Journal: International Journal of Neuropsychopharmacology

Article Title: Identification of Phosphodiesterase-7A (PDE7A) as a Novel Target for Reducing Ethanol Consumption in Mice

doi: 10.1093/ijnp/pyae032

Figure Lengend Snippet: Pharmacological inhibition of PDE7A attenuated ethanol responsiveness and consumption in female mice. (A) The locomotor trajectories of female mice were recorded at 5 minutes after i.p. injection of saline or EtOH or EtOH (i.p. 2.5g/kg) with BRL50481 (i.p. 5 mg/kg). (B) Left, locomotor activity of C57BL/6J female mice for 15 minutes before injecting saline or EtOH or EtOH with BRL50481 and 40 minutes after. The shaded box highlights that the locomotor activity is required to be analyzed by minutes (1-way ANOVA, time = 20, 25, 30, 35, and 40; F (2,21) = 0.64, 1.65, 5.65, 0.06, and 4.91; * P < .05, ** P < .01, *** P < .001). Right, locomotor activity of female mice analyzed in minute (1-way ANOVA, F (2,21) = 1.10, P = .0379). (C) BRL50481 was administered at a dose of 5 mg/kg at 2 hours before EtOHl injection (i.p., 3.5 g/kg). (D) Time required for the loss of righting reflex following administration of EtOH or EtOH combined with BRL50481 in female mice ( t test, P > .05). (E) Time required for the recovery of righting reflex following administration of EtOH or EtOH combined with BRL50481 in female mice ( t test, P = .0152). (F) Consumption of 20% EtOH in male and female mice of the DID experiment ( t test, day = 3,4; * P < .05, ** P < .01). (G) 2BC paradigm used to examine the effect of BRL50481 on EtOH consumption and preference rate in female mice. (H–I) The daily EtOH intake and preference rate for 10% EtOH in female mice treated with BRL50481 over a period of 7 days ( t test, * P < .05, ** P < .01). (J) The BEC for 10% ethanol on the seventh day of injecting BRL50481 in C57BL/6J female mice ( t test, P = .0009). (K–L) Paired t test for analyzing EtOH intake and preference rate after BRL50481 administration for 7 days in female mice (Wilcoxon test, * P < .05). (M–N) Paired t test to analyze EtOH intake and preference rate in female mice administrated with the vehicle (Wilcoxon test, P > .05). N = 8 mice for each group. All data shown are means ± SEM.

Article Snippet: BRL50481, purchased from MCE (Cat# HY-109586, Monmouth Junction, NJ, USA), was dissolved in saline containing 1% dimethyl sulfoxide (DMSO) (Cat# PWL064, Mei Lun, China).

Techniques: Inhibition, Injection, Saline, Activity Assay

Journal: International Journal of Neuropsychopharmacology

Article Title: Identification of Phosphodiesterase-7A (PDE7A) as a Novel Target for Reducing Ethanol Consumption in Mice

doi: 10.1093/ijnp/pyae032

Figure Lengend Snippet: PDE7A inhibitor regulated the drinking behavior by the cAMP-PKA/Epac2 pathways and downstream phosphorylation of CREB and ERK. (A–B) The protein expression level of PDE7A in the striatum of female mice was quantified via western-blot analysis following the 2BC test (1-way ANOVA, F (2,12) = 5.16, * P < .05). (C) ELISA was used to measure the cAMP level in the striatum (1-way ANOVA, F (2,12) = 6.16, * P < .05). (D–F) The protein levels of pPKA, PKA, pCREB, and CREB in the striatum of female mice were quantified following exposure to ethanol and treatment with BRL50481 (1-way ANOVA, pPKA/PKA, F (2,12) = 6.36; pCREB/CREB, F (2,12) = 6.98; **** P < .0001). (G–I) The protein levels of Epac2, pERK, and ERK in the striatum of female mice were quantified following exposure to EtOH and treatment with BRL50481 (1-way ANOVA, Epac2, F (2,12) = 1.69; pERK/ERK, F (2,12) = 7.4; * P < .05, **** P < .0001). N = 6 mice for each group. All data shown are means ± SEM, the ratio of PDE7A to the respective GAPDH in each lane was calculated for statistical analysis.

