Journal: International Journal of Molecular Sciences

Article Title: Follistatin-Like-1 (FSTL1) Is a Fibroblast-Derived Growth Factor That Contributes to Progression of Chronic Kidney Disease

doi: 10.3390/ijms22179513

Figure Lengend Snippet: Fstl1 and cognate receptor expression. ( A ) Heatmap with unsupervised hierarchical cluster analysis of Fstl1 , Dip2a , Cd14 , and Tlr4 genes in kidneys of 4 and 7 week old Col4a3 -/- and wild-type mice ( n = 8 per group). ( B – E ) Graphical representation of microarray expression in 4 week old wild type versus 4 week old Col4a3 -/- mice. ( F – I ) Graphical representation of microarray expression in 7 week old wild type versus 7 week old Col4a3 -/- mice. Values are mean ± SEM, and significance was defined as a p value of < 0.05 by Student’s t tests.

Article Snippet: The mouse TaqMan Gene Expression Assays that were used include: Mm00433371_m1; Fstl1 , Mm00445273_m1; Tlr4 , Mm01150153_m1; Dip2a , Mm00441242_m1; Ccl2, Mm00443258_m1; Tnfa, Mm00725412_s1; Acta2 , Mm01178820_m1; Tgfb1 , Mm00801666_g1; Col1a1 , Mm01256744_m1; Fn1 .

Techniques: Expressing, Microarray

Journal: International Journal of Molecular Sciences

Article Title: Follistatin-Like-1 (FSTL1) Is a Fibroblast-Derived Growth Factor That Contributes to Progression of Chronic Kidney Disease

doi: 10.3390/ijms22179513

Figure Lengend Snippet: Fstl1 expression and localization in UUO. ( A – C ) mRNA levels for Fstl1 and its putative receptors ( Tlr4 and Dip2a ) were determined by quantitative polymerase chain reaction in kidneys of C57B6 mice (sham n = 4, UUO n = 5). Values are the mean ± SEM (black bars). p values were determined by Student’s t test, and significance was defined as a p value of < 0.05. ( D ) Light microscopic images at three magnifications of RNASCOPE ® Fstl1 localization in sham-operated mice (upper three panels) and mice subjected to UUO for 7 days (lower three panels).

Article Snippet: The mouse TaqMan Gene Expression Assays that were used include: Mm00433371_m1; Fstl1 , Mm00445273_m1; Tlr4 , Mm01150153_m1; Dip2a , Mm00441242_m1; Ccl2, Mm00443258_m1; Tnfa, Mm00725412_s1; Acta2 , Mm01178820_m1; Tgfb1 , Mm00801666_g1; Col1a1 , Mm01256744_m1; Fn1 .

Techniques: Expressing, Real-time Polymerase Chain Reaction, RNAscope

Journal: Cancer Cell

Article Title: GKAP Acts as a Genetic Modulator of NMDAR Signaling to Govern Invasive Tumor Growth

