Nam-Kuk Kim

Comparisons of longissimus muscle metabolic enzymes and muscle fiber types in Korean and western pig breeds

We compared differentially expressed genes and muscle fiber types in the longissimus muscles of Korean native pigs (KNP) and the western meat-producing breeds Landrace and Yorkshire. The KNP breed exhibited a higher muscle fat content and more red meat color as determined by the a(∗) (redness) value (P<0.01) and b(∗) (yellowness) value (P<0.05) compared to the western breeds. Using differential display RT-PCR, we detected two genes that were differentially expressed in skeletal muscle among the pig breeds. These genes were identified as NADH dehydrogenase subunit 1 and ATPase subunit 6 by cloning and sequencing analysis. Both of these genes are involved oxidative phosphorylation and therefore energy metabolism. The genes were more highly expressed in the KNP breed than in the other breeds, indicating that KNPs exhibit more oxidative metabolism than do the western breeds. We also analyzed the mRNA levels of myosin heavy-chain isoforms such as type I (oxidative), type IIb (glycolytic), and types IIa and IIx (intermediate) fibers using real-time RT-PCR. The mRNA levels of oxidative and intermediate fibers were elevated in the KNP breed, whereas the glycolytic fibers were more highly expressed in the Landrace and Yorkshire pigs. These results suggest that the elevated expression of the oxidation-related metabolism genes NADH dehydrogenase and ATPase is related to meat quality as indicated by a higher content of oxidative fibers and muscle fat, as well as redder meat color.

Effect of fiber type on postmortem proteolysis in longissimus muscle of Landrace and Korean native black pigs.

The current study was conducted to characterize objective meat quality, fiber type and their relations to postmortem proteolysis in longissimus muscle of Landrace and Korean native black (KNP) pigs. Longissimus muscles from each 10 market-weighted male pigs were removed after conventional slaughtering and chilling procedures, and aged for 1 or 7 days at 4°C to determine WB-shear force, objective meat color, proportion of myosin heavy chain I (MyHC I), intramuscular fat content and rate of proteolysis by a proteomics approach. KNP had a significantly (p<0.05) higher content of MyHC I, and that concurred with greatly (p<0.05) higher intramuscular fat content and Hunter a(∗) value, and significantly (p<0.05) lower drip loss than those seen in Landrace. One-dimension SDS-PAGE indicated that GAPDH, troponin I and creatine kinase were prominent proteolytic products during chiller ageing. By applying a gel-based proteome analysis, 26 proteins were identified, which showed different degradation properties during ageing between the breeds. Biopsied sample revealed that myosin regulatory light chain 2, myosin light chain isotype v/sb, fatty acid-binding protein and albumin were expressed at a greatly higher level for KNP, but their relation to fiber type (or genetic background) is unclear. It was particular noticeable that different actin isoforms showed various degradation behavior during ageing time.

Heat shock protein B1 and its regulator genes are negatively correlated with intramuscular fat content in the longissimus thoracis muscle of Hanwoo (Korean cattle) steers.

In previous proteomic studies, heat shock protein β 1 (HSPB1) was detected as a candidate protein related to meat quality in cattle. This study sought to determine if its gene expression was associated with intramuscular fat content in the longissimus thoracis muscle of Korean cattle (Hanwoo). Tissue from two groups of 10 steers each, low-marbling (mean intramuscular fat content, 7.4 ± 1.5%) and high-marbling (23.5 ± 2.8%), were used for immunoblotting, real-time PCR, and statistical analyses. HSPB1 expression in both mRNA and protein was shown to be negatively related to intramuscular fat content (P < 0.05). Pathway analysis found two genes, TNF receptor superfamily member 6 (FAS) and angiotensinogen (AGT), that were regulators of the HSPB1 gene. The expression of the two genes showed a negative correlation with intramuscular fat content (P < 0.05). These results suggest that HSPB1, FAS, and AGT may be good candidate genes associated with intramuscular fat content in the longissimus muscle of Korean cattle.

