Application and prospects of organoid-on-a-chip in research on the intestinal mucosal barrier

Abstract

The integrity of the intestinal mucosal barrier is essential for maintaining normal gut physiology, and its disruption is associated with a wide range of conditions, including trauma-and burn-related intestinal injury, which remain difficult to manage clinically. Intestinal organoid-on-a-chip systems have emerged as advanced in vitro models that reproduce key features of the intestinal microenvironment and physiological function. These systems have shown promise for studying mucosal injury and repair, assessing therapeutic strategies, and supporting translational research. This review summarizes the basic principles of intestinal organoid-on-a-chip technology and examines its use in modeling intestinal barrier function, inflammatory responses, drug screening, regenerative approaches, and trauma-related barrier repair. It also reviews recent progress in preclinical studies, considers potential applications in gastrointestinal research, and discusses current technical challenges, particularly those related to scalability and reproducibility. Future directions for the development of next-generation systems are also outlined. With the continued integration of advances across disciplines, these platforms may provide useful tools for studying and treating disorders involving the intestinal mucosal barrier, especially in the context of trauma and burns.

Keywords

Intestinal mucosal barrier / Intestinal organoids / Organ-on-a-chip / Organoid-on-a-chip / Gut–organ-axis-on-a-chip / Intestinal injury / Intestinal barrier repair

References

[1] Sylvestre M, Di Carlo SE, Peduto L. Stromal regulation of the intestinal barrier. Mucosal Immunol 2023; 16: 221-31. https://doi.org/10.1016/j.mucimm.2023.01.006.

[2] Untersmayr E, Brandt A, Koidl L, Bergheim I. The intestinal barrier dysfunction as driving factor of inflammaging. Nutrients 2022; 14: 949. https://doi.org/10.3390/nu14050949.

[3] Mules TC, Inns S, Le Gros G. Helminths’ therapeutic potential to treat intestinal barrier dysfunction. Allergy 2023; 78: 2892-905. https://doi.org/10.1111/all.15812.

[4] Kartjito MS, Yosia M, Wasito E, Soloan G, Agussalim AF, Basrowi RW. Defining the relationship of gut microbiota, immunity, and cognition in early life-a narrative review. Nutrients 2023; 15: 2642. https://doi.org/10.3390/nu15122642.

[5] Matar A, Damianos JA, Jencks KJ, Camilleri M. Intestinal barrier impairment, preservation, and repair: an update. Nutrients 2024; 16: 3494. https://doi.org/10.3390/nu16203494.

[6] Macura B, Kiecka A, Szczepanik M. Intestinal permeability disturbances: causes, diseases and therapy. Clin Exp Med 2024; 24: 232. https://doi.org/10.1007/s10238-024-01496-9.

[7] Damianos J, Abdelnaem N, Camilleri M . Gut goo: physiology, diet, and therapy of intestinal mucus and biofilms in gastrointestinal health and disease. Clin Gastroenterol Hepatol Off Clin Pract J Am Gastroenterol Assoc 2025; 23: 205-15. https://doi.org/10.1016/j.cgh.2024.09.007.

[8] Liu L, Yue Q, Chen J, Liu H, Zeng X. Intestinal injury signaling pathway in sepsis. Front Immunol 2025; 16: 1620965. https://doi.org/10.3389/fimmu.2025.1620965.

[9] Ge P, Luo Y, Okoye CS, Chen H, Liu J, Zhang G, et al. Intestinal barrier damage, systemic inflammatory response syndrome, and acute lung injury: a troublesome trio for acute pancreatitis. Biomed Pharmacother Biomedecine Pharmacother 2020; 132: 110770. https://doi.org/10.1016/j.biopha.2020.110770.

[10] Wang M, Li R, Sheng S, Dong Z, Bai L, Wang X, et al. Combination therapy using intestinal organoids and their extracellular vesicles for inflammatory bowel disease complicated with osteoporosis. J Orthop Translat 2025; 53: 26-36. https://doi.org/10.1016/j.jot.2025.05.008.

[11] Riedel S, Pheiffer C, Johnson R, Louw J, Muller CJF. Intestinal barrier function and immune homeostasis are missing links in obesity and type 2 diabetes development. Front Endocrinol 2021; 12: 833544. https://doi.org/10.3389/fendo.2021.833544.

[12] Kim S, Goel R, Kumar A, Qi Y, Lobaton G, Hosaka K, et al. Imbalance of gut microbiome and intestinal epithelial barrier dysfunction in patients with high blood pressure. Clin Sci Lond Engl 1979; 2018: 701-18. https://doi.org/10.1042/CS20180087.

[13] Abdullahi A, Amini-Nik S, Jeschke MG. Animal models in burn research. Cell Mol LIFE Sci CMLS 2014; 71: 3241-55. https://doi.org/10.1007/s00018-014-1612-5.

[14] Kumari M, Alam K, Kaity S, Sah SK, Ravichandiran V, Roy S. Fabrication of multilayer heterogeneous cell assembly for pathophysiologically relevant 3D in-vitro IBD disease model for high throughput drug screening. Biofabrication 2025; 17: 2015. https://doi.org/10.1088/1758-5090/adc50e.

[15] Marín-Sáez J, Lopez-Ruiz R, Faria MA, Ferreira IMPLVO, Garrido FA. A comprehensive study on the digestion, absorption, and metabolization of tropane alkaloids in human cell models. J Hazard Mater 2024; 480: 136192. https://doi.org/10.1016/j.jhazmat.2024.136192.

[16] Hosic S, Lake W, Stas E, Koppes R, Breault DT, Murthy SK, et al. Cholinergic activation of primary human derived intestinal epithelium does not ameliorate TNF-α induced injury. Cell Mol Bioeng 2020; 13: 487-505. https://doi.org/10.1007/s12195-020-00633-0.

[17] Rudolph SE, Longo BN, Tse MW, Houchin MR, Shokoufandeh MM, Chen Y, et al. Crypt-villus scaffold architecture for bioengineering functional human intestinal epithelium. ACS Biomater Sci Eng 2022; 8: 4942-55. https://doi.org/10.1021/acsbiomaterials.2c00851.

[18] Mitrofanova O, Nikolaev M, Xu Q, Broguiere N, Cubela I, Camp JG, et al. Bioengineered human colon organoids with in vivo-like cellular complexity and function. Cell Stem Cell 2024; 31: 1175-1186.e7. https://doi.org/10.1016/j.stem.2024.05.007.

[19] Fey C, Truschel T, Nehlsen K, Damigos S, Horstmann J, Stradal T, et al. Enhancing pre-clinical research with simplified intestinal cell line models. J Tissue Eng 2024; 15: 20417314241228949. https://doi.org/10.1177/20417314241228949.

[20] Li X-G, Chen M-X, Zhao S-Q, Wang X-Q. Intestinal models for personalized medicine: from conventional models to microfluidic primary intestine-on-a-chip. Stem Cell Rev Rep 2022; 18: 2137-51. https://doi.org/10.1007/s12015-021-10205-y.

[21] Wu L, Ai Y, Xie R, Xiong J, Wang Y, Liang Q. Organoids/organs-on-a-chip: new frontiers of intestinal pathophysiological models. Lab Chip 2023; 23: 1192-212. https://doi.org/10.1039/d2lc00804a.

[22] Chelakkot C, Ghim J, Ryu SH. Mechanisms regulating intestinal barrier integrity and its pathological implications. Exp Mol Med 2018; 50: 1-9. https://doi.org/10.1038/s12276-018-0126-x.

[23] Stolfi C, Maresca C, Monteleone G, Laudisi F. Implication of intestinal barrier dysfunction in gut dysbiosis and diseases. Biomedicines 2022; 10: 289. https://doi.org/10.3390/biomedicines10020289.

[24] Yuan S, Wang Q, Li J, Xue J-C, Li Y, Meng H, et al. Inflammatory bowel disease: an overview of chinese herbal medicine formula-based treatment. Chin Med 2022; 17: 74. https://doi.org/10.1186/s13020-022-00633-4.

[25] Wang X, Ding C, Li H-B. The crosstalk between enteric nervous system and immune system in intestinal development, homeostasis and diseases. Sci China Life Sci 2024; 67: 41-50. https://doi.org/10.1007/s11427-023-2376-0.

[26] Li B, Yeganeh M, Lee D, Chusilp S, Balsamo F, Ganji N, et al. Exploring the complex pathophysiology of necrotizing enterocolitis in preterm neonates. Annu Rev Pathol 2025; 21: 37-58. https://doi.org/10.1146/annurev-pathmechdis-070224-014223.

[27] Chen S, Li T, Zhao P, Liang P, Huang X, Song L, et al. Stachyose alleviates high-fat diet-induced obesity via browning of white adipose tissue and modulation of gut microbiota. Curr Res Food Sci 2025; 10: 101081. https://doi.org/10.1016/j.crfs.2025.101081.

