Engineering 3D approaches to model the dynamic microenvironments of cancer bone metastasis

Abstract

Cancer metastasis to bone is a three-dimensional (3D), multistep, dynamic process that requires the sequential involvement of three microenvironments, namely, the primary tumour microenvironment, the circulation microenvironment and the bone microenvironment. Engineered 3D approaches allow for a vivid recapitulation of in vivo cancerous microenvironments in vitro, in which the biological behaviours of cancer cells can be assessed under different metastatic conditions. Therefore, modelling bone metastasis microenvironments with 3D cultures is imperative for advancing cancer research and anti-cancer treatment strategies. In this review, multicellular tumour spheroids and bioreactors, tissue engineering constructs and scaffolds, microfluidic systems and 3D bioprinting technology are discussed to explore the progression of the 3D engineering approaches used to model the three microenvironments of bone metastasis. We aim to provide new insights into cancer biology and advance the translation of new therapies for bone metastasis.

Oncology: Modelling bone metastasis in 3-dimensions

Better 3D models are needed to understand the entire process of bone cancer metastasis, if new treatments are to be developed. Bone metastasis is a major complication of several common cancers, yet its biology is poorly understood - in part, because conventional cell culture systems have failed to replicate the 3D microenvironment of the body, where cancer cells dynamically interact with healthy cells and the extracellular matrix. To address this, 3D models of the primary tumour microenvironment, circulation microenvironment and bone microenvironment are needed, suggest Han Qiao and Tingting Tang at Shanghai Jiao Tong University School of Medicine in China. They describe promising 3D approaches, including multicellular tumour spheroids, tissue engineering constructs, microfluidic systems, and 3D bioprinting, but stress that none of these currently recapitulates the entire process of metastasis in a single culture system.

References

[1] Yan W et al. Suppressive effects of plumbagin on invasion and migration of breast cancer cells via the inhibition of STAT3 signaling and down-regulation of inflammatory cytokine expressions. Bone Res., 2013, 1:362-370

[2] Baier SR, Wan Y. MicroRNA exert macro-effects on cancer bone metastasis. Curr. Osteoporos. Rep., 2016, 14:163-169

[3] Wu B et al. Atypical skeletal manifestations of rickets in a familial hypocalciuric hypercalcemia patient. Bone Res., 2017, 5:17001

[4] Grinnell F. Fibroblast biology in three-dimensional collagen matrices. Trends Cell Biol., 2003, 13:264-269

[5] Peela N et al. A three dimensional micropatterned tumor model for breast cancer cell migration studies. Biomaterials, 2016, 81:72-83

[6] Chen SH et al. PLGA/TCP composite scaffold incorporating bioactive phytomolecule icaritin for enhancement of bone defect repair in rabbits. Acta Biomater., 2013, 9:6711-6722

[7] Fazilaty H, Mehdipour P. Genetics of breast cancer bone metastasis: a sequential multistep pattern. Clin. Exp. Metastas-., 2014, 31:595-612

[8] Paget S. The distribution of secondary growths in cancer of the breast. 1889. Cancer Metastas-. Rev., 1989, 8:98-101

[9] Douglas Hanahan JF. Patterns and emerging mechanisms of the. Cell, 1996, 86:353-364

[10] Auguste P et al. Molecular mechanisms of tumor vascularization. Crit. Rev. Oncol. Hematol., 2005, 54:53-61

[11] Thiery JP, Sleeman JP. Complex networks orchestrate epithelial-mesenchymal transitions. Nat. Rev. Mol. Cell Biol., 2006, 7:131-142

[12] Chambers AF, Groom AC, MacDonald IC. Dissemination and growth of cancer cells in metastatic sites. Nat. Rev. Cancer, 2002, 2:563-572

[13] Adjei IM, Blanka S. Modulation of the tumor microenvironment for cancer treatment: a biomaterials approach. J. Func. Biomater., 2015, 6:81-103

