Journal: International Journal of Molecular Medicine

Article Title: Adropin attenuates pancreatitis-associated lung injury through PPARγ phosphorylation-related macrophage polarization

doi: 10.3892/ijmm.2023.5298

Figure Lengend Snippet: Excessive M1 macrophage polarization is observed in the Adro-KO + L-Arg group. (A) Western blot analysis of lung tissue form the Adro-KO + L-Arg group. (B) Co-expression of CD68 and iNOS in lung tissue in L-Arg group determined using immunofluorescence. (C) Co-expression of CD68 and CD206 of lung tissue in the L-Arg group determined using immunofluorescence. (D) iNOS mRNA level in lung tissue from the Adro-KO + L-Arg group (n≥8). (E) CD68 mRNA level in lung tissue from the Adro-KO + L-Arg group (n≥7). (F) CD163 mRNA level in lung tissue from the Adro-KO + L-Arg group (n≥8). (G) Arg-1 mRNA level in lung tissue from the Adro-KO + L-Arg group (n≥8). (H) Quantitative analysis of immunofluorescence (CD68 + iNOS) staining (n≥5). (I) Quantitative analysis of immunofluorescence (CD68; CD206) staining in macrophages (n≥9). (J) Quantitative analysis of the western blots (PPARγ; n≥6); (K) Quantitative analysis of the western blots (PPARγ Ser273; n≥6). (L) Quantitative analysis of the western blots (PPARγ Ser112; n≥6). * P<0.05. L-Arg, L-arginine; NS, normal saline; Adro-KO, adropin knockout; iNOS, inducible nitric oxide synthase.

Article Snippet: The primary antibodies were used at 4°C overnight, and the antibody included Adropin (1:1,000, cat. no. PA5-72781, Thermo Fisher Scientific, Inc.); GADPH (1:10,000; cat. no. AC001, ABclonal); peroxisome proliferator-activated receptor γ (PPARγ; 1:1,000, cat. no. bsm-52220R, BIOSS); p-PPARγ Ser112 (1:1,000, cat. no. bs-3737R, BIOSS); p-PPARγ Ser273 (1:1,000, cat. no. bs-2875R, BIOSS), caspase-3 (1:1,000, cat. no. YC0006, ImmunoWay Biotechnology Company) and PARP1 (1:1,000, cat. no. A0942, ABclonal).

Techniques: Western Blot, Expressing, Immunofluorescence, Staining, Saline, Knock-Out

Journal: International Journal of Molecular Medicine

Article Title: Adropin attenuates pancreatitis-associated lung injury through PPARγ phosphorylation-related macrophage polarization

doi: 10.3892/ijmm.2023.5298

Figure Lengend Snippet: Adropin exogenous supplement induces M2 macrophage polarization. (A) Western blot analysis of lung tissue from the Adro-KO + L-Arg + Adro (34−76) group. (B) Quantitative analysis of the western blots (PPARγ, PPARγ Ser112, PPARγ Ser273) (n≥5). (C) Co-expression of CD68 and iNOS in lung tissue from the Adro-KO + L-Arg+Adro (34−76) group. (D) Co-expression of CD68 and iNOS in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group. (E) Ratio of CD68/DAPI of lung in Adro-KO+ L-Arg+Adro (34−76) group(n≥3); (F) The ratio of iNOS/DAPI in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥3). (G) Ratio of iNOS/CD68 in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥3). (H) Ratio of CD68/DAPI in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥3). (I) Ratio of CD206/DAPI in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥3). (J) Ratio of CD206/CD68 in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥3). (K) CD163 mRNA level in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥6). (L) Arg-1 mRNA level in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥6). (M) iNOS mRNA level in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥6). (N) CD86 mRNA level level in lung tissue from the Adro-KO + L-Arg + Adro (34−76) group (n≥6). * P<0.05 and *** P<0.001. L-Arg, L-arginine; Adro-KO, adropin knockout; NS, normal saline; iNOS, inducible nitric oxide synthase; Arg-1, arginase 1.

Article Snippet: The primary antibodies were used at 4°C overnight, and the antibody included Adropin (1:1,000, cat. no. PA5-72781, Thermo Fisher Scientific, Inc.); GADPH (1:10,000; cat. no. AC001, ABclonal); peroxisome proliferator-activated receptor γ (PPARγ; 1:1,000, cat. no. bsm-52220R, BIOSS); p-PPARγ Ser112 (1:1,000, cat. no. bs-3737R, BIOSS); p-PPARγ Ser273 (1:1,000, cat. no. bs-2875R, BIOSS), caspase-3 (1:1,000, cat. no. YC0006, ImmunoWay Biotechnology Company) and PARP1 (1:1,000, cat. no. A0942, ABclonal).

Techniques: Western Blot, Expressing, Knock-Out, Saline

Journal: Journal of medicinal chemistry

Article Title: A novel N-substituted valine derivative with unique PPARγ binding properties and biological activities

doi: 10.1021/acs.jmedchem.0c01555

Figure Lengend Snippet: Effect of 7j on PPARγ phosphorylation. (A) Percentage of in vitro PPARγ Ser273 phosphorylation by CDK5 in the presence of 0.1 μM of 7j or Rosi. (B) Phosphorylation of PPARγ Ser273 in 3T3-L1 adipocyte incubated for 60 minutes with 5 μM of Rosi, 7j or Roscovitine (Rsv) before TNFα stimulation (50 ng/ml; 90 minutes). Two independent experiments are shown. Phosphospecific antibodies were from Rockland or New England Peptide (NEP). Arrow heads indicate phosphorylated PPARγ and PPARγ2. (C) RT-PCR analysis of the expression levels of a selection of genes known to be regulated by CDK5-dependent phosphorylation of PPARγ. Values are means ± SD (n = 5) expressed relative to the mean of control. **p < 0.01, ***p < 0.001 vs. control (one-way ANOVA followed by Dunnett’s post hoc test).

Article Snippet: PPARγ phospho Ser273 antibodies were from Rockland (Limerick, PA, USA) or custom produced by New England Peptide (Gardner, MA, USA).

Techniques: In Vitro, Incubation, Reverse Transcription Polymerase Chain Reaction, Expressing, Selection