Journal: Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research

Article Title: Transfer, analysis and reversion of the fibrous dysplasia cellular phenotype in human skeletal progenitors

doi: 10.1359/jbmr.091036

Figure Lengend Snippet: Table III

Article Snippet: PDE10A , hCG34059 , Hs01098924_m1.

Techniques:

Journal: Neurobiology of disease

Article Title: Inhibition of CXCR2 enhances CNS remyelination via modulating PDE10A/cAMP signaling pathway.

doi: 10.1016/j.nbd.2023.105988

Figure Lengend Snippet: Fig. 3. PDE10A regulated differentiation and maturation in primary cultured OPCs. Primary OPCs were maintained for 5 days in differentiation medium containing 0.1 μM and 1 μM TAK-063 or DMSO as control. (A) Confocal results of A2B5 expression in primary cultured OPCs. (B) Confocal results of MBP expression in primary cultured OPCs. Cells were stained with A2B5 (green), MBP (red) and DAPI (blue). (Scale bar = 50 μm). (C, D) Western blot bands and statistical analysis of PDGFRα, MBP, PLP1, MOG and MOBP expression. Primary cultured OPCs were transfected with PDE10A or control cDNA for 6 h, and then incubated with SB225002 at 1 μM for another 24 h. (E) Western blot bands of PDGFRα and MBP in PDE10A overexpressed cells. (F, G) Relative value of PDGFRα and MBP intensity normalized by β-actin. Results are shown as the mean ± SD form at least five experiments, and the confocal assay was performed three independent times with duplication. *P < 0.05, **P < 0.01 and ***P < 0.001 vs. Control cells, &&&P < 0.01 vs. OPC overexpressed PDE10A. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: The blots were incubated with primary antibodies overnight at 4 ◦C, including CXCR2 (A3301), PLP1 (A14251), MAG (A16914), MOG (A5353), Caspr (A3721), SOX10 (A8655) (1:1000), β-actin (AC026, 1:100000) (Abclonal, China), MOBP (12472–1-AP) (1:500, Proteintech, China), PDE10A (sc-515,023), PDGFRα (sc-398,206) (1:500, Santa Cruze, USA), MBP (Ab40390), OLIG2 (Ab109186) (1:1000, Abcam, USA), pERK1/2 (4695 s), ERK1/2 (4370 s), Lamin B1 (13435 s) (1:1000, Cell signaling technology, USA).

Techniques: Cell Culture, Control, Expressing, Staining, Western Blot, Transfection, Incubation, Confocal Assay

Journal: Neurobiology of disease

Article Title: Inhibition of CXCR2 enhances CNS remyelination via modulating PDE10A/cAMP signaling pathway.

doi: 10.1016/j.nbd.2023.105988

Figure Lengend Snippet: Fig. 4. Inhibition of PDE10A promoted remyelination in EB intoxicated rats. MS model rats were induced with EB injection into right lateral ventricle and then treated with TAK-063 (2 mg/kg) for 14 days. (A) Rotarod test of rats treated with TAK-063 (n = 10). (B, D) LFB staining and statistical analysis of demyelinated area in the rat corpus callosum. (C, E) Expression and calculation of the mean IOD of MBP in the rat corpus callosum. (F) Western blot assay of PDGFRα, MBP, MOG, MOBP and Caspr expression in the rat corpus callosum. (G) Relative value of PDGFRα, MBP, MOG, MOBP and Caspr intensity normalized by β-actin. Results are presented as the mean ± SD from at least four rats. **P < 0.01 and ***P < 0.001 vs. Control rats, ##P < 0.01 and ###P < 0.001 vs. EB-injected rats.

Article Snippet: The blots were incubated with primary antibodies overnight at 4 ◦C, including CXCR2 (A3301), PLP1 (A14251), MAG (A16914), MOG (A5353), Caspr (A3721), SOX10 (A8655) (1:1000), β-actin (AC026, 1:100000) (Abclonal, China), MOBP (12472–1-AP) (1:500, Proteintech, China), PDE10A (sc-515,023), PDGFRα (sc-398,206) (1:500, Santa Cruze, USA), MBP (Ab40390), OLIG2 (Ab109186) (1:1000, Abcam, USA), pERK1/2 (4695 s), ERK1/2 (4370 s), Lamin B1 (13435 s) (1:1000, Cell signaling technology, USA).

