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Journal of Chinese Integrative Medicine ›› 2007, Vol. 5 ›› Issue (6): 634-641.doi: 10.3736/jcim20070607

Special Issue: Traditional Chinese Medicine

• Original Experimental Research • Previous Articles     Next Articles

Establishment of Caco-2 cell monolayer model and standard operation procedure for assessing intestinal absorption of chemical components of traditional Chinese medicine

Xiu-wei Yang, Xiao-da Yang, Ying Wang, Lian Ma, Yue Zhang, Xiao-gai Yang, Kui Wang   

  1. State Key Laboratory of Natural and Biomimetic Drugs,School of Pharmaceutical Sciences,Peking University,Beijing 100083,China
  • Online:2007-11-20 Published:2007-11-15

Objective: To establish Caco-2 (a human colon adenocarcinoma cell line) cell monolayer model and the standard operation procedure for studying and assessing intestinal absorption of chemical components of traditional Chinese medicine.Methods: Caco-2 cell monolayer model was established and evaluated by morphology feature using scanning electron microscope, inverted microscope and transepithelial electrical resistance (TEER) assay. Additionally, the model was further tested for the activity of alkaline phosphatase and the apparent permeability (Papp) of standard compounds, i.e. propranolol and atenolol, which were the control substances for high and poor transcellular transport marker, respectively.Results: The integrality of cell monolayer, cell differentiation (reflected by expression of alkaline phosphatase and cell monolayer morphology), and the Papp value of standard compounds in the established Caco-2 cell model were satisfactory. All parameters tested were in good agreement with those reported in the literature.Conclusion: The established Caco-2 cell model can be used to study the intestinal absorption of orally administrated chemical components of traditional Chinese medicine and their absorption mechanism.

Key words: Caco-2 cells, Chinese herbal drugs, intestinal absorption, biological transport

CLC Number: 

  • R348

Figure 1

Diagram of Transwell and transport experiment"

Figure 2

Apical side of Caco-2 monolayer (scanning electron micrograph)"

Figure 3

Characteristic microvilli of apical side of the differentiated Caco-2 monolayer (transmission electron micrograph)"

Figure 4

Tight junction-desmosome of Caco-2 monolayer (transmission electron micrograph)"