[28] Chen X, Zhang P, Zhang Y, Fan S, Wei Y, Yang Z, et al. Potential effect of glutamine in the improvement of intestinal stem cell proliferation and the alleviation of burn-induced intestinal injury via activating YAP: a preliminary study. Nutrients 2023; 15: 1766. https://doi.org/10.3390/nu15071766.

[29] He W, Wang Y. Intestinal barrier dysfunction in severe burn injury. Burns Trauma 2019; 7: 24. https://doi.org/10.1186/s41038-019-0162-3.

[30] Breugelmans T, Oosterlinck B, Arras W, Ceuleers H, De Man J, Hold GL, et al. The role of mucins in gastrointestinal barrier function during health and disease. Lancet Gastroenterol Hepatol 2022; 7: 455-71. https://doi.org/10.1016/S2468-1253(21)00431-3.

[31] Okumura R, Takeda K. The role of the mucosal barrier system in maintaining gut symbiosis to prevent intestinal inflammation. Semin Immunopathol 2024; 47: 2. https://doi.org/10.1007/s00281-024-01026-5.

[32] Kashihara H, Tanaka H, Kitamata M, Shiratsuchi G, Katsuno T, Tsukita K, et al. Functional landscape of mechanistic diversity in 27 claudin family members at tight junctions. Sci Adv 2025; 11: eadx7431. https://doi.org/10.1126/sciadv.adx7431.

[33] Naydenov NG, Hu G, Robak D, Zafar A, Makhmudova K, Lechuga S, et al. The septin cytoskeleton is a novel regulator of intestinal epithelial barrier integrity and mucosal inflammation. JCI. Insight 2025; 10: e191538. https://doi.org/10.1172/jci.insight.191538.

[34] Martins-Gomes C, Nunes FM, Silva AM. Natural products as dietary agents for the prevention and mitigation of oxidative damage and inflammation in the intestinal barrier. Antioxidants (Basel) 2024; 13: 65. https://doi.org/10.3390/antiox13010065.

[35] Zhang M, Liu Q, Meng H, Duan H, Liu X, Wu J, et al. Ischemia-reperfusion injury: molecular mechanisms and therapeutic targets. Signal Transduct Target Ther 2024; 9: 12. https://doi.org/10.1038/s41392-023-01688-x.

[36] Chen Y, Han B, Guan X, Du G, Sheng B, Tang X, et al. Enteric fungi protect against intestinal ischemia-reperfusion injury via inhibiting the SAA1-GSDMD pathway. J Adv Res 2024; 61: 223-37. https://doi.org/10.1016/j.jare.2023.09.008.

[37] Huang Y, Feng Y, Wang Y, Wang P, Wang F, Ren H. Severe burn-induced intestinal epithelial barrier dysfunction is associated with endoplasmic reticulum stress and autophagy in mice. Front Physiol 2018; 9: 441. https://doi.org/10.3389/fphys.2018.00441.

[38] Palrasu M, Kakar K, Marudamuthu A, Hamida H, Thada S, Zhong Y, et al. AhR activation transcriptionally induces anti-microbial peptide alpha-defensin 1 leading to reversal of gut microbiota dysbiosis and colitis. Gut Microbes 2025; 17: 2460538. https://doi.org/10.1080/19490976.2025.2460538.

[39] Zha X, Su S, Wu D, Zhang P, Wei Y, Fan S, et al. The impact of gut microbiota changes on the intestinal mucus barrier in burned mice: a study using 16S rRNA and metagenomic sequencing. Burns Trauma 2023; 11: tkad056. https://doi.org/10.1093/burnst/tkad056.

[40] Peterson LW, Artis D. Intestinal epithelial cells: regulators of barrier function and immune homeostasis. Nat Rev Immunol 2014; 14: 141-53. https://doi.org/10.1038/nri3608.

[41] Shi N, Li N, Duan X, Niu H. Interaction between the gut microbiome and mucosal immune system. Mil Med Res 2017; 4: 14. https://doi.org/10.1186/s40779-017-0122-9.

[42] Hays KE, Pfaffinger JM, Ryznar R. The interplay between gut microbiota, short-chain fatty acids, and implications for host health and disease. Gut Microbes 2024; 16: 2393270. https://doi.org/10.1080/19490976.2024.2393270.

[43] Liu P, Wang Y, Yang G, Zhang Q, Meng L, Xin Y, et al. The role of short-chain fatty acids in intestinal barrier function, inflammation, oxidative stress, and colonic carcinogenesis. Pharmacol Res 2021; 165: 105420. https://doi.org/10.1016/j.phrs.2021.105420.

[44] Wu W-JH, Kim M, Chang L-C, Assie A, Saldana-Morales FB, Zegarra-Ruiz DF, et al. Interleukin-1β secretion induced by mucosa-associated gut commensal bacteria promotes intestinal barrier repair. Gut Microbes 2022; 14: 2014772. https://doi.org/10.1080/19490976.2021.2014772.

[45] Bègue H, Ducreux A, Paradis T, Loiselet A, Mourer T, Lucchi G, et al. Candida albicans releases a peptide from the Rbt1 protein to promote its invasion into the gut epithelium. Gut Microbes 2025; 17: 2573038. https://doi.org/10.1080/19490976.2025.2573038.

[46] Earley ZM, Akhtar S, Green SJ, Naqib A, Khan O, Cannon AR, et al. Burn injury alters the intestinal microbiome and increases gut permeability and bacterial translocation. PLoS One 2015; 10: e0129996. https://doi.org/10.1371/journal.pone.0129996.

[47] You X, Niu L, Fu J, Ge S, Shi J, Zhang Y, et al. Bidirectional regulation of the brain-gut-microbiota axis following traumatic brain injury. Neural Regen Res 2025; 20: 2153-68. https://doi.org/10.4103/NRR.NRR-D-24-00088.

[48] Sato T, Vries RG, Snippert HJ, van de Wetering M, Barker N, Stange DE, et al. Single Lgr5 stem cells build crypt-villus structures in vitro without a mesenchymal niche. Nature 2009; 459: 262-5. https://doi.org/10.1038/nature07935.

[49] Lancaster MA, Renner M, Martin C-A, Wenzel D, Bicknell LS, Hurles ME, et al. Cerebral organoids model human brain development and microcephaly. Nature 2013; 501: 373-9. https://doi.org/10.1038/nature12517.

[50] Verstegen MMA, Coppes RP, Beghin A, De Coppi P, Gerli MFM, de Graeff N, et al. Clinical applications of human organoids. Nat Med 2025; 31: 409-21. https://doi.org/10.1038/s41591-024-03489-3.

[51] Birtele M, Lancaster M, Quadrato G. Modelling human brain development and disease with organoids. Nat Rev Mol Cell Biol 2025; 26: 389-412. https://doi.org/10.1038/s41580-024-00804-1.

[52] Beumer J, Clevers H. Cell fate specification and differentiation in the adult mammalian intestine. Nat Rev Mol Cell Biol 2021; 22: 39-53. https://doi.org/10.1038/s41580-020-0278-0.

[53] Simpson HL, Smits E, Moerkens R, Wijmenga C, Mooiweer J, Jonkers IH, et al. Human organoids and organ-on-chips in coeliac disease research. Trends Mol Med 2025; 31: 117-37. https://doi.org/10.1016/j.molmed.2024.10.003.

[54] Kim S, Min S, Choi YS, Jo S-H, Jung JH, Han K, et al. Tissue extracellular matrix hydrogels as alternatives to matrigel for culturing gastrointestinal organoids. Nat Commun 2022; 13: 1692. https://doi.org/10.1038/s41467-022-29279-4.

[55] Kaur S, Kaur I, Rawal P, Tripathi DM, Vasudevan A. Non-matrigel scaffolds for organoid cultures. Cancer Lett 2021; 504: 58-66. https://doi.org/10.1016/j.canlet.2021.01.025.

[56] Hughes CS, Postovit LM, Lajoie GA. Matrigel: a complex protein mixture required for optimal growth of cell culture. Proteomics 2010; 10: 1886-90. https://doi.org/10.1002/pmic.200900758.

[57] Gnecco JS, Brown A, Buttrey K, Ives C, Goods BA, Baugh L, et al. Organoid co-culture model of the human endometrium in a fully synthetic extracellular matrix enables the study of epithelial-stromal crosstalk. Med N Y N 2023; 4: 554-579.e9. https://doi.org/10.1016/j.medj.2023.07.004.

[58] Gjorevski N, Sachs N, Manfrin A, Giger S, Bragina ME, Ordóñez-Morán P, et al. Designer matrices for intestinal stem cell and organoid culture. Nature 2016; 539: 560-4. https://doi.org/10.1038/nature20168.

