46 3, (2019) Korean J. Poult. Sci. Vol.46, No.3, (2019) Effects of Dietary

Size: px

Start display at page:

Download "46 3, (2019) Korean J. Poult. Sci. Vol.46, No.3, (2019) Effects of Dietary"

승희 선우
5 years ago
Views:

46 3, 161 171 (2019) Korean J. Poult. Sci. Vol.46, No.3, 161 171 (2019) https://doi.org/10.5536/kjps.2019.46.3.161 161 1 1 1 1 2 2 Effects of Dietary Zinc Supplements on the Antioxidant Indicators

1 46 3, (2019) Korean J. Poult. Sci. Vol.46, No.3, (2019) Effects of Dietary Zinc Supplements on the Antioxidant Indicators and the Expression of Zinc Transport Genes in Korean Native Chicks Dong-Gyung Jeon 1, Min-Jeong Kim 1, Il-Gyu Yoon 1, Ho-Sung Ahn 1, Sea-Hwan Sohn 2 and In-Surk Jang 2 1 Student, Department of Animal Science & Biotechnology, Gyeongnam National University of Science and Technology, Jinju 52725, Republic of Korea 2 Professor, Department of Animal Science & Biotechnology, Gyeongnam National University of Science and Technology, Jinju 52725, Republic of Korea ABSTRACT Four-week-old male Korean native chicks (KNC) were assigned to 3 groups with 6 replicates (8 birds/replicate) in each group: a basal diet (CON, 100 ppm of Zn), basal diet fortified with 50 ppm of Zn with zinc oxide (ZnO), or basal diet fortified with 50 ppm of Zn with Zn-methionine (ZnM). Immediately after a 4-week-feeding trial, 6 birds per group were used to evaluate the effects of zinc supplements on antioxidant indicators and the mrna expression of zinc transport genes. The nitrogen components, lipid peroxidation, and total antioxidant status in blood were not influenced by Zn fortified diets. However, the ZnM group showed a significant (P<0.05) increase in uric acid levels than those in the ZnO group. In the small intestine, superoxide dismutase (SOD) and glutathione peroxidase (GPX) activities, and malondialdehyde (MDA) level were unaffected by zinc supplements. The activity of glutathione S-transferase (GST) was significantly (P<0.05) enhanced by Zn-methionine supplementation. In the liver, the activity of GST was significantly (P<0.05) increased by Zn-methionine supplement without affecting SOD, GPX, and MDA levels. With respect to the mrna expression of zinc transport genes, the ZnM group displayed a strong tendency for increases in intestinal ZnT-1 (P=0.09) and ZnT-5 (P=0.06) levels, compared to those in the CON group. Moreover, the ZnM group showed a tendency (P=0.10) for up-regulation of hepatic metallothionein mrna as compared with the CON group. In conclusion, the Zn-fortified diet with 50 ppm of Zn-methionine helped to improve GST activity and Zn transport gene expression in the small intestine or liver of KNC. (Key words: zinc oxide, Zn-methionine, antioxidant enzymes, Zn transport genes) (Zn) 300 (metalloenzyme), (Prasad, 2014). 40(NRC, 1994) 50 ppm(korea Feeding Standard for Poultry, 2017) ZnO ZnSO 4 (Huang et al., 2007).,, ppm (Abd El-Wahab et al., 2013; Navidshad et al., 2016). (Liu et al., 2015). Zn-methionine To whom correspondence should be addressed : isjang@gntech.ac.kr

2 162 : (Bun et al., 2011; He et al., 2019). (Reactive oxygen species; ROS) DNA, (Song et al., 2009). ROS. ROS, (Yang et al., 2017). ROS DNA DNA (Alam and Kelleher, 2012; Prasad, 2014). Cu/Zn superoxide dismutase(sod) (Liochev and Fridovich, 2000). Bun et al.(2011) (29.6 ppm) (20, ppm), SOD,. Zn ROS (Zhou et al., 2005)., (Zinc transporters; ZnT) (Nishito and Kambe, 2018). metallothionein(mt) (Nishito and Kambe, 2018). Zn MT,, thiol (SH group) (Hamer, 1986; Kimura and Kambe, 2016). MT ROS (Tupe et al., 2010). MT Cu/ZnSOD, (Cabre et al., 2001). (100 ppm) 50 ppm Zn-methionine,. (IACUC). 4 (, ±4.50 g) 144 (CON), (ZnO, 50 ppm) (Znmethionine, 50 ppm) (ZnM) 3 6 ( ), 8. (ZnO) Zn-methionine ( Zn-120, Avalia, Zinpro Corp, USA) (100 ppm, ) 50 ppm 4. (2007 )., Table 1, 91 ppm.,.,. ( ) m 2 /, Table 1. Nutrient composition of a basal diet Items Nutrient composition ME (Mcal/kg) 2,890 Crude protein (%) Crude fat (%) 3.70 Crude fiber (%) 3.08 Lysine (%) 1.06 Ca (%) 1.14 P (%) 0.57 Zn 1 (ppm) 1 Analyzed values. 91 ppm

3 Jeon et al. : Effects of Zinc on Antioxidant Indicators and Zn Transport Genes , 6 1 (n=6). 8, sodium heparin. 3,000 rpm ( ). 3 grass slide. 5,000 rpm(4, 10 ) (Vision, VS-15,000 CF, Korea). mrna 70. total protein, albumin, blood urea nitrogen(bun) uric acid (Mindray, BS-120, Mindry Bio Medical Electronics Co., Shnzhen, China). total antioxidant status(tas) Randox kit(nx 2332, Randox Lab, Antrim, UK) (600 nm). chromogen 1: nm., 600 nm TAS. cytosol microsome Kupfer Levin(1972). 1 g 5 ml 0.25 M sucrose (0.25 M, sucrose, M MgCl 2, M KCl M CaCl 2 ph 7.4) (Omni tissue homogenizer, Omni Int. NW, USA). ( M sucrose, M MgCl M CaCl 2 ), 10,000 rpm 15. cytosol, 1 ml 1.15% KCl microsome 70. cytosol superoxide dismutase(sod) sigma ki(sigma assay kit 19160, Sigma-Aldrich, St. Louis, MO, USA). xanthine oxidase, ELISA reader(450 nm, V Max, Molecular Devices, CA, USA). kit. SOD 50% 1 unit. Glutathione peroxidase(gpx) Tappel et al. (1978) (0.1 mm NADPH, glutathione reductase 1 unit/ml, reduced glutathione, 0.25 mm; ph 7.4) 5 37 incubator, H 2 O nm. GPX unit mg protein 1 NADPH nm. Glutathione S-transferase(GST) Habig et al.(1974) 1 mm reduced glutathione chlorodinitrobenzene(cdnb), 340 nm, 1 unit mg protein 1 CDNB μm. Microsome thiobarbituric acid malondialdehyde(mda) (532 nm) (Bidlack and Tappel, 1973). (specific activity) mg. BCA (Pierce Biotechnology, Rockford, IL, USA) 560 nm ELISA reader. (ZnT-1 ZnT-5) metallothionein(mt) mrna. total RNA HiGene Total RNA Prep Kit(Biofact, Cat. No. RP , Daejeon, Korea). 30 mg 350 µl RB 5 ul protein K(10 ng/ml) ,000 rpm (4 ) 3 tube 100% 14,000 rpm(4 )

4 164 : 30. column RNA 500 μl 14,000 rpm(4 ) 2. Column 50 ul RNAease free water, total RNA (Gene Quant pro, Amersham, NJ, USA) 260/230 nm. cdna 1 µg total RNA 1.0 mg oligo dt(promega Co., Wisconsin, USA) 70 5, 4 5 (reverse transcription). 5 first strand buffer, 2.5 mm dntp, 0.1 M DTT, superscript III RT , 90 10, mrna SYBR green supermix(bio-rad, CA, USA) real-time quantitative PCR(Bio-Rad, MyiQ TM, CA, USA). ZnT-1, ZnT-5 MT primer Table 2, β-actin Livak & Schmittgen(2001) ΔCt 2 Ct. mrna SAS package program(sas, 1996) General Linear Model(GLM). 95% Tukey. (ZnO) Zn-methionine 4 4 Table 3. total protein, albumin, blood urea nitrogen(bun). (uric acid) Zn-methionine, ZnO (P<0.05). (TAS) (MDA). 70% (Park and Kweon, 2013). (Wayner et al., 1987). (Halliwell and Gutteridge, 1990). hydroxyl radicals bilirubin, A E (Wayner et al., 1987)., Znmethionine., ( ) free radical (Tate et al., 1999)., NADPH oxidase (Prasad, 2014). Yang et al.(2017) Zn palygorskite (80 ppm) 20, 40, ppm 3,, SOD. Table 2. Primer sequences used for real-time PCR Genes Primer sequences Size ZnT-1 ZnT-5 Metallothionein β-actin Forward: 5'-TGC GAG TGC CTT CTT CCT-3' Reward: 5'-AAG GAG CTG TCA GGT CTG TAA T-3' Forward: 5'-ATG CTG TTG TGG GAT GTA-3' Reward: 5'-TTG TCT TGG CTG GTC CTC-3' Forward: 5'-AAG GGC TGT GTC TGC AAG GA-3' Reward: 5'-CTT CAT CGG TAT GGA AGG TAC AAA-3' Forward: 5'-GGA GAT TAC TGC CCT GGC TCC TA-3' Reward: 5'-GAC TCA TCG TAC TCC TGC TTG CTG-3' 131 bp 159 bp 164 bp 150 bp