[1] 杨秀伟 . 基于体内过程的中药有效成分和有效效应物质的发现策略[J]. 中国中药杂志, 2007,32(5):365-370
Yang XW . Discovery strategy for effective and active constituents of Chinese material medica based on the processes of metabolism and disposition in intra-body[J]. Zhongguo Zhong Yi Yao Za Zhi, 2007,32(5):365-370
[2] 杨秀伟 . 基于体内过程的中药毒性成分和毒性效应物质的发现策略[J]. 中华中医药杂志, 2007,22(2):67-72
Yang XW . Discovery strategy for toxic-invited constituents and toxic-effective substances on Chinese material medica based on the processes of metabolism and disposition in intra-body[J]. Zhong Hua Zhong Yi Yao Za Zhi, 2007,22(2):67-72
[3] 杨秀伟 . 中药实验医学研究中的关键基础科学问题[J]. 中西医结合学报, 2005,3(2):154-159
Yang XW . Key foundational science problem in experimental medicine study of Chinese materia medica: ascertainment of active and toxic constituents in Chinese materia medica[J]. Zhong Xi Yi Jie He Za Zhi, 2005,3(2):154-159
[4] 杨秀伟 . 中药成分的肠内和肝内行为与中药有效成分的确定[J]. 中草药, 2004,35(增刊):1-5
Yang XW . Relationship between confirmation of Chinese materia medica bioactive constituents and intra-intestinal and intra-hepatic behavior of Chinese materia medica active constituents[J]. Zhong Cao Yao, 2004,35(Suppl):1-5
[5] 杨秀伟, 郝美荣, 服部征雄 . 中药成分代谢分析[M]. 北京: 中国医药科技出版社,2003.
Yang XW, Hao MR Hattori M, , . Metabolite analysis for chemical constituents of traditional Chinese medicines[M]. Beijing: China Medical-Pharmaceutical Science & Technology Publishion House, 2003.
[6] 周昕 . 药物吸收体外研究方法的概述[J]. 中国临床药理学与治疗学, 2006,11(4):379-384
Zhou X . Study on in vitro models of drug absorption[J]. Zhongguo Lin Chuang Yao Li Xue Yu Zhi Liao Xue, 2006,11(4):379-384
[7] Turowski GA, Rashid Z, Hong F , et al. Glutamine modulates phenotype and stimulates proliferation in human colon cancer cell lines[J]. Cancer Res, 1994,54(22):5974-5980
doi: 10.1002/1097-0142(19941115)74:10<2885::AID-CNCR2820741023>3.0.CO;2-P pmid: 7954430
[8] Rashid Z, Basson MD . Topoisomerase II inhibition differentially modulates Caco-2 intestinal epithelial cell phenotype[J]. Biochem Biophy Res Comm, 1996,219(1):82-88
[9] Yee SY . In vitro permeability across Caco-2 cells(colonic) can predict in vivo(small intestinal) absorption in man-fact or myth[J]. Pharm Res, 1997,14(6):763-766
[10] Chong S, Dando SA, Morrison RA . Evaluation of biocoat intestinal epithelium differentiation environment(3-day cultured Caco-2 cells) as an absorption screening model with improved productivity[J]. Pharm Res, 1997,14(12):1835-1837
[11] Gao JN, Hugger ED, Beck-Westermeyer MS , et al.Estimating intestinal mucosal permeation of compounds using Caco-2 cell monolayer. In ‘Current Protocols in Pharmacology.[M].Chichester:John Wiley & Sons, 2000.
[12] Walle UK, French KL, Walgren RA , et al. Transport of genistein-7-glucoside by human intestinal Caco-2 cells: Potential role for MRP2[J]. Res Commun Mol Pathol Pharmacol, 1999,103(1):45-46
doi: 10.1159/000028443 pmid: 10440570
[13] Tian XJ, Yang XD, Wang K , et al. The efflux of flavonoids morin, isorhamnetin-3-O-rutinoside and diosmetin-7-O-beta-D-xylopyranosyl-(1-6) -beta-D-glucopyranoside in the human intestinal cell line caco-2[J]. Pharm Res, 2006,23(8):1721-1728
[14] Yang XW, Guo QM, Wang Y , et al. Intestinal permeability of antivirus constituents from the fruits of Eucalyptus globulus Labill. in Caco-2 cell model[J]. Bioorg Med Chem Lett, 2007,17(4):1107-1111
[15] 田莉, 杨秀伟, 王莹 , 等. 顺式-和反式-阿霍烯在Caco-2细胞模型中的体外摄取、转运和外排特性[J]. 药学学报, 2007,42(1):87-92
Tian L, Yang XW, Wang Y , et al. Characteristics of uptake, transport and efflux of Z- and E-ajoenes in Caco-2 cell monolayers in vitro[J]. Yao Xue Xue Bao, 2007,42(1):87-92
[16] Yang XG, Yuan Lan, Wang K , et al. Comparison of intestinal absorption of two insulinmimic vanadyl complexes using Caco-2 monolayers as model system[J]. Chin Sci Bull, 2003,48(9):876-881
