Journal: Experimental Hematology

Article Title: Synergistic action of dual IGF1/R and MEK inhibition sensitizes childhood acute lymphoblastic leukemia (ALL) cells to cytotoxic agents and involves downregulation of STAT6 and PDAP1

doi: 10.1016/j.exphem.2018.04.002

Figure Lengend Snippet: Predicted interactions of “synergy” genes STAT6 and PDAP1 . (A) Heat map showing that STAT6 and PDAP1 are highly differentially downregulated by AG1204 + U0126. (B) Pearson correlation coefficient analyses from 20 microarrays indicating that STAT6 and PDAP1 expression are highly codependent. (C) STAT6 and PDAP1 display common patterns of expression c-dependency with specific “synergy” genes highlighted in bold text. (Pearson coefficient r values: below 0.4 = weak; 0.4–0.6 = modest; above 0.6 = marked codependency). (D) Ingenuity network analysis predicting interaction of PDAP1 and STAT6 in a STAT6–ERK–NF-κB regulatory network seeded with “synergy” genes that might be involved in apoptosis resistance (green = downregulated; red = upregulated). * p ≤ 0.05; ** p ≤ 0.005; *** p ≤ 0.0005.

Article Snippet: Antibodies included IGF1Rβ (#3027), phospho-IGF1Rβ (Tyr1131)/insulin receptor β (Tyr1146) (#3021), ERK1/2 (137F5), phospho-ERK1/2 (Thr202/Tyr204) (D13.14.4E), PDAP1 (#4300), procaspase 7 (D2Q3L), cleaved caspase 3 (5A1E) (Cell Signaling Technology, Danvers, MA, USA), STAT6 (ab44718) (Abcam, Cambridge, MA, USA), and PARP1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Expressing

Journal: Experimental Hematology

Article Title: Synergistic action of dual IGF1/R and MEK inhibition sensitizes childhood acute lymphoblastic leukemia (ALL) cells to cytotoxic agents and involves downregulation of STAT6 and PDAP1

doi: 10.1016/j.exphem.2018.04.002

Figure Lengend Snippet: Independent qRT-PCR validation of the most differentially altered ‘synergy’ genes, STAT6 and PDAP1 , in ALL employed in the microarray analysis as well as others treated under the same conditions.

Article Snippet: Antibodies included IGF1Rβ (#3027), phospho-IGF1Rβ (Tyr1131)/insulin receptor β (Tyr1146) (#3021), ERK1/2 (137F5), phospho-ERK1/2 (Thr202/Tyr204) (D13.14.4E), PDAP1 (#4300), procaspase 7 (D2Q3L), cleaved caspase 3 (5A1E) (Cell Signaling Technology, Danvers, MA, USA), STAT6 (ab44718) (Abcam, Cambridge, MA, USA), and PARP1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Quantitative RT-PCR, Biomarker Discovery, Microarray

Journal: Experimental Hematology

Article Title: Synergistic action of dual IGF1/R and MEK inhibition sensitizes childhood acute lymphoblastic leukemia (ALL) cells to cytotoxic agents and involves downregulation of STAT6 and PDAP1

doi: 10.1016/j.exphem.2018.04.002

Figure Lengend Snippet: Cellular consequences of siRNA-mediated knockdown of synergy genes. (A) Western blot showing siRNA mediated knockdown of STAT6 and PDAP1 in HeLa cells. (B) siRNA-mediated knockdown of STAT6 results in a significant loss in cell viability, whereas PDAP1 knockdown has no impact after 1 week. (C) siRNA-mediated knockdown of STAT6 causes a 20% increase in cells in the G 1 phase of the cell cycle, indicating decreased proliferation, whereas PDAP1 knockdown has no impact on the cell cycle in HeLa cells. (D) Effect on loss of cell viability by STAT6 knockdown leads to an additive effect with daunorubicin after 72hours of treatment, whereas PDAP1 knockdown is synergistic with daunorubicin at more than one dose in HeLa cells. * p ≤ 0.05; ** p ≤ 0.005.

