staining was performed as previously described respectively., The primary antibodies used for this study included: rat anti-mouse CD4 from BD Pharmingen (San Jose, CA); rabbit anti-mouse IL-9 from Aviva Systems Biology (San Diego, CA); rabbit anti-mouse IL-9R, IL-33, ZO-1, claudin 1 and occludin from Thermo Fisher Scientific (Waltham, MA); rabbit anti-mouse PU.1, NFATc1, or NFATc2 from Proteintech Group (Rosemont, IL); and rabbit anti-mouse ST2, E-cadherin from Novus Biologicals (Centennial CO).

Techniques: Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Expressing, Immunohistochemistry, Double Staining, Isolation

Journal: Mucosal immunology

Article Title: IL-33/ST2/IL-9/IL-9R Signaling Disrupts Ocular Surface Barrier in Allergic Inflammation

doi: 10.1038/s41385-020-0288-4

Figure Lengend Snippet: EAC model was induced in ST2 −/− mice using the same method. The ocular surface clinical signs were observed through the slit lamp microscope (A). Th2 cytokines (IL-4, IL-5 and IL-13) mRNA expressions were evaluated by RT-qPCR in the conjunctiva (Conj), cornea and CLN in EAC-Balb/c mice and EAC- ST2 −/− mice, compared with PBS control (B). The protein of Th2 cytokines (IL-4, IL-5 and IL-13) in the conjunctiva (Conj) and CLN of EAC-Balb/c mice and EAC- ST2 −/− mice, compared with PBS group (C). OGD staining in the cornea was carried out and scored in EAC model of ST2 −/− mice (D, E). ZO-1, claudin 1, occludin and E-cadherin were detected in the corneal and conjunctival epithelial cells in EAC model and PBS control in ST2 −/− mice through whole-mount IF staining with quantification (F. G). Results shown are the Mean ± SD (n=5). *P<0.05; **P<0.01; ***P<0.001, compared with PBS controls. Bar: 20μm.

Article Snippet: IHC or IF staining was performed as previously described respectively., The primary antibodies used for this study included: rat anti-mouse CD4 from BD Pharmingen (San Jose, CA); rabbit anti-mouse IL-9 from Aviva Systems Biology (San Diego, CA); rabbit anti-mouse IL-9R, IL-33, ZO-1, claudin 1 and occludin from Thermo Fisher Scientific (Waltham, MA); rabbit anti-mouse PU.1, NFATc1, or NFATc2 from Proteintech Group (Rosemont, IL); and rabbit anti-mouse ST2, E-cadherin from Novus Biologicals (Centennial CO).

Techniques: Microscopy, Quantitative RT-PCR, Control, Staining

Journal: Mucosal immunology

Article Title: IL-33/ST2/IL-9/IL-9R Signaling Disrupts Ocular Surface Barrier in Allergic Inflammation

doi: 10.1038/s41385-020-0288-4

Figure Lengend Snippet: In ST2 −/− mice, IL-9 (A), IL-9R (D), SPI 1, NFATc1 and NFATc2 (F) mRNA expression in the conjunctiva (Conj), cornea and/or CLN of EAC model was evaluated by RT-PCR. IL-9 protein expression in the conjunctiva (Conj) and CLN of EAC mice was detected by ELISA (B) and IF staining (C). IL-9R protein expression in the ocular surface (E) and PU.1, NFATc1 NFATc2 protein expression in CLN (G) were shown by IHC staining and IF staining respectively. Results shown are the Mean ± SD (n=5). Bar: 20μm.

Article Snippet: IHC or IF staining was performed as previously described respectively., The primary antibodies used for this study included: rat anti-mouse CD4 from BD Pharmingen (San Jose, CA); rabbit anti-mouse IL-9 from Aviva Systems Biology (San Diego, CA); rabbit anti-mouse IL-9R, IL-33, ZO-1, claudin 1 and occludin from Thermo Fisher Scientific (Waltham, MA); rabbit anti-mouse PU.1, NFATc1, or NFATc2 from Proteintech Group (Rosemont, IL); and rabbit anti-mouse ST2, E-cadherin from Novus Biologicals (Centennial CO).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Staining, Immunohistochemistry

Journal: Mucosal immunology

Article Title: IL-33/ST2/IL-9/IL-9R Signaling Disrupts Ocular Surface Barrier in Allergic Inflammation

doi: 10.1038/s41385-020-0288-4

Figure Lengend Snippet: The ocular surface of ST2 −/− mice was topically administrated by rmIL-9 at 50 ng/ml, three times per day for 3 days, with PBS eyedrop as control. (A) OGD staining in the cornea from ST2 −/− mice, which were untreated, treated with PBS or rmIL-9; (B-D). ZO-1, claudin 1, occludin and E-cadherin in the corneal (B) and conjunctival (C) epithelia of ST2 −/− mice treated with rmIL-9 or PBS control were detected by whole-mount IF staining with quantification (D). Results shown are the Mean ± SD (n=5). *P<0.05, **P<0.01; ***P<0.001, compared with PBS controls. Bar: 20μm.

Article Snippet: IHC or IF staining was performed as previously described respectively., The primary antibodies used for this study included: rat anti-mouse CD4 from BD Pharmingen (San Jose, CA); rabbit anti-mouse IL-9 from Aviva Systems Biology (San Diego, CA); rabbit anti-mouse IL-9R, IL-33, ZO-1, claudin 1 and occludin from Thermo Fisher Scientific (Waltham, MA); rabbit anti-mouse PU.1, NFATc1, or NFATc2 from Proteintech Group (Rosemont, IL); and rabbit anti-mouse ST2, E-cadherin from Novus Biologicals (Centennial CO).

