• BMB Reports
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• v.40 no.4
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• pp.459-466
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• 2007

The cDNA sequence of the Japanese flounder (Paralychthys olivaceus) IgD has been previously reported (GenBank accession no. AB052658) and this was followed by the detection of IgD mRNA expression in some flounder organ tissues. However, it has not been determined whether the flounder IgD gene is virtually expressed into IgD protein. To characterize the flounder immunoglobulins utilized in elucidating the mechanism, evolution and diversity of the flounder immune system, antibodies specific to IgD and IgM were necessary. In the present study, partial flounder recombinant IgD (rIgD), IgM (rIgM) and the conserved regions of IgD and IgM (rCIg) were produced by cloning the cDNA sequence using isotype specific primers which were designed to produce unique fragments of IgD and IgM specific amino acid sequences. The production of recombinant Igs was ascertained by SDS-gel electrophoresis and immunoblot analysis using anti-T7$\cdot}$Taq antibody. The produced recombinant Igs were purified using affinity columns, and used as immunogens. Antibodies specific to the isotype of flounder Igs were generated by immunizing rabbits with rfIgs and the antibodies produced were identified by enzyme-linked immunosorbent assay (ELISA) and immunoblotting. Specificities of the generated antibodies were evaluated by testing cross-reactivity between recombinant IgM and IgD. By ELISA, rabbit antibodies against the rfIgD fragment (anti-rfIgD) failed to recognize any kind of flounder serum Igs, whereas respective antibodies against rfCIg (anti-rfCIg) and rfIgM fragments (anti-rfIgM) reacted with serum Igs. Likewise, in immunoblot assays, though anti-rfIgD did not, both anti-rfCIg and anti-rfIgM bound with the ~85 kd flounder IgM heavy chain. By flow cytometry analysis, anti-rfCIg, anti-rfIgD and anti-rfIgM reacted with 6%, 3% and 6.5% of cells, respectively, suggesting that flounder IgD is not secreted in serum but expressed on flounder B-like cell surfaces as in mammals. Antibodies produced against recombinant flounder Igs could be used to develop sandwich assay systems for detecting flounder Igs and for further investigating the flounder immune system.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.9
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• pp.1217-1226
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• 2011

Finding alternative feeds, such as sesame hulls and Prosopis juliflora species can attenuate difficulties of feed shortage and reduce the cost of animal feed in arid and semi-arid countries. Thirty-two Black male kids with similar initial weights (BW = $16.7{\pm}0.80\;kg$) and $120{\pm}5\;d$ of age, were used to evaluate the effect of replacing barley grains and soybean meal with Prosopis juliflora pods (PJP) and sesame hulls (SH) on growth performance, digestibility and carcass and meat characteristics. Kids were equally divided into four dietary treatment groups for an 84-d fattening period. Treatment diets had similar crude protein (CP) and metabolizable energy (ME). The treatment groups were: (T1) no PJP nor SH, (T2) 10% PJP and 20% SH, (T3) 15% PJP and 15% SH, and (T4) 20% PJP and 10% SH. A tendency was detected (p<0.08) for dry matter (DM), crude protein (CP), organic matter (OM) and neutral detergent fiber (NDF) intakes that were greater for T2 than T3 and T4 while T1 was not different from all other treatment groups. Ether extract (EE) intake was the greatest (p<0.05) for T2 and the lowest for T1. Acid detergent fiber (ADF) intake was greater (p<0.05) for T2 than T1 while T3 and T4 were intermediate. Final live weight, average daily gain (ADG) and feed conversion ratio (FCR) were comparable among different treatment groups. Digestibility of DM, OM, CP, NDF and ADF were similar among all treatment groups, however, EE digestibility was the lowest (p<0.05) for T1 when compared to other treatments. In addition, nitrogen intake, nitrogen in urine and retained and retention percentages were similar among all treatment groups. However, N loss in feces was higher (p<0.05) for T2 than T3 and T4 while T1 was intermediate. No differences were observed among treatment groups with respect to fasting live weight, hot and cold carcass weights, dressing-out percentages, mesenteric fat, visceral organs, carcass cuts percentages and carcass linear dimensions. No differences were also observed for dissected loin, leg, rack and shoulder tissues except in the total bone % for loin cuts and in the meat to bone ratio for rack cuts. T3 has the greatest total bone % and the lowest meat to bone ratio when compared to all other treatment groups. No differences were observed between treatment groups in all quality characteristics of the longissimus muscle. The present study demonstrates the potential of using PJP and SH for growing kids without adverse effects on growth performance, carcass characteristics and meat quality.