[59] Ye S, Boeter JWB, Mihajlovic M, van Steenbeek FG, van Wolferen ME, Oosterhoff LA, et al. A chemically defined hydrogel for human liver organoid culture. Adv Funct Mater 2020; 30: 2000893. https://doi.org/10.1002/adfm.202000893.

[60] Jia Z, Wang Z. Photo-crosslinking hydrogel based on porcine small intestinal submucosa decellularized matrix/fish collagen/GelMA for culturing small intestinal organoids and repairing intestinal defects. Int J Mol Sci 2025; 26: 663. https://doi.org/10.3390/ijms26020663.

[61] Wijnakker JJAPM, Lim S, Schreurs R, Faria JF, Korving J, Begthel H, et al. Invasin-functionalized PIC hydrogels enable long-term 3D culture of epithelial organoids. Proc Natl Acad Sci U S A 2025; 122: e2507500122. https://doi.org/10.1073/pnas.2507500122.

[62] Farin HF, Jordens I, Mosa MH, Basak O, Korving J, Tauriello DVF, et al. Visualization of a short-range wnt gradient in the intestinal stem-cell niche. Nature 2016; 530: 340-3. https://doi.org/10.1038/nature16937.

[63] Shi M, Fu P, Bonventre JV, McCracken KW. Directed differentiation of ureteric bud and collecting duct organoids from human pluripotent stem cells. Nat Protoc 2023; 18: 2485-508. https://doi.org/10.1038/s41596-023-00847-2.

[64] Wang Y, Hou Q, Wu Y, Xu Y, Liu Y, Chen J, et al. Methionine deficiency and its hydroxy analogue influence chicken intestinal 3-dimensional organoid development. Anim Nutr Zhongguo Xu Mu Shou Yi Xue Hui 2022; 8: 38-51. https://doi.org/10.1016/j.aninu.2021.06.001.

[65] Hayashi R, Okubo T, Kudo Y, Ishikawa Y, Imaizumi T, Suzuki K, et al. Generation of 3D lacrimal gland organoids from human pluripotent stem cells. Nature 2022; 605: 126-31. https://doi.org/10.1038/s41586-022-04613-4.

[66] Xu H, Jiao D, Liu A, Wu K. Tumor organoids: applications in cancer modeling and potentials in precision medicine. J Hematol Oncol J Hematol Oncol 2022; 15: 58. https://doi.org/10.1186/s13045-022-01278-4.

[67] Zhong G, Lin R, Jiang T, Huo H, Jiang J, Mu P, et al. Low dosage exposure of deoxynivalenol and copper synergistically enhanced the intestinal toxicity. Environ Int 2025; 205: 109891. https://doi.org/10.1016/j.envint.2025.109891.

[68] Rubert J, Schweiger PJ, Mattivi F, Tuohy K, Jensen KB, Lunardi A. Intestinal organoids: a tool for modelling diet-microbiome-host interactions. Trends Endocrinol Metab TEM 2020; 31: 848-58. https://doi.org/10.1016/j.tem.2020.02.004.

[69] Kim R, Sung JH. Recent advances in gut-and gut-organ-axis-on-a-chip models. Adv Healthc Mater 2024; 13: e2302777. https://doi.org/10.1002/adhm.202302777.

[70] Zhao Y, Landau S, Okhovatian S, Liu C, Lu RXZ, Lai BFL, et al. Integrating organoids and organ-on-a-chip devices. Nat Rev Bioeng 2024; 2: 588-608. https://doi.org/10.1038/s44222-024-00207-z.

[71] Özkan A, LoGrande NT, Feitor JF, Goyal G, Ingber DE. Intestinal organ chips for disease modelling and personalized medicine. Nat Rev Gastroenterol Hepatol 2024; 21: 751-73. https://doi.org/10.1038/s41575-024-00968-3.

[72] Huang Y, Liu T, Huang Q, Wang Y. From organ-on-a-chip to human-on-a-chip: a review of research progress and latest applications. ACS Sens 2024; 9: 3466-88. https://doi.org/10.1021/acssensors.4c00004.

[73] Farhang Doost N, Srivastava SK . A comprehensive review of organ-on-a-chip technology and its applications. Biosensors 2024; 14: 225. https://doi.org/10.3390/bios14050225.

[74] Thakar RG, Fenton KN. Bioethical implications of organ-on-a-chip on modernizing drug development. Artif Organs 2023; 47: 1553-8. https://doi.org/10.1111/aor.14620.

[75] Frey N, Sönmez UM, Minden J, LeDuc P. Microfluidics for understanding model organisms. Nat Commun 2022; 13: 3195. https://doi.org/10.1038/s41467-022-30814-6.

[76] Monteduro AG, Rizzato S, Caragnano G, Trapani A, Giannelli G, Maruccio G. Organs-on-chips technologies-a guide from disease models to opportunities for drug development. Biosens Bioelectron 2023; 231: 115271. https://doi.org/10.1016/j.bios.2023.115271.

[77] Orlova VV, Nahon DM, Cochrane A, Cao X, Freund C, van den Hil F, et al. Vascular defects associated with hereditary hemorrhagic telangiectasia revealed in patient-derived isogenic iPSCs in 3D vessels on chip. Stem Cell Rep 2022; 17: 1536-45. https://doi.org/10.1016/j.stemcr.2022.05.022.

[78] Wu J, Fang H, Zhang J, Yan S. Modular microfluidics for life sciences. J Nanobiotechnology 2023; 21: 85. https://doi.org/10.1186/s12951-023-01846-x.

[79] Quintard C, Tubbs E, Jonsson G, Jiao J, Wang J, Werschler N, et al. A microfluidic platform integrating functional vascularized organoids-on-chip. Nat Commun 2024; 15: 1452. https://doi.org/10.1038/s41467-024-45710-4.

[80] Chen H, Luo Z, Lin X, Zhu Y, Zhao Y. Sensors-integrated organ-on-a-chip for biomedical applications. Nano Res 2023; 16: 10072-10099. https://doi.org/10.1007/s12274-023-5651-9.

[81] Dabbagh Moghaddam F, Anvar A, Ilkhani E, Dadgar D, Rafiee M, Ranjbaran N, et al. Advances in engineering immune-tumor microenvironments on-a-chip: integrative microfluidic platforms for immunotherapy and drug discovery. Mol Cancer 2025; 24: 271. https://doi.org/10.1186/s12943-025-02479-4.

[82] Zhu C, Wang K, Lv Y, Liu S, Shen Y, Zhang Y. Immobilization protocols of nanozyme with different morphologies in microfluidic chips for biosensing. ACS Appl Mater Interfaces 2025; 17: 57967-75. https://doi.org/10.1021/acsami.5c14204.

[83] Khetani S, Yong KW, Alhasadi MF, Hamedzadeh A, Shahsavari L, Rafiei A, et al. A programmable, 3D neuron-on-chip platform integrating near real-time biosensing and multiaxial loading for mechanobiological injury profiling. Adv Sci Weinh Baden-Wurtt Ger 2025; 13: e10309. https://doi.org/10.1002/advs.202510309.

[84] Cao UMN, Zhang Y, Chen J, Sayson D, Pillai S, Tran SD. Microfluidic organ-on-a-chip: a guide to biomaterial choice and fabrication. Int J Mol Sci 2023; 24: 3232. https://doi.org/10.3390/ijms24043232.

[85] Petta D, D’Amora U, D’Arrigo D, Tomasini M, Candrian C, Ambrosio L, et al. Musculoskeletal tissues-on-a-chip: role of natural polymers in reproducing tissue-specific microenvironments. Biofabrication 2022; 14: 2001. https://doi.org/10.1088/1758-5090/ac8767.

[86] Liu J, Du C, Chen J, Tang B, Liu S, Tan J, et al. Hydrogel microspheres empowering organ-on-a-chip systems: innovations and applications. Small Weinh Bergstr Ger 2025; 21: e2504563. https://doi.org/10.1002/smll.202504563.

[87] Sharma A, Sances S, Workman MJ, Svendsen CN. Multi-lineage human iPSC-derived platforms for disease modeling and drug discovery. Cell Stem Cell 2020; 26: 309-29. https://doi.org/10.1016/j.stem.2020.02.011.

[88] Lall D, Workman MJ, Sances S, Ondatje BN, Bell S, Lawless G, et al. An organ-chip model of sporadic ALS using iPSC-derived spinal cord motor neurons and an integrated blood-brain-like barrier. Cell Stem Cell 2025; 32: 1139-1153.e7. https://doi.org/10.1016/j.stem.2025.05.015.

[89] Carter RD, Purnama U, Castro-Guarda M, Montes-Aparicio CN, Chandran A, Mbasu R, et al. Systems biology and functional assessments of human iPSC-cardiomyocyte models of insulin resistance capture key hallmarks of diabetic cardiomyopathy. Diabetes 2025; 74: 1929-45. https://doi.org/10.2337/db25-0204.