Article Snippet: BRL50481, purchased from MCE (Cat# HY-109586, Monmouth Junction, NJ, USA), was dissolved in saline containing 1% dimethyl sulfoxide (DMSO) (Cat# PWL064, Mei Lun, China).

Techniques: Phospho-proteomics, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Theranostics

Article Title: The peroxisome proliferator-activated receptor agonist rosiglitazone specifically represses tumour metastatic potential in chromatin inaccessibility-mediated FABP4-deficient gastric cancer

doi: 10.7150/thno.66814

Figure Lengend Snippet: Identification of CADM3 as a function-related target of FABP4. (A) Volcano plot (left panel) and lollipop chart (right panel) showing the correlations between FABP4 the differentially expressed genes identified by RNA-seq. (B) The results of real-time PCR verifying that the mRNA levels of FABP4 and CADM3 were significantly positively correlated in 36 GC samples. (C) Representative images of FABP4 and CADM3 staining in tissue microarrays are presented (left panel), and CADM3 expression in various FABP4 groups of the internal cohort (right panel) was calculated. (D) Scatter plots showing the correlations between FABP4 and CADM3 expression levels in the TCGA-STAD dataset. (E) Peak signals indicating that FABP4 did not directly bind to the CADM3 promoter region. (F) Gene set enrichment analysis of RNA-seq data and the PPAR signalling pathway is presented. (G) MGC-803 cells were pretreated with or without fenofibrate (50 μM), GW0742 (10 μM), and rosiglitazone (20 μM) for 24 h. The cells were used for real-time PCR to detect the effects of various PPAR subtype agonists on CADM3 transcription. The data are presented as the mean ± SD (** P < 0.01).

Article Snippet: One week after the transplantation, the mice were treated daily with DMSO (Solarbio, Beijing, China) or rosiglitazone (10 mg/kg or 20 mg/kg; MedChemExpress, Shanghai, China) via intraperitoneal injection.

Techniques: RNA Sequencing, Real-time Polymerase Chain Reaction, Staining, Expressing

Journal: Theranostics

Article Title: The peroxisome proliferator-activated receptor agonist rosiglitazone specifically represses tumour metastatic potential in chromatin inaccessibility-mediated FABP4-deficient gastric cancer

doi: 10.7150/thno.66814

Figure Lengend Snippet: FABP4 promotes PPAR-γ-mediated CADM3 transcription. (A) A coimmunoprecipitation assay was performed to analyse the interaction between FABP4 and PPAR-γ in GC cells. (B) The results of immunofluorescence staining indicating that FABP4 facilitates the translocation of PPAR-γ from the cytoplasm to the nucleus. Scale bars, 20 μm. (C) The protein levels of FABP4 and PPAR-γ in GC cells measured by western blotting. (D) MGC-803 cells with or without altered FABP4 expression were treated with 25 μg/mL cycloheximide at indicated time points. (E) Stably transfected BGC-823 and MGC-803 cells were treated with or without MG132 (10 μM) for 6 h. The protein levels of PPAR-γ were analysed by western blotting. (F-G) MGC-803 cells were collected for real-time PCR or western blotting detection of time- and dose-dependent effects of rosiglitazone on CADM3 expression. (H) The PPRE region of the CADM3 promoter sequence is presented. (I-J) Dual-luciferase reporter assays were performed to investigate the effect of FABP4 and PPAR-γ on CADM3 transcription. The data are presented as the mean ± SD (* P < 0.05; ** P < 0.01; *** P < 0.001). CHX, cycloheximide. Rosi, rosiglitazone. TSS, transcription start site. PPRE, peroxisome proliferator responsive element.