• Nutrition Research and Practice
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• v.9 no.1
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• pp.22-29
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• 2015

BACKGROUND/OBJECTIVES: Recently, anthocyanins have been reported to have various biological activities. Furthermore, anthocyanin-rich purple corn extract (PCE) ameliorated insulin resistance and reduced diabetes-associated mesanginal fibrosis and inflammation, suggesting that it may have benefits for the prevention of diabetes and diabetes complications. In this study, we determined the anthocyanins and non-anthocyanin component of PCE by HPLC-ESI-MS and investigated its anti-diabetic activity and mechanisms using C57BL/KsJ db/db mice. MATERIALS/METHODS: The db/db mice were divided into four groups: diabetic control group (DC), 10 or 50 mg/kg PCE (PCE 10 or PCE 50), or 10 mg/kg pinitol (pinitol 10) and treated with drugs once per day for 8 weeks. During the experiment, body weight and blood glucose levels were measured every week. At the end of treatment, we measured several diabetic parameters. RESULTS: Compared to the DC group, Fasting blood glucose levels were 68% lower in PCE 50 group and 51% lower in the pinitol 10 group. Furthermore, the PCE 50 group showed 2-fold increased C-peptide and adiponectin levels and 20% decreased HbA1c levels, than in the DC group. In pancreatic islets morphology, the PCE- or pinitol-treated mice showed significant prevention of pancreatic ${\beta}$-cell damage and higher insulin content. Microarray analyses results indicating that gene and protein expressions associated with glycolysis and fatty acid metabolism in liver and fat tissues. In addition, purple corn extract increased the phosphorylation of AMP-activated protein kinase (AMPK) and decreased phosphoenolpyruvate carboxykinase (PEPCK), glucose 6-phosphatase (G6pase) genes in liver, and also increased glucose transporter 4 (GLUT4) expressions in skeletal muscle. CONCLUSIONS: Our results suggested that PCE exerted anti-diabetic effects through protection of pancreatic ${\beta}$-cells, increase of insulin secretion and AMPK activation in the liver of C57BL/KsJ db/db mice.

• BMB Reports
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• v.38 no.1
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• pp.71-76
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• 2005

Ceruloplasmin (CP) is the major plasma antioxidant and copper transport protein. Monoclonal antibodies (mAbs) against human CP were produced and characterized. A total of five hybridoma cell lines were established (CP2, CP10, CP20, CP25, CP30). From the epitope mapping analysis, two subgroups of mAbs recognize different peptide fragments were identified. When the purified CP was incubated with the mAbs, the ferroxidase activity of CP was inhibited up to a maximum 57%. Immunoblotting with various tissue homogenates indicated that all the mAbs specifically recognize a single protein band of 130 kDa. They also appear to be extensively cross-reactive among different mammalian including human and avian sources. These results demonstrated that only one type of immunologically similar CP is present in all of the mammalian tissues including human. The CP mAbs could be of great benefit to design the diagnostic kit for CP-related diseases such as Wilson's disease.

• BMB Reports
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• v.38 no.6
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• pp.668-675
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• 2005

A full-length cDNA encoding taxadiene synthase (designated as TmTXS), which catalyzes the first committed step in the Taxol biosynthetic pathway, was isolated from young leaves of Taxus media by rapid amplification of cDNA ends (RACE). The full-length cDNA of TmTXS had a 2586 bp open reading frame (ORF) encoding a protein of 862 amino acid residues. The deduced protein had isoelectric point (pI) of 5.32 and a calculated molecular weight of about 98 kDa, similar to previously cloned diterpene cyclases from other Taxus species such as T. brevifolia and T. chinenisis. Sequence comparison analysis showed that TmTXS had high similarity with other members of terpene synthase family of plant origin. Tissue expression pattern analysis revealed that TmTXS expressed strongly in leaves, weak in stems and no expression could be detected in fruits. This is the first report on the mRNA expression profile of genes encoding key enzymes involved in Taxol biosynthetic pathway in different tissues of Taxus plants. Phylogenetic tree analysis showed that TmTXS had closest relationship with taxadiene synthase from T. baccata followed by those from T. chinenisis and T. brevifolia. Expression profiles revealed by RT-PCR under different chemical elicitor treatments such as methyl jasmonate (MJ), silver nitrate (SN) and ammonium ceric sulphate (ACS) were also compared for the first time, and the results revealed that expression of TmTXS was all induced by the tested three treatments and the induction effect by MJ was the strongest, implying that TmTXS was high elicitor responsive.