[90] Mochida T, Miyoshi M, Kakinuma S, Shimizu T, Tsuchiya J, Watakabe K, et al. Crosstalk via ICAM-1 enhances supportive phenotype of stellate cells and drives hepatocyte proliferation in iPSC-derived hepatic organoids. Stem Cell Rep 2025; 20: 102642. https://doi.org/10.1016/j.stemcr.2025.102642.

[91] Kasendra M, Tovaglieri A, Sontheimer-Phelps A, Jalili-Firoozinezhad S, Bein A, Chalkiadaki A, et al. Development of a primary human small intestine-on-a-chip using biopsy-derived organoids. Sci Rep 2018; 8: 2871. https://doi.org/10.1038/s41598-018-21201-7.

[92] Lindner M, Laporte A, Block S, Elomaa L, Weinhart M. Physiological shear stress enhances differentiation, mucus formation and structural 3D organization of intestinal epithelial cells in vitro. Cells 2021; 10: 2062. https://doi.org/10.3390/cells10082062.

[93] Lee KK, McCauley HA, Broda TR, Kofron MJ, Wells JM, Hong CI. Human stomach-on-a-chip with luminal flow and peristaltic-like motility. Lab Chip 2018; 18: 3079-85. https://doi.org/10.1039/c8lc00910d.

[94] Lamers MM, van der Vaart J, Knoops K, Riesebosch S, Breugem TI, Mykytyn AZ, et al. An organoid-derived bronchioalveolar model for SARS-CoV-2 infection of human alveolar type II-like cells. EMBO J 2021; 40: e105912. https://doi.org/10.15252/embj.2020105912.

[95] Wang Z, Zhang Y, Li Z, Wang H, Li N, Deng Y. Microfluidic brain-on-a-chip: from key technology to system integration and application. Small Weinh Bergstr Ger 2023; 19: e2304427. https://doi.org/10.1002/smll.202304427.

[96] Campisi M, Shin Y, Osaki T, Hajal C, Chiono V, Kamm RD. 3D self-organized microvascular model of the human blood-brain barrier with endothelial cells, pericytes and astrocytes. Biomaterials 2018; 180: 117-29. https://doi.org/10.1016/j.biomaterials.2018.07.014.

[97] Zhi Y, Zhu Y, Wang J, Zhao J, Zhao Y. Cortical organoid-on-a-chip with physiological hypoxia for investigating tanshinone IIA-induced neural differentiation. Res Wash DC 2023; 6: 273. https://doi.org/10.34133/research.0273.

[98] Ma C, Banan Sadeghian R, Negoro R, Fujimoto K, Araoka T, Ishiguro N, et al. Protocol to develop a proximal tubule-on-chip model based on hiPSC-derived kidney organoids for functional analysis of renal transporters. STAR Protoc 2025; 6: 103777. https://doi.org/10.1016/j.xpro.2025.103777.

[99] Brandenberg N, Hoehnel S, Kuttler F, Homicsko K, Ceroni C, Ringel T, et al. High-throughput automated organoid culture via stem-cell aggregation in microcavity arrays. Nat Biomed Eng 2020; 4: 863-74. https://doi.org/10.1038/s41551-020-0565-2.

[100] Shrestha S, Lekkala VKR, Acharya P, Kang S-Y, Vanga MG, Lee M-Y. Reproducible generation of human liver organoids (HLOs) on a pillar plate platform via microarray 3D bioprinting. Lab Chip 2024; 24: 2747-61. https://doi.org/10.1039/d4lc00149d.

[101] Fumadó Navarro J, Crilly S, Chan WK, Browne S, Mason JO, Vallejo-Giraldo C, et al. Cerebral organoids with integrated endothelial networks emulate the neurovascular unit and mitigate core necrosis. Adv Sci Weinh Baden-Wurtt Ger 2025; 12: e07256. https://doi.org/10.1002/advs.202507256.

[102] Yu T, Yang Q, Peng B, Gu Z, Zhu D. Vascularized organoid-on-a-chip: design, imaging, and analysis. Angiogenesis 2024; 27: 147-72. https://doi.org/10.1007/s10456-024-09905-z.

[103] Li B, Tang Y, Huang Z, Ma L, Song J, Xue L. Synergistic innovation in organ-on-a-chip and organoid technologies: reshaping the future of disease modeling, drug development and precision medicine. Protein. Cell 2025;: 1-18. https://doi.org/10.1093/procel/pwaf058.

[104] Ramadan Q, Jing L. Characterization of tight junction disruption and immune response modulation in a miniaturized caco-2/U937 coculture-based in vitro model of the human intestinal barrier. Biomed Microdevices 2016; 18: 11. https://doi.org/10.1007/s10544-016-0035-5.

[105] Sibilio S, Mennella R, Gregorio VD, Rocca AL, Urciuolo F, Imparato G, et al. A novel membrane-on-chip guides morphogenesis for the reconstruction of the intestinal crypt-villus axis. Biofabrication 2024; 16: 5019. https://doi.org/10.1088/1758-5090/ad6599.

[106] Jang Y, Kim H, Oh J. An array of carbon nanofiber Bundle_Based 3D in vitro intestinal microvilli for mimicking functional and physical activities of the small intestine. Small Weinh Bergstr Ger 2024; 20: e2404842. https://doi.org/10.1002/smll.202404842.

[107] Floor E, Su J, Chatterjee M, Kuipers ES, IJssennagger N, Heidari F, et al. Development of a caco-2-based intestinal mucosal model to study intestinal barrier properties and bacteria-mucus interactions. Gut Microbes 2025; 17: 2434685. https://doi.org/10.1080/19490976.2024.2434685.

[108] Sontheimer-Phelps A, Chou DB, Tovaglieri A, Ferrante TC, Duckworth T, Fadel C, et al. Human colon-on-a-chip enables continuous in vitro analysis of colon mucus layer accumulation and physiology. Cell Mol Gastroenterol Hepatol 2020; 9: 507-26. https://doi.org/10.1016/j.jcmgh.2019.11.008.

[109] Almalla A, Alzain N, Elomaa L, Richter F, Scholz J, Lindner M, et al. Hydrogel-integrated millifluidic systems: advancing the fabrication of mucus-producing human intestinal models. Cells 2024; 13: 1080. https://doi.org/10.3390/cells13131080.

[110] Kharaghani D, DeLoid GM, He P, Swenor B, Bui TH, Zuverza-Mena N, et al. Toxicity and absorption of polystyrene micro-nanoplastics in healthy and crohn’s disease human duodenum-chip models. J Hazard Mater 2025; 490: 137714. https://doi.org/10.1016/j.jhazmat.2025.137714.

[111] Feile A, Wegner VD, Raasch M, Mosig AS . Immunocompetent intestine-on-chip model for analyzing gut mucosal immune responses. J Vis Exp JoVE 2024; 207: e66603. https://doi.org/10.3791/66603.

[112] Gijzen L, Marescotti D, Raineri E, Nicolas A, Lanz HL, Guerrera D, et al. An intestine-on-a-chip model of plug-and-play modularity to study inflammatory processes. SLAS Technol 2020; 25: 585-97. https://doi.org/10.1177/2472630320924999.

[113] Min S, Than N, Shin YC, Hu G, Shin W, Ambrosini YM, et al. Live probiotic bacteria administered in a pathomimetic leaky gut chip ameliorate impaired epithelial barrier and mucosal inflammation. Sci Rep 2022; 12: 22641. https://doi.org/10.1038/s41598-022-27300-w.

[114] Penarete-Acosta D, Mittal M, Chakraborty S, Han A, Jayaraman A. Interplay between dietary fiber, macrophages and colonocytes in a microfluidic model of host-microbiota interactions in colorectal cancer. Lab Chip 2025; 25: 5482-96. https://doi.org/10.1039/d5lc00052a.

[115] Grassart A, Malardé V, Gobaa S, Sartori-Rupp A, Kerns J, Karalis K, et al. Bioengineered human organ-on-chip reveals intestinal microenvironment and mechanical forces impacting shigella infection. Cell Host Microbe 2019; 26: 435-444.e4. https://doi.org/10.1016/j.chom.2019.08.007.

[116] Trbojević-Stanković JB, Milićević NM, Milosević DP, Despotović N, Davidović M, Erceg P, et al. Morphometric study of healthy jejunal and ileal mucosa in adult and aged subjects. Histol Histopathol 2010; 25: 153-8. https://doi.org/10.14670/HH-25.153.

[117] Yi B, Shim KY, Ha SK, Han J, Hoang H-H, Choi I, et al. Three-dimensional in vitro gut model on a villi-shaped collagen scaffold. BioChip J 2017; 11: 219-31. https://doi.org/10.1007/s13206-017-1307-8.