Article Snippet: Antibodies included IGF1Rβ (#3027), phospho-IGF1Rβ (Tyr1131)/insulin receptor β (Tyr1146) (#3021), ERK1/2 (137F5), phospho-ERK1/2 (Thr202/Tyr204) (D13.14.4E), PDAP1 (#4300), procaspase 7 (D2Q3L), cleaved caspase 3 (5A1E) (Cell Signaling Technology, Danvers, MA, USA), STAT6 (ab44718) (Abcam, Cambridge, MA, USA), and PARP1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Knockdown, Western Blot

Journal: Experimental Hematology

Article Title: Synergistic action of dual IGF1/R and MEK inhibition sensitizes childhood acute lymphoblastic leukemia (ALL) cells to cytotoxic agents and involves downregulation of STAT6 and PDAP1

doi: 10.1016/j.exphem.2018.04.002

Figure Lengend Snippet: Sensitization of HeLa cells by siRNA silencing of ‘synergy’ candidate genes, STAT6 and PDAP1. Graphs show the decrease in cell proliferation caused by STAT6 knockdown protects HeLa cells from vincristine-induce killing whereas PDAP1 knockdown has no effect compared with scrambled siRNA, following 72h treatment in vitro .

Article Snippet: Antibodies included IGF1Rβ (#3027), phospho-IGF1Rβ (Tyr1131)/insulin receptor β (Tyr1146) (#3021), ERK1/2 (137F5), phospho-ERK1/2 (Thr202/Tyr204) (D13.14.4E), PDAP1 (#4300), procaspase 7 (D2Q3L), cleaved caspase 3 (5A1E) (Cell Signaling Technology, Danvers, MA, USA), STAT6 (ab44718) (Abcam, Cambridge, MA, USA), and PARP1 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).

Techniques: Knockdown, In Vitro

Journal: PLoS ONE

Article Title: Proteomic analysis of rat colonic mucosa following acupuncture treatment for irritable bowel syndrome with diarrhea

doi: 10.1371/journal.pone.0273853

Figure Lengend Snippet: Important shared DE proteins in two comparing groups.

Article Snippet: After being sealed with 5% bovine serum albumin at room temperature for 2 hours, the membrane was incubated overnight with anti-ATP5a1 (1:2000, proteintech) and anti-BPNT1(1:2000, proteintech) at 4°C, and the membrane was hybridized with secondary antibodies bound to horseradish peroxidase.

Techniques: Polymer, Control

Journal: PLoS ONE

Article Title: Proteomic analysis of rat colonic mucosa following acupuncture treatment for irritable bowel syndrome with diarrhea

doi: 10.1371/journal.pone.0273853

Figure Lengend Snippet: Important MCL clusters of DEPs.

Article Snippet: After being sealed with 5% bovine serum albumin at room temperature for 2 hours, the membrane was incubated overnight with anti-ATP5a1 (1:2000, proteintech) and anti-BPNT1(1:2000, proteintech) at 4°C, and the membrane was hybridized with secondary antibodies bound to horseradish peroxidase.

Techniques: Membrane, Generated, Activity Assay, Functional Assay, Transferring, Gene Expression, Activation Assay, Phospho-proteomics, Clinical Proteomics, Produced, Cell Surface Receptor Assay, Transduction

Journal: PLoS ONE

Article Title: Proteomic analysis of rat colonic mucosa following acupuncture treatment for irritable bowel syndrome with diarrhea

doi: 10.1371/journal.pone.0273853

Figure Lengend Snippet: The protein level of ATP5a1 and BPNT1 were analyzed by western blot. A. Quantitative analysis of ATP5a1 and BPNT1 expression in colonic mucosa (each group, n = 4, ANOVA). B. All values were expressed as the mean ± SD, **P<0.01: compared with Group C; ##P<0.01: compared with Group M; ns: no significant. Bonferroni’s post hoc test was used for multiple comparisons.

Article Snippet: After being sealed with 5% bovine serum albumin at room temperature for 2 hours, the membrane was incubated overnight with anti-ATP5a1 (1:2000, proteintech) and anti-BPNT1(1:2000, proteintech) at 4°C, and the membrane was hybridized with secondary antibodies bound to horseradish peroxidase.

Techniques: Western Blot, Expressing

Journal: Cancer Science

Article Title: Pancreatic ductal adenocarcinoma cells regulated the gemcitabine‐resistance function of CAFs by LINC00460

doi: 10.1111/cas.15547

Figure Lengend Snippet: LINC00460 could directly bind with PDAP1. (A) FISH assay showed the sub‐cellular location of LINC00460 in CFPAC‐1. (U6: nucleus location positive control, 18S: cytoplasm control) The subcellular location of LINC00460 is mainly in cytoplasm, followed by the nucleus. (B) LINC00460 expression in the nucleus and cytoplasm fractions by qRT‐PCR were consistent with FISH results. Data from three independent experiments were expressed as mean ± SD. (C) RNA‐pull‐down‐MS results showed 291 proteins have the possibility of binding to LINC00460 in the intersection of three PDAC cell lines. (D) Volcano plot of RNA‐pull‐down‐MS revealed that PDAP1 (Log 10 [iBAQ] = 9.10) interacts with LINC00460. The red font indicates PDAP1. (E) RNA immunoprecipitation (RIP)‐qPCR assay detects the coupling of PDAP1 and LINC00460. (F) Predicted structure of the LINC00460 determined by RNAfold software and the prediction of LINC00460/PDAP1 interaction probabilities using RPIseq website. (G) Immunoblot of PDAP1 in RNA pull‐down extracts with different LINC00460 fragment sequences (1–237, 238–747, and 748–913).

