• Journal of Animal Environmental Science
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• v.3 no.1
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• pp.1-12
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• 1997

This study was conducted to investigate the combined effects of VFA composition of rumen fluid and heat exposure (30${\pm}$2$^{\circ}C$) on the general clinical view and insulin secretory response to glucose in sheep. The total infusion of nutrients was examined in sheep via the technique of continuous alimentation. Four adult Suffolk sheep fitted with a permanent ruminal cannula and a simple T-shaped duodenal cannula were used. A peristaltic pump was used to infuse the solutions of volatile fatty acid triglycerides (VFA-TG) consisting of 70 triacetin : 20 tripropionin : 10 tributyrin (low propionin division: LP) and 50 triacetin : 40 tripropionin : 10 tributyrin (high propionin division: HP) on the basis of energy and minerals into the rumen, and casein solution into the duodenum. The effects of heat exposure and type of the levels of VFA-TG solutions on the insulin secretory response to glucose in sheep were investigated by using hyperglycemic clamp (HGC) technique. The results obtained are summarized as follows: 1. During the heat exposure (latter half of the infusion period), respiration rate, heart rate and rectal temperature increased (P＜0.01, P＜0.01, P＜0.05), but the levels of VFA-TG solutions (LP and HP division) did not affect the general clinical view except for the heart rate. 2. In the HGC technique, glucose infusion rate (GIR) and mean plasma insulin increments (MPII) tended to be ower in the heat exposure than in the thermoneutral environment, but no significant difference was found among the treatments. GIR and MPII remained unchanged between the levels of VFA-TG solutions. 3. In the HGC technique, ratio of MPII to GIR (MPII/GIR) which represents pancreatic ${\beta}$-cell response to glucose stimulation remained unchanged among the treatments.

• Journal of Nutrition and Health
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• v.33 no.7
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• pp.697-702
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• 2000

The objective of this study was to determine the extent to which the compensatory nutrition regimen modulates lactation performance and milk protein gene expression in the first and second lactation cycles. Female rats(28 days of age) were assigned to 1)control ad libitum ; 2) stari-step compensatory nutrition(SSCN) regimen an alternating 3-2-3-4-week schedule beginning with an energy restriction diet(40% restriction) for 3 weeks followed by the control diet(ad libitum) for 2 weeks and then alternating another 3-4 week feeding regimen. The SSCN rats were received an overall 20% energy restriction(average from all stair-step periods) compared with the conventionally fed control group. Rats were bred during the first week of the second realimentation. All pups were weaned on day 21 of lactation. About 1 week after weaning all dams were mated for the second pregnancy. Mammary tissues were obtained from pregnant and lactating rats during the first and second lactation cycles. During these lactation cycles the SSCN group had a 11% increase in average lactation performance over that of control. The SSCN group had significantly increased levels of milk protein gene($\alpha$- and $\beta$-casein) expression in mammary tissues during the first lactation cycle compared with those of the control group. During the second lactation period the levels of milk protein gene expression in lactating mammary tissues of the SSCN group were also higher than those of the control group. These results suggest that the effects of compensatory growth imposed at an early age extend to the second lactation cycle with regard to increased lactation performance and milk protein gene expression.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.60-60
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• 2001

Human thrombopoietin (hTPO) is a cytokine that plays a central role in megakaryopoiesis by influencing on the development and maturation of megakaryocyte and platelet production. To induce hTPO production in the mammary gland, expression vector was constructed by combining the promoter of bovine beta-casein gene, cDNA of hTPO and neomycine resistance gene for transfection into fibroblasts. Bovine fibroblast cells derived from female ear skin were transfercted with the expression vector using Lipofectamine (Life Technology, NY). Transected cells resistant to G4l8 treatment (600 $\mu\textrm{g}$/$m\ell$) were recovered and colony formation was initiated at 13 days. The colonies with about 1 cm diameter were picked and analysed by PCR. Single transfected cells were individually transferred to enucleated oocytes. After electrofusion, the reconstructed embryos were exposed to calcium ionophore (5uM) for 5 min followed by treatment with 6-DMAP (2.5 mM) for 4h. The nuclear transfer embryos were cultured in CRlaa medium at 38.5C, 5% $CO_2$ for 7 days. Twenty three of 29 (79.3%) colonies were proved to be hTPO transfectants by PCR. The colonies were further passaged and used to produce transgenic embryos using nuclear transfer. Cleavage and developmental rates of reconstructed embryos to the blastocyst stage were 65.1% and 39.4%, respectively Of 22 blastocysts that developed from reconstructed embryos with the transfected cell, 20 embryos (90.9%) were positive for hTPO by using PCR analysis. The results suggest that somatic cell nuclear transfer is efficient for production of transgenic embryos.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.1
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• pp.19-26
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• 2017