[118] Costello CM, Hongpeng J, Shaffiey S, Yu J, Jain NK, Hackam D, et al. Synthetic small intestinal scaffolds for improved studies of intestinal differentiation. Biotechnol Bioeng 2014; 111: 1222-32. https://doi.org/10.1002/bit.25180.

[119] Wang Y, Gunasekara DB, Reed MI, DiSalvo M, Bultman SJ, Sims CE, et al. A microengineered collagen scaffold for generating a polarized crypt-villus architecture of human small intestinal epithelium. Biomaterials 2017; 128: 44-55. https://doi.org/10.1016/j.biomaterials.2017.03.005.

[120] Altay G, Tosi S, García-Díaz M, Martínez E. Imaging the cell morphological response to 3D topography and curvature in engineered intestinal tissues. Front Bioeng Biotechnol 2020; 8: 294. https://doi.org/10.3389/fbioe.2020.00294.

[121] Gustafsson JK, Johansson MEV. The role of goblet cells and mucus in intestinal homeostasis. Nat Rev Gastroenterol Hepatol 2022; 19: 785-803. https://doi.org/10.1038/s41575-022-00675-x.

[122] Ojo BA, VanDussen KL, Rosen MJ. The promise of patient-derived colon organoids to model ulcerative colitis. Inflamm Bowel Dis 2022; 28: 299-308. https://doi.org/10.1093/ibd/izab161.

[123] Ren J, Huang S. Intestinal organoids in inflammatory bowel disease: advances, applications, and future directions. Front Cell Dev Biol 2025; 13: 1517121. https://doi.org/10.3389/fcell.2025.1517121.

[124] Hahn S, Han IW, Shin SH, Kim G, Kim JH. Modeling diabetic intestinal organoids: aspects of rapid gut barrier disruption. Biochem Biophys Res Commun 2025; 760: 151730. https://doi.org/10.1016/j.bbrc.2025.151730.

[125] Huang S, Zhang S, Chen L, Pan X, Wen Z, Chen Y, et al. Lipopolysaccharide induced intestinal epithelial injury: a novel organoids-based model for sepsis in vitro. Chin Med J 2022; 135: 2232-9. https://doi.org/10.1097/CM9.0000000000002348.

[126] Zhang M, Huang X, Zhang Y, Yu M, Yuan X, Xu Y, et al. Gut microbial metabolite butyrate suppresses hepatocellular carcinoma growth via CXCL11-dependent enhancement of natural killer cell infiltration. Gut Microbes 2025; 17: 2519706. https://doi.org/10.1080/19490976.2025.2519706.

[127] Nearing JT, Douglas GM, Hayes MG, MacDonald J, Desai DK, Allward N, et al. Microbiome differential abundance methods produce different results across 38 datasets. Nat Commun 2022; 13: 342. https://doi.org/10.1038/s41467-022-28034-z.

[128] Wang H, Li X, Shi P, You X, Zhao G. Establishment and evaluation of on-chip intestinal barrier biosystems based on microfluidic techniques. Mater Today Bio 2024; 26: 101079. https://doi.org/10.1016/j.mtbio.2024.101079.

[129] Kollmann C, Buerkert H, Meir M, Richter K, Kretzschmar K, Flemming S, et al. Human organoids are superior to cell culture models for intestinal barrier research. Front Cell Dev Biol 2023; 11: 1223032. https://doi.org/10.3389/fcell.2023.1223032.

[130] Palamà MEF, Aiello M, Borka G, Furci J, Parodi I, Firpo G, et al. A dynamic double-flow gut-on-chip model for predictive absorption studies in vitro. Adv Mater Technol 2025; 10: 2401661. https://doi.org/10.1002/admt.202401661.

[131] Li Y, Tang W, Guo M. The cell as matter: connecting molecular biology to cellular functions. Matter 2021; 4: 1863-91. https://doi.org/10.1016/j.matt.2021.03.013.

[132] Tang W, Huang J, Pegoraro AF, Zhang JH, Tang Y, Kotton DN, et al. Topology and nuclear size determine cell packing on growing lung spheroids. Phys Rev X 2025; 15: 11067. https://doi.org/10.1103/physrevx.15.011067.

[133] Virumbrales-Muñoz M, Ayuso JM, Gong MM, Humayun M, Livingston MK, Lugo-Cintrón KM, et al. Microfluidic lumen-based systems for advancing tubular organ modeling. Chem Soc Rev 2020; 49: 6402-42. https://doi.org/10.1039/d0cs00705f.

[134] Li Z, Hui J, Yang P, Mao H. Microfluidic organ-on-a-chip system for disease modeling and drug development. Biosensors 2022; 12: 370. https://doi.org/10.3390/bios12060370.

[135] Wang X, Stefanello ST, Shahin V, Qian Y. From mechanoelectric conversion to tissue regeneration: translational progress in piezoelectric materials. Adv Mater Deerfield Beach Fla 2025; 37: 2417564. https://doi.org/10.1002/adma.202417564.

[136] Pérez-González C, Ceada G, Greco F, Matejčić M, Gómez-González M, Castro N, et al. Mechanical compartmentalization of the intestinal organoid enables crypt folding and collective cell migration. Nat Cell Biol 2021; 23: 745-57. https://doi.org/10.1038/s41556-021-00699-6.

[137] Xue S-L, Yang Q, Liberali P, Hannezo E. Mechanochemical bistability of intestinal organoids enables robust morphogenesis. Nat Phys 2025; 21: 608-17. https://doi.org/10.1038/s41567-025-02792-1.

[138] Baghdadi MB, Houtekamer RM, Perrin L, Rao-Bhatia A, Whelen M, Decker L, et al. PIEZO-dependent mechanosensing is essential for intestinal stem cell fate decision and maintenance. Science 2024; 386: eadj7615. https://doi.org/10.1126/science.adj7615.

[139] Bao L, Cui X, Wang X, Wu J, Guo M, Yan N, et al. Carbon nanotubes promote the development of intestinal organoids through regulating extracellular matrix viscoelasticity and intracellular energy metabolism. ACS Nano 2021; 15: 15858-73. https://doi.org/10.1021/acsnano.1c03707.

[140] Zhou Y, Wang F, Zhao R, Huang T, Su W, Wen Y, et al. Advanced strategies in organoid/organ-on-a-chip for food safety and nutrition. Food Res Int 2025; 219: 117011. https://doi.org/10.1016/j.foodres.2025.117011.

[141] Wang H, Ning X, Zhao F, Zhao H, Li D. Human organoids-on-chips for biomedical research and applications. Theranostics 2024; 14: 788-818. https://doi.org/10.7150/thno.90492.

[142] Yilmaz EG, Hacıosmanoğlu N, Jordi SBU, Yilmaz B, Inci F. Revolutionizing IBD research with on-chip models of disease modeling and drug screening. Trends Biotechnol 2025; 43: 1062-78. https://doi.org/10.1016/j.tibtech.2024.10.002.

[143] Villablanca EJ, Selin K, Hedin CRH. Mechanisms of mucosal healing: treating inflammatory bowel disease without immunosuppression? Nat Rev Gastroenterol Hepatol 2022; 19: 493-507. https://doi.org/10.1038/s41575-022-00604-y.

[144] Lin J, Wei Y, Gu X, Liu M, Wang M, Zhou R, et al. Nanotherapeutics-mediated restoration of pancreatic homeostasis and intestinal barrier for the treatment of severe acute pancreatitis. J Control Release Off J Control Release Soc 2025; 377: 93-105. https://doi.org/10.1016/j.jconrel.2024.11.022.

[145] Huang Y, Zhou J, Wang S, Xiong J, Chen Y, Liu Y, et al. Indoxyl sulfate induces intestinal barrier injury through IRF1-DRP1 axis-mediated mitophagy impairment. Theranostics 2020; 10: 7384-400. https://doi.org/10.7150/thno.45455.

[146] Zhao L, Huang Y, Ye Z, Chen W, Zhang N, Wen Z, et al. Short-chain fatty acids attenuate sepsis-induced gut dysbiosis and hippocampal neuroinflammation via NLRP6 inflammasome activation in mice. Int J Surg Lond Engl 2025; 112: 570-582. https://doi.org/10.1097/JS9.0000000000003554.

[147] Lechuga S, Braga-Neto MB, Naydenov NG, Rieder F, Ivanov AI. Understanding disruption of the gut barrier during inflammation: should we abandon traditional epithelial cell lines and switch to intestinal organoids? Front Immunol 2023; 14: 1108289. https://doi.org/10.3389/fimmu.2023.1108289.

[148] Günther C, Winner B, Neurath MF, Stappenbeck TS. Organoids in gastrointestinal diseases: from experimental models to clinical translation. Gut 2022; 71: 1892-908. https://doi.org/10.1136/gutjnl-2021-326560.

