anti phospho igf1r  (Cell Signaling Technology Inc)

Journal: The Journal of Biological Chemistry

Article Title: Downregulation of ClC-3 chloride channels in dorsal root ganglia neurons contributes to bone metastasis-induced pain

doi: 10.1016/j.jbc.2026.111268

Figure Lengend Snippet: Knockdown of Igf1r in DRG neurons disrupts intrathecal IGF1-induced activation of AKT signaling as well as the upregulated expression of HDAC2, the decreased expression of ClC-3 protein and pain hypersensitivity in na9;ve rats . A and B , RT-qPCR analysis and western blotting analysis of IGF1R mRNA and protein abundance in L4/5 DRG tissues from sham and BCP rats at 14 days after tumor cells inoculation. n = 6–7 rats per group. C , representative images showing the colocalization of IGF1R, HDAC2, and ClC-3 in DRG neurons. The scale bar represents 25 μm. D – F , western blotting analysis of p-AKT ( E ) and AKT ( F ) protein abundance in ipsilateral L4/5 DRG tissues obtained from IGF1-treated naïve rats that received intrathecal LV-shIGF1R and the control LV-GFP, respectively (n = 4 rats per group). D , representative blots are shown. G – I , western blotting analysis of HDAC2 ( H ) and ClC-3 ( I ) protein abundance in ipsilateral L4/5 DRG tissues obtained from IGF1-treated naïve rats that received intrathecal LV-shIGF1R and the control LV-GFP, respectively (n = 4 rats per group). G , representative blots are shown. J and K , assessment of ipsilateral PWT and PWL for IGF1-treated naïve rats that received intrathecal LV-shIGF1R and the control LV-GFP (n = 10 rats per group). L , assessment of animal’s locomotor function before and after intrathecal drug administration (n = 10 rats per group). Data are presented as mean ± SEM. ∗ p < 0.05, ∗∗∗ p < 0.001, ns, not significant, unpaired t test for ( A )–( B ); one-way ANOVA followed by Dunnett's post hoc test for ( E )–( I ) two-way ANOVA with Sidak’s post hoc test for ( J )–( L ). BCP, bone cancer pain; DRG, dorsal root ganglion; HDAC2, histone deacetylase 2; IGF1, insulin-like growth factor 1; LV-GFP, lentivirus expressing green fluorescent protein; p-AKT, phosphorylated AKT; PWL, paw withdrawal latency; PWT, paw withdrawal threshold; RT-qPCR, real-time quantitative PCR.

Article Snippet: The membranes were blocked with 5% nonfat milk in TBST [20 mM Tris–HCl (pH 7.5), 150 mM NaCl, and 0.05% Tween 20] for 60 min at room temperature and then incubated with the following primary antibodies at 4 °C overnight: rabbit anti-ClC-3 antibody (1:1000, Abcam), rabbit anti-HDAC2 antibody (1:1000, Merck Millipore), rabbit anti-IGF1 antibody (1:1000, Abcam), rabbit anti-IGF1R antibody (1:1000, Cell Signaling Technology),rabbit anti-p-AKT antibody (1:1000, Cell Signaling Technology), rabbit anti-AKT antibody (1:2000, Cell Signaling Technology), mouse anti-β-actin (1:2000, Santa Cruz Biotechnology).

Techniques: Knockdown, Activation Assay, Expressing, Quantitative RT-PCR, Western Blot, Quantitative Proteomics, Control, Histone Deacetylase Assay, Real-time Polymerase Chain Reaction

Journal: The Journal of Biological Chemistry

Article Title: Downregulation of ClC-3 chloride channels in dorsal root ganglia neurons contributes to bone metastasis-induced pain