We investigated whether betulin affects the gene expression, secretion and proteolytic activity of matrix metalloproteinase-3 (MMP-3) in primary cultured rabbit articular chondrocytes, as well as in vivo production of MMP-3 in the rat knee joint to evaluate the potential chondroprotective effect of betulin. Rabbit articular chondrocytes were cultured and reverse transcription-polymerase chain reaction (RT-PCR) was used to measure interleukin-$1{\beta}$ ($IL-1{\beta}$)-induced gene expression of MMP-3, MMP-1, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs-4 (ADAMTS-4), ADAMTS-5 and type II collagen. Effect of betulin on IL-$1{\beta}$-induced secretion and proteolytic activity of MMP-3 was investigated using western blot analysis and casein zymography, respectively. Effect of betulin on MMP-3 protein production was also examined in vivo. The results were as follows: (1) betulin inhibited the gene expression of MMP-3, MMP-1, MMP-13, ADAMTS-4, and ADAMTS-5, but increased the gene expression of type II collagen; (2) betulin inhibited the secretion and proteolytic activity of MMP-3; (3) betulin suppressed the production of MMP-3 protein in vivo. These results suggest that betulin can regulate the gene expression, secretion, and proteolytic activity of MMP-3, by directly acting on articular chondrocytes.

• The Korean Journal of Nuclear Medicine
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• v.35 no.5
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• pp.313-323
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• 2001

Purpose: The purpose of this study was to search activated genes that could be related to radiation adaptive response (RAR) induced by low-level radiation from $^{99m}Tc$ in human cell lines. Methods: We used gene discovery array (GDA) and representational difference analysis (RDA) methods. $^{99m}Tc$-pertechnetate was added to $2{\times}106/mL$ NC-37 cells (human lymphoblastic cells) to make concentrations ranging from 148 MBq/mL to 148 Bq/mL by serial 10 fold dilutions. After 44 hours, 2 Gy gamma irradiation was given to them using a Cs-137 cell irradiator. Results: As compared to the control (Con) group to which no $^{99m}Tc$ was added, those cells to which 148 and 14.8 KBq of $^{99m}Tc$ were added showed significantly lower damage to chromosomes, which was evaluated by metaphase analysis. Cells with 148 KBq $^{99m}Tc$ (T148 group) showed most significant protection. Activated genes in the T148 group as compared to Con group were evaluated by GDA and GDA methods. GDA revealed genes of casein kinase 2 (CK2) beta chain, immunoglobulins (lg), human leukocyte antigen (HLA)-B, and two novel genes. Twenty RAR related clones were selected by RDA method. The size of those genes was from 234 to 603 base pairs. Conclusions: RAR was induced by low dose irradiation from $^{99m}Tc$ in NC-37 cell lines. Genes related to the response included CK2, lg, HLA-B in human lymphoblastic cell lines.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.85-85
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• 2003