5 Jeon et al. : Effects of Zinc on Antioxidant Indicators and Zn Transport Genes 165 Table 3. Effect of dietary Zn supplements on blood biochemical and antioxidant components in Korean native chick Item Treatment * Significance 1 CON ZnO ZnM (P-value) Total protein (g/dl) 3.64± ± ± Albumin (g/dl) 1.21± ± ± Uric acid (mg/dl) 4.60±0.68 ab 4.00±0.73 b 5.13±0.50 a 0.05 Blood urea nitrogen (mg/dl) 0.98± ± ± Lipid peroxidation (mm MDA/mg protein) 0.70± ± ± Total antioxidant status (mm/l) 1.23± ± ± * CON (Control), ZnO (Zinc oxide, 50 ppm), and ZnM (Zn-methionine, 50 ppm). 1 Significance (P-value) indicated the treatment P-value of Proc GLM procedure. a,b Values (mean±sd, n=6) with different superscripts differ significantly (P<0.05) among treatments. TAS. 50 ppm uric acid,. Zn-methionine, Fig SOD Fig. 1. Effects of dietary Zn supplements on the specific activity of antioxidant enzymes (A, SOD; B, GPX; and C, GST) and lipid peroxidation level (MDA, D) in the small intestinal mucosa of Korean native chick. * CON (Control), ZnO (Zinc oxide, 50 ppm), and ZnM (Zn-methionine, 50 ppm). Significance (P-value) indicated the treatment P-value of Proc GLM procedure. a,b Values (mean±sd, n=6) with different superscripts differ significantly (P<0.05) among treatments.

166 : Fig. 2. Effects of dietary Zn supplements on the specific activity of antioxidant enzymes (A, SOD; B, GPX; and C, GST) and lipid peroxidation level (MDA, D) in the liver of Korean native chick.

6 166 : Fig. 2. Effects of dietary Zn supplements on the specific activity of antioxidant enzymes (A, SOD; B, GPX; and C, GST) and lipid peroxidation level (MDA, D) in the liver of Korean native chick. * CON (Control), ZnO (Zinc oxide, 50 ppm), and ZnM (Zn-methionine, 50 ppm). Significance (P-value) indicated the treatment P-value of Proc GLM procedure. a,b Values (mean±sd, n=6) with different superscripts differ significantly (P<0.05) among treatments. (P=0.07), SOD GPX (MDA) (Fig. 1). GST, ZnM (P<0.05). SOD GPX Zn-methionine 50 ppm. GST Zn-methionine (P<0.05) (Fig. 2). (MDA)., (Espinosa-Diez et al., 2015). SOD, GPX, GST catalase, thiol ( glutathione, MT ), C, E (Surai, 2003). Cu/Zn SOD O 2 H 2 O 2 (Liochev and Fridovich, 2000). GPX H 2 O 2 H 2 O (Tappel et al., 1978). GST (Hayes et al., 2005). Zn. Liu et al.(2015) (0 ppm) (60, ppm) (ZnSO 4 Zn-amino acid), SOD.,, 60 ppm (Liu et al., 2015). Bun et al.(2011) (29.6 ppm)