Article Snippet: One week after the transplantation, the mice were treated daily with DMSO (Solarbio, Beijing, China) or rosiglitazone (10 mg/kg or 20 mg/kg; MedChemExpress, Shanghai, China) via intraperitoneal injection.

Techniques: Co-Immunoprecipitation Assay, Immunofluorescence, Staining, Translocation Assay, Western Blot, Expressing, Stable Transfection, Transfection, Real-time Polymerase Chain Reaction, Sequencing, Luciferase

Journal: Theranostics

Article Title: The peroxisome proliferator-activated receptor agonist rosiglitazone specifically represses tumour metastatic potential in chromatin inaccessibility-mediated FABP4-deficient gastric cancer

doi: 10.7150/thno.66814

Figure Lengend Snippet: Rosiglitazone can reverse the metastatic ability of FABP4-deficient GC cells in vitro and in vivo . (A-B) Stably transfected MGC-803 cells were treated with siPPAR-γ and rosiglitazone (20 μM). The protein levels of FABP4, PPAR-γ and CADM3 were determined by western blotting. The migration and invasion of GC cells were determined by the Transwell assays. (C) Scheme of experimental design for generation of a lung metastasis model. FABP4-deficient BGC-823 cells were transplanted into the tail veins of BALB/c nude mice, followed by daily treatment with DMSO or rosiglitazone (10 mg/kg or 20 mg/kg) (5 mice per group). (D-E) Representative images of mice and quantified results of bioluminescence imaging are presented. (F-G) The lungs were collected for H&E staining, and the number of metastatic nodules was determined. Scale bars, 1 mm. (H) Scheme of experimental design for generation of a liver metastasis model. FABP4-deficient BGC-823 cells were transplanted into the spleens of BALB/c nude mice, followed by daily treatment with DMSO or rosiglitazone (10 mg/kg or 20 mg/kg) (8 mice per group). (I-J) The livers were collected for H&E staining, and the number of metastatic nodules was determined. Scale bars, 1 mm. (K) The survival of BALB/c nude mice was monitored. The data are presented as the mean ± SD (* P < 0.05; ** P < 0.01; *** P < 0.001). Rosi, rosiglitazone.

Article Snippet: One week after the transplantation, the mice were treated daily with DMSO (Solarbio, Beijing, China) or rosiglitazone (10 mg/kg or 20 mg/kg; MedChemExpress, Shanghai, China) via intraperitoneal injection.

Techniques: In Vitro, In Vivo, Stable Transfection, Transfection, Western Blot, Migration, Imaging, Staining

Journal: Theranostics

Article Title: The peroxisome proliferator-activated receptor agonist rosiglitazone specifically represses tumour metastatic potential in chromatin inaccessibility-mediated FABP4-deficient gastric cancer

doi: 10.7150/thno.66814

Figure Lengend Snippet: Potential antitumour effect of rosiglitazone in patients with low FABP4 expression. (A) The proportion and distribution of the number of patients with diabetes and patients receiving rosiglitazone treatment in GC patients of the internal cohort. (B) Analysis of recurrence and longitudinal survival status of FABP4-negative patients who did or did not receive rosiglitazone treatment. (C) Analysis of survival data from FABP4-positive patients who did or did not receive rosiglitazone treatment. Rosi, rosiglitazone.

Article Snippet: One week after the transplantation, the mice were treated daily with DMSO (Solarbio, Beijing, China) or rosiglitazone (10 mg/kg or 20 mg/kg; MedChemExpress, Shanghai, China) via intraperitoneal injection.