Gene expression profiling of metabolism-related genes between top round and loin muscle of Korean cattle (Hanwoo).

Using differential display reverse transcriptase polymerase chain reaction, we detected 11 differentially expressed genes between top round and loin muscle in Korean cattle (Hanwoo). In the loin muscle, the lightness (L*) value (P<0.01) and marbling fat content (P<0.01), which are important factors in determining meat quality, were higher than in top round muscle. Three of the 11 genes were validated as significant genes between two types of muscle by real-time polymerase chain reaction (P<0.05). To determine whether the three genes were associated with meat quality traits, a regression analysis was preformed. The result demonstrated that two genes (NADH dehydrogenase 2 and cytochrome oxidase III), which are involved in oxidative phosphorylation in mitochondria, were significantly correlated with marbling fat content in the loin muscle (P<0.01), while two genes were not significant with marbling fat content in top round muscle. No significant effects for two genes on other meat quality traits such as meat color (redness and yellowness value), Warner-Bratzler shear force, and water-holding capacity were detected in this study.

Gene Expression Patterns Associated with Peroxisome Proliferator-activated Receptor (PPAR) Signaling in the Longissimus dorsi of Hanwoo (Korean Cattle)

Adipose tissue deposited within muscle fibers, known as intramuscular fat (IMF or marbling), is a major determinant of meat quality and thereby affects its economic value. The biological mechanisms that determine IMF content are therefore of interest. In this study, 48 genes involved in the bovine peroxisome proliferator-activated receptor signaling pathway, which is involved in lipid metabolism, were investigated to identify candidate genes associated with IMF in the longissimus dorsi of Hanwoo (Korean cattle). Ten genes, retinoid X receptor alpha, peroxisome proliferator-activated receptor gamma (PPARG), phospholipid transfer protein, stearoyl-CoA desaturase, nuclear receptor subfamily 1 group H member 3, fatty acid binding protein 3 (FABP3), carnitine palmitoyltransferase II, acyl-Coenzyme A dehydrogenase long chain (ACADL), acyl-Coenzyme A oxidase 2 branched chain, and fatty acid binding protein 4, showed significant effects with regard to IMF and were differentially expressed between the low- and high-marbled groups (p<0.05). Analysis of the gene co-expression network based on Pearson’s correlation coefficients identified 10 up-regulated genes in the high-marbled group that formed a major cluster. Among these genes, the PPARG-FABP4 gene pair exhibited the strongest correlation in the network. Glycerol kinase was found to play a role in mediating activation of the differentially expressed genes. We categorized the 10 significantly differentially expressed genes into the corresponding downstream pathways and investigated the direct interactive relationships among these genes. We suggest that fatty acid oxidation is the major downstream pathway affecting IMF content. The PPARG/RXRA complex triggers activation of target genes involved in fatty acid oxidation resulting in increased triglyceride formation by ATP production. Our findings highlight candidate genes associated with the IMF content of the loin muscle of Korean cattle and provide insight into the biological mechanisms that determine adipose deposition within muscle.

Expression Profiles of Triacylglycerol Biosynthesis Genes on Fattening Stages in Hanwoo

Muscle fat content including intramuscular fat content(IMF) is the most important parameter in meat quality of cattle. Triacylglycerol is the major component of fat and is found in adipocyte and skeletal muscle. In present study, we carried out the determined of triacylglycerol contents and mRNA expression patterns of genes related with triacylglycerol biosynthesis such as ACSS2, GPAT, MGAT and DGAT in Hanwoo longissimus muscle using the real-time PCR. The triacylglycerol contents were continuously increased during the fattening stages. Interestingly, the contents of triacylglycerol were 7.4 fold higher(0.34 vs. 2.51, P

Association Study Between the Polymorphisms of Exostosin-1 Gene and Economic Traits in Hanwoo

The aim of this study was to identify the polymorphism on exostosin-1 (EXT1) gene and to associate with economic traits in Hanwoo (Korean cattle). We sequenced for detection of single nucleotide polymorphism (SNP) with 24 unrelated individuals and identified four SNPs (T272196A, C272359T, G290964A and A302092G). Relationship between the genotypes of 583 Hanwoo individuals by PCR-RFLP and economic traits were analyzed by general linear model. In EXT1 gene, there were four SNPs associated with economic traits such as eye muscle area breeding value, marbling score breeding value, backfat and thickness breeding value (p<0.05 to p<0.01). In conclusion, this study indicates an important role of EXT1 gene in determining the meat quality or economic characteristics in Hanwoo.