Techniques: Inhibition, Injection, Staining, Expressing, Western Blot, Control

Journal: Frontiers in Systems Neuroscience

Article Title: Traumatic Brain Injury Upregulates Phosphodiesterase Expression in the Hippocampus

doi: 10.3389/fnsys.2016.00005

Figure Lengend Snippet: PDE expression changes after TBI in the hippocampus. (A,C,E) Representative western blots of the ipsilateral hippocampus analyzed for PDE1A, 1B, 1C, 3A, 8A, 8B, and 10A levels. Each of the corresponding β-actin western blots are shown below the PDE western blots. (B,D,F) Densitometry results. Significant increases in PDE1A were observed at 30 min, 1 h and 6 h post-injury. PDE10A decreased at 1 h post-injury. n = 6/group, ∗ p < 0.05, ∗∗ p < 0.01 vs. Sham; one-way ANOVA with Tukey’s HSD correction for multiple comparisons.

Article Snippet: Proteins were transferred to Immobilon-P membranes (EMD Millipore) and membranes were incubated with the following primary antibodies: β-actin (AC-15, 1:10,000, Sigma–Aldrich), PDE1A (sc-50480, 1:4,000, Santa Cruz Biotechnology), PDE1B (ab14600, 1:500, Abcam; ), PDE1C (sc-67323, 1:500, Santa Cruz Biotechnology; ), PDE3A (sc-20792, 1:250, Santa Cruz Biotechnology; ), phospho-PDE4A (GTX14610, 1:2,000, GeneTex), PDE4A5 (ab42094, 1:2,000, Abcam; ), PDE4A8 (GTX14606, 1:1,000, GeneTex), PDE4B (sc-25812, 1:500, Santa Cruz Biotechnology; ), PDE4D (sc-25814, 1:500, Santa Cruz Biotechnology; ), phospho-PDE4D (ab59212, 1:1,000, Abcam), PDE8A (sc-30059, 1:500, Santa Cruz Biotechnology; ), PDE8B (sc-17234, 1:500, Santa Cruz Biotechnology; ), and PDE10A (sc-67298, 1:250, Santa Cruz Biotechnology; ).

Techniques: Expressing, Western Blot

Journal: Journal of Neurotrauma

Article Title: Intravenous Infusion of Mesenchymal Stem Cells Alters Motor Cortex Gene Expression in a Rat Model of Acute Spinal Cord Injury

doi: 10.1089/neu.2018.5793

Figure Lengend Snippet: The neuron-restrictive silencer factor (NRSF) binding domains in some “behaviorally-associated differentially expressed genes (DEGs).” Genomic loci of NRSF binding domains within 20 kb from the transcription start of the genes including (A) Kcnip2 (Base genome: human chromosome: chr10 103,561,373–103,626,222), (B) Scn3b (Base genome: human chromosome: chr11 123,481,778–123,540,867), and (C) Pde10a (Base genome: human chromosome: chr6 165,727,008–166,409,519) (red arrows) are shown. The binding motifs of NRSF are indicated with red characters.

Article Snippet: TaqMan ® Universal Master Mix II with Uracil-N glycosylase (UNG), and TaqMan ® Gene Expression assays were purchased from Thermo Fisher Scientific Inc. ( Gapdh , Rn01775763_g1; Kcnip2 , Rn01411450_g1; lipid phosphate phosphatase-related protein type 4 [ Lppr4 ], Rn01522267_m1; Pde10a , Rn00673152_m1; calcium binding protein 7 [ Cabp7 ], Rn01443564_m1; plakophilins-2 [ Pkp2 ], Rn01404502_m1; Scn3b , Rn01422019_m1; nuclear receptor binding protein 2 [ Nrbp2 ], Rn01505756_m1; Sik1 , Rn00572495_m1; Fos , Rn02396759_m1; Dusp1 , Rn00678341_g1; Btg2 , Rn00568504_m1; Ier2 , Rn02531674_s1; Cyr61 , Rn00580055_m1; Apold1 , Rn02131262_s1; Id2, Rn01495280_m1). qRT-PCR was performed in triplicate using PRISM7500 with 7500 software v2.3 (Thermo Fisher Scientific Inc.).

Techniques: Binding Assay

Journal: PLoS ONE

Article Title: HPRT-Deficiency Dysregulates cAMP-PKA Signaling and Phosphodiesterase 10A Expression: Mechanistic Insight and Potential Target for Lesch-Nyhan Disease?

doi: 10.1371/journal.pone.0063333

Figure Lengend Snippet: ( A&B ) Immuno-blot and quantification analysis for various PDEs that can affect cAMP/PKA signaling. Data show that the expression of PDE1C, PDE4B, and PDE7B are similar between control (parent) and HPRT-deficient (mutant) MN9D cell lines. Expression of PDE10A protein is significantly increased in HPRT-deficient cells. Error bars represent mean ± SEM of duplicate measurements of two independent experiments (n = 4). The asterisk (*) represents the statistical significant between the control (open bars) and HPRT-deficient (closed bars) MND9 cells (p<0.05 t-test).

