The experiment was conducted to study the effects of total flavones of Hippophae rhamnoides (TFH) and quercetin on alkaline phosphatase (ALP) activity in osteoblasts of arbor acres (AA) broilers’ embryo in vitro and its mechanisms of action. Cells were cultured in basal culture medium, basal culture medium supplemented with β-estradiol (E2), TFH and quercetin for 24 h, respectively; after treated by estrogen receptor (ER) antagonist for 2 h, the cells were cultured for another 24 h in basal culture medium, basal culture medium supplemented with TFH and quercetin, respectively; after treated with extracellular signal-regulated kinase (ERK) inhibitor for 2 h, the cells were cultured for another 24 h in basal culture medium, basal culture medium supplemented with TFH and quercetin, respectively; the cells were collected for the analysis of ALP activity. Cells were cultured in basal culture medium supplemented with TFH or quercetin, and collected at six time points (0, 2, 5, 15, 30, 60 min) for the determination of phosphorylated extracellular-regulated kinase (p-ERK) concentration by ELASE. Cells were cultured in basal culture medium, basal culture medium supplemented with E2, TFH or quercetin for 30 min, respectively; after treated by ER antagonist for 15 min, the cells were cultured for 30 min in basal culture medium, basal culture medium supplemented with E2, TFH and quercetin, respectively; cells were collected for the determination of p-ERK concentration by ELASE. The results showed as follows: 1) culture medium supplemented with TFH or quercetin significantly increased the ALP activity (P<0.05), and the effects of which were totally prevented by ER antagonist and MEK inhibitor (P<0.05); 2) the phosphorylation level of ERK were significantly increased in culture medium supplemented with TFH after 5, 15 min incubation and with quercetin after 2, 5, 15, 30, 60 min incubation (P<0.05); 3) culture medium supplemented with TFH or quercetin significantly increased the phosphorylation level of ERK, the effects of which were totally prevented by ER antagonist (P<0.05). The results indicate that TFH and quercetin can significantly increase the ALP activity and ERK phosphorylation level, exert the estrogen-like effect, and may adjust the ALP activity via ERK and ER pathways. ［Chinese Journal of Animal Nutrition, 2011, 23（8）：1378 -1385］
LIU Hongzhu,LI Yao,CHEN Xin,ZHAO Wei,ZHANG Lixin,OUYANG Wenwen,LIU Hongnan
. Mechanisms of Action for Total Flavones of Hippophae Rhamnoides and Quercetin on Alkaline Phosphatase Activity in Chicken Osteoblasts[J]. Chinese Journal of Animal Nutrition, 2011
, 23(08)
: 1378
-1385
.
DOI: 10.3969/j.issn.1006-267x.2011.08.017
［1］史琳娜,苏宜香.大豆异黄酮类对去卵巢大鼠骨丢失的影响[J].动物营养学报,2010,22(2):113-117.
[2]范媛媛.大豆异黄酮对体外培养成骨细胞作用的实验研究[D].硕士学位论文.天津:天津医科大学,2007.
[3]潘永梅.异黄酮类植物雌激素对大鼠成骨细胞生物活性影响的实验研究[D].硕士学位论文.石家庄:河北医科大学,2006.
[4]张力,郑中朝,田青,等.大豆异黄酮对樱桃谷鸭钙磷代谢的影响[J].甘肃农业大学学报,2006,41(6):1-5.
[5]高锦明,张鞍灵,李芸生,等.沙棘黄酮化学研究的进展[J].沙棘,1998,11(2):34-40.
[6]李垚,王宝东,王艳波,等.沙棘黄酮对AA肉鸡生长性能和内分泌的影响[J].中国家禽,2008,30(2):13-15.
[7]陈鑫,李垚,赵伟,等.沙棘叶总黄酮对AA肉鸡饲粮中钙磷利用率和胫骨发育的影响[J].中国饲料,2010(8):30-33.
[8]IVANOVA T V, IVANOV V N, NADEZHDINA E S. Transcription factors NF-kappaB and AP-1/c-fos in cell response to nocodazol[J]. Membrane & Cell Biology, 2001, 14(6):727-741.
[9]赵二劳,张敬,吕晶,等.可见与紫外光度法测定沙棘叶中总黄酮的比较[J].沙棘,2008,21(2):22-23.
[10]张建峰.鸡IGF-Ⅰ对成骨细胞功能的影响及其基因多态性与蛋鸡骨代谢相关性的研究[D].博士学位论文.南京:南京农业大学,2002.
[11]RODAN G A. Introduction to bone biology bone[J]. Bone, 1992, 13 (Suppl. 1):S3-S6.
[12]李勇,季晖,李萍.淫羊藿总黄酮对体外培养成骨细胞的影响[J].中国药科大学学报,2002,33(1):48-50.
[13]卓丽玲,顾建红,赵瑞英,等.乳鼠成骨细胞的培养及鉴定[J].中国兽医科学,2007,37(10):901-904.
[14]邓益锋,陈秀霞,侯加法.雌激素鸡胚成骨细胞增殖、凋亡及其ALP活性的影响[C]//中国畜牧兽医学会2008年学术年会暨第六届全国畜牧兽医青年科技工作者学术研讨会论文集.北京：中国畜牧兽医学会,2008.
[15]PROUILLET C, MAZIERE J C, MAZIERE C, et al. Stimulatory effect of naturally occurring flavonols quercetin and kaempferol on alkaline phosphatase activity in MG-63 human osteoblasts through ERK and estrogen receptor pathway[J]. Biochemical Pharmacology, 2004, 67(7):1307-1313.
[16]KANNO S, HIRANO S, KAYAMA F. Effects of phytoestrogens and environmental estrogens on osteoblastic differentiation in MC3T3-E1 cells[J]. Toxicology, 2004, 196(1):137-145.
[17]KUIPER G G, LEMMEN J G, CARLSSON B, et al. Interaction of estrogenic chemicals and phytoestrogens with receptor beta[J]. Endocrinology, 1998, 139(10):4252-4263.
[18]HSU Y L, LIANG H L, HUNG C H, et al. Syringetin, a flavonoid derivative in grape and wine, induces human osteoblast differentiation through bone morphogenetic protein-2/extracellular signal-regulated kinase 1/2 pathway[J]. Molecular Nutrition & Food Research, 2009, 53(11):1452-1461.
[19]ZHANG Z, MAIER B, SANTEN R J, et al. Membrane association of estrogen receptor alpha mediates estrogen effect on MAPK activation[J]. Biochemical and Biophysical Research Communications, 2002, 28(5):926-933.
[20]PEVERALI F A, BASDRA E K, PAPAVASSILIOU A G. Stretch-mediated activation of selective MAPK subtypes and potentiation of AP-1 binding in human osteoblastic cells[J]. Molecular Medicine, 2001, 7(1):68-78.