[149] Mennella R, Sibilio S, Urciuolo F, Imparato G, Netti PA. Flow-induced mechano-modulation of intestinal permeability on chip. Mater Today Bio 2025; 33: 101951. https://doi.org/10.1016/j.mtbio.2025.101951.

[150] Wang X, Zhu Y, Cheng Z, Zhang C, Liao Y, Liu B, et al. Emerging microfluidic gut-on-a-chip systems for drug development. Acta Biomater 2024; 188: 48-64. https://doi.org/10.1016/j.actbio.2024.09.012.

[151] Keuper-Navis M, Eslami Amirabadi H, Donkers J, Walles M, Poller B, Heming B, et al. Intestinal cells-on-chip for permeability studies. Micromachines 2024; 15: 1464. https://doi.org/10.3390/mi15121464.

[152] Papamichail L, Koch LS, Veerman D, Broersen K, van der Meer AD. Organoids-on-a-chip: microfluidic technology enables culture of organoids with enhanced tissue function and potential for disease modeling. Front Bioeng Biotechnol 2025; 13: 1515340. https://doi.org/10.3389/fbioe.2025.1515340.

[153] Malik M, Steele SA, Mitra D, Long CJ, Hickman JJ. Trans-epithelial/endothelial electrical resistance (TEER): current state of integrated TEER measurements in organ-on-a-chip devices. Curr Opin Biomed Eng 2025; 34: 100588. https://doi.org/10.1016/j.cobme.2025.100588.

[154] Holzreuter MA, Segerink LI. Innovative electrode and chip designs for transendothelial electrical resistance measurements in organs-on-chips. Lab Chip 2024; 24: 1121-34. https://doi.org/10.1039/d3lc00901g.

[155] Guo Y, Luo R, Wang Y, Deng P, Song T, Zhang M, et al. SARS-CoV-2 induced intestinal responses with a biomimetic human gut-on-chip. Sci Bull 2021; 66: 783-93. https://doi.org/10.1016/j.scib.2020.11.015.

[156] Morelli M, Savova MV, Queiroz K, Harms AC, Hankemeier T. Cytokine-induced barrier dysfunction and lipid signaling in a gut-on-chip model. FASEB J Off Publ Fed Am Soc Exp Biol 2025; 39: e71059. https://doi.org/10.1096/fj.202501685R.

[157] Huang J, Xu Z, Jiao J, Li Z, Li S, Liu Y, et al. Microfluidic intestinal organoid-on-a-chip uncovers therapeutic targets by recapitulating oxygen dynamics of intestinal ischemia/reperfusion injury. Bioact Mater 2023; 30: 1-14. https://doi.org/10.1016/j.bioactmat.2023.07.001.

[158] Hu W, Wang Y, Han J, Zhang W, Chen J, Li X, et al. Microfluidic organ-on-a-chip models for the gut-liver axis: from structural mimicry to functional insights. Biomater Sci 2025; 13: 1624-56. https://doi.org/10.1039/d4bm01273a.

[159] Gough A, Soto-Gutierrez A, Vernetti L, Ebrahimkhani MR, Stern AM, Taylor DL. Human biomimetic liver microphysiology systems in drug development and precision medicine. Nat Rev Gastroenterol Hepatol 2021; 18: 252-68. https://doi.org/10.1038/s41575-020-00386-1.

[160] Mehta V, Karnam G, Madgula V. Liver-on-chips for drug discovery and development. Mater Today Bio 2024; 27: 101143. https://doi.org/10.1016/j.mtbio.2024.101143.

[161] Lucchetti M, Aina KO, Grandmougin L, Jäger C, Pérez Escriva P, Letellier E, et al. An organ-on-chip platform for simulating drug metabolism along the gut-liver axis. Adv Healthc Mater 2024; 13: e2303943. https://doi.org/10.1002/adhm.202303943.

[162] Alqahtani S, Bukhari I, Albassam A, Alenazi M. An update on the potential role of intestinal first-pass metabolism for the prediction of drug-drug interactions: the role of PBPK modeling. Expert Opin Drug Metab Toxicol 2018; 14: 625-34. https://doi.org/10.1080/17425255.2018.1482277.

[163] Guo Y, Xie Y, Qin J. A generic pump-free organ-on-a-chip platform for assessment of intestinal drug absorption. Biotechnol J 2024; 19: e2300390. https://doi.org/10.1002/biot.202300390.

[164] Song Y, Li Y, Hu W, Li F, Sheng H, Huang C, et al. Luminol-conjugated cyclodextrin biological nanoparticles for the treatment of severe burn-induced intestinal barrier disruption. Burns Trauma 2024; 12: tkad054. https://doi.org/10.1093/burnst/tkad054.

[165] Ding X, Xu N, Zhang W, Wang P. Integrated microfluidic three-organ chip for real-time toxicity analysis of fluorotelomer alcohols in the gut-vascular-nerve axis. Lab Chip 2025; 25: 6170-6. https://doi.org/10.1039/d5lc00631g.

[166] Azar J, Bahmad HF, Daher D, Moubarak MM, Hadadeh O, Monzer A, et al. The use of stem cell-derived organoids in disease modeling: an update. Int J Mol Sci 2021; 22: 7667. https://doi.org/10.3390/ijms22147667.

[167] Pat Y, Yazici D, Zeyneloglu C, Babayev H, Ardicli S, Garci-Sanchez A, et al. Cellular stress, inflammation and barrier damage in gut epithelial cells caused by aspartame. Allergy 2025; 81: 884-901. https://doi.org/10.1111/all.70095.

[168] Rodrigues DB, Failla ML. Intestinal cell models for investigating the uptake, metabolism and absorption of dietary nutrients and bioactive compounds. Curr Opin Food Sci 2021; 41: 169-79. https://doi.org/10.1016/j.cofs.2021.04.002.

[169] Wang X, Xia Y, Wang J, Zhang Y, Lv P, Wang Y. Intestinal organoids as advanced modeling platform for food research: a review. Crit Rev Food Sci Nutr 2025; 66: 1463-1481. https://doi.org/10.1080/10408398.2025.2546510.

[170] Bein A, Fadel CW, Swenor B, Cao W, Powers RK, Camacho DM, et al. Nutritional deficiency in an intestine-on-a-chip recapitulates injury hallmarks associated with environmental enteric dysfunction. Nat Biomed Eng 2022; 6: 1236-47. https://doi.org/10.1038/s41551-022-00899-x.

[171] Nie H-Y, Ge J, Huang G-X, Liu K-G, Yue Y, Li H, et al. New insights into the intestinal barrier through “gut-organ” axes and a glimpse of the microgravity’s effects on intestinal barrier. Front Physiol 2024; 15: 1465649. https://doi.org/10.3389/fphys.2024.1465649.

[172] Wu X, Wang S, Li M, Li J, Shen J, Zhao Y, et al. Conditional reprogramming: next generation cell culture. Acta Pharm Sin B 2020; 10: 1360-81. https://doi.org/10.1016/j.apsb.2020.01.011.

[173] Tang X-Y, Wu S, Wang D, Chu C, Hong Y, Tao M, et al. Human organoids in basic research and clinical applications. Signal Transduct Target Ther 2022; 7: 168. https://doi.org/10.1038/s41392-022-01024-9.

[174] Sugimoto S, Kobayashi E, Fujii M, Ohta Y, Arai K, Matano M, et al. An organoid-based organ-repurposing approach to treat short bowel syndrome. Nature 2021; 592: 99-104. https://doi.org/10.1038/s41586-021-03247-2.

[175] Sampaziotis F, Muraro D, Tysoe OC, Sawiak S, Beach TE, Godfrey EM, et al. Cholangiocyte organoids can repair bile ducts after transplantation in the human liver. Science 2021; 371: 839-46. https://doi.org/10.1126/science.aaz6964.

[176] Ozaki A, Sakai D, Mandai M. hPSC-based treatment of retinal diseases-current progress and challenges. Adv Drug Deliv Rev 2025; 221: 115587. https://doi.org/10.1016/j.addr.2025.115587.

[177] Hou Y, Cao C, Guo J, Du P, Li D, Yuan P, et al. From the laboratory to the clinic: differentiation, disease modeling and transplantation of cardiac organoids. Adv Healthc Mater 2025; 15: e02698. https://doi.org/10.1002/adhm.202502698.

[178] Zhou C, Li Z, Lu K, Liu Y, Xuan L, Mao H, et al. Advances in human organs-on-chips and applications for drug screening and personalized medicine. Fundam Res 2025; 5: 1258-72. https://doi.org/10.1016/j.fmre.2023.12.019.

[179] Ingber DE. Human organs-on-chips for disease modelling, drug development and personalized medicine. Nat Rev Genet 2022; 23: 467-91. https://doi.org/10.1038/s41576-022-00466-9.

[180] Tilg H, Adolph TE, Trauner M. Gut-liver axis: pathophysiological concepts and clinical implications. Cell Metab 2022; 34: 1700-18. https://doi.org/10.1016/j.cmet.2022.09.017.