Article Snippet: 2×10 5 HPRT-deficient MN9D cells in 6 well-plate were exposed to 80 pmole of control (scramble siRNA) or PDE10A siRNA (directed to mouse PDE10A) from Santa cruz biotechnology and transfected according to the established transfection protocol ( http://datasheets.scbt.com/siRNA_protocol.pdf ).

Techniques: Expressing, Control, Mutagenesis

Journal: PLoS ONE

Article Title: HPRT-Deficiency Dysregulates cAMP-PKA Signaling and Phosphodiesterase 10A Expression: Mechanistic Insight and Potential Target for Lesch-Nyhan Disease?

doi: 10.1371/journal.pone.0063333

Figure Lengend Snippet: ( A & B ), immuno-blot and quantification analysis of p-Syn (Ser9), data show that the lower expression of phospho-PKA substrate and p-Syn (Ser9) in response to forskolin treatment in HPRT-deficient MN9D cells is restored in the presence of Papaverine (200 µM). Error bars represent mean ± SEM of duplicate measurements of two independent experiments (n = 4). The asterisks (*) represent statistical significance between forskolin treated cells without papaverine treatment (p<0.05, t-test). ( C & D ), immuno-blot and quantification analysis of PDE10A expression after transfection of HPRT-deficient MN9D cells with siRNA directed to the mouse PDE10 gene. The data show lower expression PDE10A protein in cells transfected with siRNA-PDE10A relative to cells transfected with scramble control siRNA (siRNA-CTL). Error bars represent mean ± SEM of duplicate measurements carried out independently twice (n = 4). The asterisks (*) represent the statistical significant (p<0.05, t-test) between the control (open bar, siRNA-CTL) and siRNA-PDE10A transfected cells (closed bar). This significant reduction of PDE10A protein contributes to the enhanced expression of phospho-PKA-substrates including p-Syn observed in ( E & F ). Error bars represent mean ± SEM of duplicate measurements of two independent experiments (n = 4). The asterisk (*) represents the statistical significant (p<0.05, t-test) between the siRNA-CTL and siRNA-PDE10A HPRT-deficient transfected MND9 cells treated with forskolin.

Article Snippet: 2×10 5 HPRT-deficient MN9D cells in 6 well-plate were exposed to 80 pmole of control (scramble siRNA) or PDE10A siRNA (directed to mouse PDE10A) from Santa cruz biotechnology and transfected according to the established transfection protocol ( http://datasheets.scbt.com/siRNA_protocol.pdf ).

Techniques: Expressing, Transfection, Control

Journal: Cell Reports Methods

Article Title: Antibody-assisted selective isolation of Purkinje cell nuclei from mouse cerebellar tissue

doi: 10.1016/j.crmeth.2024.100816

Figure Lengend Snippet: qPCR quantification in SCA7 PC nuclei unveils transcriptional changes in both known and new disease-associated genes (A) qPCR quantification of selected disease-linked genes in PC nuclei isolated from SCA7 and WT mouse cerebella. n = 4 cerebella for each group; 4,000 nuclei were sorted per animal. (B) qPCR quantification of Pde1c , Pde4d , Pde9a , and Pde10a PC nuclei isolated from SCA7 and WT mouse cerebella. n = 7 cerebella for each group; 4,000 nuclei were sorted per animal. Data are represented as mean ± SEM.

Article Snippet: Pde10a TaqMan Assay , Thermo Fisher Scientific , Cat#: Mm00449329_m1.

Techniques: Isolation

Journal: Cell Reports Methods

Article Title: Antibody-assisted selective isolation of Purkinje cell nuclei from mouse cerebellar tissue

doi: 10.1016/j.crmeth.2024.100816

Figure Lengend Snippet:

Article Snippet: Pde10a TaqMan Assay , Thermo Fisher Scientific , Cat#: Mm00449329_m1.