doi: 10.1016/j.ccell.2018.02.011

Figure Lengend Snippet: Differential GKAP Expression between the C57/BL6 and C3HeB/Fe Genetic Backgrounds Is Associated with a Differential NMDAR Pathway Activity In Vitro (A) qRT-PCR of Dlgap1 mRNA (upper) and western blot for GKAP protein expression (lower) in mPanNET tumor-derived cancer cell lines (βTC-B6 and βTC-C3H) or primary tumors that arose in RIP1Tag2 transgenic mice inbred into the B6 and C3H backgrounds, respectively. ∗ p < 0.05; ∗∗ p < 0.01 (n = 3 individual tumors/genetic background; n = 3 independent RNA extraction/cell line). (B) qRT-PCR analysis of FACS-sorted cell types from primary tumors derived from B6 mice. Cells were sorted from pools of multiple PanNETs isolated from two mice. One-way ANOVA, Dunnett multiple comparisons test was used when cancer cells were compared with all other populations (p < 0.0001 in all comparisons). (C) Upper panel: a region within the Dlgap1 gene sequence containing a SNP site, as shown in red. Putative HSF1 binding domains (p < 0.004) are shown by the green circles. Lower panel: ChIP-qPCR for the Dlgap1 SNP site after immunoprecipitation with an anti-HSF1 antibody. The βmaj (β globin, Hbb-b1 ) promoter region was used as negative control. Mann-Whitney test: ∗ p = 0.02 (n = 4, two batches of cell lysates per cell line, and two qPCR/batch). (D) Western blot for HSF1 and GKAP in βTC-B6 cells. Expression levels were normalized to GAPDH and small interfering RNA (siRNA) control (n = 3 independent experiments). (E) In vitro invasion assay of βTC-B6 and βTC-C3H cells, under either static or flow conditions. Two-way ANOVA, Bonferroni multiple comparisons test: n.s, not significant; ∗∗∗ p < 0.001 (n = 4 independent assays for static condition; n = 6–9 for flow condition). (F) Glutamate secretion by βTC-B6 and βTC-C3H cells under static and flow conditions, sampled from invasion assays. Two-way ANOVA, Bonferroni multiple comparisons test: ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; n.s., not significant (n = 3 invasion assay devices/condition/cell line). All bar graphs represent the mean ± SEM. See also Figure S1 .

Article Snippet: DLGAP1 (mouse) Taqman Probe , Thermo Fisher Scientific , # 4331182, Assay ID Mm00510688_m1.

Techniques: Expressing, Activity Assay, In Vitro, Quantitative RT-PCR, Western Blot, Derivative Assay, Transgenic Assay, RNA Extraction, Isolation, Sequencing, Binding Assay, ChIP-qPCR, Immunoprecipitation, Negative Control, MANN-WHITNEY, Small Interfering RNA, Control, Invasion Assay

Journal: Cancer Cell

Article Title: GKAP Acts as a Genetic Modulator of NMDAR Signaling to Govern Invasive Tumor Growth

doi: 10.1016/j.ccell.2018.02.011

Figure Lengend Snippet: High GKAP Expression Is Associated with Increased NMDAR Pathway Activity In Vivo (A) qRT-PCR evaluation for the NMDAR subunits GluN1 ( Grin1 ), GluN2b ( Grin2b ), and the scaffold protein GKAP ( Dlgap1 ) in PanNET tumors from the two genetic backgrounds. Mean ± SEM. Mann-Whitney test was used to compare the expression of each gene in the B6 and C3H tumors. ∗ p = 0.0175 (qRT-PCR: n = 7 tumors/7 mice/background). (B) Western blot of GluN2b and GKAP in PanNETs from B6 and C3H backgrounds. After normalization, the one-column t test was used for comparison, hypothetical value = 1; ∗ p = 0.01; n.s., not significant (mean ± SEM; n = 4 tumors/4 mice). (C) Immunohistochemistry (IHC) analysis of large T oncoprotein, GluN2b, and p-GluN2b in PanNET tumor tissue sections. Images are representative of >50 tumors from >10 RIP1Tag2 mice/background. S, spleen; LN, lymph node. (D) MK801 treatment in RIP1Tag2 mice. Cohorts of seven to nine mice were used for control (saline treated) and MK801 treatment in each genetic background; mean ± SEM. Mann-Whitney test: ∗∗ p < 0.01; n.s., not significant.

Article Snippet: DLGAP1 (mouse) Taqman Probe , Thermo Fisher Scientific , # 4331182, Assay ID Mm00510688_m1.

Techniques: Expressing, Activity Assay, In Vivo, Quantitative RT-PCR, MANN-WHITNEY, Western Blot, Comparison, Immunohistochemistry, Control, Saline

Journal: Cancer Cell

Article Title: GKAP Acts as a Genetic Modulator of NMDAR Signaling to Govern Invasive Tumor Growth

doi: 10.1016/j.ccell.2018.02.011

Figure Lengend Snippet:

Article Snippet: DLGAP1 (mouse) Taqman Probe , Thermo Fisher Scientific , # 4331182, Assay ID Mm00510688_m1.

