ABSTRACT: The miRNA Transcriptome directly Reflect the Physiological and Biochemical Differences between Red, White and Intermediate Muscle Fiber Types
Project description:MicroRNAs (miRNAs) are small non-coding RNAs that can regulate the expression of their target genes at post-transcriptional level. Skeletal muscle comprises different �ber types that can be broadly classified as red, intermediate and white fiber types. Recently, a set of miRNAs were identified to be expressed on a fiber type-specific manner between red and white fiber types. Nonetheless, an integral explanation of the miRNA transcriptome differences in all three fiber types is long overdue. Herein, we collected 15 kinds of porcine skeletal muscles from different anatomic locations, which were then clearly divided into red, white and intermediate fiber type based on the hierarchical clustering analysis for the ratios of myosin heavy chain (MHC) isoforms. We further illustrated that three muscles, which typically represented each muscle fiber type (i.e. red: peroneal longus (PL), intermediate: psoas major muscle (PMM), white: longissimus dorsi muscle (LDM)), have distinct metabolic patterns of mitochondrial and glycolytic enzyme levels. In addition, we constructed the small RNA libraries for PL, PMM and LDM using deep sequencing approach. Results showed that, the differentially expressed miRNAs were mainly enriched in PL and played a vital role in myogenesis and energy metabolism. Overall, this comprehensive analysis will contribute to a better understanding of the miRNA regulatory mechanism for the phenotypic diversity of skeletal muscles. 3 sample
Project description:MicroRNAs (miRNAs) are small non-coding RNAs that can regulate the expression of their target genes at post-transcriptional level. Skeletal muscle comprises different fiber types that can be broadly classified as red, intermediate and white fiber types. Recently, a set of miRNAs were identified to be expressed on a fiber type-specific manner between red and white fiber types. Nonetheless, an integral explanation of the miRNA transcriptome differences in all three fiber types is long overdue. Herein, we collected 15 kinds of porcine skeletal muscles from different anatomic locations, which were then clearly divided into red, white and intermediate fiber type based on the hierarchical clustering analysis for the ratios of myosin heavy chain (MHC) isoforms. We further illustrated that three muscles, which typically represented each muscle fiber type (i.e. red: peroneal longus (PL), intermediate: psoas major muscle (PMM), white: longissimus dorsi muscle (LDM)), have distinct metabolic patterns of mitochondrial and glycolytic enzyme levels. In addition, we constructed the small RNA libraries for PL, PMM and LDM using deep sequencing approach. Results showed that, the differentially expressed miRNAs were mainly enriched in PL and played a vital role in myogenesis and energy metabolism. Overall, this comprehensive analysis will contribute to a better understanding of the miRNA regulatory mechanism for the phenotypic diversity of skeletal muscles.
Project description:The introduction of long oligonucleotide-based probes has led to many comparative studies of these two platforms in mammals, but remains to be performed for pig. Further, the characteristics of global gene expression in diverse porcine muscle tissues have not been yet been fully established. This is the first global gene expression study of a collection of nine porcine muscle tissues consisting of cardiac and various skeletal muscle types using both cDNA-based and long oligonucleotide-based microarray platforms. The expression profiles from the two platforms agree in differentiating between cardiac, skeletal red, skeletal intermediate and skeletal white muscle types by producing almost identical hierarchical expression clusters. The clusters from both platforms also reveal that gene expression profiles of the skeletal intermediate type are more similar to the red type than to the white type. Analysis of the ability to identify differentially expressed genes based on gene set analysis and GO term integration show a platform overlap of at least 80% for the cardiac-skeletal comparisons and a platform overlap of at least 58% for the skeletal red-white comparisons. Interestingly, the oligonucleotide platform was always able to identify more GO terms associated with differential expression than the cDNA platform. We further examined the skeletal red-white expression differences and found many GO biological processes that are known to be associated with these phenotypes including calcium ion transport, glycolysis, fatty acid beta-oxidation and muscle contraction. In addition, the expression of genes involved in differentiation between