一种GLP-1过表达肠类器官构建的方法

doi:10.16288/j.yczz.20-423

遗传 ›› 2021, Vol. 43 ›› Issue (7): 694-703.doi: 10.16288/j.yczz.20-423

一种GLP-1过表达肠类器官构建的方法

曾之扬¹(), 陆佳微¹, 曹希雅¹, 王芯悦^2,³, 李大力¹()

1. 华东师范大学生命科学学院,上海市调控生物学重点实验室,上海 200241
2. 安徽理工大学医学院,淮南 232000
3. 上海交通大学附属第六人民医院南院消化内科,上海 201499

收稿日期:2021-03-01 修回日期:2021-04-14 出版日期:2021-07-20 发布日期:2021-07-20
通讯作者: 李大力 E-mail:52161300029@stu.ecnu.edu.cn;dlli@bio.ecnu.edu.cn
作者简介:曾之扬,博士,博后,研究方向：发育生物学。E-mail: 52161300029@stu.ecnu.edu.cn
基金资助:
国家重点研发计划编号(2019YFA0110802)

A method for constructing GLP-1 overexpression intestinal organoids

Zhiyang Zeng¹(), Jiawei Lu¹, Xiya Cao¹, Xinyue Wang^2,³, Dali Li¹()

1. Shanghai Key Laboratory of Regulatory Biology, School of Life Sciences, East China Normal University, Shanghai 200241, China
2. School of Medicine, Anhui University of Science and Technology, Huainan 232000, China
3. Department of Gastroenterology, South Campus of Sixth People's Hospital, Shanghai Jiaotong University, Shanghai 201499, China

Received:2021-03-01 Revised:2021-04-14 Online:2021-07-20 Published:2021-07-20
Contact: Li Dali E-mail:52161300029@stu.ecnu.edu.cn;dlli@bio.ecnu.edu.cn
Supported by:
Supported by the National Key R&D Program of China No(2019YFA0110802)

摘要/Abstract

摘要：

胰高血糖素样肽1 (glucagon-like peptide 1, GLP-1)作为一种肠促胰岛素,主要由肠道L细胞分泌,由于其能够有效促进胰岛素的释放从而降低血糖,因此GLP-1及其类似物在2型糖尿病的治疗上具有良好的应用前景。本研究优化了慢病毒感染类器官的方法,利用该方法成功构建了GLP-1过表达的小鼠小肠类器官(organoids)。结果显示该类器官分泌的GLP-1能够有效地提高野生型及糖尿病小鼠的葡萄糖耐受能力。因此,本研究构建的GLP-1过表达类器官可以为2型糖尿病的治疗提供一种新的策略。

关键词: 胰高血糖素样肽1, 2型糖尿病, 小肠类器官

Abstract:

As a potent insulinotrophic hormone, glucagon-like peptide 1 (GLP-1) is mainly secreted by intestinal L cells, which can effectively promote the release of insulin and thus reduce blood glucose. Therefore, GLP-1 and its analogs have a good prospect in the treatment of type 2 diabetes. In this study, we constructed mouse intestinal organoids that overexpress GLP-1 by optimizing the GLP-1 lentivirus infection method. We found that supernatants secreted by the GLP-1 overexpression organoids effectively enhanced glucose tolerance in wild-type and diabetic mouse. Thus, the GLP-1 overexpression organoids built in this study may provide a novel strategy for the treatment of type 2 diabetes.

Key words: GLP-1, type 2 diabetes, intestinal organoids

曾之扬, 陆佳微, 曹希雅, 王芯悦, 李大力. 一种GLP-1过表达肠类器官构建的方法[J]. 遗传, 2021, 43(7): 694-703.

Zhiyang Zeng, Jiawei Lu, Xiya Cao, Xinyue Wang, Dali Li. A method for constructing GLP-1 overexpression intestinal organoids[J]. Hereditas(Beijing), 2021, 43(7): 694-703.

图/表 5

图1

图2

图3

图4

图5

参考文献 32

[1]	Kieffer TJ, Habener JF. The glucagon-like peptides. Endocr Rev, 1999, 20(6):876-913. pmid: 10605628
[2]	Nauck MA, Homberger E, Siegel EG, Allen RC, Eaton RP, Ebert R, Creutzfeldt W. Incretin effects of increasing glucose loads in man calculated from venous insulin and C-peptide responses. J Clin Endocrinol Metab, 1986, 63(2):492-498. doi: 10.1210/jcem-63-2-492
[3]	Perley MJ, Kipnis DM. Plasma insulin responses to oral and intravenous glucose: studies in normal and diabetic sujbjects. J Clin Invest, 1967, 46(12):1954-1962. pmid: 6074000
[4]	Meier JJ, Nauck MA, Kranz D, Holst JJ, Deacon CF, Gaeckler D, Schmidt WE, Gallwitz B. Secretion, degradation, and elimination of glucagon-like peptide 1 and gastric inhibitory polypeptide in patients with chronic renal insufficiency and healthy control subjects. Diabetes, 2004, 53(3):654-662. doi: 10.2337/diabetes.53.3.654
[5]	Hui HX, Farilla L, Merkel P, Perfetti R. The short half-life of glucagon-like peptide-1 in plasma does not reflect its long-lasting beneficial effects. Eur J Endocrinol, 2002, 146(6):863-869.
[6]	Kumar M, Hunag Y, Glinka Y, Prud'homme GJ, Wang Q. Gene therapy of diabetes using a novel GLP-1/IgG1-Fc fusion construct normalizes glucose levels in db/db mice. Gene Ther, 2007, 14(2):162-172. pmid: 16943856
[7]	Soltani N, Kumar M, Glinka Y, Prud'homme GJ, Wang Q,. In vivo expression of GLP-1/IgG-Fc fusion protein enhances beta-cell mass and protects against streptozotocin-induced diabetes. Gene Ther, 2007, 14(12):981-988. pmid: 17410180
[8]	Shao JW, Xue S, Yu GL, Yu YH, Yang XP, Bai Y, Zhu SC, Yang LF, Yin JL, Wang YD, Liao SY, Guo SW, Xie MQ, Fussenegger M, Ye HF. Smartphone-controlled optogenetically engineered cells enable semiautomatic glucose homeostasis in diabetic mice. Sci Transl Med , 2017, 9(387): eaal2298. doi: 10.1126/scitranslmed.aal2298
[9]	Ye HF, Daoud-El Baba M, Peng RW, Fussenegger M. A synthetic optogenetic transcription device enhances blood- glucose homeostasis in mice. Science, 2011, 332(6037):1565-1568. doi: 10.1126/science.1203535
[10]	Clevers H. Modeling development and disease with organoids. Cell, 2016, 165(7):1586-1597. doi: S0092-8674(16)30729-2 pmid: 27315476
[11]	Sato T, Stange DE, Ferrante M, Vries RG, Van Es JH, Van den Brink S, Van Houdt WJ, Pronk A, Van Gorp J, Siersema PD, Clevers H. Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett's epithelium. Gastroenterology, 2011, 141(5):1762-1772. doi: 10.1053/j.gastro.2011.07.050
[12]	Sato T, Vries RG, Snippert HJ, van de Wetering M, Barker N, Stange DE, van Es JH, Abo A, Kujala P, Peters PJ, Clevers H. Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche. Nature , 2009, 459(7244):262-265. doi: 10.1038/nature07935
[13]	Petersen N, Reimann F, Bartfeld S, Farin HF, Ringnalda FC, Vries RGJ, van den Brink S, Clevers H, Gribble FM, de Koning EJP. Generation of L cells in mouse and human small intestine organoids. Diabetes, 2014, 63(2):410-420. doi: 10.2337/db13-0991 pmid: 24130334
[14]	Goldspink DA, Lu VB, Miedzybrodzka EL, Smith CA, Foreman RE, Billing LJ, Kay RG, Reimann F, Gribble FM. Labeling and characterization of human GLP-1-secreting L-cells in primary ileal organoid culture. Cell Rep, 2020, 31(13):107833. doi: S2211-1247(20)30814-7 pmid: 32610134
[15]	Lund ML, Sorrentino G, Egerod KL, Kroone C, Mortensen B, Knop FK, Reimann F, Gribble FM, Drucker DJ, de Koning EJP, Schoonjans K, Bäckhed F, Schwartz TW, Petersen N. L-Cell differentiation is induced by bile acids through GPBAR1 and paracrine GLP-1 and serotonin signaling. Diabetes, 2020, 69(4):614-623. doi: 10.2337/db19-0764
[16]	Goldspink DA, Lu VB, Billing LJ, Larraufie P, Tolhurst G, Gribble FM, Reimann F. Mechanistic insights into the detection of free fatty and bile acids by ileal glucagon-like peptide-1 secreting cells. Mol Metab, 2018, 7:90-101. doi: S2212-8778(17)30750-0 pmid: 29167062
[17]	Yui S, Nakamura T, Sato T, Nemoto Y, Mizutani T, Zheng X, Ichinose S, Nagaishi T, Okamoto R, Tsuchiya K, Clevers H, Watanabe M. Functional engraftment of colon epithelium expanded in vitro from a single adult Lgr5 + stem cell. Nat Med, 2012, 18(4):618-623. doi: 10.1038/nm.2695
