基于高通量测序技术的牡蛎壳粉缓解骨质疏松症的研究

石秋月 侯付景 韩姣姣 李菁菁 陈菊 苏秀榕

石秋月,侯付景,韩姣姣,等. 基于高通量测序技术的牡蛎壳粉缓解骨质疏松症的研究[J]. 食品工业科技,2021,42(18):372−379. doi:  10.13386/j.issn1002-0306.2020120189
引用本文: 石秋月,侯付景,韩姣姣,等. 基于高通量测序技术的牡蛎壳粉缓解骨质疏松症的研究[J]. 食品工业科技,2021,42(18):372−379. doi:  10.13386/j.issn1002-0306.2020120189
SHI Qiuyue, HOU Fujing, HAN Jiaojiao, et al. Research on the Alleviation of Osteoporosis by Oyster Shell Powder Based on High-throughput Sequencing Technology[J]. Science and Technology of Food Industry, 2021, 42(18): 372−379. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2020120189
Citation: SHI Qiuyue, HOU Fujing, HAN Jiaojiao, et al. Research on the Alleviation of Osteoporosis by Oyster Shell Powder Based on High-throughput Sequencing Technology[J]. Science and Technology of Food Industry, 2021, 42(18): 372−379. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2020120189

基于高通量测序技术的牡蛎壳粉缓解骨质疏松症的研究

doi: 10.13386/j.issn1002-0306.2020120189
基金项目: 国家海洋经济创新发展区域示范项目(2013710)
详细信息
    作者简介:

    石秋月(1995−),女,硕士研究生,研究方向:食品加工与安全,E-mail:2685373194@qq.com

    通讯作者:

    苏秀榕(1956−),女,博士,教授,研究方向:食品科学与工程,E-mail:suxiurong@nbu.edu.cn

  • 中图分类号: Q93

Research on the Alleviation of Osteoporosis by Oyster Shell Powder Based on High-throughput Sequencing Technology

  • 摘要: 目的:探究牡蛎壳粉对糖皮质激素性骨质疏松小鼠的缓解作用。方法:将ICR雌性小鼠随机分为对照组、模型组、碳酸钙组和牡蛎壳粉组,肌肉注射地塞米松(1 mg/kg/d)构建骨质疏松小鼠模型,分析股骨微结构、血清钙、粪便钙、股骨钙、骨转换指标和肠道菌群变化。结果:牡蛎壳粉能显著提高骨密度和骨小梁厚度(P<0.05)并降低骨小梁分离度和结构模型指数,改善骨微结构,显著降低血清钙含量(P<0.001)。牡蛎壳粉也能显著升高血清中碱性磷酸酶酶活(P<0.001),降低抗酒石酸酸性磷酸酶酶活(P<0.01);显著升高骨形成标记物碱性磷酸酶和骨保护素的基因转录(P<0.05),降低骨吸收标记物抗酒石酸酸性磷酸酶和硬骨素的基因转录(P<0.05)。牡蛎壳粉还能改变小鼠肠道菌群结构,增加约氏乳杆菌、产粪甾醇真细菌等的丰度。嗜酸乳酸杆菌、豚鼠乳杆菌、罗伊氏乳杆菌、产酸拟杆菌与骨形成指标显著负相关,而与骨吸收指标显著正相关。结论:牡蛎壳粉能够抑制骨矿物质流失,提高骨密度和骨小梁厚度,促进骨形成指标而抑制骨吸收指标,改善肠道菌群,从而有效缓解骨质疏松。
  • 图  1  牡蛎壳粉的粒径分布图

    Figure  1.  Particle size distribution of OSP

    图  2  小鼠的骨股micro CT图像

    Figure  2.  Micro CT images of the femur in mice

    图  3  小鼠骨组织形态计量学指标

    Figure  3.  Bone tissue morphology index in mice

    注:模型组与对照组相比,#:P<0.05;两个处理组与模型组相比,*:P<0.05;图6同。

    图  4  小鼠的骨转换标记物的酶活

    Figure  4.  Enzyme activity of bone turnover markers in mice

    注:模型组与对照组相比,#:P<0.05,##:P<0.01;两个处理组与模型组相比,**:P<0.01,***:P<0.001。

    图  5  小鼠的骨转换标记物相关基因表达

    Figure  5.  Gene expression related to bone turnover markers in mice

    注:模型组与对照组相比,#:P<0.05,##:P<0.01,###:P<0.001;两个处理组与模型组相比,*:P<0.05,**:P<0.01。

    图  6  小鼠肠道菌群Chao1指数及Shannon指数

    Figure  6.  Chao1 index and Shannon index of gut microbiota in mice

    图  7  各组小鼠的PCoA图

    Figure  7.  PCoA of each group of mice

    图  8  小鼠肠道菌群门和种的变化趋势

    Figure  8.  Changes in the phyla and species of mice gut microbiota

    图  9  每组小鼠特有的OTUs

    Figure  9.  Specific OTUs to each group of mice

    图  10  小鼠肠道菌群优势菌种与骨转换标记物的相关性分析

    Figure  10.  Correlation analysis of dominant species of gut microbiota and bone turnover markers in mice

