Fig 1: Abi1 modulates phosphorylation and activation of Jak2 and STAT3(A and B) Abi1 KD attenuates the PDGF-induced Jak2 (A) and STAT3 (B) phosphorylation in HASM cells. Data are means ± SEM. n = 5. Two-way ANOVA was used for statistical analysis.(C) Abi1 deficiency inhibits the PDGF-induced Jak2 kinase activity. Data are means ± SEM. n = 5. Two-way ANOVA was used for statistical analysis.(D) The reporter activity of STAT3 is reduced by Abi1 KD. Data are means ± SEM. n = 5. Two-way ANOVA was used for statistical analysis.(E and F) Overexpression of Abi1 in HASM cells enhances Jak2 kinase activity and STAT3 activity. Data are means ± SEM. n = 5. The t test was used for statistical analysis. *p < 0.05; **p < 0.01. Data normalization is calculated as follows: Data of treated samples/Data of control samples.
Fig 2: Abi1 regulates STAT3 nuclear import in smooth muscle cells(A) Cells with perinuclear STAT3 localization are considered “untranslocated cells.” Cells with concentrated STAT3 distribution in the nucleus are considered “translocated cells.” The white arrow points to the nucleus without STAT3 translocation. The green arrows point to the nuclei with STAT3 translocation.(B) PDGF stimulation induces STAT3 translocation to the nucleus. Data are means ± SEM. n = 5.(C) Abi1 KD inhibits the PDGF-induced STAT3 nuclear import. Ctrl and Abi1 KD cells were treated with PDGF for 2 h followed by immunostaining. Data are means ± SEM. n = 4. Two-way ANOVA was used for statistical analysis.(D) Overexpression (OE) of Abi1 is sufficient to induce STAT3 nuclear translocation. Cells were infected with lentivirus encoding the constructs for 2 days followed by staining. Data are means ± SEM. n = 4. The t test was used for statistical analysis. **p < 0.01. The percentage of cells with translocated STAT3 was calculated as follows: numbers of cells with translocated STAT3/numbers of total cells observed × 100.Scale bar, 10 µm
Fig 3: Abi1 knockdown inhibits the activity of Jak2/STAT3 and the proliferation in Abi1-up HASM cells(A and B) The basal and the PDGF-induced activity of Jak2 (A) and STAT3 (B) is higher in Abi1-up cells. Data are means ± SEM. n = 4 for each group. One-way ANOVA was used for statistical analysis.(C and D) Abi1 KD inhibits the basal and the PDGF-induced activity of Jak2 (C) and STAT3 (D) in Abi1-up cells. Data are means ± SEM. n = 4 for each group. Two-way ANOVA was used for statistical analysis(E and F) Abi1 KD reduced the PDGF-induced cell numbers (E) and BrdU incorporation (F). Data are means ± SEM. n = 4 for each group. The t test was used for statistical analysis. *p < 0.05. Data normalization is calculated as follows: Data of treated samples/Data of control samples.
Fig 4: Proposed mechanismStimulation with growth factors such as PDGF induces spatial distribution of Abi1, which facilitates the activation of the Jak2/STAT3 pathway. The association of Abi1 with Jak2 renders the receptor in active state. Abi1 is upregulated in ~50% of asthmatic airway smooth muscle cultures, promoting the activation of Jak2 and STAT3, hyperplasia, and airway smooth muscle (ASM) thickening.
Fig 5: IHC for the protein levels of Ki67, pY705-STAT3, and c-Myc in the tumor tissues. (A) Ki67, (B) pY705-STAT3, (C) c-Myc. Scale bar = 100 μm. Data are expressed as mean ± SD. n = 3. * p < 0.05 vs. vehicle.
Supplier Page from Abcam for Recombinant Human STAT3 protein (His tag)