Techniques: Expressing

Journal: Molecular Pain

Article Title: Antidepressants inhibit P2X 4 receptor function: a possible involvement in neuropathic pain relief

doi: 10.1186/1744-8069-5-20

Figure Lengend Snippet: Effect of antidepressants on the ATP-evoked [Ca 2+ ] i response via rat P2X 4 receptors . Three rounds of ATP stimulation (30 μM, 20 sec) induced [Ca 2+ ] i response in a reproducible fashion (a). Effect of pretreatment of cells with EGTA (500 μM, 10 min) (b) or with paroxetine (10 μM, 10 min) (c) on [Ca 2+ ] i response evoked by the second ATP stimulation. Traces indicate 340/380 fura-2 emission ratios averaged from (a) 48 cells, (b) 66 cells and (c) 18 cells obtained from each representative experiment. Effect of pretreatment of cells with TNP-ATP, antidepressants and anticonvulsants (10 μM, 10 min) on the ATP-evoked [Ca 2+ ] i response via rat P2X 4 receptors (d). Paroxetine dose-dependently inhibited the [Ca 2+ ] i response via rat P2X 4 receptors with an IC 50 value of 2.45 μM for 30 μM ATP stimulation (e). ATP dose-response curve was generated in the presence of increasing concentrations of paroxetine (f). Data are means ± SEM of at least 10 cells (d, e and f).

Article Snippet: Carbamazepine (Sigma-Aldrich) and paroxetine hydrochloride (Toronto Research Chemicals Inc.) were dissolved in 0.1% dimethyl sulfoxide (DMSO) in BSS.

Techniques: Generated

Journal: Molecular Pain

Article Title: Antidepressants inhibit P2X 4 receptor function: a possible involvement in neuropathic pain relief

doi: 10.1186/1744-8069-5-20

Figure Lengend Snippet: Effect of antidepressants on ATP-evoked [Ca 2+ ] i response via human P2X 4 receptors . Effect of pretreatment of cells with antidepressants (10 μM, 10 min) on the ATP-evoked [Ca 2+ ] i response via human P2X 4 receptors (a). Paroxetine dose-dependently inhibited the ATP-evoked [Ca 2+ ] i response via human P2X 4 receptors with an IC 50 value of 1.87 μM for 30 μM ATP stimulation (b). ATP dose-response curve was generated in the presence of increasing concentrations of paroxetine (c). Data are means ± SEM of at least 3 cells (a, b and c).

Article Snippet: Carbamazepine (Sigma-Aldrich) and paroxetine hydrochloride (Toronto Research Chemicals Inc.) were dissolved in 0.1% dimethyl sulfoxide (DMSO) in BSS.

Techniques: Generated

Journal: Molecular Pain

Article Title: Antidepressants inhibit P2X 4 receptor function: a possible involvement in neuropathic pain relief

doi: 10.1186/1744-8069-5-20

Figure Lengend Snippet: Effect of antidepressants on ATP-evoked currents in 1321N1 cells expressing rat P2X 4 receptors and primary microglia . Paroxetine (10 μM, 10 min) inhibited ATP (30 μM, 10 sec) evoked currents in rat P2X 4 receptor-expressed 1321N1 cells (a and b) and primary micloglia (c and d) (*** p < 0.001; paroxetine vs. vehicle, unpaired t-test). Data are means ± SEM of more than three separate experiments.

Article Snippet: Carbamazepine (Sigma-Aldrich) and paroxetine hydrochloride (Toronto Research Chemicals Inc.) were dissolved in 0.1% dimethyl sulfoxide (DMSO) in BSS.

Techniques: Expressing

Journal: Molecular Pain

Article Title: Antidepressants inhibit P2X 4 receptor function: a possible involvement in neuropathic pain relief

doi: 10.1186/1744-8069-5-20

Figure Lengend Snippet: Effect of intrathecal administration of paroxetine on the decrease in the PWT after nerve injury . A significant antiallodynic effect was observed following intrathecal administration of paroxetine at doses of 3 nmol and 10 nmol at day 7 (a), and 3 nmol at day 14 (b) after nerve injury (** p < 0.01 and * p < 0.05; paroxetine 3 nmol vs. control, # p < 0.05; paroxetine 10 nmol vs. control by a Dunn's multiple comparison test after a Kruskal-Wallis test). Data are means ± SEM of 5–8 rats.