7 Jeon et al. : Effects of Zinc on Antioxidant Indicators and Zn Transport Genes 167 Zn (0, 20, ppm), catalase, SOD 20 ppm. Zhao et al.(2014) (60 ppm, ZnO) (20, ppm), ppm SOD., Bun et al.(2011) (oocyst). Rao et al.(2016) 40 ppm, SOD,., Zn-methionine GST,., Mansour et al.(2017) GST., GST (Jamakala and Rani, 2015). (40 ppm), SOD, GST,. Zn-methionine, (ZnT-1, ZnT-5) MT mrna Table 4 5. ZnT-1(8.62, P=0.09) ZnT-5 (7.3, P=0.06) mrna Zn-methionine,. MT mrna (Table 4). (ZnT-1, ZnT-5) MT mrna, ZnO (2.47 ) ZnM (4.12 ) MT mrna (P=0.10) (Table 5)., SOD, MT (Kimura and Kambe, 2016). (ZnT-1 ZnT-5) MT mrna. Zn (ZnT-1, ZnT-5 ) (ZIP) 2 (Nishito and Kambe, 2018). ZnT-1 Table 4. Effects of dietary Zn supplements on the mrna expression of Zn transporters (ZnT-1 and ZnT-5) and metallothionein (MT) in small intestinal mucosa of Korean native chick Items Treatment * Significance 1 CON ZnO ZnM ΔCt 2 ΔΔCt ΔCt 2 ΔΔCt ΔCt 2 ΔΔCt (P-value) ZnT ± ± ± ZnT ± ± ± MT 9.47± ± ± * CON (Control), ZnO (Zinc oxide, 50 ppm), and ZnM (Zn-methionine, 50 ppm). 1 Significance (P-value) indicated the treatment P-value of Proc GLM procedure. The values (mean±sd, n=6) are Ct, which is represented as the Ct of each target gene corrected by Ct of the control gene (β-actin). The fold difference in the relative expression of the target gene was calculated as the 2 Ct.

8 168 : Table 5. Effects of dietary Zn supplements on the mrna expression of Zn transporters (ZnT-1 and ZnT-5) and metallothionein (MT) in the liver of Korean native chick Items Treatment CON ZnO ZnM ΔCt 2 -ΔΔCt ΔCt 2 -ΔΔCt ΔCt 2 -ΔΔCt Significance 1 (P-value) ZnT ± ± ± ZnT ± ± ± MT 3.90± ± ± * CON (Control), ZnO (Zinc oxide, 50 ppm) and ZnM (Zn-methionine, 50 ppm). 1 Significance (P-value) indicated the treatment P-value of Proc GLM procedure. The values (mean±sd, n=6) are Ct, which is represented as the Ct of each target gene corrected by Ct of the control gene (β-actin). The fold difference in the relative expression of the target gene was calculated as the 2 Ct. (McMahon and Cousins, 1998; Yu et al., 2008). ZnT-5 (Cragg et al., 2005). MT (Nishito and Kambe, 2018). MT, 23 33% thiol (Pool) (Hamer, 1986; Ruttkay-Nedecky et al., 2013). Tang et al.(2015), ZnSO 4 Zn zeolite clinoptilolite(zn-zcp) 80 ppm ZnT-1, ZnT-2 ZnT-5 MT mrna. Wang et al.(2019) 80 ppm Zinc pectin oligosaccharides MT ZnT-1 mrna. Li et al.(2015) (80 ppm, ZnSO 4 ) zeolite clinoptilolite ppm 80.5 ppm ZnT-1 MT mrna. He et al.(2019) 90 ppm ZnSO 4 ZnT-5, ZnT-1. Liu et al.(2015) Zn ( vs. ) (0, 60, ppm) MT mrna 0 ppm, 60 ppm. 100 ppm 50 ppm (ZnT-1 ZnT-5) MT mrna. Zn, Zn,. Zn-methionine,.. Zn (100 ppm, ) (ZnO) Zn-methionine(ZnM) 50 ppm 3 4. total protein, albumin, blood urea nitrogen, uric acid ZnM ZnO