[18]	Fukuda M, Mizutani T, Mochizuki W, Matsumoto T, Nozaki K, Sakamaki Y, Ichinose S, Okada Y, Tanaka T, Watanabe M, Nakamura T. Small intestinal stem cell identity is maintained with functional Paneth cells in heterotopically grafted epithelium onto the colon. Genes Dev, 2014, 28(16):1752-1757. doi: 10.1101/gad.245233.114
[19]	O'Rourke KP, Loizou E, Livshits G, Schatoff EM, Baslan T, Manchado E, Simon J, Romesser PB, Leach B, Han T, Pauli C, Beltran H, Rubin MA, Dow LE, Lowe SW. Transplantation of engineered organoids enables rapid generation of metastatic mouse models of colorectal cancer. Nat Biotechnol, 2017, 35(6):577-582. doi: 10.1038/nbt.3837 pmid: 28459450
[20]	Koo BK, Stange DE, Sato T, Karthaus W, Farin HF, Huch M, van Es JH, Clevers H. Controlled gene expression in primary Lgr5 organoid cultures. Nat Methods, 2011, 9(1):81-83.
[21]	Andersson-Rolf A, Fink J, Mustata RC, Koo BK. A video protocol of retroviral infection in primary intestinal organoid culture. J Vis Exp, 2014, (90):e51765.
[22]	Jang HJ, Kokrashvili Z, Theodorakis MJ, Carlson OD, Kim BJ, Zhou J, Kim HH, Xu XR, Chan SL, Juhaszova M, Bernier M, Mosinger B, Margolskee RF, Egan JM. Gut-expressed gustducin and taste receptors regulate secretion of glucagon-like peptide-1. Proc Natl Acad Sci USA, 2007, 104(38):15069-15074. doi: 10.1073/pnas.0706890104
[23]	Gorboulev V, Schürmann A, Vallon V, Kipp H, Jaschke A, Klessen D, Friedrich A, Scherneck S, Rieg T, Cunard R, Veyhl-Wichmann M, Srinivasan A, Balen D, Breljak D, Rexhepaj R, Parker HE, Gribble FM, Reimann F, Lang F, Wiese S, Sabolic I, Sendtner M, Koepsell H. Na(+)-D- glucose cotransporter SGLT1 is pivotal for intestinal glucose absorption and glucose-dependent incretin secretion. Diabetes, 2012, 61(1):187-196. doi: 10.2337/db11-1029 pmid: 22124465
[24]	Reimann F, Gribble FM. Glucose-sensing in glucagon-like peptide-1-secreting cells. Diabetes, 2002, 51(9):2757-2763. pmid: 12196469
[25]	Parker HE, Adriaenssens A, Rogers G, Richards P, Koepsell H, Reimann F, Gribble FM. Predominant role of active versus facilitative glucose transport for glucagon-like peptide-1 secretion. Diabetologia, 2012, 55(9):2445-2455. doi: 10.1007/s00125-012-2585-2 pmid: 22638549
[26]	Sonne DP, Rehfeld JF, Holst JJ, Vilsboll T, Knop FK. Postprandial gallbladder emptying in patients with type 2 diabetes: potential implications for bile-induced secretion of glucagon-like peptide 1. Eur J Endocrinol, 2014, 171(4):407-419. doi: 10.1530/EJE-14-0309
[27]	Baggio LL, Drucker DJ. Biology of incretins: GLP-1 and GIP. Gastroenterology, 2007, 132(6):2131-2157. doi: 10.1053/j.gastro.2007.03.054
[28]	Holman RR, Bethel MA, Mentz RJ, Thompson VP, Lokhnygina Y, Buse JB, Chan JC, Choi J, Gustavson SM, Iqbal N, Maggioni AP, Marso SP, Öhman P, Pagidipati NJ, Poulter N, Ramachandran A, Zinman B, Hernandez AF, Group ES. Effects of once-weekly exenatide on cardiovascular outcomes in type 2 diabetes. N Engl J Med, 2017, 377(13):1228-1239. doi: 10.1056/NEJMoa1612917
[29]	Wadden TA, Hollander P, Klein S, Niswender K, Woo V, Hale PM, Aronne L, Investigators NN. weight maintenance and additional weight loss with liraglutide after low-calorie- diet-induced weight loss: the scale maintenance randomized study. Int J Obes (Lond), 2013, 37(11):1443-1451. doi: 10.1038/ijo.2013.120
[30]	Nauck MA, Niedereichholz U, Ettler R, Holst JJ, Orskov C, Ritzel R, Schmiegel WH. Glucagon-like peptide 1 inhibition of gastric emptying outweighs its insulinotropic effects in healthy humans. Am J Physiol, 1997, 273(5):E981-E988.
[31]	Müller TD, Finan B, Bloom SR, D'Alessio D, Drucker DJ, Flatt PR, Fritsche A, Gribble F, Grill HJ, Habener JF, Holst JJ, Langhans W, Meier JJ, Nauck MA, Perez-Tilve D, Pocai A, Reimann F, Sandoval DA, Schwartz TW, Seeley RJ, Stemmer K, Tang-Christensen M, Woods SC, DiMarchi RD, Tschöp MH. Glucagon-like peptide 1 (GLP-1). Mol Metab, 2019, 30:72-130. doi: S2212-8778(19)30913-5 pmid: 31767182
[32]	Parsons GB, Souza DW, Wu H, Yu D, Wadsworth SG, Gregory RJ, Armentano D. Ectopic expression of glucagon- like peptide 1 for gene therapy of type II diabetes. Gene Ther, 2007, 14(1):38-48. pmid: 16929351