doi: 10.1016/j.jbc.2026.111268

Figure Lengend Snippet: Knockdown of Igf1r in DRG neurons impairs HDAC2-mediated transcriptional repression of Clc-3 gene, reduces neuronal excitability and attenuates pain hypersensitivity in bone metastasis model rats . A – E , western blotting analysis of p-AKT ( B ), AKT ( C ), HDAC2 ( D ) and ClC-3 ( E ) abundance in ipsilateral L4/5 DRG tissues from intrathecal LV-GFP and LV-shIGF1R treated rats at 14 days after tumor cells inoculation. n = 4 to 6 rats per group. A , representative blots are shown. F – H , electrophysiological analyses of neuronal excitability in ipsilateral L4/5 DRG neurons of bone metastasis model rats that received intrathecal LV-GFP and LV-shIGF1R, recorded at 14 days after tumor cells inoculation. F , representative neuronal action potentials evoked by a large depolarizing current pulse (1-s, 2-fold AP rheobase) are shown. The scale bar represents 20 mV, 100 ms. G and H , plots of threshold potential and rheobase are shown. n = 10∼20 cells from three rats per group. I and J , assessment of ipsilateral PWT ( I ) and PWL ( J ) from intrathecal LV-GFP and LV-shIGF1R treated rats after tumor cells inoculation. n = 10 rats per group. K , assessment of animal’s locomotor function by inclined-plate test, compared before and after drug administration. n = 10 rats per group. Data are presented as mean ± SEM. ∗ p < 0.05, ∗∗∗ p < 0.001, ns, not significant, unpaired t test for ( B )–( H ); two-way ANOVA with Sidak’s post hoc test for ( I )–( K ). DRG, dorsal root ganglion; HDAC2, histone deacetylase 2; LV-GFP, lentivirus expressing green fluorescent protein; p-AKT, phosphorylated AKT; PWL, paw withdrawal latency; PWT, paw withdrawal threshold.

Article Snippet: The membranes were blocked with 5% nonfat milk in TBST [20 mM Tris–HCl (pH 7.5), 150 mM NaCl, and 0.05% Tween 20] for 60 min at room temperature and then incubated with the following primary antibodies at 4 °C overnight: rabbit anti-ClC-3 antibody (1:1000, Abcam), rabbit anti-HDAC2 antibody (1:1000, Merck Millipore), rabbit anti-IGF1 antibody (1:1000, Abcam), rabbit anti-IGF1R antibody (1:1000, Cell Signaling Technology),rabbit anti-p-AKT antibody (1:1000, Cell Signaling Technology), rabbit anti-AKT antibody (1:2000, Cell Signaling Technology), mouse anti-β-actin (1:2000, Santa Cruz Biotechnology).

Techniques: Knockdown, Western Blot, Histone Deacetylase Assay, Expressing

Journal: Biomedicines

Article Title: Single-Cell Analysis Links C7+ Cancer-Associated Fibroblasts with Incomplete Resection in Platinum-Sensitive Relapsed Ovarian Cancer

doi: 10.3390/biomedicines13123011

Figure Lengend Snippet: IGF-I inhibits anoikis and promotes migration in ovarian cancer cells. ( A , B ) Western blot analysis showing the knockout efficiency of IGF1R in COV318 ( A ) and SKOV3 ( B ) cells. ( C ) Following IGF-I treatment, anoikis levels in IGF1R knockout (KO) COV318 cells were significantly increased compared to the negative control (NC) group. ( D ) Quantitative analysis of anoikis in COV318 cells. ( E ) IGF1R knockout SKOV3 cells exhibited significantly elevated anoikis levels after IGF-I treatment compared with the NC group. ( F ) Quantitative analysis of anoikis in SKOV3 cells. ( G , H ) Scratch assay demonstrating that IGF-I treatment failed to promote migration in IGF1R knockout COV318 ( G ) and SKOV3 ( H ) cells. Scale bar, 200 μm. ( I , J ) Quantitative analysis of migration rate in COV318 ( I ) and SKOV3 ( J ) cells from scratch assay. ( K , L ) Trans-well migration assay further confirmed that IGF-I inhibited migration of IGF1R knockout COV318 ( K ) and SKOV3 ( L ) cells. Scale bar, 100 μm. ( M , N ) Quantitative analysis of Trans-well migration in COV318 ( M ) and SKOV3 ( N ) cells. Significance thresholds were defined as * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: Membranes were blocked and subsequently incubated overnight at 4 °C with primary antibodies against IGF1R (#9750, Cell Signaling Technology).

Techniques: Migration, Western Blot, Knock-Out, Negative Control, Wound Healing Assay