Article Snippet: Thereafter, immunoblottings were performed with below primary antibodies against PDAP1 (#15081‐1‐AP, Proteintech), PDGFA (#203911, Abcam), PDGFR (#32570, Abcam), a PDGFR Activation Antibody Sampler Kit (#12651, CST), β‐actin (#4970, CST), and GAPDH (#60004‐1‐Ig, Proteintech) in TBST buffer containing 5% BSA overnight.

Techniques: Positive Control, Control, Expressing, Quantitative RT-PCR, Binding Assay, RNA Immunoprecipitation, Software, Western Blot

Journal: Cancer Science

Article Title: Pancreatic ductal adenocarcinoma cells regulated the gemcitabine‐resistance function of CAFs by LINC00460

doi: 10.1111/cas.15547

Figure Lengend Snippet: LINC00460 could regulate the proliferation of CAFs via the PDAP1/PDGFA/PDGFR pathway. (A) qRT‐PCR assay detected the mRNA level of PDAP1 and PDGFA after transduced LINC00460 shRNAs. (B) Western blot assay detected the protein level of PDAP1 and PDGFA after transduced LINC00460 shRNAs. (C) The concentration of PDGFA in the medium supernatant of CFAPC‐1 and PANC‐1 after silencing LINC00460 was measured by ELISA assay; values represent means ± SD of six replicate samples in each group. (D) Western blot assay validated KO‐PDAP1 cell lines. (E) Western blot assay showed the expression level of PDGFA in PANC‐1 cell line (KO‐PDAP1) after transduction of LINC00460 shRNAs. (F) ELISA assay showed the concentration of PDGFA in medium supernatant of PANC‐1 cell line (KO‐PDAP1) after transduction of LINC00460 shRNAs. Values represent means ± SD of six replicate samples in each group. (G) Western blot assay showed PDGFR‐β expression in eight pairs of PDAC CAFs and tumor cells. (H) Cell viability showed the different proliferation rate of CAFs after co‐cultured with the supernatant of CFPAC‐1 and PANC‐1. (I) Cell viability showed that crenolanib can reverse the proliferation of CAFs caused by silencing LINC00460. (J) Western blot analysis of the PDGFR‐β pathway activated genes in after transduced LIINC00460 shRNAs whose co‐cultured with CFPAC‐1 or PANC‐1 supernatant. (K) Western blot shows crenolanib can reverse PDGFR‐β pathway activation.

Article Snippet: Thereafter, immunoblottings were performed with below primary antibodies against PDAP1 (#15081‐1‐AP, Proteintech), PDGFA (#203911, Abcam), PDGFR (#32570, Abcam), a PDGFR Activation Antibody Sampler Kit (#12651, CST), β‐actin (#4970, CST), and GAPDH (#60004‐1‐Ig, Proteintech) in TBST buffer containing 5% BSA overnight.

Techniques: Quantitative RT-PCR, Western Blot, Concentration Assay, Enzyme-linked Immunosorbent Assay, Expressing, Transduction, Cell Culture, Activation Assay

Journal: Cancer Science

Article Title: Pancreatic ductal adenocarcinoma cells regulated the gemcitabine‐resistance function of CAFs by LINC00460

doi: 10.1111/cas.15547

Figure Lengend Snippet: Proposed schematic model illustrating the interaction between LINC00460 and PDAP1, leading to the increase of CAF proliferation, which could cause chemotherapy resistance.

Article Snippet: Thereafter, immunoblottings were performed with below primary antibodies against PDAP1 (#15081‐1‐AP, Proteintech), PDGFA (#203911, Abcam), PDGFR (#32570, Abcam), a PDGFR Activation Antibody Sampler Kit (#12651, CST), β‐actin (#4970, CST), and GAPDH (#60004‐1‐Ig, Proteintech) in TBST buffer containing 5% BSA overnight.

Techniques: