Journal: Oncology Letters

Article Title: PDGF-BB regulates the transformation of fibroblasts into cancer-associated fibroblasts via the lncRNA LURAP1L-AS1/LURAP1L/IKK/IκB/NF-κB signaling pathway

doi: 10.3892/ol.2021.12798

Figure Lengend Snippet: PDGF-BB induced the formation of the CAF phenotype in hOMF. (A) The cells were treated with PDGF-BB in a dose-dependent manner (10, 20, 30 ng/ml) in 10% fetal bovine serum for 72 h for three passages. The cells were subjected to western blot analysis with antibodies against CAF markers α-SMA, FAP-α, PDGFR-β and p-PDGFR-β. β-tubulin served as a loading control and sample loading was 20 µg. (B) Fluorescence microscopy of hOMF stained with α-SMA, FAP-α and PDGFR-β (probed with a primary and a secondary antibody). Cells were counterstained with DAPI. Data were expressed as means ± SEM (n=3). *P<0.05 vs. hOMF, # P<0.05 vs. 10 ng/ml PDGF-BB. PDGF, platelet-derived growth factor; CAF, cancer-associated fibroblast; hOMF, human oral mucosa (p3) 500K fibroblast; α-SMA, α-smooth muscle actin; FAP-α, fibroblast activation protein-α; PDGFR-β, platelet-derived growth factor receptor-β; p, phosphorylated.

Article Snippet: The following antibodies were used in the present study: α-SMA (cat. no. ab5694; 1:1,000), FAP-α (cat. no. ab53066; 1:500), IKKα (cat. no. ab32041; 1:1,000), IκBα (cat. no. ab32518; 1:1,000), NF-κB p65 (cat. no. ab16502; 1:1,000), p-NF-κBp65 (p-S536; cat. no. ab86299; 1:1,000), GAPDH (cat. no. ab181602; 1:10,000), β-tubulin (cat. no. ab179511; 1:1,000) and β-actin (cat. no. ab8227; 1:1,000) were obtained from Abcam; LURAP1L (cat. no. PA5-55072; 1:1,000) was obtained from Thermo Fisher Scientific, Inc., and PDGFR-β (cat. no. bs-0232R; 1:1,000) and p-PDGFR-β (Tyr740; cat. no. bs-3323R; 1:1,000) were obtained from BIOSS.

Techniques: Western Blot, Fluorescence, Microscopy, Staining, Derivative Assay, Activation Assay

Journal: Oncology Letters

Article Title: PDGF-BB regulates the transformation of fibroblasts into cancer-associated fibroblasts via the lncRNA LURAP1L-AS1/LURAP1L/IKK/IκB/NF-κB signaling pathway

doi: 10.3892/ol.2021.12798

Figure Lengend Snippet: Effects of the PDGFR-β inhibitor, CP-673451 on LURAP1L-AS1-LURAP1L/IKK/IκB/NF-κB signaling and CAF programming. (A) Reverse transcription-quantitative PCR analysis of LURAP1L and LURAP1L-AS1 expression levels. (B) Western blot analysis of FAP-α, α-SMA, IKKα, IκBα, NF-κB p65 and p-p65 expression following overexpression of LURAP1L-AS1 and treatment with PDGF-BB. *P<0.05 vs. PDGF-BB. PDGF, platelet-derived growth factor; LURAP1L, leucine-rich adaptor protein 1-like; LURAP1L-AS1, LURAP1L antisense RNA 1; IKKα, IκB kinase α; IKK, I-κB kinase; IκBα, nuclear factor of κ light polypeptide gene enhancer in B-cells inhibitor α; NF-κB, nuclear factor-κB; CAF, cancer-associated fibroblasts; p, phosphorylated; α-SMA, α-smooth muscle actin; FAP-α, fibroblast activation protein-α.

Article Snippet: The following antibodies were used in the present study: α-SMA (cat. no. ab5694; 1:1,000), FAP-α (cat. no. ab53066; 1:500), IKKα (cat. no. ab32041; 1:1,000), IκBα (cat. no. ab32518; 1:1,000), NF-κB p65 (cat. no. ab16502; 1:1,000), p-NF-κBp65 (p-S536; cat. no. ab86299; 1:1,000), GAPDH (cat. no. ab181602; 1:10,000), β-tubulin (cat. no. ab179511; 1:1,000) and β-actin (cat. no. ab8227; 1:1,000) were obtained from Abcam; LURAP1L (cat. no. PA5-55072; 1:1,000) was obtained from Thermo Fisher Scientific, Inc., and PDGFR-β (cat. no. bs-0232R; 1:1,000) and p-PDGFR-β (Tyr740; cat. no. bs-3323R; 1:1,000) were obtained from BIOSS.

Techniques: Real-time Polymerase Chain Reaction, Expressing, Western Blot, Over Expression, Derivative Assay, Activation Assay

Journal: Drug Design, Development and Therapy

Article Title:

miR-370-3p Alleviates Ulcerative Colitis-Related Colorectal Cancer in Mice Through Inhibiting the Inflammatory Response and Epithelial-Mesenchymal Transition

doi: 10.2147/dddt.s238124

Figure Lengend Snippet: Figure 3 Effect of miR-370-3p on inflammatory response and TLR4 signaling pathway in AOM/DSS-induced mice. (A) The expression of TNF-α, IL-1β, and IL-6 in colonic tissues of mice from the Sham group, UC-CRC group, Ad-control group, and Ad-miR-370-3p group was detected by qRT-PCR. (B) The production of TNF-α and PGE2 in the lysate of the tumor colon tissues were detected by ELISA. (C) The expression of TLR4 and COX-2 in the tumor colon tissues from each group was evaluated by using IHC staining. Scale bar=50 μm. (D) The expression levels of TLR4, COX-2, and pEGFR in the tumor colon tissues from each group were determined by Western blot, and the relative band intensity was analyzed.The comparison among groups was performed by One-way ANOVA. Data are presented as mean ± SD. ##p<0.01 compared with the Sham group, **p<0.01 compared with the Ad-control group.

Article Snippet: After blocking with 5% skim milk (Sangon Biotech, Shanghai China) at room temperature for 1 h, the membranes were incubated with corresponding primary antibodies incuding TLR4 (1:1000, proteintech, Wuhan, China), COX-2 (1:1000, proteintech, Wuhan, China), pEGFR (phosphorylated epidermal growth factor receptor, 1:1000, Affinity, Changzhou, Jiangsu, China), β-catenin (1:5000, proteintech, Wuhan, China), p53 (1:3000, proteintech, Wuhan, China), ki67 (1:1000, Affinity, Changzhou, Jiangsu, China), E-cadherin (1:10,000, proteintech, Wuhan, China), N-cadherin (1:5000, proteintech, Wuhan, China), Vimentin (1:5000, proteintech, Wuhan, China) and GAPDH (1:10,000, proteintech, Wuhan, China) overnight at 4° C. Then the membranes were washed by TBST and incubated with appropriate secondary HRPconjugated goat anti-rabbit or goat anti-mouse antibodies (1:3000, Solarbio, Beijing, China) for 1 h at 37 ° C. The proteins were visualized by using ECL Western Blotting Substrate (Solarbio, Beijing, China).

Techniques: Expressing, Control, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Immunohistochemistry, Western Blot, Comparison

Journal: Cancer cell

Article Title: Overcoming Resistance to Dual Innate Immune and MEK Inhibition Downstream of KRAS

doi: 10.1016/j.ccell.2018.08.009

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit monoclonal anti-phospho IGF1R β (Tyr1135) , Cell signaling technology , Cat# 3918S; RRID: AB_10548764.

Techniques: Control, Recombinant, Enzyme-linked Immunosorbent Assay, Cell Viability Assay, shRNA, Negative Control, Software, Binding Assay, Expressing

Journal: Biochemical and biophysical research communications

Article Title: Rebound pathway overactivation by cancer cells following discontinuation of PI3K or mTOR inhibition promotes cancer cell growth.

doi: 10.1016/j.bbrc.2019.04.044

Figure Lengend Snippet: Fig. 4. Blocking IGF-1R prevents drug washout mediated cancer cell growth. (A) Washout of PI3K or mTOR inhibitors increases IGF-1Rb phosphorylation. HT-29 cells were treated for six hours with DMSO, BYL719 (10 mM) (BYL), BKM-120 (500 nM) (BKM), PP242 (5 mM) or Ku0063794 (5 mM) (Ku). Following washout, cells were incubated for six hours, and cell lysates were collected and analyzed for IGF-1Rb phosphorylation and actin. (B) IGF-1R inhibition with NVP-AEW541 blocks AKT phosphorylation mediated by washout of PP242 or BKM-120. HT-29 cells were treated with DMSO, PP242 (5 mM) or BKM-120 (500 mM) for 6 h. Following drug washout, cells were treated as indicated with sapitinib (2 mM), NVP- AEW541 (1 mM) (NVP-AEW) or erlotinib (1 mM) for six hours, and AKT phosphorylation was assessed by Western blot in cell lysates. (C) NVP-AEW541 inhibits cancer cell pro- liferation induced by washout of PP242 or BKM-120. HT-29 cells were treated with DMSO, PP242 (5 mM) or BKM-120 (500 nM) for six hours. Following drug washout, cells were incubated with DMSO or NVP-AEW541 (1 mM) for an additional 48 h. Cell proliferation was assessed with an MTS proliferation assay. Columns: Mean cell proliferation of three independent experiments expressed as percentage of DMSO treated cancer cells. Bars: SD. White columns: DMSO treated cells, grey columns: cells treated with PP242, and black columns: cells treated with BKM-120. *p < 0.05 compared to control cells.

Article Snippet: Anti-phospho-AKT (#4060), anti-AKT (#2920), and anti phospho-IGF-1Rb (#3024) antibodies were from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Blocking Assay, Phospho-proteomics, Incubation, Inhibition, Western Blot, Proliferation Assay, Control

Journal: Journal of Biological Chemistry

Article Title: Silencing of Insulin-like Growth Factor-binding Protein-2 in Human Glioblastoma Cells Reduces Both Invasiveness and Expression of Progression-associated Gene CD24

doi: 10.1074/jbc.m609567200

Figure Lengend Snippet: FIGURE 2. Modulation of IGF/IGF-IR signaling caused by knockdown of IGFBP-2. The IGFBP-2-knockdown cells (IGFBP-2 KD) and control cells were treated with the indicated concentrations of IGF-I or IGF-II in serum-free con- dition, and extents of phosphorylation of IGF-IR (p-IGF-IR), Akt (p-Akt), and ERK1/2 (p-Erk1/2) were analyzed by immunoblot and a representative result of U251 was shown. The band intensity was measured and the ratio of phos- phorylated protein to the corresponding total protein was calculated.

Article Snippet: The following primary antibodies were used: anti-human IGFBP-2 mouse monoclonal antibody (C-10; Santa Cruz Biotechnology, Santa Cruz, CA), anti- -actinmousemonoclonal antibody (AC-74; Sigma), anti-phosphoERK1/2 rabbit monoclonal antibody (Thr185/Tyr187, clone AW39;Upstate Cell Signaling Solutions, Lake Placid, NY), antiphospho-IGF-IR rabbit polyclonal antibody (Tyr1131), antiIGF-IR rabbit polyclonal antibody, anti-phospho-Akt mouse monoclonal antibody (Ser473), anti-Akt rabbit polyclonal antibody, anti-phospho-ERK1/2 rabbit monoclonal antibody (Thr185/Tyr187, clone AW39; Upstate Cell Signaling Solutions, Lake Placid, NY), anti-ERK1/2 rabbit polyclonal antibody, anticaspase-3, and cleaved capase-3 (Asp175) rabbit polyclonal antibodies, and anti-heat shock protein 70 rabbit polyclonal antibody (Cell Signaling Technology, Danvers, MA).

Techniques: Knockdown, Control, Phospho-proteomics, Western Blot

Journal: Blood Advances

Article Title: Prevalence and impact of diabetes on survival of patients with multiple myeloma in different racial groups

doi: 10.1182/bloodadvances.2023010815

Figure Lengend Snippet: Progression of myeloma xenografts is faster in a mouse model of type 2 diabetes than without diabetes. (A) Representative MM1.S tumor xenograft growth trajectories from Rag1 −/− (Rag wild-type [WT]), and Rag1 −/− /MKR (Rag MKR) male mice; n = 3 to 5 per group. (B) Plasma insulin concentrations in Rag1 −/− and Rag1 −/− /MKR male mice; n = 3 to 5 per group. (C) Representative western blot analysis of MM1.S tumor xenograft protein lysates from Rag1 −/− and Rag1 −/− /MKR mice, as indicated. (D-E) Quantification of total insulin receptor expression corrected for β actin, and S6rp phosphorylation, relative to total S6rp protein levels. Results are expressed as relative difference to that of Rag1 −/− mice. (F) Representative western blot analysis of MM1.S tumor cell protein lysates from with and without insulin stimulation, as indicated. (G-I) Quantification of pIR/IGF-1R, pAkt, and pS6rp relative to total protein levels; n = 3 per group. ∗ P < .05 between groups; ∗∗∗ P < .001, as indicated.

Article Snippet: Primary antibodies and dilutions used were as follows: anti–phosphorylated IGF-1Rβ (Tyr1150/1151)/phosphorylated insulin receptor β (pIRβ) (Tyr1135/1136) (#3024, 1:1000, Cell Signaling Technology [CST], Danvers, MA), total IGF-1R (1:1000, #3027, CST), total IRβ (1:200, C-19, Santa Cruz Biotechnology, Santa Cruz, Dallas, TX), phosphorylated Akt (pAkt) (Ser473) (1:1000, #9271, CST), total Akt (1:2000, #2920, CST), phosphorylated S6 ribosomal protein (pS6rp) (Ser235/236) (1:1000, #2211, CST), total S6 ribosomal protein (1:1000, #2317, CST), and β-actin (1:10 000, A228, Sigma-Aldrich).

Techniques: Clinical Proteomics, Western Blot, Expressing, Phospho-proteomics