    表  1  基因引物序列(5'→3')

    Table  1.   Target genes primer sequence (5'→3')

    基因名称引物序列
    管家基因(β-ActinF- TGTGATGGTGGGAATGGG
    R- TGCCAGATCTTCTCCATGTC
    碱性磷酸酶(ALP)F- TAACACCAACGCTCAGGTCC
    R- TGGATGTGACCTCATTGCCC
    抗酒石酸酸性磷酸酶(TRACP)F- TTCAGGACGAGAACGGTGTG
    R- CCAGGGAGTCCTCAGATCCA
    骨保护素(OPG)F- ACGGAGACACAGCTCACAAG
    R- CAGGCTCTCCATCAAGGCAA
    硬骨素(sclerostin)F- GGTAGTGAACAGACTCCGGC
    R- GGCGGTCTTCAAGCCATACT
    下载: 导出CSV

    表  2  牡蛎壳粉的矿物质元素组成

    Table  2.   Mineral composition of OSP

    矿物质元素牡蛎壳粉(g/kg)
    Ca36.90±0.31
    P0.67±0.03
    Sr0.23±0.03
    Na0.644±0.053
    Mg0.988±0.091
    Zn0.008±0.001
    Fe0.402±0.036
    Pb0.00097±0.000042
    下载: 导出CSV