Human lactoferrin (hLF) is an 80 kDa iron-binding glycoprotein that is expressed in high concentration in milk and in lesser amount in the secondary or specific granules of neutrophils and in plasma, LF is classically considered to be related to the binding, transport, and storage of iron. The transgenic mice carrying the human hLF gene in conjunction with the bovine $\beta$-casein promoter produced the human hLF in their milk during lactation. To screen transgenic mice, PCR was carried out using chromosomal DNA extracted from tail or toe tissues. In this study, stability of germ line transmission and expression of hLF were monitored up to generation Fl7 of a transgenic line. When female mouse of generation F9 was crossbred with normal male, generation F9 to Fl7 mice showed similar transmission rates ($66.0 \pm 12.57%, 42.0 \pm 14.98%, 72.2 \pm 25.45%, 50.0 \pm 16.70%, 65.7 \pm 6.45%, 48.6 \pm 14.65%, 54 1 \pm 18 11%, 57.8 \pm 16.16% and 48.6 \pm 20.66$, respectively), implying that the hLF gene can be transmitted stably up to long term generation in the transgenic mice For ELISA analysis, hLF expression levels were determined with an hLF ELISA kit in accordance with the supplier's protocol. Expression levels of human hLF from milk of generation F9 to Fl3 mice were $ 3.2 \pm 0.69 mg/ml, 3.1 \pm 0.81 mg/ml, 4.6 \pm 1.38 mg/ml, 3.1 \pm 0.42 mg/ml, and 4.5 \pm 1,48 mg/ml$, respectively. These expression levels were lower than that of founder (6.6 mg/$m\ell$) mouse. We concluded that transgenic mice faithfully passed the transgene on their progeny and successively secreted target proteins into their milk through several generations.

• Parasites, Hosts and Diseases
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• v.56 no.2
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• pp.135-145
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• 2018

Due to the critical location and physiological activities of the retinal pigment epithelial (RPE) cell, it is constantly subjected to contact with various infectious agents and inflammatory mediators. However, little is known about the signaling events in RPE involved in Toxoplasma gondii infection and development. The aim of the study is to screen the host mRNA transcriptional change of 3 inflammation-related gene categories, PI3K/Akt pathway regulatory components, blood vessel development factors and ROS regulators, to prove that PI3K/Akt or mTOR signaling pathway play an essential role in regulating the selected inflammation-related genes. The selected genes include PH domain and leucine- rich-repeat protein phosphatases (PHLPP), casein kinase2 (CK2), vascular endothelial growth factor (VEGF), pigment epithelium-derived factor (PEDF), glutamate-cysteine ligase (GCL), glutathione S-transferase (GST), and NAD(P)H: quinone oxidoreductase (NQO1). Using reverse transcription polymerase chain reaction (RT-PCR) and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), we found that T. gondii up-regulates PHLPP2, $CK2{\beta}$, VEGF, GCL, GST and NQO1 gene expression levels, but down-regulates PHLPP1 and PEDF mRNA transcription levels. PI3K inhibition and mTOR inhibition by specific inhibitors showed that most of these host gene expression patterns were due to activation of PI3K/Akt or mTOR pathways with some exceptional cases. Taken together, our results reveal a new molecular mechanism of these gene expression change dependent on PI3K/Akt or mTOR pathways and highlight more systematical insight of how an intracellular T. gondii can manipulate host genes to avoid host defense.

• Proceedings of the KSAR Conference
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• 2003.06a
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• pp.46-46
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• 2003

Interleukin-10 (IL-10) is a homodimeric protein with a wide spectrum of anti-inflammatory and immune activities. It inhibits cytokine production and expression of immune surface molecules in various cell types. The transgenic mice carrying the human IL-10 gene in conjunction with the bovine $\beta$-casein promoter produced the human IL-10 in milk during lactation. Transgenic mice were generated using a standard method as described previously. To screen transgenic mice, PCR was carried out using chromosomal DNA extracted from tail or toe tissues with a primer set. In this study, stability of germ line transmission and expression of IL-10 gene integrated into host chromosome were monitored up to generation F15 of a transgenic line. When female mouse of generation F9 was crossbred with normal male, generation F9 to F15 mice showed similar transmission rates (66.0$\pm$20.13%, 61.5$\pm$16.66%, 41.1$\pm$8.40%, 40.7$\pm$20.34%, 61.3$\pm$10.75%, 49.2$\pm$18.82%, and 43.8$\pm$25.91%, respectively), implying that the IL-10 gene can be transmitted stably up to long term generation in the transgenic mice. For ELISA analysis, IL-10 expression levels were determined with an hIL-10 ELISA and a mIL-10 ELISA kit in accordance with the supplier's protocol. Expression levels of human IL-10 from milk of generation F9 to F13 mice were 3.6$\pm$1.20 mg/ml, 4.2$\pm$0.93 mg/ml, 5.7$\pm$1.46 mg/ml, 6.3$\pm$3.46 mg/ml, and 6.8$\pm$4.52 mg/ml, respectively. These expression levels are higher than in generation F1 (1.6 mg/ml) mice. We concluded that transgenic mice faithfully passed the transgene on their progeny and successively secreted target proteins into their milk through several generations, although there was a little fluctuation in the transmission frequency and expression level between the generations.