9 Jeon et al. : Effects of Zinc on Antioxidant Indicators and Zn Transport Genes 169 (P<0.05).., superoxide dismutase(sod), glutathione peroxidase(gpx), glutathione S-transferase (GST) ZnM (P<0.05). SOD GPX, GST ZnM (P<0.05). ZnT-1(8.62 P=0.09) ZnT-5(7.3, P=0.06) mrna ZnM,. mrna, ZnM MT mrna 4.12 (P=0.10). Zn-methionine (50 ppm) GST, mrna. (RAIC) (2018 ),. Dong-Gyung Jeon Min-Jeong Kim Il-Gyu Yoon Ho-Sung Ahn Sea-Hwan Sohn In-Surk Jang Abd El-Wahab A, Radko D, Kamphues J 2013 High dietary levels of biotin and zinc to improve health of foot pads in broilers exposed experimentally to litter with critical moisture content. Poult Sci 92(7): Alam S, Kelleher SL 2012 Cellular mechanisms of zinc dysregulation: a perspective on zinc homeostasis as an etiological factor in the development and progression of breast cancer. Nutrients 4(8): Bidlack WR, Tappel AL 1973 Damage to microsomal membrane by lipid peroxidation. Lipid 8(4): Bun SD, Guo YM, Guo FC, Ji FJ, Cao H 2011 Influence of organic zinc supplementation on the antioxidant status and immune responses of broilers challenged with Eimeria tenella. Poult Sci 90(8): Cabre M, Camps J, Ferre N, Paternain JL, Joven J 2001 The antioxidant and hepatoprotective effects of zinc are related to hepatic cytochrome P450 depression and metallothionein induction in rats with experimental cirrhosis. Int J Vitam Nutr Res 71(4): Cragg RA, Phillips SR, Piper JM, Varma JS, Campbell FC, Mathers JC, Ford D 2005 Homeostatic regulation of zinc transporters in the human small intestine by dietary zinc supplementation. Gut 54(4): Espinosa-Diez C, Miguel V, Mennerich D, Kietzmann T, Sanchez-Perez P, Cadenas S, Lamas S 2015 Antioxidant responses and cellular adjustments to oxidative stress. Redox Biol 6: Habig WH, Pabst MJ, Jakoby WB 1974 Glutathionine S transferase: the first enzymatic steps in mercapturic acid formation. J Biol Chem 249(22): Halliwell B, Gutteridge JM 1990 The antioxidants of human extracellular fluids. Arch Biochem Biophys 280(1):1-8. Hamer DH 1986 Metallothionein. Annu Rev Biochem 55: Hayes JD, Flanagan JU, Jowsey IR 2005 Glutathione transferases. Annu Rev Pharmacol Toxicol 45: He B, Bortoluzzi C, King WD, Graugnard D, Dawson KA, Applegate TJ 2019 Zinc source influences the gene expression of zinc transporters in jejunum and cecal tonsils during broiler challenge with Eimeria maxima and Clostridium perfringens. Poult Sci 98(3): Huang YL, Lu L, Luo XG, Liu B 2007 An optimal dietary zinc level of broiler chicks fed a corn-soybean meal diet. Poult Sci 86(12): Jamakala O, Rani UA 2015 Amelioration effect of zinc and