• Clinical and Experimental Reproductive Medicine
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• v.33 no.4
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• pp.253-263
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• 2006

Objective: Identification of the regulatory mechanism for arrest and initiation of primordial follicular growth is crucial for female fertility. Previously, we found 15 expressed sequence tags (ESTs) that were specifically abundant in the day-S-subtracted cDNA library and that the B357 clone was novel. The present study was conducted to obtain the whole sequence of the novel gene including B357 and to characterize its mRNA and protein expression in mouse ovary and testis. Methods: The extended sequence of the 2,965-bp cDNA fragment for the clone B357 was named ${\underline{5}}-{\underline{d}}ay-{\underline{o}}vary-{\underline{s}}pecific\;gene-{\underline{1}}$ (5DOS1) and submitted to GenBank (accession number ${\underline{AY751521}}$). Expression of 5DOS1 was characterized in both female and male gonads at various developmental stages by Northern blotting, real-time RT-PCR, in situ hybridization, Western blotting, and immunohistochemistry. Results: The 5DOS1 transcript was highly expressed in the adult testis, brain, and muscle as compared to the other tissues. In the ovary, the 5DOS1 transcript was detected in all oocytes from primordial to antral follicles, and highly expressed at day 5 after birth and decreased thereafter. In contrast, expression of 5DOS1 showed a gradual increase during testicular development and its expression was limited to various stages of male germ cells except spermatogonia. Conclusions: This is the first report on the expression and characterization of the 5DOS1 gene in the mouse gonads. Further functional analysis of the 5DOS1 protein will be required to predict its role in gametogenesis.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.6
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• pp.2753-2758
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• 2014

Background: GC-binding factor 2 (GCF2) is a transcriptional regulator that represses transcriptional activity of the epidermal growth factor receptor (EGFR) by binding to a specific GC-rich sequence in the EGFR gene promoter. In addition to this function, GCF2 has also been identified as a tumor-associated antigen and regarded as a potentially valuable serum biomarker for early human hepatocellular carcinoma (HCC) diagnosis. GCF2 is high expressed in most HCC tissues and cell lines including HepG2. This study focused on the influence of GCF2 on cell proliferation and apoptosis in HepG2 cells. Materials and Methods: GCF2 expression at both mRNA and protein levels in HepG2 cells was detected with reverse transcription (RT) PCR and Western blotting, respectively. RNA interference (RNAi) technology was used to knock down GCF2 mRNA and protein expression. Afterwards, cell viability was analyzed with a Cell Counting Kit-8 (CCK-8), and cell apoptosis and caspase 3 activity by flow cytometry and with a Caspase 3 Activity Kit, respectively. Results: Specific down-regulation of GCF2 expression caused cell growth inhibition, and increased apoptosis and caspase 3 activity in HepG2 cells. Conclusions: These primary results suggest that GCF2 may influence cell proliferation and apoptosis in HepG2 cells, and also provides a molecular basis for further investigation into the possible mechanism at proliferation and apoptosis in HCC.

• Microbiology and Biotechnology Letters
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• v.5 no.4
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• pp.177-184
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• 1977