Techniques: Control, Staining

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: mRNA expression level of ST2 and FoxP3 in the adenomas and CRC tissues. The mRNA expression level of ST2 determined by real-time PCR was significantly increased in the adenoma tissues ( grey bar in A ) compared to the normal controls ( white bar in A , P < 0.05). It was also significantly higher in the CRC tissues ( black bar in A ) than in the normal controls ( white bar in A , P < 0.05) but slightly lower than that in the adenoma tissues ( grey bar in A ). As seen in B , the expression level of FoxP3 mRNA showed an increasing trend along the adenoma-carcinoma sequence. Increased expression of FoxP3 mRNA was started from the adenoma stage ( grey bar in B ) and even higher in the CRC stage ( black bar in B ) compared to the normal controls ( white bar in B , both P < 0.05). NS: no significant, P > 0.05.

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Sequencing

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: Photographic representations and density analysis of ST2-positive cells and Foxp3-positive Tregs in normal colorectal, adenoma and CRC tissues examined with immunohistochemistry (IHC). IHC images in first panel of Fig. 2 showed that ST2-immunoreactivity (IR) in the control tissues was observed in both the epithelium ( green arrowhead in A ) and lamina propria ( blue arrow in A ). In the adenoma and CRC tissues, increased ST2-IR was observed in both the adenoma ( green arrowhead in inserted image in B )/CRC epithelium ( arrowhead in inserted image in C ) and adenoma ( blue arrow in B )/CRC stroma ( blue arrow in C ). As seen in second panel of Fig. 2, low density of Foxp3-positive Tregs were observed in control lamina propria ( blue arrow in D ). However, dense Foxp3-positive Tregs were found in both the adenoma ( blue arrow in E )/CRC stroma, ( blue arrow in F ), and some were infiltrated into adenoma/CRC epithelium (green arrowhead in inserted images in E,F ). ( A–F : IHC, counterstained with hematoxylin, original magnification 200×).

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques: Immunohistochemistry, Control

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: Density and phenotypical analysis of ST2-positive cells and FoxP3-positive Tregs in the adenoma/CRC stroma. Semiquantitative data showed that the scores of ST2-positive cells ( A,B , epithelium and stroma respectively) and FoxP3-positive Tregs ( C,D , epithelium and stroma respectively) in both the adenoma and CRC were increased. The density of ST2-positive cells and FoxP3-positive Trges were even higher in the adenoma epithelium ( A,C , grey bar) and that in the CRC epithelium ( A,C , black bar). Further phenotypic analysis of ST2-positive cells ( green color in E,F ) in the adenoma/CRC stroma defined that most positive cells were colocalized with CD3-positive labelled ( red color in E ) T lymphocytes ( white arrow point in E ) and some were SMA-alpha-labelled ( red color in F ) myofibroblasts ( white arrow point in F ). ( E , F , confocal images, original magnification 200×; counterstaining was not applied).

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques:

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: Spearman’s R coefficient analysis of correlation between ST2 and FoxP3 at mRNA and cellular levels in the adenoma/CRC microenvironment.

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques:

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: Clinicopathological analysis of ST2-positive cells and FoxP3-positive Tregs in patients with adenomas and CRCs. Further clinicopathological analysis revealed that increased densities of ST2-positive cells in the adenoma stroma ( A ) correlated with the degree of dysplasia, patients with high grade of dysplasia (HGD) tend to have a higher density of ST2-positive stromal cells than those with low grade of dysplasia (LGD). The density of ST2-positive cells in the CRC stroma correlates with TNM stages and node involvement ( B ), and the density of FoxP3-positive Tregs in the CRC stroma correlated with node involvement ( C ). NS: no significant, P > 0.05.

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques:

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: The Kaplan–Meier analysis of overall survival differences in CRC patients with differing density scores of ST2-positive cells and FoxP3-positive Tregs. The Kaplan–Meier analysis revealed that the density scores of ST2-positive cells ( A ) and FoxP3-positive Tregs ( B ) in the CRC stroma predicate the overall survival time in patients with CRC. CRC patients with high densities of ST2-positive cells and FoxP3-positive Tregs in the tumor stroma did have a significantly shorter overall survival time compared with those with low densities of ST2-positive cells and FoxP3-positive Tregs.

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques:

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: Photographic representations of ST2 expressed in FoxP3-positive Tregs in the adenoma stroma. Double immunohistochemistry images revealed that many FoxP3-labelled ( red color in A ) Tregs expressed ST2-IR ( brown color in B ) in a represent adenoma section. ( A,B , double immunohistochemistry images, original magnification 400×; counterstaining was hematoxylin).

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques: Immunohistochemistry

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: A schematic representation of the potential role of IL-33 functional receptor ST2 in contributing to the development of colorectal cancer.

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques: Functional Assay

Journal: Scientific Reports

Article Title: ST2 and regulatory T cells in the colorectal adenoma/carcinoma microenvironment: implications for diseases progression and prognosis

doi: 10.1038/s41598-020-62502-0

Figure Lengend Snippet: Primer/probe sequences of house-keeping gene, ST2 and FoxP3 for quantitative real-time PCR.

Article Snippet: The following primary antibodies were used: rabbit anti-ST2 polyclonal antibody (working dilution 1:100; Thermo Fisher Scientific, USA) and mouse anti-FoxP3 monoclonal antibody (working dilution 1:100, Abcam, UK).

Techniques: Sequencing