Article Snippet: Carbamazepine (Sigma-Aldrich) and paroxetine hydrochloride (Toronto Research Chemicals Inc.) were dissolved in 0.1% dimethyl sulfoxide (DMSO) in BSS.

Techniques: Control, Comparison

Journal: Molecular Pain

Article Title: Antidepressants inhibit P2X 4 receptor function: a possible involvement in neuropathic pain relief

doi: 10.1186/1744-8069-5-20

Figure Lengend Snippet: Effect of intrathecal co-administration of 5-HT receptor blockers with paroxetine on the decrease in the PWT after nerve injury . No significant change was observed by intrathecal co-administration of 100 nmol WAY100635, 30 nmol ketanserin or 30 nmol ondansetron with 3 nmol paroxetine compared with 3 nmol paroxetine alone. ( p > 0.05 by a Dunn's multiple comparison test after a Friedman test.) Data are means ± SEM of 5–6 rats.

Article Snippet: Carbamazepine (Sigma-Aldrich) and paroxetine hydrochloride (Toronto Research Chemicals Inc.) were dissolved in 0.1% dimethyl sulfoxide (DMSO) in BSS.

Techniques: Comparison

Journal: Molecular Pain

Article Title: Antidepressants inhibit P2X 4 receptor function: a possible involvement in neuropathic pain relief

doi: 10.1186/1744-8069-5-20

Figure Lengend Snippet: Effect of spinal 5-HT depletion on the antiallodynic effect of paroxetine . 5-HT immunoreactivity in the dorsal horn of the spinal cord nine days after intrathecal injection of either saline or 5,7-dihydroxytryptamine (a). A marked reduction in the number of 5-HT immunoreactive fibres was observed in the dorsal horn of the L5 spinal cord after 5,7-DHT treatment compared with the saline-treated group. No significant change in the antiallodynic effect of paroxetine was observed in the 5,7-DHT-treated group compared with the saline-treated group (b). ( p > 0.05 by a Dunn's multiple comparison test after a Friedman test.) Data are means ± SEM of 5–6 rats.

Article Snippet: Carbamazepine (Sigma-Aldrich) and paroxetine hydrochloride (Toronto Research Chemicals Inc.) were dissolved in 0.1% dimethyl sulfoxide (DMSO) in BSS.

Techniques: Injection, Saline, Comparison

Journal: Microbiology Spectrum

Article Title: Plasmodium falciparum GAP40 Plays an Essential Role in Merozoite Invasion and Gametocytogenesis

doi: 10.1128/spectrum.01434-23

Figure Lengend Snippet: Mutations of phosphorylation sites do not affect the localization of GFP-tagged PfGAP40 proteins to the parasite periphery. (A) Schematic illustrating PfGAP40 with phosphorylation sites that have been identified by various studies ( , , , ). The phosphorylation sites are marked with lines, with the sites investigated in this study highlighted with red lines and sites not investigated highlighted in black. TM, transmembrane domain. (B) Illustration depicting the strategy to mutate S370, S372, and S376 sites of PfGAP40 to alanine and aspartic acid using selection-linked integration (SLI) method. The location of PCR primers used for genotyping are indicated; hDHFR, human dihydrofolate reductase; yDHODH, yeast dihydroorotate dehydrogenase; GFP, green fluorescent protein; FKBP, FK506-binding protein. (C) Diagnostic PCR of PfGAP40 phosphorylation mutants. The expected sizes of the PCR products that represent the integration of the native locus (p1×p3: WT, 1,721 bp; mutant, null) and the modified locus (p1×p2: WT, null; mutant, 2,868 bp) are indicated. (D) PCR products encompassing the modified locus were sequenced. An image of the chromatogram reveals successful editing of the locus of interest (marked with black boxes). (E) Western blot analysis of the transgenic mutants. The lysates were probed with anti-GFP antibody, which recognized a band of 132.5 kDa, corresponding to the GFP-2×FKBP-PfGAP40 fusion protein in all three mutants (GFP::WT, GFP::3S-to-D, and GFP::3S-to-A). The blots were also probed with an anti-aldolase antibody to serve as a loading control. (F) IFA analysis of matured schizonts and merozoites from three GFP::PfGAP40 mutants (GFP::WT, GFP::3S-to-D, and GFP::3S-to-A). Specific GFP-2×FKBP-PfGAP40 fusion proteins were probed with anti-PfGAP45 (green) and anti-GFP (red) antibodies. BF, bright field. Nuclei are stained with DAPI (blue). Scale bar: 5 μm.