[181] Meerschaert KA, Chiu IM. The gut-brain axis and pain signalling mechanisms in the gastrointestinal tract. Nat Rev Gastroenterol Hepatol 2025; 22: 206-21. https://doi.org/10.1038/s41575-024-01017-9.

[182] Zhao Y, Yu C, Zhang J, Yao Q, Zhu X, Zhou X. The gut-skin axis: emerging insights in understanding and treating skin diseases through gut microbiome modulation (review). Int J Mol Med 2025; 56: 210. https://doi.org/10.3892/ijmm.2025.5651.

[183] Yang T, Richards EM, Pepine CJ, Raizada MK. The gut microbiota and the brain-gut-kidney axis in hypertension and chronic kidney disease. Nat Rev Nephrol 2018; 14: 442-56. https://doi.org/10.1038/s41581-018-0018-2.

[184] Liu Q, Ruan K, An Z, Li L, Ding C, Xu D, et al. Updated review of research on the role of the gut microbiota and microbiota-derived metabolites in acute pancreatitis progression and inflammation-targeted therapy. Int J Biol Sci 2025; 21: 1242-58. https://doi.org/10.7150/ijbs.108858.

[185] Ohara TE, Hsiao EY. Microbiota-neuroepithelial signalling across the gut-brain axis. Nat Rev Microbiol 2025; 23: 371-84. https://doi.org/10.1038/s41579-024-01136-9.

[186] Ramasamy B, Sharma DK, Callary SA, Ramadass B, Solomon LB, Atkins GJ. Role of gut microbiota disruption in prosthetic joint infection: a scoping review. Lancet Microbe 2025; 6: 101193. https://doi.org/10.1016/j.lanmic.2025.101193.

[187] Zhang Y, Liu Y, Liang X, Wen Y, Zhao J, He Y, et al. Intestinal barrier in chronic gut and liver diseases: pathogenesis and therapeutic targets. Acta Pharm Sin B 2025; 15: 5515-36. https://doi.org/10.1016/j.apsb.2025.08.028.

[188] Kang SG, Choi YY, Mo SJ, Kim TH, Ha JH, Hong DK, et al. Effect of gut microbiome-derived metabolites and extracellular vesicles on hepatocyte functions in a gut-liver. Nano Converg 2023; 10: 5. https://doi.org/10.1186/s40580-022-00350-6.

[189] Marin TM, de Carvalho IN, Rocco SA, Basei FL, de Carvalho M, de Almeida GK, et al. Acetaminophen absorption and metabolism in an intestine/liver microphysiological system. Chem Biol Interact 2019; 299: 59-76. https://doi.org/10.1016/j.cbi.2018.11.010.

[190] Liu H, Yin G, Kohlhepp MS, Schumacher F, Hundertmark J, Hassan MIA, et al. Dissecting acute drug-induced hepatotoxicity and therapeutic responses of steatotic liver disease using primary mouse liver and blood cells in a liver-on-a-chip model. Adv Sci Weinh Baden-Wurtt Ger 2024; 11: e2403516. https://doi.org/10.1002/advs.202403516.

[191] Kim M-H, van Noort D, Sung JH, Park S. Organ-on-a-chip for studying gut-brain interaction mediated by extracellular vesicles in the gut microenvironment. Int J Mol Sci 2021; 22: 13513. https://doi.org/10.3390/ijms222413513.

[192] Góralska J, Raźny U, Polus A, Dziewońska A, Gruca A, Zdzienicka A, et al. Enhanced GIP secretion in obesity is associated with biochemical alteration and miRNA contribution to the development of liver steatosis. Nutrients 2020; 12: 476. https://doi.org/10.3390/nu12020476.

[193] Mulay AR, Hwang J, Kim D-H. Microphysiological blood-brain barrier systems for disease modeling and drug development. Adv Healthc Mater 2024; 13: e2303180. https://doi.org/10.1002/adhm.202303180.

[194] Zhang Y, Lu S-M, Zhuang J-J, Liang L-G. Advances in gut-brain organ chips. Cell Prolif 2024; 57: e13724. https://doi.org/10.1111/cpr.13724.

[195] Fakhri Bafghi MS, Khoshnam Rad N, Roostaei G, Nikfar S, Abdollahi M. The reality of modeling irritable bowel syndrome: progress and challenges. Expert Opin Drug Discov 2025; 20: 433-45. https://doi.org/10.1080/17460441.2025.2481264.

[196] Ahn K, Park H-S, Choi S, Lee H, Choi H, Hong SB, et al. Differentiating visceral sensory ganglion organoids from induced pluripotent stem cells. Nat Methods 2024; 21: 2135-46. https://doi.org/10.1038/s41592-024-02455-8.

[197] Kim NY, Lee HY, Choi YY, Mo SJ, Jeon S, Ha JH, et al. Effect of gut microbiota-derived metabolites and extracellular vesicles on neurodegenerative disease in a gut-brain axis chip. Nano Converg 2024; 11: 7. https://doi.org/10.1186/s40580-024-00413-w.

[198] Lyu Z, Park J, Kim K-M, Jin H-J, Wu H, Rajadas J, et al. A neurovascular-unit-on-a-chip for the evaluation of the restorative potential of stem cell therapies for ischaemic stroke. Nat Biomed Eng 2021; 5: 847-63. https://doi.org/10.1038/s41551-021-00744-7.

[199] Yan M, Man S, Sun B, Ma L, Guo L, Huang L, et al. Gut liver brain axis in diseases: the implications for therapeutic interventions. Signal Transduct Target Ther 2023; 8: 443. https://doi.org/10.1038/s41392-023-01673-4.

[200] Li M, Duan W, Hao X, Li S, Wang C, Liang Y, et al. Effects of esketamine on electrophysiology and metabolic reprogramming in brain organoids: insights into antidepressant mechanisms. Mol Psychiatry 2025; 30: 6107-18. https://doi.org/10.1038/s41380-025-03198-4.

[201] Lee HR, Sung JH. Multiorgan-on-a-chip for realization of gut-skin axis. Biotechnol Bioeng 2022; 119: 2590-601. https://doi.org/10.1002/bit.28164.

[202] Ko B, Son J, In Won J, Kang BM, Choi CW, Kim R, et al. Gut microbe-skin axis on a chip for reproducing the inflammatory crosstalk. Lab Chip 2025; 25: 2609-19. https://doi.org/10.1039/d4lc01010h.

[203] Brandauer K, Schweinitzer S, Lorenz A, Krauß J, Schobesberger S, Frauenlob M, et al. Advances of dual-organ and multi-organ systems for gut, lung, skin and liver models in absorption and metabolism studies. Lab Chip 2025; 25: 1384-403. https://doi.org/10.1039/d4lc01011f.

[204] Wang J, Zhang H, Qu Y, Yang Y, Xu S, Ji Z, et al. An eighteen-organ microphysiological system coupling a vascular network and excretion system for drug discovery. Microsyst Nanoeng 2025; 11: 89. https://doi.org/10.1038/s41378-025-00933-3.

[205] Sajin D, Moonshi SS, Cha H, Akther F, Zhang J, Nguyen N-T, et al. Multi-organ-on-a-chip: the gut and inflammatory diseases. ACS Biomater Sci Eng 2025; 11: 5330-42. https://doi.org/10.1021/acsbiomaterials.5c00208.

[206] Ziaka M, Exadaktylos A. Exploring the lung-gut direction of the gut-lung axis in patients with ARDS. Crit Care Lond Engl 2024; 28: 179. https://doi.org/10.1186/s13054-024-04966-4.

[207] Rae B, Bood V, Dijk H-J, Vasse GF, Melgert BN, Nagelkerke A, et al. Interorgan communication between lung and colorectal epithelial cells studied using a novel multi-organ-on-chip system. Compr Physiol 2025; 15: e70051. https://doi.org/10.1002/cph4.70051.

[208] Giordano L, Mihaila SM, Eslami Amirabadi H, Masereeuw R. Microphysiological systems to recapitulate the gut-kidney axis. Trends Biotechnol 2021; 39: 811-23. https://doi.org/10.1016/j.tibtech.2020.12.001.

[209] Mendes M, Morais AS, Carlos A, Sousa JJ, Pais AC, Mihăilă SM, et al. Organ-on-a-chip: quo vademus? Applications and regulatory status. Colloids Surf B Biointerfaces 2025; 249: 114507. https://doi.org/10.1016/j.colsurfb.2025.114507.

[210] Thorel L, Perréard M, Florent R, Divoux J, Coffy S, Vincent A, et al. Patient-derived tumor organoids: a new avenue for preclinical research and precision medicine in oncology. Exp Mol Med 2024; 56: 1531-51. https://doi.org/10.1038/s12276-024-01272-5.