Techniques: Recombinant, Reverse Transcription, TaqMan Assay, Microscopy, Western Blot, Software

Journal: Journal of Cheminformatics

Article Title: Structure‐based identification of dual ligands at the A 2A R and PDE10A with anti‐proliferative effects in lung cancer cell‐lines

doi: 10.1186/s13321-021-00492-5

Figure Lengend Snippet: The Structures of the identified PDE10A inhibitors with the potential to bind to the A 2A R, and their pharmacology at PDE10A. a Virtual screening protocol. b Chemical structures for the six compounds identified in the in silico screen, literature references, compound IDs (used here) and pIC 50 for PDE10A inhibition. c Concentration-response curves generated for 1–6 and CGS21680 at PDE10A. Data is the mean of six individual replicates ± SEM. d Representative binding modes proposed for the triazoloquinazolines 1 and 4 docked to the PDE10A crystal structure (PDB ID: 4DDL). Yellow spheres indicate lipophilic contacts. Aromatic interactions are illustrated by purple disks and hydrogen bond acceptors are shown as red arrows. Tyr 683 is part of the “selectivity pocket” of PDE10A , and its interaction through a hydrogen bond with the thioether of both compounds could explain their subtype selectivity

Article Snippet: A PDE10A assay kit (BPS Bioscience, San Diego, CA) was used to test the PDE10A inhibition of compounds 1 – 6 as described in the manufactures protocol.

Techniques: In Silico, Inhibition, Concentration Assay, Generated, Binding Assay

Journal: Journal of Cheminformatics

Article Title: Structure‐based identification of dual ligands at the A 2A R and PDE10A with anti‐proliferative effects in lung cancer cell‐lines

doi: 10.1186/s13321-021-00492-5

Figure Lengend Snippet: Characterisation of ligands targeting A 2A R/PDE10A using a NanoBRET-based ligand binding assay. a Kinetic binding curve of CA200645 at Nluc-A 2A R expressed HEK293T cells. After 19 minutes association with 40 nM CA200645, CGS21680 was injected to give a final concentration of 10 µM in order to displace the fluorescent probe. The curve was fit into “association then dissociation” model built in Prism 8.4.3. b Competition of CA200645 (300 nM) by reference compounds including CGS21680, NECA, and isoprenaline at equilibrium. c Competitive binding curves of triazoloquinazolines in correspond to of 300 nM CA200645. Both curves (panel B and C) were fitted using the “one-site Ki” equation where K D and concentration of hot ligand were set to 65 nM and 300 nM, respectively. Data points are the mean ± SEM from 3–27 repeats performed in duplicate. (D) The summary of binding affinities (pK i ) of tested ligands. pK i values were calculated from inhibition of CA200645 binding at equilibrium to Nluc-A 2A R-expressed HEK293T cells. # Cmpd 5 did not fully displace binding of CA200645 under condition tested. Statistical significance (* p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001) compared to CGS21680 was determined by one-way ANOVA with Dunnett’s post-test

Article Snippet: A PDE10A assay kit (BPS Bioscience, San Diego, CA) was used to test the PDE10A inhibition of compounds 1 – 6 as described in the manufactures protocol.

Techniques: Ligand Binding Assay, Binding Assay, Injection, Concentration Assay, Inhibition

Journal: Journal of Cheminformatics

Article Title: Structure‐based identification of dual ligands at the A 2A R and PDE10A with anti‐proliferative effects in lung cancer cell‐lines

doi: 10.1186/s13321-021-00492-5

Figure Lengend Snippet: Lung squamous cell carcinoma and adenocarcinoma cells display increasing sensitivity to compounds , 3–5 in terms of proliferation, dependent upon combined A 2A R and PDE10A expression. Lung squamous cell carcinoma cells (LK-2 and H520) and lung adenocarcinoma cells (H1792 and H1563 were subjected to semi-quantitative RT-PCR analysis to determine expression of the A R, A 2A R, A 2B R, A 3 R and PDE 10A, data represented relative to GAPDH expression, ± SEM of 3 individual replicates. Further, each cell line was stimulated with CGS21680 or compounds , 3–6 for 30 minutes and cAMP levels determined. Data represented relative to the response obtained upon stimulation with 100 µM Forskolin, ± SEM of 4–8 individual replicates. Additionally, all cells were stimulated with CGS21680, or , 3–6 for 72 hours and cell number determined using CCK-8. Data represented as a percentage of the cell number present after treatment with 1 % DMSO, ± SEM of 4–8 replicates. (B) Correlation plot for pEC 50 of each compounds ability to stimulate cAMP production vs. its pIC 50 for inhibiting proliferation. Data represented ± SEM. (C) Proliferation factor (pIC50 x span anti-proliferative Additional file : Table S5) calculated for , 3–6 , CGS21680s and forskolin at LK-2, H520, H1792 and H1563 cells. Bars represent the mean I max ± SEM, whilst individual data points are shown as a scatter plot.

Article Snippet: A PDE10A assay kit (BPS Bioscience, San Diego, CA) was used to test the PDE10A inhibition of compounds 1 – 6 as described in the manufactures protocol.

Techniques: Expressing, Quantitative RT-PCR, CCK-8 Assay