slow (red) and fast (white) muscle types such as MYBPC1, TNNI1, TNNT3 and ATP2A1 is highly regulated between red and white type muscles. Not previously shown to be associated with this tissue difference we found the biological process post-Golgi vesicle-mediated transport. Similar results were obtained with GO classes cellular components and molecular functions. Using the gene expression profiles from the oligonucleotide platform we presented and applied an approach for predicting alternatively spliced transcripts across cardiac and skeletal type muscles. One of the predicted transcripts from the gene named UBE2C has been shown to have six transcript variants in human, but they are not expressed in a cardiac-skeletal specific manner as we have observed here. We speculate that one of our oligonucleotide probes for this gene is able to detect only three of these variants whereas the other detects all six variants and that these variants are expressed in a cardiac-skeletal muscle specific manner. These results supports some of the advantages of using oligonucleotide-based microarray platforms for global gene expression profiling and the observed differences in gene expression among muscle tissues contribute to the understanding of the molecular processes behind porcine muscle biology. Keywords: tissue comparison, platform comparison
Project description:The introduction of long oligonucleotide-based probes has led to many comparative studies of these two platforms in mammals, but remains to be performed for pig. Further, the characteristics of global gene expression in diverse porcine muscle tissues have not been yet been fully established. This is the first global gene expression study of a collection of nine porcine muscle tissues consisting of cardiac and various skeletal muscle types using both cDNA-based and long oligonucleotide-based microarray platforms. The expression profiles from the two platforms agree in differentiating between cardiac, skeletal red, skeletal intermediate and skeletal white muscle types by producing almost identical hierarchical expression clusters. The clusters from both platforms also reveal that gene expression profiles of the skeletal intermediate type are more similar to the red type than to the white type. Analysis of the ability to identify differentially expressed genes based on gene set analysis and GO term integration show a platform overlap of at least 80% for the cardiac-skeletal comparisons and a platform overlap of at least 58% for the skeletal red-white comparisons. Interestingly, the oligonucleotide platform was always able to identify more GO terms associated with differential expression than the cDNA platform. We further examined the skeletal red-white expression differences and found many GO biological processes that are known to be associated with these phenotypes including calcium ion transport, glycolysis, fatty acid beta-oxidation and muscle contraction. In addition, the expression of genes involved in differentiation between slow (red) and fast (white) muscle types such as MYBPC1, TNNI1, TNNT3 and ATP2A1 is highly regulated between red and white type muscles. Not previously shown to be associated with this tissue difference we found the biological process post-Golgi vesicle-mediated transport. Similar results were obtained with GO classes cellular components and molecular functions. Using the gene expression profiles from the oligonucleotide platform we presented and applied an approach for predicting alternatively spliced transcripts across cardiac and skeletal type muscles. One of the predicted transcripts from the gene named UBE2C has been shown to have six transcript variants in human, but they are not expressed in a cardiac-skeletal specific manner as we have observed here. We speculate that one of our oligonucleotide probes for this gene is able to detect only three of these variants whereas the other detects all six variants and that these variants are expressed in a cardiac-skeletal muscle specific manner. These results supports some of the advantages of using oligonucleotide-based microarray platforms for global gene expression profiling and the observed differences in gene expression among muscle tissues contribute to the understanding of the molecular processes behind porcine muscle biology. Keywords: tissue comparison, platform comparison Muscle tissue samples from heart (HEA), Vastus intermedius (VIN), Infraspinatus (ISP), Supraspinatus (SSP), Biceps femoris (BFE), Longissimus dorsi (LDO), Semimembranosus (SME), Semitendinosus (STE) and Triceps brachii (TBR). A common reference sample was constructed by combining all samples. The exact same tissue samples were used for expression profiling with cDNA microarrays and 70-mer long oligonucleotide microarrays.