Techniques: Produced, Microarray, Recombinant, shRNA, Software, Plasmid Preparation, Staining

Journal: OncoImmunology

Article Title: CD200 and CD200R1 are differentially expressed and have differential prognostic roles in non-small cell lung cancer

doi: 10.1080/2162402x.2020.1746554

Figure Lengend Snippet: Figure 4. Evaluation of CD200R1 functions with CD200R1 knockdown and CD200Fc administration. (a) Western blots in the left part showing the representing protein levels after CD200R1 knockdown with siRNA in PC9 and H358 cells, respectively. Bar graphs on the right part show western blotting quantification of pAKT/AKT and pERK/ERK in the siRNA1 and siRNA2 groups relative to those in negative controls (NCs). The data represent the mean ± SD, N = 4. *P <.05 and **P <.001 vs. NC (one-way ANOVA). (b–c) Effect of CD200R1 knockdown with siRNA on cell proliferation in PC9 and H358 cells as analyzed by CCK-8 assays. The negative control (NC) was scramble RNA-transfected cells. The data represent the mean ± SD, N = 5. *P <.05 and **P <.001 vs. NC (one-way ANOVA). (d) Effect of CD200Fc treatment on cell proliferation in PC9 cells as analyzed by CCK-8 assays. (e) Effect of CD200Fc treatment on endogenous mRNA expression levels of immune markers in PC9 cells as analyzed by RT-qPCR. Gene expression was normalized to the expression of GAPDH and is shown relative to vehicle control expression. The data represent the mean ± SD, N = 3. *P <.05 and **P <.001 vs. vehicle (Student’s t-test).

Article Snippet: To investigate the binding of CD200 to CD200R1, we used recombinant human CD200Fc protein (cat no. 2724-CD; R&D Systems, Minneapolis, MN, USA).

Techniques: Knockdown, Western Blot, CCK-8 Assay, Negative Control, Transfection, Expressing, Quantitative RT-PCR, Gene Expression, Control

Journal: OncoImmunology

Article Title: CD200 and CD200R1 are differentially expressed and have differential prognostic roles in non-small cell lung cancer

doi: 10.1080/2162402x.2020.1746554

Figure Lengend Snippet: Figure 5. Enriched gene profiles in tumors with high CD200R1 expression and differentially-expressed genes in response to CD200Fc administration as assessed by cDNA microarray. (a) Volcano plots showing the significantly overexpressed genes among tumors with high CD200R1 expression using online RNA sequencing data (NSCLC, TCGA, Provisional) including 230 adenocarcinomas (ADCs), and 501 squamous cell carcinomas (SCCs). The overexpressed genes in high CD200R1-expressing tumors are surrounded by dashed lines in the volcano plots, and these were additionally analyzed based on GSEA Investigation gene set analysis using the hallmark gene set. (b) Log2 fold expression changes of the 35 most strongly up- and downregulated genes in PC9 cells treated with CD200Fc versus expression in cells treated with vehicle (N = 2). (c) GSEA analysis comparing up- and downregulated cancer hallmark gene sets and oncogenic signature gene sets in PC9 cells treated with CD200Fc versus expression in cells treated with vehicle. (d–e) Expression of certain genes differentially-expressed upon CD200Fc administration in PC9 cells based on validation by RT-qPCR. Gene expression was normalized to the expression of GAPDH and is shown relative to the vehicle-treated control expression. The data represent the mean ± SD, N = 3. *P <.05 and **P <.001 vs. vehicle (Student’s t-test).

Article Snippet: To investigate the binding of CD200 to CD200R1, we used recombinant human CD200Fc protein (cat no. 2724-CD; R&D Systems, Minneapolis, MN, USA).

Techniques: Expressing, Microarray, RNA Sequencing, Biomarker Discovery, Quantitative RT-PCR, Gene Expression, Control