Arylsulfatase catalyzes the release of SO$\sub$4//sup2/- from sulfate esters of simple phenols. Arylsolfatase occurs widely in animal tissues and in microorganisms including soil bacteria. Its widespread distribution suggests that it has a rather fundamental function and environmental meaning. It has been shown previously that arylsulfatase of Klebsiella was purified and characterized. A condition of arylsulfatase synthesis was tested with several strains of Serratia. Serratia marcescens could not utilize some sugars, such as xylose, rhamnose, glucosamine and arabinose hut glucose and mannitol as a sole carbon source. However, arylsulfatase synthesis was repressed by glucose but not by mannitol. The enzyme synthesis was repressed ob inorganic sulfate and methionine, and this repression was relieved by addition of tyramine. Arylsulfatase of S. marcescen was purified by fractionation with ammonium sulfate and followed by chromatographies on DEAE-Cellulose CM-Cellulose, and DEAE-Sephadex A-25. The molecular weight of arylsulfatase was determined to be 46,000 by SDS-Gel electrophoresis and 49,000 by Sephadex G-100 column chromatography. The enzyme showed some different properties with that of K. aerogenes. The activity was maximum at pH 6.8. The Km and Vmax values for p-nitrophenyl sulfate were 2.5${\times}$10$\^$－4/ M and 20 nmoles/min/mg protein, respectively. The enzyme showed high activities toward phenyl sulfate, ο-and p-nitro phenyl sulfates, and p-nitrocatechol sulfate. The inhibition of enzyme was strongly affected by hydroxylamine, inorganic fluoride, sulfide and phosphate, but by inorganic sulfate. Like Klebsiella arylsulfatase, tyramine, octopamine, and dopamine gave signifcant inhibitory effect.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.21 no.8
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• pp.417-428
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• 2020

Housekeeping genes are expressed in cells of all organisms and perform basic cellular functions such as energy generation, substance synthesis, cell death, and cell defense. Accordingly, the expression levels of housekeeping genes are relatively constant, and thus they are used as reference genes in gene expression studies, such as protein expression and mRNA expression analysis of target genes. However, the levels of expression of these genes may be different among various tissues or cells and may change under certain circumstances. Therefore, it is important to select the best reference gene for specific gene expression research by exploring the stability of housekeeping gene expression. This review summarizes housekeeping genes found in humans, chickens, pigs, and rats in the literature and estimates expression stability using geNorm, NormFinder, and BestKeeper software. The most suitable reference housekeeping gene can selected based on expression stability according to the experimental conditions of the gene expression study and can thus be applied to data normalization.

• Journal of Nutrition and Health
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• v.36 no.5
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• pp.459-469
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• 2003

We examined the effects of calcium intake levels on mineral metabolism during pregnancy using calcium-deficient young adult rats. Five week-old female Sprague-Dawley rats were fed normal Ca (NCa, 0.5%) and low Ca (LCa, 0.15%) diets for five weeks (pre-pregnancy). The low Ca intake group was then divided into three groups and fed low Ca (0.15%), normal Ca (0.5%) and high Ca (1.5%) diets for 3 weeks (pregnancy). All of the rats were mated with normal male rats. The control group was fed a consistently normal Ca (0.5%) diet during pre-pregnancy and pregnancy. On the day after delivery, dams and their pups were sacrificed. We measured total protein, albumin, alkaline phosphatase and mineral content in serum and weight, length, breaking force, ash and mineral content in the femur and lumbar (L2-L4) of the dams. Whole body mineral content was measured in the pups. There was no difference in weight gain and food intake among the groups. Serum total protein and albumin were in the normal range but a little lower during pregnancy. High Ca supplementation decreased serum Mg and Fe during pregnancy. Weight, ash and Ca of the femur and lumbar significantly decreased in rats fed a chronically low Ca diet during pre-pregnancy and pregnancy. Calcium supplementation levels were above normal during pregnancy and increased the bone weight and breaking force of rats fed the low Ca diet during pre-pregnancy. However, Ca supplementation did not increase the ash and Ca contents in the bones. High Ca supplementation during pregnancy significantly decreased Mg in the bones and increased Ca and P in the kidneys. Ash content of pups from dams fed the chronically low Ca diet decreased but there was no difference in whole body Ca among the groups. Mg and Fe in the whole body decreased in pups from dams fed the high Ca diet. Pregnancy performance was reduced in dams fed the low Ca diet. These results suggest that above normal Ca supplementation levels during pregnancy restored maternal bone status to some extent in rats fed the chronically low Ca diet. The same could not be said for mineral content. Also, high Ca supplementation during pregnancy may deteriorate mineral composition in bones and other tissues. Therefore, more detailed research is needed to facilitate sound recommendations on appropriate calcium intake during pregnancy. (Korean J Nutrition 36(5): 459∼469, 2003)