A microfluidic cell culture device (μFCCD) to culture epithelial cells with physiological and morphological properties that mimic those of the human intestine

  • Meiying Chi
  • Banya Yi
  • Seunghan Oh
  • Dong-June Park
  • Jong Hwan Sung
  • Sungsu Park


Physiological and morphological properties of the human intestine cannot be accurately mimicked in conventional culture devices such as well plates and petri dishes where intestinal epithelial cells form a monolayer with loose contacts among cells. Here, we report a novel microfluidic cell culture device (μFCCD) that can be used to culture cells as a human intestinal model. This device enables intestinal epithelial cells (Caco-2) to grow three-dimensionally on a porous membrane coated with fibronectin between two polydimethylsiloxane (PDMS) layers. Within 3 days, Caco-2 cells cultured in the μFCCD formed villi- and crypt-like structures with small intercellular spaces, while individual cells were tightly connected to one another through the expression of the tight junction protein occludin, and were covered with a secreted mucin, MUC-2. Caco-2 cells cultured in the μFCCD for 3 days were less susceptible to bacterial attack than those cultured in transwell plates for 21 days. μFCCD-cultured Caco-2 cells also displayed physiologically relevant absorption and paracellular transport properties. These results suggest that our intestinal model more accurately mimics the morphological and physiological properties of the intestine in vivo than the conventional transwell culture model.


Caco-2 μFCCD Intestine Mucin-2 Microorganisms 



This work was supported by a grant from the Korea Science and Engineering Foundation (KOSEF) funded by the Korea government (MOST) (#2012-0001138), and by a grant from the Public Welfare & Safety Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (#2012R1A2A2A01012221 and #2012-0006522). JHS and BL acknowledges support from Hongik University Research Fund.


