Project description:Background: Treatment and morbidity control of schistosomiasis relies on a single drug, praziquantel. Therefore, there is a pressing need to explore the effects of PZQ on the parasites at the molecular level and to pursue alternative and/or or synergistic drugs against schistosomiasis. Methodology: We used a custom-designed Schistosoma mansoni oligo-microarray to explore the effects of a sublethal dose of praziquantel on S. mansoni adult worms’ gene expression. We used functional analysis of gene interaction networks to identify differentially expressed genes that are known targets of other drugs already tested in humans for diverse disease conditions. Omeprazole, a proton pump inhibitor drug that is widely prescribed, was tested in combination with praziquantel. We comparatively assessed the efficacy of praziquantel or omeprazole alone and in combination against S. mansoni adult worms in vitro over a period of 120 hours. Principal Findings: We identified sets of genes that were affected by praziquantel on both paired and unpaired females, however with opposite gene expression patterns (up-regulated in paired and down-regulated in unpaired females), indicating that the transcriptomics changes induced by praziquantel are heavily influenced by the mating status. We also identified genes that were similarly affected by praziquantel in males and females. The use of sublethal doses of praziquantel together with omeprazole showed a significantly enhanced worm mortality in vitro compared to praziquantel alone, thus evidencing a synergic effect. Conclusions: Functional analysis of gene interaction networks is an important approach to identify genes whose expression is affected by one drug and that are at the same time known targets of other drugs already tested in humans for diverse disease conditions, thus pointing the latter as possible synergic drugs. Combined treatment with omeprazol increases the efficacy of praziquantel against S. mansoni adult worms in vitro, and additional in vivo studies are warranted.

Project description:Background: Treatment and morbidity control of schistosomiasis relies on a single drug, praziquantel. Therefore, there is a pressing need to explore the effects of PZQ on the parasites at the molecular level and to pursue alternative and/or or synergistic drugs against schistosomiasis. Methodology: We used a custom-designed Schistosoma mansoni oligo-microarray to explore the effects of a sublethal dose of praziquantel on S. mansoni adult worms’ gene expression. We used functional analysis of gene interaction networks to identify differentially expressed genes that are known targets of other drugs already tested in humans for diverse disease conditions. Omeprazole, a proton pump inhibitor drug that is widely prescribed, was tested in combination with praziquantel. We comparatively assessed the efficacy of praziquantel or omeprazole alone and in combination against S. mansoni adult worms in vitro over a period of 120 hours. Principal Findings: We identified sets of genes that were affected by praziquantel on both paired and unpaired females, however with opposite gene expression patterns (up-regulated in paired and down-regulated in unpaired females), indicating that the transcriptomics changes induced by praziquantel are heavily influenced by the mating status. We also identified genes that were similarly affected by praziquantel in males and females. The use of sublethal doses of praziquantel together with omeprazole showed a significantly enhanced worm mortality in vitro compared to praziquantel alone, thus evidencing a synergic effect. Conclusions: Functional analysis of gene interaction networks is an important approach to identify genes whose expression is affected by one drug and that are at the same time known targets of other drugs already tested in humans for diverse disease conditions, thus pointing the latter as possible synergic drugs. Combined treatment with omeprazol increases the efficacy of praziquantel against S. mansoni adult worms in vitro, and additional in vivo studies are warranted. Two-condition experiment, PZQ-treated vs. untreated . Biological replicates: 2 with dye-swap

Project description:Omeprazole stimulated gene expression in rats Samples and RNA preparation: The series is composed of 9 hybridizations for analysis of differentially expressed genes in the gastric mucosa of omeprazole dosed vs. control rats. Nine rats (200-250 g) were given (by gavage) 400 micromol/kg of the proton pump inhibitor omeprazole (Astra AB, Gothenburg, Sweden) in Methocel (Dow Corning, Midland, MI) daily for 10 weeks. Seven rats received vehicle only. After 10 weeks the stomachs were removed for sampling of full-thickness corpus from the upper part of the greater curvature. Total RNA extraction was performed first by ultrasentrifugation on a cesium chloride cushion and then using TRIZOL Reagent (GIBCO BRL Life Technologies, New York, NY). Labeling and hybridization: Total RNA from 7 untreated rats were pooled and used as a common control. Three micrograms of total RNA from the control pool, and three micrograms of total RNA from each omeprazole dosed rat were reverse transcribed and labeled with capture sequences for subsequent attachment of Cy3- and Cy5-attached dendrimer, respectively, using 3DNA Array 350 Expression Array Detection kit as described in the manufacturer's protocols (Genisphere, Montvale, NJ). The labeling reactions were also spiked with exogenous control RNAs (Stratagene) at ratios of 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 0.5, 0.33, 0.25 and 0.125 for control spike 1, 2, 3, 4, 5, 6, 8, 9, 10 and 7, respectively. The slides were pre-washed in 2xSSC, 0.2% SDS at 55oC for 20 min, then 0.2x SSC at room temperature for 3 min, and deionized sterile filtered H2O at room temperature for 2 min. The Cy3 and Cy5 labeled samples (combined volume 9 microliter) were mixed with 25 microliter of the hybridization solution containing 0.25 M NaPO4, 1 mM EDTA, 4.5% SDS, 1 x SSC, 2 x Denhardt's solution and 2 microliter LNA dT Blocking reagent (Genisphere). A total hybridization volume of 50 microliter was added onto the array and covered with 24 x 60 mm cover slip. The array was assembled into humidified hybridization chamber (Corning) and hybridized at 60 oC for 18 h by submerging in a water bath. Post-hybridization washes were done once in 2 x SSC, 0.2% SDS at 55 oC for 15 min, then in 2 x SSC at room temperature for 15 min, and finally with 0.2 x SSC at room temperature for 15 min. After washing the fluorescent 3DNA reagent was hybridized to the cDNA at 60 oC for 3 h as described in the manufacturer's protocols. After dendrimer hybridization the slides were washed as described above. Scanning, image analysis and data processing: Arrays were scanned at 10 micrometer resolution with a ScanArray Express HT scanner (Packard BioSciences, Billerica, MA). The microarrays were analyzed using GenePixâ¢ Pro 4.1 (Axon Instruments, Inc., Union City, CA). All subsequent statistical analyses were performed using the statistical package R, R Development Core Team. A signal intensity based criterion was applied to remove the spots with median spot signal intensity less than 200. A spot area based criterion was applied to remove the spots with the area less than 50 pixels in either channel. Spots with more than 40% of the pixels saturated in either channel were also removed. Print tip-dependent normalization was applied to the data in order to compensate for systematic errors. The normalized ratios of the duplicated spots were averaged. Further analysis was done on the log2 transformed ratios. In order to assess the significance of up- or downregulation of the genes, tests for differential expression were performed using moderated T-tests, as implemented in the Limma R package of Smyth.