[211] Li J, Li Y, Song G, Wang H, Zhang Q, Wang M, et al. Revolutionizing cardiovascular research: human organoids as a beacon of hope for understanding and treating cardiovascular diseases. Mater Today Bio 2025; 30: 101396. https://doi.org/10.1016/j.mtbio.2024.101396.

[212] Probst C, Schneider S, Loskill P. High-throughput organ-on-a-chip systems: current status and remaining challenges. Curr Opin Biomed Eng 2018; 6: 33-41. https://doi.org/10.1016/j.cobme.2018.02.004.

[213] Schröter J, Deininger L, Lupse B, Richter P, Syrbe S, Mikut R, et al. A large and diverse brain organoid dataset of 1,400 cross-laboratory images of 64 trackable brain organoids. Sci Data 2024; 11: 514. https://doi.org/10.1038/s41597-024-03330-z.

[214] Ravi K, Manoharan TJM, Wang K-C, Pockaj B, Nikkhah M. Engineered 3D ex vivo models to recapitulate the complex stromal and immune interactions within the tumor microenvironment. Biomaterials 2024; 305: 122428. https://doi.org/10.1016/j.biomaterials.2023.122428.

[215] Man Y, Liu Y, Chen Q, Zhang Z, Li M, Xu L, et al. Organoids-on-a-chip for personalized precision medicine. Adv Healthc Mater 2024; 13: e2401843. https://doi.org/10.1002/adhm.202401843.

[216] Tong M, Huang G, Zhuang S, Yu X, Hu D, Lin W, et al. Robotic micromanipulation for patterned and complex organoid biofabrication. Sci Adv 2025; 11: eadz0808. https://doi.org/10.1126/sciadv.adz0808.

[217] Diosdi A, Toth T, Harmati M, Istvan G, Schrettner B, Hapek N, et al. HCS-3DX, a next-generation AI-driven automated 3D-oid high-content screening system. Nat Commun 2025; 16: 8897. https://doi.org/10.1038/s41467-025-63955-5.

[218] Hong Y, Li L, Yan L, Bai L, Su J, Zhang X. Parabiosis, assembloids, organoids (PAO). Adv Sci Weinh Baden-Wurtt Ger 2025; 13: e11671. https://doi.org/10.1002/advs.202511671.

[219] Huang P, Lan H, Liu B, Mo Y, Gao Z, Ye H, et al. Transformative laboratory medicine enabled by microfluidic automation and artificial intelligence. Biosens Bioelectron 2025; 271: 117046. https://doi.org/10.1016/j.bios.2024.117046.

[220] Wang Y, Jeon H. 3D cell cultures toward quantitative high-throughput drug screening. Trends Pharmacol Sci 2022; 43: 569-81. https://doi.org/10.1016/j.tips.2022.03.014.

[221] Duarte LC, Figueredo F, Chagas CLS, Cortón E, Coltro WKT. A review of the recent achievements and future trends on 3D printed microfluidic devices for bioanalytical applications. Anal Chim Acta 2024; 1299: 342429. https://doi.org/10.1016/j.aca.2024.342429.

[222] Murata D, Arai K, Nakayama K. Scaffold-free bio-3D printing using spheroids as “bio-inks” for tissue (re-)construction and drug response tests. Adv Healthc Mater 2020; 9: e1901831. https://doi.org/10.1002/adhm.201901831.

[223] Sharma C, Raza MA, Purohit SD, Pathak P, Gautam S, Corridon PR, et al. Cellulose-based 3D printing bio-inks for biomedical applications: a review. Int J Biol Macromol 2025; 305: 141174. https://doi.org/10.1016/j.ijbiomac.2025.141174.

[224] Fatimi A, Okoro OV, Podstawczyk D, Siminska-Stanny J, Shavandi A. Natural hydrogel-based bio-inks for 3D bioprinting in tissue engineering: a review. Gels Basel Switz 2022; 8: 179. https://doi.org/10.3390/gels8030179.

[225] Fukushima T, Togasaki K, Hamamoto J, Emoto K, Ebisudani T, Mitsuishi A, et al. An organoid library unveils subtype-specific IGF-1 dependency via a YAP-AP1 axis in human small cell lung cancer. Nat Cancer 2025; 6: 874-91. https://doi.org/10.1038/s43018-025-00945-y.

[226] Yue SSK, Tong Y, Siu HC, Ho SL, Law SYK, Tsui WY, et al. Divergent lineage trajectories and genetic landscapes in human gastric intestinal metaplasia organoids associated with early neoplastic progression. Gut 2025; 74: 522-38. https://doi.org/10.1136/gutjnl-2024-332594.

[227] Issing C, Menche C, Richter MR, Mosa MH, von der Grün J, Fleischmann M, et al. Head and neck tumor organoid biobank for modelling individual responses to radiation therapy according to the TP53/HPV status. J Exp Clin Cancer Res CR 2025; 44: 85. https://doi.org/10.1186/s13046-025-03345-3.

[228] Dayton TL, Alcala N, Moonen L, den Hartigh L, Geurts V, Mangiante L, et al. Druggable growth dependencies and tumor evolution analysis in patient-derived organoids of neuroendocrine neoplasms from multiple body sites. Cancer Cell 2023; 41: 2083-2099.e9. https://doi.org/10.1016/j.ccell.2023.11.007.

[229] Beumer J, Geurts MH, Lamers MM, Puschhof J, Zhang J, van der Vaart J, et al. A CRISPR/Cas9 genetically engineered organoid biobank reveals essential host factors for coronaviruses. Nat Commun 2021; 12: 5498. https://doi.org/10.1038/s41467-021-25729-7.

[230] Geurts MH, de Poel E, Amatngalim GD, Oka R, Meijers FM, Kruisselbrink E, et al. CRISPR-based adenine editors correct nonsense mutations in a cystic fibrosis organoid biobank. Cell Stem Cell 2020; 26: 503-510.e7. https://doi.org/10.1016/j.stem.2020.01.019.

[231] Tindle C, Fonseca AG, Taheri S, Katkar GD, Lee J, Maity P, et al. A living organoid biobank of patients with crohn’s disease reveals molecular subtypes for personalized therapeutics. Cell Rep Med 2024; 5: 101748. https://doi.org/10.1016/j.xcrm.2024.101748.

[232] Nakanishi A, Toyama S, Onozato D, Watanabe C, Hashita T, Iwao T, et al. Effects of human induced pluripotent stem cell-derived intestinal organoids on colitis-model mice. Regen Ther 2022; 21: 351-61. https://doi.org/10.1016/j.reth.2022.08.004.

[233] Shin YJ, Safina D, Zheng Y, Levenberg S. Microvascularization in 3D human engineered tissue and organoids. Annu Rev Biomed Eng 2025; 27: 473-98. https://doi.org/10.1146/annurev-bioeng-103023-115236.

[234] de Jongh D, Massey EK, VANGUARD consortium, Bunnik EM. Organoids: a systematic review of ethical issues. Stem Cell Res Ther 2022; 13: 337. https://doi.org/10.1186/s13287-022-02950-9.

[235] Weidema J, de Vries M, Mummery C, de Graeff N. The ethical aspects of human organ-on-chip models: a mapping review. Stem Cell Rep 2025; 20: 102686. https://doi.org/10.1016/j.stemcr.2025.102686.

[236] Pașca SP, Arlotta P, Campbell P, Charo A, Evans JH, Farahany N, et al. The need for a global effort to attend to human neural organoid and assembloid research. Science 2025; 390: 574-7. https://doi.org/10.1126/science.aeb1510.

[237] Chen HI, Wolf JA, Blue R, Song MM, Moreno JD, Ming G-L, et al. Transplantation of human brain organoids: revisiting the science and ethics of brain chimeras. Cell Stem Cell 2019; 25: 462-72. https://doi.org/10.1016/j.stem.2019.09.002.

[238] Ding L, Xiao Z, Gong X, Peng Y. Knowledge graphs of ethical concerns of cerebral organoids. Cell Prolif 2022; 55: e13239. https://doi.org/10.1111/cpr.13239.

[239] Fujimori K, Yamanaka S, Shimada K, Matsui K, Kawagoe S, Kuroda T, et al. Generation of human-pig chimeric renal organoids using iPSC technology. Commun Biol 2024; 7: 1278. https://doi.org/10.1038/s42003-024-06986-w.

[240] van Daal M, de Kanter A-FJ, Bredenoord AL, de Graeff N. Personalized 3D printed scaffolds: the ethical aspects. New Biotechnol 2023; 78: 116-22. https://doi.org/10.1016/j.nbt.2023.10.006.

[241] Schäffers OJM, Gribnau J, van Rijn BB, Bunnik EM. Ethical considerations for advancing research using organoid models derived from the placenta. Hum Reprod Update 2025; 31: 392-401. https://doi.org/10.1093/humupd/dmaf007.