编辑推荐

Metrics

www.chinagene.cn
备案号：京ICP备09063187号-4
总访问:,今日访问:,当前在线:

一种GLP-1过表达肠类器官构建的方法

A method for constructing GLP-1 overexpression intestinal organoids

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

图/表 5

参考文献 32

相关文章 15

编辑推荐

Metrics

[1]	吕承安, 王若然, 孟卓贤. 2型糖尿病进程中胰岛β细胞功能变化的分子机制[J]. 遗传, 2022, 44(10): 840-852.
[2]	曹岚, 李志强, 师咏勇, 刘赟. 端粒长度与2型糖尿病：孟德尔随机化研究与多基因风险评分分析[J]. 遗传, 2020, 42(9): 882-888.
[3]	王玉琢, 张一鸣, 董晓莲, 王学才, 朱建福, 王娜, 江峰, 陈跃, 姜庆五, 付朝伟. 2型糖尿病易感基因SNP位点对生活方式干预降低血糖应答效果的修饰效应[J]. 遗传, 2020, 42(5): 483-492.
[4]	弓弦,张超,伊利亚斯·艾萨,时瑛,杨雪唯,努尔斯曼古丽奥斯曼,关亚群,徐书华. 2型糖尿病易感候选基因在世界不同人群中的多样性比较分析[J]. 遗传, 2016, 38(6): 543-559.
[5]	张君, 张望强, 丁毓磊, 许彭, 王婷婷, 徐文静, 陆环, 刘宗智, 谢建新. 腹部脂肪组织APN基因DNA甲基化及mRNA表达与维吾尔族T2DM的相关性[J]. 遗传, 2015, 37(3): 269-275.
[6]	吴日,马超,李晓丹,段会坤,姬艳丽,王宇,姜苹哲,王海松,屠培培,李淼,尼钢钢,马百成,李明刚. 长效促胰岛素降糖酵母的构建及其对糖尿病模型小鼠的治疗效果[J]. 遗传, 2015, 37(2): 183-191.
[7]	汤琳琳刘琼步世忠徐雷艇王钦文麦一峰段世伟. 2型糖尿病环境因素与DNA甲基化的研究进展[J]. 遗传, 2013, 35(10): 1143-1152.
[8]	蒲连美，南楠，杨泽，金泽宁. SUMO4基因多态性与2型糖尿病的关系[J]. 遗传, 2012, 34(3): 315-325.
[9]	汤晓丽，邓立彬，李桂林，刘双梅，林加日，谢金燕，刘俊，孔繁君，梁尚栋. 2型糖尿病早期大鼠外周神经节基因表达谱分析[J]. 遗传, 2012, 34(2): 198-207.
[10]	王敏，彭婵，屈亚莉，黄青阳. 中国汉族人群ENPP1基因K121Q多态与2型糖尿病的关联及Meta分析[J]. 遗传, 2010, 32(8): 808-816.
[11]	陈芳建，俞红，樊璠，吕建新. 线粒体基因组D-Loop区基因多态性与2型糖尿病的相关性[J]. 遗传, 2009, 31(3): 265-272.
[12]	苏燕，彭姝彬，李智琼，黄青阳. PPARGC1A基因Thr394Thr/Gly482Ser多态性与2型糖尿病的关联研究[J]. 遗传, 2008, 30(3): 304-308.
[13]	杜纪坤，黄青阳，李守华，熊国梅. 脂蛋白脂酶基因HindⅢ酶切多态性与2型糖尿病的关联研究[J]. 遗传, 2007, 29(8): 929-929―933.
[14]	程祖建，杨滨，刘奇才，江凌，谢海花，欧启水. 一个2型糖尿病家系中新发现的线粒体DNA G7444A 突变分析[J]. 遗传, 2007, 29(4): 433-437.
[15]	杜纪坤，黄青阳. 脂蛋白脂酶基因的研究进展[J]. 遗传, 2007, 29(1): 8-16.