Article Snippet: The transgenic parasites were obtained using 2.5 nM WR99210 (MCE) and subsequently treated with 1.5 μM DSM1 (a dihydroorotate dehydrogenase inhibitor) until a stably growing parasite population was obtained.

Techniques: Phospho-proteomics, Selection, Binding Assay, Diagnostic Assay, Mutagenesis, Modification, Western Blot, Transgenic Assay, Control, Staining

Journal: Neurobiology of Pain

Article Title: ATP-gated potassium channels contribute to ketogenic diet-mediated analgesia in mice

doi: 10.1016/j.ynpai.2023.100138

Figure Lengend Snippet: Activation of SUR1-Containing K ATP Channels Mimics the Antinociceptive Effect of a Ketogenic Diet. Capsaicin increased nociceptive events ( A, C ) and increased time engaged in nocifensive behavior ( B, D ) in chow-fed mice. Intraplantar injection of diazoxide one hour before capsaicin injection was sufficient to rescue capsaicin-evoked nociception ( A-B ), while intraplantar injection of levcromakalim (levcro) offered only modest protection ( C-D ). ( A-D ) N-way ANOVA (n = 8/group), *** denotes the effect of noxious stimulus: p < 0.005, ΨΨΨ denotes the effect of K ATP channel openers: p < 0.005, + denotes the effect of stimulus-K ATP openers interaction: p < 0.05, +++the effect of stimulus-K ATP openers interaction: p < 0.005.

Article Snippet: Diazoxide (115 ng, Sigma) or levcromakalim (573 ng, MedChemExpress) were delivered in a 10 μL intraplantar injection 1 h before capsaicin (20 μg) to assess whether K ATP channel activation could prevent capsaicin-evoked nociception.

Techniques: Activation Assay, Injection

Journal: Neurobiology of Pain

Article Title: ATP-gated potassium channels contribute to ketogenic diet-mediated analgesia in mice

doi: 10.1016/j.ynpai.2023.100138

Figure Lengend Snippet: Diazoxide Reduces p-ERK + Cell Number in the Spinal Dorsal Horn of Mice Receiving Intraplantar Capsaicin . ( A-D ) Intraplantar capsaicin increased the number of p-ERK + cells ( A, C , white arrowheads ) in the spinal dorsal horn of mice. ( A-B ) Prior injection of diazoxide prevented spinal neuronal activation, whereas prior injection of levcromakalim (levcro, C-D ) did not affect increased p-ERK + cell number. ( B, C ) N-way ANOVA (n = 3/group), *** denotes the effect of noxious stimulus: p < 0.005, ΨΨΨ denotes the effect of K ATP channel openers: p < 0.005, + denotes the effect of stimulus-K ATP openers interaction: p < 0.05, +++ denotes the effect of stimulus-K ATP openers interaction: p < 0.005.

Article Snippet: Diazoxide (115 ng, Sigma) or levcromakalim (573 ng, MedChemExpress) were delivered in a 10 μL intraplantar injection 1 h before capsaicin (20 μg) to assess whether K ATP channel activation could prevent capsaicin-evoked nociception.

Techniques: Injection, Activation Assay