Project description:Isolation, Morphological, Physiological, Biochemical and Molecular Characterization of Actinobacteria Members from White and Red Soil Samples
Project description:In meat animal production, favourable meat traits such as color and, in the pig in particular, tenderness have been found to closely associate with the greater abundance of red or highly oxidative fibres. Red muscles possess higher lipid concentration (intra- and inter-fibre fat) which is associated with more tender and juicy meat. In addition, individuals with muscles that are abundant in oxidative type I fibres are associated with favourable metabolic health, and are less likely to predispose to obesity and insulin resistance. Collectively, understanding the molecular processes that govern the expression of specific fiber types and the phenotypic characteristics of muscles is important in agricultural and medical fields. In this study, a genome-wide investigation of the porcine differential expression between two red (soleus, SE) and white (longissimus dorsi, LD) muscle was conducted using the Affymetrix GeneChip® Porcine Genome Array containing oligonucleotides representing approximately 24123 transcripts from 20201 S. scrofa genes Three meishan gilts from the same litter were slaughtered at the 150 days of age by electrical stunning and exsanguination, in compliance with national regulations applied in commercial slaughtering. Immediately after slaughter, two muscles with different locations, functions, and biochemical properties were sampled: the longissimus at the last rib level, a fast twitch glycolytic muscle involved in voluntary movements of the back, and the deep portion of the SE, a oxydolytic muscle.
Project description:We procured PBMCs whole blood from five HC preterm infants and five preterm infants with BPD. PBMCs were extracted using a density gradient centrifugation method. Initially, 10ml of peripheral blood was mixed with an equal volume of physiological saline, then carefully layered onto Ficoll solution (T10124, from Shangbao Biotech Co., Ltd., Shanghai). After centrifugation at 2,000 rpm for 20 minutes, the cells stratified due to differences in density, with PBMCs positioned between the red blood cells and plasma. Subsequently, the intermediate layer containing PBMCs was gently collected, washed several times with physiological saline to remove residual medium and red blood cells, and finally, PBMCs were isolated and collected through centrifugation.
Project description:Background: skeletal muscle is a complex, versatile tissue composed of a variety of functionally diverse fiber types. Although the biochemical, structural and functional properties of myofibers have been the subject of intense investigation for the last decades, understanding molecular processes regulating fiber type diversity is still complicated by the heterogeneity of cell types present in the whole muscle organ. Methodology/Principal Findings: we have produced a first catalogue of genes expressed in mouse slow-oxidative (type 1) and fast-glycolytic (type 2B) fibers through transcriptome analysis at the single fiber level (microgenomics). Individual fibers were obtained from murine soleus and EDL muscles and initially classified by myosin heavy chain isoform content. Gene expression profiling on high density DNA oligonucleotide microarrays showed that both qualitative and quantitative improvements were achieved, compared to results with standard muscle homogenate. First, myofiber profiles were virtually free from non-muscle transcriptional activity. Second, thousands of muscle-specific genes were identified, leading to a better definition of gene signatures in the two fiber types as well as the detection of metabolic and signaling pathways that are differentially activated in specific fiber types. Several regulatory proteins showed preferential expression in slow myofibers. Discriminant analysis revealed novel genes that could be useful for fiber type functional classification. Conclusions/Significance: as gene expression analyses at the single fiber level significantly increased the resolution power, this innovative approach would allow a better understanding of the adaptive transcriptomic transitions occurring in myofibers under physiological and pathological conditions.
Project description:In meat animal production, favourable meat traits such as color and, in the pig in particular, tenderness have been found to closely associate with the greater abundance of red or highly oxidative fibres. Red muscles possess higher lipid concentration (intra- and inter-fibre fat) which is associated with more tender and juicy meat. In addition, individuals with muscles that are abundant in oxidative type I fibres are associated with favourable metabolic health, and are less likely to predispose to obesity and insulin resistance. Collectively, understanding the molecular processes that govern the expression of specific fiber types and the phenotypic characteristics of muscles is important in agricultural and medical fields. In this study, a genome-wide investigation of the porcine differential expression between two red (soleus, SE) and white (longissimus dorsi, LD) muscle was conducted using the Affymetrix GeneChip® Porcine Genome Array containing oligonucleotides representing approximately 24123 transcripts from 20201 S. scrofa genes
Project description:White adipose tissue is a major site of energy storage and plays a noteworthy role in physiological homeostasis and metabolic disease. Different types of fat have been demonstrated to exhibit distinct metabolic and inflammatory profiles and are differentially associated with disease risk. It is unclear whether these differences are intrinsic to the pre-differentiated stage. We address this challenging question using RNA sequencing on clonally-expanded human white preadipocyte cell lines. We have identified differences in gene expression that are separate from the process of browning and beiging.