  1. M. Andrianifahanana, N. Moniaux, S.K. Batra, Regulation of mucin expression: mechanistic aspects and implications for cancer and inflammatory diseases. Biochim. Biophys. Acta 1765(2), 189–222 (2006)Google Scholar
  2. P. Artursson, K. Palm, K. Luthman, Caco-2 monolayers in experimental and theoretical predictions of drug transport. Adv. Drug Deliv. Rev. 46(1–3), 27–43 (2001)CrossRefGoogle Scholar
  3. M.D. Basson, Paradigms for mechanical signal transduction in the intestinal epithelium. Category: molecular, cell, and developmental biology. Digestion 68(4), 217–225 (2003)CrossRefGoogle Scholar
  4. J.M. Biazik, K.A. Jahn, Y. Su, Y.N. Wu, F. Braet, Unlocking the ultrastructure of colorectal cancer cells in vitro using selective staining. World J. Gastroenterol. 16(22), 2743–2753 (2010)CrossRefGoogle Scholar
  5. X.D. Bu, N. Li, X.Q. Tian, P.L. Huang, Caco-2 and LS174T cell lines provide different models for studying mucin expression in colon cancer. Tissue Cell 43(201), 201–206 (2011)CrossRefGoogle Scholar
  6. J.C. Byrd, R.S. Bresalier, Mucins and mucin binding proteins in colorectal cancer. Cancer Metastasis Rev. 23(1–2), 77–99 (2004)CrossRefGoogle Scholar
  7. M. Dickson, J.P. Gagnon, Key factors in the rising cost of new drug discovery and development. Nat. Rev. Drug Discov. 3(5), 417–429 (2004)CrossRefGoogle Scholar
  8. L.C. Duffy, Interactions mediating bacterial translocation in the immature intestine. J. Nutr. 130(2S Suppl), 432S–436S (2000)Google Scholar
  9. M.B. Esch, J.H. Sung, J. Yang, C. Yu, J. Yu, J.C. March, M.L. Shuler, On chip porous polymer membranes for integration of gastrointestinal tract epithelium with microfluidic ‘body-on-a-chip’ devices. Biomed. Microdevices 14(5), 895–906 (2012)CrossRefGoogle Scholar
  10. G.J. Feldman, J.M. Mullin, M.P. Ryan, Occludin: structure, function and regulation. Adv. Drug Deliv. Rev. 57(6), 883–917 (2005)CrossRefGoogle Scholar
  11. B.E. Goodman, Insights into digestion and absorption of major nutrients in humans. Adv. Physiol. Educ. 34(20), 44–53 (2010)CrossRefGoogle Scholar
  12. B.M. Gumbiner, Structure, biochemistry, and assembly of epithelial tight junctions. Am. J. Physiol. 253(6 Pt 1), C749–C758 (1987)Google Scholar
  13. B.M. Gumbiner, Breaking through the tight junction barrier. J. Cell Biol. 123(6 Pt 2), 1631–1633 (1993)CrossRefGoogle Scholar
  14. P. Guo, A.M. Weinstein, S. Weinbaum, A hydrodynamic mechanosensory hypothesis for brush border microvilli. Am J Physiol Renal Physiol. 279(4), F698–F712 (2000)Google Scholar
  15. J. Hansen, L. Andrew, R. Ruedy, M. Sato, Potential climate impact of mount pinatubo eruption. Geophys. Res. Lett. 19(2), 215–218 (1992)CrossRefGoogle Scholar
  16. D.H. Huh, Y.S. Torisawa, G.A. Hamilton, H.J. Kim, D.E. Ingber, Microengineered physiological biomimicry: organs-on-chips. Lab Chip 12(12), 2156–2164 (2012)CrossRefGoogle Scholar
  17. Y. Imura, Y. Asano, K. Sato, E. Yoshimura, A microfluidic system to evaluate intestinal absorption. Anal. Sci. 25, 1403–1407 (2009)CrossRefGoogle Scholar
  18. T. Ishikawa, T. Sato, G. Mohit, Y. Imai, T. Yamaguchi, Transport phenomena of microbial flora in the small intestine with peristalsis. J. Theor. Biol. 279(1), 63–73 (2011)CrossRefGoogle Scholar
  19. K. Izumikawa, Y. Hirakata, T. Yamaguchi, H. Takemura, S. Maesaki, K. Tomono, S. Igimi, M. Kaku, Y. Yamada, S. Kohno, S. Kamihira, Escherichia coli O157 interactions with human intestinal Caco-2 cells and the influence of fosfomycin. J. Antimicrob. Chemother. 42(3), 341–347 (1998)CrossRefGoogle Scholar
  20. Y. Jin, Y. Takegahara, Y. Sugawara, T. Matsumura, Y. Fujinaga, Disruption of the epithelial barrier by botulinum haemagglutinin (HA) proteins - differences in cell tropism and the mechanism of action between HA proteins of types A or B, and HA proteins of type C. Microbiology 155(Pt 1), 35–45 (2009)CrossRefGoogle Scholar
  21. J.A. Kiernan, Histological and histochemical methods: theory and practice (Butterworth Heinemann, Oxford, 1999). x, 502 p Google Scholar
  22. H.J. Kim, D.E. Ingber, Gut-on-a-Chip microenvironment induces human intestinal cells to undergo villus differentiation. Integr. Biol. 5(9), 1130–1140 (2013)CrossRefGoogle Scholar
  23. H.J. Kim, D. Huh, G. Hamilton, D.E. Ingber, Human gut-on-a-chip inhabited by microbial flora that experiences intestinal peristalsis-like motions and flow. Lab Chip 12(12), 2165–2174 (2012)CrossRefGoogle Scholar
  24. S.H. Kim, J.W. Lee, I. Choi, Y.C. Kim, J.B. Lee, J.H. Sung, A microfluidic device with 3-d hydrogel villi scaffold to simulate intestinal absorption. J. Nanosci. Nanotechnol. 13(11), 7220–7228 (2013)CrossRefGoogle Scholar