Project description:Omeprazole stimulated gene expression in rats Samples and RNA preparation: The series is composed of 9 hybridizations for analysis of differentially expressed genes in the gastric mucosa of omeprazole dosed vs. control rats. Nine rats (200-250 g) were given (by gavage) 400 micromol/kg of the proton pump inhibitor omeprazole (Astra AB, Gothenburg, Sweden) in Methocel (Dow Corning, Midland, MI) daily for 10 weeks. Seven rats received vehicle only. After 10 weeks the stomachs were removed for sampling of full-thickness corpus from the upper part of the greater curvature. Total RNA extraction was performed first by ultrasentrifugation on a cesium chloride cushion and then using TRIZOL Reagent (GIBCO BRL Life Technologies, New York, NY). Labeling and hybridization: Total RNA from 7 untreated rats were pooled and used as a common control. Three micrograms of total RNA from the control pool, and three micrograms of total RNA from each omeprazole dosed rat were reverse transcribed and labeled with capture sequences for subsequent attachment of Cy3- and Cy5-attached dendrimer, respectively, using 3DNA Array 350 Expression Array Detection kit as described in the manufacturer's protocols (Genisphere, Montvale, NJ). The labeling reactions were also spiked with exogenous control RNAs (Stratagene) at ratios of 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 0.5, 0.33, 0.25 and 0.125 for control spike 1, 2, 3, 4, 5, 6, 8, 9, 10 and 7, respectively. The slides were pre-washed in 2xSSC, 0.2% SDS at 55oC for 20 min, then 0.2x SSC at room temperature for 3 min, and deionized sterile filtered H2O at room temperature for 2 min. The Cy3 and Cy5 labeled samples (combined volume 9 microliter) were mixed with 25 microliter of the hybridization solution containing 0.25 M NaPO4, 1 mM EDTA, 4.5% SDS, 1 x SSC, 2 x Denhardt's solution and 2 microliter LNA dT Blocking reagent (Genisphere). A total hybridization volume of 50 microliter was added onto the array and covered with 24 x 60 mm cover slip. The array was assembled into humidified hybridization chamber (Corning) and hybridized at 60 oC for 18 h by submerging in a water bath. Post-hybridization washes were done once in 2 x SSC, 0.2% SDS at 55 oC for 15 min, then in 2 x SSC at room temperature for 15 min, and finally with 0.2 x SSC at room temperature for 15 min. After washing the fluorescent 3DNA reagent was hybridized to the cDNA at 60 oC for 3 h as described in the manufacturer's protocols. After dendrimer hybridization the slides were washed as described above. Scanning, image analysis and data processing: Arrays were scanned at 10 micrometer resolution with a ScanArray Express HT scanner (Packard BioSciences, Billerica, MA). The microarrays were analyzed using GenePix™ Pro 4.1 (Axon Instruments, Inc., Union City, CA). All subsequent statistical analyses were performed using the statistical package R, R Development Core Team. A signal intensity based criterion was applied to remove the spots with median spot signal intensity less than 200. A spot area based criterion was applied to remove the spots with the area less than 50 pixels in either channel. Spots with more than 40% of the pixels saturated in either channel were also removed. Print tip-dependent normalization was applied to the data in order to compensate for systematic errors. The normalized ratios of the duplicated spots were averaged. Further analysis was done on the log2 transformed ratios. In order to assess the significance of up- or downregulation of the genes, tests for differential expression were performed using moderated T-tests, as implemented in the Limma R package of Smyth. Keywords: repeat sample

Dataset Information

Expression data from young and senescent HCAECs treated with proton pump inhibitors (omeprazole and lansoprazole)

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure