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RNA sequencing reveals a slow to fast muscle fiber type transition after olanzapine infusion in rats.

Lynch CJ, Xu Y, Hajnal A, Salzberg AC, Kawasawa YI - PLoS ONE (2015)

Bottom Line: Understanding how SGAs affect the skeletal muscle transcriptome could elucidate approaches for mitigating these side effects.Thus these effects could contribute to the altered body composition and metabolic disease olanzapine causes.A potential interventional strategy is implicated because aerobic exercise, in contrast to resistance exercise, can oppose such slow to fast fiber transitions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Molecular Physiology, College of Medicine, Penn State University, Hershey, Pennsylvania, 17033, United States of America.

ABSTRACT
Second generation antipsychotics (SGAs), like olanzapine, exhibit acute metabolic side effects leading to metabolic inflexibility, hyperglycemia, adiposity and diabetes. Understanding how SGAs affect the skeletal muscle transcriptome could elucidate approaches for mitigating these side effects. Male Sprague-Dawley rats were infused intravenously with vehicle or olanzapine for 24h using a dose leading to a mild hyperglycemia. RNA-Seq was performed on gastrocnemius muscle, followed by alignment of the data with the Rat Genome Assembly 5.0. Olanzapine altered expression of 1347 out of 26407 genes. Genes encoding skeletal muscle fiber-type specific sarcomeric, ion channel, glycolytic, O2- and Ca2+-handling, TCA cycle, vascularization and lipid oxidation proteins and pathways, along with NADH shuttles and LDH isoforms were affected. Bioinformatics analyses indicate that olanzapine decreased the expression of slower and more oxidative fiber type genes (e.g., type 1), while up regulating those for the most glycolytic and least metabolically flexible, fast twitch fiber type, IIb. Protein turnover genes, necessary to bring about transition, were also up regulated. Potential upstream regulators were also identified. Olanzapine appears to be rapidly affecting the muscle transcriptome to bring about a change to a fast-glycolytic fiber type. Such fiber types are more susceptible than slow muscle to atrophy, and such transitions are observed in chronic metabolic diseases. Thus these effects could contribute to the altered body composition and metabolic disease olanzapine causes. A potential interventional strategy is implicated because aerobic exercise, in contrast to resistance exercise, can oppose such slow to fast fiber transitions.

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Comparison of RNA-Seq to QT-RTPCR for selected gastrocnemius muscle genes affected by olanzapine infusion.Genes significantly affected by olanzapine infusion based on RNA-Seq were selected for QT-RTPCR analysis using TaqMan gene expression assays using the same preparation of RNA. Genes were selected so that different FKPM size bins (1–10: OSTN; 10–99: OSTN, Casq2, Lpl; 100–999: Lpl, Atp2a2, Tnni1, 1000–9000: Tnni, Pvalb; >9000: Tpm1) were represented. The bars show mean± SE. Statistical significance was determined using the DEGSeq R package, which takes into consideration the variability of all of the genes analyzed. QT-RTPCR differences were analyzed using Student’s t-test. Asterisk symbols indicate a significant difference compared to control, *: p <0.05, **: p<0.01, ***: p<0.001)
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pone.0123966.g001: Comparison of RNA-Seq to QT-RTPCR for selected gastrocnemius muscle genes affected by olanzapine infusion.Genes significantly affected by olanzapine infusion based on RNA-Seq were selected for QT-RTPCR analysis using TaqMan gene expression assays using the same preparation of RNA. Genes were selected so that different FKPM size bins (1–10: OSTN; 10–99: OSTN, Casq2, Lpl; 100–999: Lpl, Atp2a2, Tnni1, 1000–9000: Tnni, Pvalb; >9000: Tpm1) were represented. The bars show mean± SE. Statistical significance was determined using the DEGSeq R package, which takes into consideration the variability of all of the genes analyzed. QT-RTPCR differences were analyzed using Student’s t-test. Asterisk symbols indicate a significant difference compared to control, *: p <0.05, **: p<0.01, ***: p<0.001)

Mentions: We next sought to compare genes with statistically different changes due to OLZ according to DEG-Seq to QT-RTPCR data. Our goal was to examine compare these two over different FKPM bin ranges (e.g., with one value below 10,10–100, 100–1000, 1000–10,000 and >10,000) compared to a QT-RTPCR (Fig 1). We endeavored to have at least one OLZ and one Control value in different bins so that more bins could be evaluated. Ostn, Pvalb and Tpm1 expression was significantly elevated by olanzapine treatment based on RNA-Seq and exhibited close proportionally elevated changes based on QT-RTPCR assays using Eef2 expression as a normalizer. Consistently, olanzapine similarly decreased the expression of Tnni1, Casq2, Lpl and Atpa2 as determined by either RNA-Seq or QT-RTPCR. The effects of olanzapine based on these two methods were highly correlated (r2 = 0.98, S1 Fig). Thus, these findings suggested that our RNA-Seq data were similarly quantitative as QT-RTPCR, albeit the QT-RTPCR data generally exhibited comparatively higher normalized standard deviations (not shown) as reflected in the size of the standard errors (Fig 1). A weakness is that we did not study QT-RTPCR of genes with lower % FKPM changes, however our conclusions in the results below about affected pathways are largely based on changes in many rather than individual genes. Furthermore, other studies have demonstrated that RNA-Seq is highly accurate for quantifying expression levels [33].


RNA sequencing reveals a slow to fast muscle fiber type transition after olanzapine infusion in rats.

Lynch CJ, Xu Y, Hajnal A, Salzberg AC, Kawasawa YI - PLoS ONE (2015)

Comparison of RNA-Seq to QT-RTPCR for selected gastrocnemius muscle genes affected by olanzapine infusion.Genes significantly affected by olanzapine infusion based on RNA-Seq were selected for QT-RTPCR analysis using TaqMan gene expression assays using the same preparation of RNA. Genes were selected so that different FKPM size bins (1–10: OSTN; 10–99: OSTN, Casq2, Lpl; 100–999: Lpl, Atp2a2, Tnni1, 1000–9000: Tnni, Pvalb; >9000: Tpm1) were represented. The bars show mean± SE. Statistical significance was determined using the DEGSeq R package, which takes into consideration the variability of all of the genes analyzed. QT-RTPCR differences were analyzed using Student’s t-test. Asterisk symbols indicate a significant difference compared to control, *: p <0.05, **: p<0.01, ***: p<0.001)
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4404103&req=5

pone.0123966.g001: Comparison of RNA-Seq to QT-RTPCR for selected gastrocnemius muscle genes affected by olanzapine infusion.Genes significantly affected by olanzapine infusion based on RNA-Seq were selected for QT-RTPCR analysis using TaqMan gene expression assays using the same preparation of RNA. Genes were selected so that different FKPM size bins (1–10: OSTN; 10–99: OSTN, Casq2, Lpl; 100–999: Lpl, Atp2a2, Tnni1, 1000–9000: Tnni, Pvalb; >9000: Tpm1) were represented. The bars show mean± SE. Statistical significance was determined using the DEGSeq R package, which takes into consideration the variability of all of the genes analyzed. QT-RTPCR differences were analyzed using Student’s t-test. Asterisk symbols indicate a significant difference compared to control, *: p <0.05, **: p<0.01, ***: p<0.001)
Mentions: We next sought to compare genes with statistically different changes due to OLZ according to DEG-Seq to QT-RTPCR data. Our goal was to examine compare these two over different FKPM bin ranges (e.g., with one value below 10,10–100, 100–1000, 1000–10,000 and >10,000) compared to a QT-RTPCR (Fig 1). We endeavored to have at least one OLZ and one Control value in different bins so that more bins could be evaluated. Ostn, Pvalb and Tpm1 expression was significantly elevated by olanzapine treatment based on RNA-Seq and exhibited close proportionally elevated changes based on QT-RTPCR assays using Eef2 expression as a normalizer. Consistently, olanzapine similarly decreased the expression of Tnni1, Casq2, Lpl and Atpa2 as determined by either RNA-Seq or QT-RTPCR. The effects of olanzapine based on these two methods were highly correlated (r2 = 0.98, S1 Fig). Thus, these findings suggested that our RNA-Seq data were similarly quantitative as QT-RTPCR, albeit the QT-RTPCR data generally exhibited comparatively higher normalized standard deviations (not shown) as reflected in the size of the standard errors (Fig 1). A weakness is that we did not study QT-RTPCR of genes with lower % FKPM changes, however our conclusions in the results below about affected pathways are largely based on changes in many rather than individual genes. Furthermore, other studies have demonstrated that RNA-Seq is highly accurate for quantifying expression levels [33].

Bottom Line: Understanding how SGAs affect the skeletal muscle transcriptome could elucidate approaches for mitigating these side effects.Thus these effects could contribute to the altered body composition and metabolic disease olanzapine causes.A potential interventional strategy is implicated because aerobic exercise, in contrast to resistance exercise, can oppose such slow to fast fiber transitions.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Molecular Physiology, College of Medicine, Penn State University, Hershey, Pennsylvania, 17033, United States of America.

ABSTRACT
Second generation antipsychotics (SGAs), like olanzapine, exhibit acute metabolic side effects leading to metabolic inflexibility, hyperglycemia, adiposity and diabetes. Understanding how SGAs affect the skeletal muscle transcriptome could elucidate approaches for mitigating these side effects. Male Sprague-Dawley rats were infused intravenously with vehicle or olanzapine for 24h using a dose leading to a mild hyperglycemia. RNA-Seq was performed on gastrocnemius muscle, followed by alignment of the data with the Rat Genome Assembly 5.0. Olanzapine altered expression of 1347 out of 26407 genes. Genes encoding skeletal muscle fiber-type specific sarcomeric, ion channel, glycolytic, O2- and Ca2+-handling, TCA cycle, vascularization and lipid oxidation proteins and pathways, along with NADH shuttles and LDH isoforms were affected. Bioinformatics analyses indicate that olanzapine decreased the expression of slower and more oxidative fiber type genes (e.g., type 1), while up regulating those for the most glycolytic and least metabolically flexible, fast twitch fiber type, IIb. Protein turnover genes, necessary to bring about transition, were also up regulated. Potential upstream regulators were also identified. Olanzapine appears to be rapidly affecting the muscle transcriptome to bring about a change to a fast-glycolytic fiber type. Such fiber types are more susceptible than slow muscle to atrophy, and such transitions are observed in chronic metabolic diseases. Thus these effects could contribute to the altered body composition and metabolic disease olanzapine causes. A potential interventional strategy is implicated because aerobic exercise, in contrast to resistance exercise, can oppose such slow to fast fiber transitions.

Show MeSH
Related in: MedlinePlus