    表  3  小鼠各部分的钙量变化

    Table  3.   Changes in the amount of calcium in each part of mice

    组别血清钙(mmol/L)粪便钙(mg/g)股骨钙(mg/g)
    对照组1.01±0.0925.97±2.83135.01±17.09
    模型组1.22±0.10###30.30±3.64#116.26±18.24
    碳酸钙组0.97±0.05***27.48±2.06127.86±17.66
    牡蛎壳粉组0.92±0.07***29.14±1.97129.06±17.81
    注:模型组与对照组相比,#:P<0.05,###:P<0.001;两个处理组与模型组相比,***:P<0.001。
    下载: 导出CSV
  • [1] Buckley L, Humphrey M B. Glucocorticoid-induced osteoporosis[J]. The New England Journal of Medicine,2018,379(26):2547−2556. doi:  10.1056/NEJMcp1800214
    [2] Frenkel B, White W, Tuckermann J. Glucocorticoid-induced osteoporosis[J]. Adv Exp Med Biol,2015,872:179−215.
    [3] 耿彬, 夏亚一. 调控骨质疏松模型小鼠的ERK5蛋白信号通路[J]. 中国组织工程研究,2021,25(2):178−185. [Geng B, Xia Y Y. Involvement of ERK5 signaling pathway in osteoporosis development in mice[J]. Journal of Clinical Rehabilitative Tissue Engineering Research,2021,25(2):178−185. doi:  10.3969/j.issn.2095-4344.2955
    [4] Yanbeiy Z A, Hansen K E. Denosumab in the treatment of glucocorticoid-induced osteoporosis: A systematic review and meta-analysis[J]. Drug Des Devel Ther,2019,13:2843−2852. doi:  10.2147/DDDT.S148654
    [5] 黄东. 糖皮质激素诱导骨质疏松症防治研究进展[J]. 临床合理用药杂志,2020,13(11):180−181. [Huang D. Research progress in prevention and treatment of glucocorticoid-induced osteoporosis[J]. Chinese Journal of Clinical Rational Drug Use,2020,13(11):180−181.
    [6] 张家国, 徐晓峰, 李明, 等. 银杏叶提取物对糖皮质激素诱导的骨质疏松症大鼠骨密度和骨生物力学的影响[J]. 中国骨质疏松杂志,2019,25(4):461−464, 471. [Zhang J G, Xu X F, Li M, et al. Effects of Ginkgo biloba extract on bone mineral density and bone biomechanics in rats with glucocorticoid-induced osteoporosis[J]. Chinese Journal of Osteoporosis,2019,25(4):461−464, 471. doi:  10.3969/j.issn.1006-7108.2019.04.007
    [7] 崔宗梅, 黄津伟, 王海艳, 等. 广东雷州半岛东部沿岸潮间带常见牡蛎的种类及其分布[J]. 海洋与湖沼,2018,49(6):1350−1357. [Cui Z M, Huang J W, Wang H Y, et al. Classification and distribution of oysters in eastern coast of Leizhou peninsula, China[J]. Oceanologia Et Limnologia Sinica,2018,49(6):1350−1357. doi:  10.11693/hyhz20180300054
    [8] 杨韵, 徐波. 牡蛎的化学成分及其生物活性研究进展[J]. 中国现代中药,2015,17(12):1345−1349. [Yang Y, Xu B. Research progress on chemical composition of oyster and its biological activity[J]. Modern Chinese Medicine,2015,17(12):1345−1349.
    [9] 蒋金来, 王令充, 吴皓, 等. 钙制剂研究进展[J]. 食品工业科技,2012,33(11):379−382, 387. [Jiang J L, Wang L C, Wu H, et al. Research progress in calcium preparation[J]. Science and Technology of Food Industry,2012,33(11):379−382, 387.
    [10] 李佳, 韩丽娜, 韦玮, 等. 牡蛎在补钙方面的研究进展[J]. 内蒙古中医药,2019,38(3):90−91. [Li J, Han L N, Wei W, et al. Research progress of oyster in calcium supplement[J]. Nei Mongol Journal of Traditional Chinese Medicine,2019,38(3):90−91.
    [11] Chen Y, Jiang Y, Liao L, et al. Inhibition of 4NQO-Induced oral carcinogenesis by dietary oyster shell calcium[J]. Integrative Cancer Therapies,2016,15(1):96−101. doi:  10.1177/1534735415596572
    [12] Wang Z, Wang K Y, Feng Y N, et al. Preparation, characterization of L-aspartic acid chelated calcium from oyster shell source and its calcium supplementation effect in rats[J]. Journal of Functional Foods,2020:75.
    [13] Lerner A, Neidhöfer S, Matthias T. The gut microbiome feelings of the brain: A perspective for non-microbiologists[J]. Microorganisms,2017,5(4):66. doi:  10.3390/microorganisms5040066
    [14] Li J, Yang M, Lu C, et al. Tuna bone powder alleviates glucocorticoid-induced osteoporosis via coregulation of the NF-κB and Wnt/β-catenin signaling pathways and modulation of gut microbiota composition and metabolism[J]. Molecular Nutrition & Food Research,2020,64(5):e1900861.
    [15] Han J, Huang Z, Tang S, et al. The novel peptides ICRD and LCGEC screened from tuna roe show antioxidative activity via Keap1/Nrf2-ARE pathway regulation and gut microbiota modulation[J]. Food Chemistry,2020,327:127094. doi:  10.1016/j.foodchem.2020.127094
    [16] Jiang Q, Lu C, Sun T, et al. Alterations of the brain proteome and gut microbiota in d-galactose-induced brain-aging mice with krill oil supplementation[J]. Journal of Agricultural and Food Chemistry,2019,67(35):9820−9830. doi:  10.1021/acs.jafc.9b03827
    [17] Han J J, Wang X, Tang S, et al. Protective effects of tuna meat oligopeptides(TMOP) supplementation on hyperuricemia and associated renal inflammation mediated by gut microbiota[J]. Faseb Journal,2020,34(4):5061−5076. doi:  10.1096/fj.201902597RR
    [18] Jiang X T, Peng X, Deng G H, et al. Illumina sequencing of 16S rRNA tag revealed spatial variations of bacterial communities in a mangrove wetland[J]. Microbial Ecology,2013,66(1):96−104. doi:  10.1007/s00248-013-0238-8
    [19] 鲍伟, 韩姣姣, 张旨轩, 等. 基于高通量测序技术对浙江传统发酵蔬菜微生物多样性的解析[J]. 食品科学,2021,42(6):178−185. [Bao W, Han J J, Zhang Z X, et al. Analysis of microbial community diversity in Zhejiang traditional fermented vegetables using high-throughput sequencing[J]. Food Science,2021,42(6):178−185.
    [20] Ibrahim N', Khamis M F, Mod Yunoh M F, et al. Targeted delivery of lovastatin and tocotrienol to fracture site promotes fracture healing in osteoporosis model: Micro-computed tomography and biomechanical evaluation[J]. PLoS One,2014,9(12):e115595. doi:  10.1371/journal.pone.0115595
    [21] Chen C, Jin D, Liu Y, et al. Trabecular bone characterization on the continuum of plates and rods using in vivo MR imaging and volumetric topological analysis[J]. Physics in Medicine and Biology,2016,61(18):N478−N496. doi:  10.1088/0031-9155/61/18/N478
    [22] 尚芬兰, 徐晶晶, 赫荣波, 等. 骨转换标志物在糖尿病中的研究进展[J]. 中国骨质疏松杂志,2017,23(4):541−547. [Shang F L, Xu J J, He R B, et al. Research progress on the application of bone turnover markers in diabetes[J]. Chinese Journal of Osteoporosis,2017,23(4):541−547. doi:  10.3969/j.issn.1006-7108.2017.04.026
    [23] 耿娟娟, 秦福芳, 张文才. NTX与2型糖尿病性骨质疏松患者血清中骨转换标志物的相关性研究[J]. 中国合理用药探索,2020,17(6):56−60. [Geng J J, Qin F F, Zhang W C. Study on the correlation between NTX and serum bone turnover markers in patients with type 2 diabetic osteoporosis[J]. Chinese Journal of Rational Drug Use,2020,17(6):56−60.
    [24] 黄长安, 喻景弈. 血清癌胚抗原相关细胞黏附分子1、骨标志物硬化蛋白、Ⅰ型前胶原氨基端延长肽及护骨素变化与绝经后骨质疏松病人骨密度变化的相关性[J]. 安徽医药,2020,24(9):1828−1831. [Huang C A, Yu J Y. Correlation between changes of serum CEACAM1, SOST, P1NP and osteoprotegerin levels and changes of bone mineral density in PMOP patients with osteoporosis[J]. Anhui Medical and Pharmaceutical Journal,2020,24(9):1828−1831. doi:  10.3969/j.issn.1009-6469.2020.09.034
    [25] Chung H, Pamp S J, Hill J A, et al. Gut immune maturation depends on colonization with a host-specific microbiota[J]. Cell,2012,149(7):1578−1593. doi:  10.1016/j.cell.2012.04.037
    [26] Yan J, Charles J F. Gut microbiome and bone: To Build, destroy, or both?[J]. Current Atherosclerosis Reports,2017,15(4):376−384.
    [27] Ohlsson C, Sjögren K. Effects of the gut microbiota on bone mass[J]. Trends in Endocrinology and Metabolism: TEM,2015,26(2):69−74. doi:  10.1016/j.tem.2014.11.004
    [28] Yan J, Herzog J W, Tsang K, et al. Gut microbiota induce IGF-1 and promote bone formation and growth[J]. Proc Natl Acad Sci U S A,2016,113(47):E7554−E7563. doi:  10.1073/pnas.1607235113
    [29] Ejtahed H S, Soroush A R, Angoorani P, et al. Gut microbiota as a target in the pathogenesis of metabolic disorders: A new approach to novel therapeutic agents[J]. Hormone and Metabolic Research,2016,48(6):349−358. doi:  10.1055/s-0042-107792
    [30] Wu T, Yang L, Jiang J, et al. Chronic glucocorticoid treatment induced circadian clock disorder leads to lipid metabolism and gut microbiota alterations in rats[J]. Life Sciences,2018,192:173−182. doi:  10.1016/j.lfs.2017.11.049
    [31] 孙梦晗, 邓敏, 白杨. 罗伊氏乳杆菌益生特性及治疗溃疡性结肠炎研究进展[J]. 现代消化及介入诊疗,2019,24(12):1493−1496. [Sun M H, Deng M, Bai Y. Research progress on the probiotic properties of Lactobacillus reuteri and the treatment of ulcerative colitis[J]. Modern Digestion & Intervention,2019,24(12):1493−1496. doi:  10.3969/j.issn.1672-2159.2019.12.035
    [32] 丁佳, 王慧艳, 何秋雯, 等. 益生菌对雌激素代谢及相关疾病的影响[J]. 食品科学,2016,37(11):248−254. [Ding J, Wang H Y, He Q W, et al. Effects of probiotics on estrogen metabolism and related disorders[J]. Food Science,2016,37(11):248−254. doi:  10.7506/spkx1002-6630-201611044
    [33] Thaiss C A, Zmora N, Levy M, et al. The microbiome and innate immunity[J]. Nature,2016,535(7610):65−74. doi:  10.1038/nature18847
    [34] Lucas S, Omata Y, Hofmann J, et al. Short-chain fatty acids regulate systemic bone mass and protect from pathological bone loss[J]. Nat Commun,2018,9(1):55. doi:  10.1038/s41467-017-02490-4
  • 加载中
图(10) / 表(3)
计量
  • 文章访问数:  45
  • HTML全文浏览量:  11
  • PDF下载量:  7
  • 被引次数: 0
出版历程
  • 收稿日期:  2020-12-23
  • 网络出版日期:  2021-08-05
  • 刊出日期:  2021-09-14

目录

    /

    返回文章
    返回

    重要通知

    《食品工业科技》编辑部携手万方数据开通学术不端专属检测通道,具体信息参见本刊动态。