[17] Zhang Y, Yang XD, Wang K , et al. The permeability and cytotoxicity of insulin-mimetic vanadium(III, IV, V)-dipicolinate complexes[J]. J Inorg Biochem, 2006,100(1):80-87
[18] Bohets H, Annaert P, Mannens G , et al. Strategies for absorption screening in drug discovery and development[J]. Curr Top Med Chem, 2001,1(5):367-383
doi: 10.2174/1568026013394886 pmid: 11899103
[19] 张正全, 陆彬 . 口服结肠定位给药系统新进展[J]. 中国药学杂志, 2000,35(1):221-223
Mu YP, Wu G, Qiao XW , et al. The transport and absorption of chiral metal domplexes d,l-[Ni(phen)3]2+ across the model of Caco-2 cell monolayers[J]. Baotou Yi Xue Yuan Xue Bao, 2000,35(1):221-223
[20] Komura H, Shigemoto Y, Kawahara I , et al. High throughput screening of pharmacokinetics and metabolism in drug discovery(III)--investigation on in- silico model for membrane permeability and CYP1A2 inhibition[J]. Yakugaku Zasshi, 2005,125(1):141-147
[21] 牟永平, 吴刚, 乔秀文 , 等. 手性金属配合物d,l-[Ni(phen)3]2+ 经Caco-2细胞单层模型的吸收转运[J]. 包头医学院学报, 2006,22(4):358-360
Mu YP, Wu G, Qiao XW , et al. The transport and absorption of chiral metal domplexes d,l-[Ni(phen)3]2+ across the model of Caco-2 cell monolayers[J]. Baotou Yi Xue Yuan Xue Bao, 2006,22(4):358-360
[22] Schmiedlin-Ren P, Thummel KE, Fisher JM , et al. Expression of enzymatically active CYP3A4 by Caco-2 cells grown on extracellular matrix-coated permeable supports in the presence of 1alpha,25-dihydroxyvitamin D3[J]. Mol Pharmacol, 1997,51(5):741-754
[23] Annaert P, Kinget R, Naesens L , et al. Transport, uptake, and metabolism of the bis(pivaloyloxymethyl)-ester prodrug of 9-(2-phosphonylmethoxyethyl)adenine in an in vitro cell culture system of the intestinal mucosa(Caco-2)[J]. Pharm Res, 1997,14(4):492-496
[24] van Gelder J, Annaert P, Naesens L , et al. Inhibition of intestinal metabolism of the antiviral ester prodrug bis(POC)-PMPA by nature-identical fruit extracts as a strategy to enhance its oral absorption: an in vitro study[J]. Pharm Res, 1999,16(7):1035-1040
[25] Bartels H, Korsiak E,Daniel H . Transport activity of the MDR1-gene product in monolayers of Caco-2 cells[J].Zeitschrift fur Gastroenterologie, 1994, 32 15-18
[26] Dibas A, Wood J, Mia AJ et al.The ATP-depleting reagent iodoacetamide induces the degradation of protein kinase C alpha(PKCalpha) in LLC-PK1 pig kidney cells[J]. Life Sci, 1997,61(17):1697-1704
[27] Tsuruo T, Iida H, Tsukagoshi S , et al. Overcoming of vincristine resistance in P388 leukemia in vivo and in vitro through enhanced cytotoxicity of vincristine and vinblastine by verapamil[J]. Cancer Res, 1981,41(5):1967-1972
[28] Roepe PD . The P-glycoprotein efflux pump: how does it transport drugs?[J]. J Membr Bio, 1997,160(3):161-175
[29] Leier I, Jedlitschky G, Buchholz U , et al. The MRP gene encodes an ATP-dependent export pump for leukotriene C4 and structurally related conjugates[J]. J Biol Chem, 1994,269(45):27807-27810
[30] Jedlitschky G, Leier I, Buchholz U . ATP-dependent transport of glutathione S-conjugates by the multidrug resistance-associated protein[J]. Cancer Res, 1994,54(18):4833-4836
[31] Barrand MA, Bargrij T, Neo SY . Multidrug resistance-associated protein: A protein distinct from P-glycoprotein involved in cytotoxic drug expulsion[J]. Gen Pharmacol, 1997,28(5):639-645
[32] Chen J, Lin H, Hu M . Absorption and metabolism of genistein and its five isoflavone analogs in the human intestinal Caco-2 model[J]. Cancer Chemother Pharmacol, 2005,55(2):159-169
[33] Nikolic D, Li Y, Chadwick LR , et al. In vitro studies of intestinal permeability and hepatic and intestinal metabolism of 8-prenylnaringenin, a potent phytoestrogen from hops(Humulus lupulus L.)[J]. Pharm Res, 2006,23(5):864-872
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