  25. S.H. Kim, M. Chi, B. Yi, S.H. Kim, S. Oh, Y. Kim, S. Park, J.H. Sung, Three-dimensional intestinal villi epithelium enhances protection of human intestinal cells from bacterial infection by inducing mucin expression. Integr. Biol. 6(12), 1122–1131 (2014)CrossRefGoogle Scholar
  26. R.G. Lentle, P.W.M. Janssen, Physical characteristics of digesta and their influence on flow and mixing in the mammalian intestine. J. Comp. Physiol. B. 178(6), 673–690 (2008)CrossRefGoogle Scholar
  27. F. Leonard, E.M. Collnot, C.M. Lehr, A three-dimensional coculture of enterocytes, monocytes and dendritic cells to model inflamed intestinal mucosa in vitro. Mol. Pharm. 7, 2103–2119 (2010)CrossRefGoogle Scholar
  28. N. Li, D. Wang, Z. Sui, X. Qi, L. Ji, X. Wang, L. Yang, Development of an improved three-dimensional in vitro intestinal mucosa model for drug absorption evaluation. Tissue Eng. Part C Methods 19(9), 708–719 (2013)CrossRefGoogle Scholar
  29. O. Lieleg, I. Vladescu, K. Ribbeck, Characterization of particle translocation through mucinhydrogels. Biophys. J. 98(9), 1782–1789 (2010)CrossRefGoogle Scholar
  30. K.M. McCarthy, I.B. Skare, M.C. Stankewich, M. Furuse, S. Tsukita, R.A. Rogers, R.D. Lynch, E.E. Schneeberger, Occludin is a functional component of the tight junction. J. Cell Sci. 109, 2287–2298 (1996)Google Scholar
  31. I. Meyvantsson, D.J. Beebe, Cell culture models in microfluidic systems. Annu. Rev. Anal. Chem. 1, 423–449 (2008)CrossRefGoogle Scholar
  32. K.W. Oh, L. Lee, B. Ahn, E.P. Furlani, Design of pressure-driven microfluidic networks using electric circuit analogy. Lab Chip 12(3), 515–545 (2012)CrossRefGoogle Scholar
  33. J. Olesen, L. Edvinsson, Basic mechanisms of headache, Amsterdam; NY, USA: Elsevier; Sole distributors for the USA and Canada, Elsevier Science Pub. Co. xxvi, 492 p (1988)Google Scholar
  34. S.P. Olesen, P.F. Davies, D.E. Clapham, Muscarinic-activated K+ current in bovine endothelial cells. Circ. Res. 62(2), 1059–1064 (1988b)CrossRefGoogle Scholar
  35. E. Panteris, P. Apostolakos, B. Galatis, Microtubule organization, mesophyll cell morphogenesis, and intercellular space formation in Adiantum capillus veneris leaflets. Protoplasma 172(2–4), 97–110 (1993)CrossRefGoogle Scholar
  36. S. Park, H.J. Chun, J.S. Jang, B. Keum, Y.S. Seo, Y.S. Kim, Y.T. Jeen, H.S. Lee, S.H. Um, C.D. Kim, H.S. Ryu, C.S. Uhm, S.J. Lee, Is intercellular space different among layers in normal esophageal mucosa? An electron microscopic study. Dig. Dis. Sci. 56(12), 3492–3497 (2011)CrossRefGoogle Scholar
  37. B.M.H. Schneeberger, Die Musikerfamilie Fürstenau: Untersuchungen zu Leben und Werk (Lit, Münster, 1992)Google Scholar
  38. J.P. Schouten, Revolution of the mystics: on the social aspects of Vīraśaivism (Kok Pharos Pub. House, Kampen, 1991). xiii, 331 p Google Scholar
  39. J.M. Staddon, L.L. Rubin, Cell adhesion, cell junctions and the blood-brain barrier. Curr. Opin. Neurobiol. 6(5), 622–627 (1996)CrossRefGoogle Scholar
  40. T.M. Straub, J.R. Hutchison, R.A. Bartholomew, C.O. Valdez, N.B. Valentine, A. Dohnalkova, R.M. Ozanich, C.J. Bruckner-Lea, Defining cell culture conditions to improve human norovirus infectivity assays. Water Sci. Technol. 67(4), 863–868 (2013)CrossRefGoogle Scholar
  41. J.H. Sung, J. Yu, D. Luo, M.L. Shuler, J.C. March, Microscale 3-D hydrogel scaffold for biomimetic gastrointestinal (GI) tract model. Lab Chip 11(3), 389–392 (2011)CrossRefGoogle Scholar
  42. J.H. Sung, M.B. Esch, J.M. Prot, C.J. Long, A. Smith, J.J. Hickman, M.L. Shuler, Microfabricated mammalian organ systems and their integration into models of whole animals and humans. Lab Chip 13(7), 1201–1212 (2013)CrossRefGoogle Scholar
  43. F.J. Van Asten, H.G. Hendriks, J.F. Koninkx, B.A. Van der Zeijst, W. Gaastra, Inactivation of the flagellin gene of Salmonella enterica serotype Enteritidis strongly reduces invasion into differentiated Caco-2 cells. FEMS Microbiol. Lett. 185(2), 175–179 (2000)CrossRefGoogle Scholar
  44. R.B. van Breemen, Y. Li, Caco-2 cell permeability assays to measure drug absorption. Expert Opin. Drug Metab. Toxicol. 1, 175–185 (2005)CrossRefGoogle Scholar
  45. P. Zanassi, M. Paolillo, A. Feliciello, E.V. Avvedimento, V. Gallo, S. Schinelli, cAMP-dependent protein kinase induces cAMP-response element-binding protein phosphorylation via an intracellular calcium release/ERK-dependent pathway in striatal neurons. J. Biol. Chem. 276(15), 11487–11495 (2001)CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of Chemistry and Nano Sciences (BK21 plus)EwhaWomans UniversitySeoulKorea
  2. 2.Department of Chemical EngineeringHongik UniversitySeoulKorea
  3. 3.Korea Food Research InstituteSeongnamKorea
  4. 4.School of Mechanical EngineeringSungkyunkwan UniversitySuwonKorea

Personalised recommendations