Gene Expression of Candidate Genes Involved in Fat Metabolism During In vitro Adipogenic Differentiation of Bovine Mesenchymal Stem Cell

Adipogenesis has been one of the most intensely studied models of cellular differentiation. During adipogenesis, differential expression of many adipogenesis related genes lead to profound changes in cellular, morphological, and physiological characteristics of the differentiating cells. The aim of the present study was to examine the expression levels of adipogenic candidate genes, cAMP early repressor (ICER), nephroblastoma over-expressed protein (NOV), heat shock protein beta 1 (HSPB1) and succinate dehydrogenase (SDH), during adipogenesis of bovine mesenchymal stem cells (BMSC). The BMSC were cultured in DMEM / low glucose medium with adipogenic inducers for 6 days and the expression of various candidate genes which seemed related to adipogenesis were measured by real-time PCR. This study showed that the expression of peroxisome proliferator activated receptor γ (PPARγ) and fatty acid binding protein 4 (FABP4) genes as adipogenic indicators were increased to 3.11 and 3.11 folds on day 6 than on day 0, respectively (p<0.05). To determine whether candidate genes were related to adipogenesis, the expression levels of ICER, NOV, HSPB1, and SDH genes were measured during adipogenesis in BMSC. Our results showed that the expression level of ICER gene was significantly increased to 4.12 folds (0.01729 vs. 0.07138; p<0.05), whereas NOV, HSPB1, and SDH genes were decreased to 2.89, 3.18 and 2.36 folds, respectively, on day 6 when compared to day 0. These results suggest that these candidate genes have stimulatory or inhibitory effects on adipogenesis in BMSC, indicating that these genes may be directly or indirectly related to the adipogenic event of adipose precursor cells. (Key words : Bovine mesenchymal stem cell, Adipogenesis, Differential expression)

Comparative Analysis of Muscle Proteome from Porcine White and Red Muscles by Two-dimensional Electrophoresis

The technique known as proteomics is useful for characterizing the protein expression pattern of a particular tissue or cell type as well as quantitatively identifying differences in the levels of individual proteins. In present study, we carried out the comparative expression patterns of white and red muscles. We used the two-dimensional electrophoresis(2-DE) for analyzing the protein expression. Proteins isolated from porcine white and red muscles were separated by 12% poly-acrylamide gel and then were detected by coomassie blue and silver staining. More than 600 protein spots were detected on each 2-DE gel. By visual analysis of the stained gel, five proteins were identified to be differentially expressed in the white vs red muscle. By database searching based on the molecular weights and pI(isoelectric point) of the five proteins, three of them were found to be most close to troponin I, T and myoglobin. However, further researche is needed for identification and functional analysis of the unidentified proteins. In conclusion, we found five proteins, which are differentially expressed in the white vs red muscle. The functional analysis of the differentially expressed proteins will provide valuable information on biochemical characteristics of the muscle type.

Association of Succinate Dehydrogenase and Triose Phosphate Isomerase Gene Expression with Intramuscular Fat Content in Loin Muscle of Korean (Hanwoo) Cattle

In a previous study, succinate dehydrogenase (SDH) and triose phosphate isomerase (TPI) were detected by 2D gel electrophoresis as differentially expressed proteins in the longissimus thoracis muscles of cattle aged between 12 and 27 months old. In the present study, we investigated the association of SDH and TPI gene expression with intramuscular fat content in 50 Hanwoo steers. The SDH gene was expressed at a 4 times higher level in the 12 month old group than in the 27 month old group (p