10 170 : iron supplementation on selected oxidative stress enzymes in liver and kidney of cadmium-treated male albino rat. Toxicol Int 22(1):1-9. Kimura T, Kambe T 2016 The functions of metallothionein and ZIP and ZnT transporters: an overview and perspective. Int J Mol Sci 17(3): Korea Feeding Standard for Poultry 2007 National Institute of Animal Science, Rural Development Administration. Kupfer D, Levin E 1972 Monooxygenase drug metabolizing activity in CaCl 2 -aggregated hepatic microsomes from rat liver. Biochem Biophys Res Commun 47(3): Li L, Li P, Chen Y, Wen C, Zhuang S, Zhou Y 2015 Zinc-bearing zeolite clinoptilolite improves tissue zinc accumulation in laying hens by enhancing zinc transporter gene mrna abundance. Anim Sci J 86(8): Liochev SI, Fridovich I 2000 Cu- and Zn-containing superoxide dismutase can act as a superoxide reductase and a superoxide oxidase. J Biol Chem 275(49): Liu ZH, Lu L, Wang RL, Lei HL, Li SF, Zhang LY, Luo XG 2015 Effects of supplemental zinc source and level on antioxidant ability and fat metabolism-related enzymes of broilers. Poult Sci 94(11): Livak KJ, Schmittgen TD 2001 Analysis of relative gene expression data using real-time quantitative PCR and the 2 ΔΔCT method. Methods 25(4): Mansour SA, Abbassy MA, Shaldam HA 2017 Zinc ameliorate oxidative stress and hormonal disturbance induced by methomyl, abamectin, and their mixture in male rats. Toxics 5(4):E McMahon RJ, Cousins RJ 1998 Mammalian zinc transporters. J Nutr 128(4): National Research Council 1994 Nutrient Requirement of Poultry. 9 th rev ed. National Academic Press. Washington, DC, USA. Navidshad B, Jabbari S, Mirzael Aghjeh Gheshlagh F 2016 The new progresses in Zn requirements of poultry. Iranian J App Anim Sci 6(4): Nishito Y, Kambe T Absorption mechanisms of iron, copper, and zinc: an overview. J Nutr Sci Vitaminol (Tokyo) 64(1):1-7 Park JH, Kweon GR 2013 Clinical applications of antioxidants. Hanyang Med Rev 33(2): Prasad AS 2014 Zinc: an antioxidant and anti-inflammatory agent: role of zinc in degenerative disorders of aging. J Trace Elem Med Biol 28(4): Rao SV, Prakash B, Raju MV, Panda AK, Kumari RK, Reddy EP 2016 Effect of supplementing organic forms of zinc, selenium and chromium on performance, antioxidant and immune responses in broiler chicken reared in tropical summer. Biol Trace Elem Res 172(2): Ruttkay-Nedecky B, Nejdl L, Gumulec J, Zitka O, Masarik M, Eckschlager T, Stiborova M, Adam V, Kizek R 2013 The role of metallothionein in oxidative stress. Int J Mol Sci 14(3): SAS 1996 User's Guide: Statistics Version 6.12 Ed. SAS Inst., Inc., Cary, NC. Song Y, Leonard SW, Traber MG, Ho E 2009 Zinc deficiency affects DNA damage, oxidative stress, antioxidant defenses, and DNA repair in rats. J Nutr 139(9): Surai PF 2003 Natural antioxidant in avian nutrition and reproduction. Pages 1-25 In: Antioxidant Systems in the Animal Body. Nottingham University Press, Thrumpton. Tang Z, Wen C, Li P, Wang T, Zhou Y 2014 Effect of zinc-bearing zeolite clinoptilolite on growth performance, nutrient retention, digestive enzyme activities, and intestinal function of broiler chickens. Biol Trace Elem Res 158(1): Tappel AL 1978 Glutathione peroxidase and hydroperoxides. Methods in Enzymol 52: Tate DJ Jr, Miceli MV, Newsome DA 1999 Zinc protects against oxidative damage in cultured human retinal pigment epithelial cells. Free Radic Biol Med 26(5-6): Tupe RS, Tupe SG, Tarwadi KV, Agte VV 2010 Effect of different dietary zinc levels on hepatic antioxidant and micronutrients indices under oxidative stress conditions. Metabolism 59(11): Wang ZC, Yu HM, Xie JJ, Cui H, Nie H, Zhang TT, Gao XH 2019 Effect of dietary zinc pectin oligosaccharides chelate on growth performance, enzyme activities, Zn accumulation, metallothionein concentration, and gene expression of Zn transporters in broiler chickens. J Anim Sci 97(5):

11 Jeon et al. : Effects of Zinc on Antioxidant Indicators and Zn Transport Genes 171 Wayner DD, Burton GW, Ingold KU, Barclay LR, Locke SJ 1987 The relative contributions of vitamin E, urate, ascorbate and proteins to the total peroxyl radical-trapping antioxidant activity of human blood plasma. Biochim Biophys Acta 924(3): Yang W, Chen Y, Cheng Y, Wen C, Zhou Y 2017 Effects of zinc bearing palygorskite supplementation on the growth performance, hepatic mineral content, and antioxidant status of broilers at early age. Asian-Aust J Anim Sci 30(7): Yu Y, Lu L, Luo XG, Liu B 2008 Kinetics of zinc absorption by in situ ligated intestinal loops of broilers involved in zinc transporters. Poult Sci 87(6): Zhao CY, Tan SX, Xiao XY, Qiu XS, Pan JQ, Tang ZX 2014 Effects of dietary zinc oxide nanoparticles on growth performance and antioxidative status in broilers. Biol Trace Elem Res 160(3): Zhou Z, Wang L, Song Z, Saari JT, McClain CJ, Kang YJ 2005 Zinc supplementation prevents alcoholic liver injury in mice through attenuation of oxidative stress. Am J Pathol 166(6): Received Jul. 11, 2019, Revised Aug. 3, 2019, Accepted Aug. 10, 2019

γ

경락경혈학회지 Vol.27, No.1, pp.87 106, 2010 Journal of Meridian & Acupoint Dept. of 1 Meridian & Acupoint, 3 Acupuncture & Moxibustion, College of Oriental Medicine, Daejeon University 2 Division of Clinical