• Korean Journal of Animal Reproduction
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• v.27 no.1
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• pp.25-34
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• 2003

Human thrombopoietin (hTPO) is a cytokine that plays a central role in megakaryopoiesis. To direct hTPO expression in the mammary gland, an expression vector was constructed by combining the promoter of bovine beta-casein gene, cDNA of hTPO and neomycin resistance gene (pBT-L neo). Fibroblast cells derived from cow's ear skin tissue were transfected with the expression vector (pBT-L neo) using Lipofectamine. Transfected cells resistant to G418 trea？nt were cultured to form the colonies for more than 2 weeks. The transformed colonies identified by PCR were further expanded prior to nuclear transfer. Reconstructed oocytes with transformed cells were electrofused, activated using calcium ionophore and 6-DMAP, and cultured in vitro for 7 days. Of 35 cell colonies analyzed by PCR, 29 colonies (82.9%) were positive for the hTPO gene. Cleavage and developmental rates to the blastocyst stage of reconstructed embryos with the transformed cells were 65.1% and 23.8%, respectively Of 29 blastocysts that developed from reconstructed embryos with the transformed cells, 27 embryos (93.1%) were transgenic. These results indicate that transgenic bovine embryos can be efficiently produced by somatic cell nuclear transfer using transformed cells.

• Journal of Nutrition and Health
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• v.2 no.4
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• pp.173-181
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• 1969

Since 1959 ${\beta}-aminoethylphosphonic$ acid was discovered in the living organism, the biosynthesis and biological functions of aminophosphonic acids have been extensively studied. The author designed and carried out this study for 14 weeks to find out the metabolic function of Ethylaminophosphonic acid (AEP) and it's utilization in the living body. Sixty rats, thirty males and thirty females aged $40{\pm}5$ days were divided into two parts, one for alanine supplemented as control group and the other for AEP as experimental group to compare metabolic pathway of ordinary amino acid with that of AEP. Both alamine and AEP group were divived into two subgroups according to the level of supplements, 0.1% and 0.2% of the diet. The major components of the diet in this study were composed of 20% casein, 72% Sugar, 4% fat, 4% salt Mixture, and all kind of Uitamins in adeguate amount. For comparision of biological values between experimental and control group in terms of body weight, uninary nitrogen, creatinine excretion and final orgam weight, there were no statically significant difference in these respects. This meant AEP could be utilized in the body as much as alanine could. Urinary phosphorus excretion was determined by developing the blue color to read on the Spectronic 20. Statistically insignificance in the urinary phosphorus excretion between experimental and control group was observed in spite of the supplementation of phosphorus of AEP for experimental group in the diet. The level of blood phosphorus was higher in experimental group than that in control group this result supported above result. In the analysis of fat and nitrogen contents in the liver, AEP group showed slightly higher than control in both respects. But it was noteworthy 0.2% AEP group in both sex were higher than 0.1% AEP in liver fat content. Histological examinal of internal organs liiver, lung, spleen, heart, kindey, adrenal and sex organs showed no changes in all groups included in this study. The group supplemented higher level of diet. by alanine 0.2% and AEP 0.2% stayed on less body weight gain and lower liver weight. This result could be interpreted that amino acid imbalanced condition was arose in the body.