[14] Douma S et al. Suppression of anoikis and induction of metastasis by the neurotrophic receptor TrkB. Nature, 2004, 430:1034-1039

[15] Weidle UH et al. Molecular mechanisms of bone metastasis. Cancer Genom. Proteom., 2016, 13:1-12

[16] Giancotti FG. Mechanisms governing metastatic dormancy and reactivation. Cell, 2013, 155:750-764

[17] Bonewald LF. The amazing osteocyte. J. Bone Miner. Res., 2011, 26:229-238

[18] Nakashima T et al. Evidence for osteocyte regulation of bone homeostasis through RANKL expression. Nat. Med., 2011, 17:1231-1234

[19] Qiao H et al. Targeting osteocytes to attenuate early breast cancer bone metastasis by theranostic upconversion nanoparticles with responsive plumbagin release. ACS Nano, 2017, 11:7259-7273

[20] Giuliani N et al. Increased osteocyte death in multiple myeloma patients: role in myeloma-induced osteoclast formation. Leukemia, 2012, 26:1391-1401

[21] Sottnik JL et al. Tumor-induced pressure in the bone microenvironment causes osteocytes to promote the growth of prostate cancer bone metastases. Cancer Res., 2015, 75:2151-2158

[22] Hensel J, Thalmann GN. Biology of bone metastases in prostate cancer. Urology, 2016, 92:6-13

[23] Buijs JT, van der Pluijm G. Osteotropic cancers: from primary tumor to bone. Cancer Lett., 2009, 273:177-193

[24] Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell, 2011, 144:646-674

[25] Alemany-Ribes M, Semino CE. Bioengineering 3D environments for cancer models. Adv. Drug Deliv. Rev., 2014, 79-80:40-49

[26] Salamanna F et al. A systematic review on in vitro 3D bone metastases models: A new horizon to recapitulate the native clinical scenario? Oncotarget, 2016, 7:44803-44820

[27] Verjans ET et al. Three-dimensional cell culture models for anticancer drug screening: worth the effort? J. Cell. Physiol., 2017, 233:2993-3003

[28] Halfter K, Mayer B. Bringing 3D tumor models to the clinic-predictive value for personalized medicine. Biotechnol. J., 2017, 12:1600295

[29] Santo VE et al. Drug screening in 3D in vitro tumor models: overcoming current pitfalls of efficacy read-outs. Biotechnol. J., 2017, 12:1600505

[30] Ravi M, Ramesh A, Pattabhi A. Contributions of 3D cell cultures for cancer research. J. Cell. Physiol., 2017, 232:2679-2697

[31] Fong EL et al. Heralding a new paradigm in 3D tumor modeling. Biomaterials, 2016, 108:197-213

[32] Duval K et al. Modeling physiological events in 2D vs. 3D cell culture. Physiol, 2017, 32:266-277

[33] Sutherland RM et al. A multi-component radiation survival curve using an in vitro tumour model. Int. J. Radiat. Biol. Relat. Stud. Phys. Chem. Med., 1970, 18:491-495

[34] Grimes DR et al. A method for estimating the oxygen consumption rate in multicellular tumour spheroids. J. R. Soc. Interface, 2014, 11:20131124

[35] Nelson CM, Inman JL, Bissell MJ. Three-dimensional lithographically defined organotypic tissue arrays for quantitative analysis of morphogenesis and neoplastic progression. Nat. Protoc., 2008, 3:674-678

[36] Boo L et al. MiRNA transcriptome profiling of spheroid-enriched cells with cancer stem cell properties in human breast MCF-7 cell line. Int. J. Biol. Sci., 2016, 12:427-445

[37] Rouhani M et al. Lithium increases radiosensitivity by abrogating DNA repair in breast cancer spheroid culture. Arch. Iran. Med., 2014, 17:352-360

[38] Ray A, Vasudevan S, Sengupta S. 6-Shogaol inhibits breast cancer cells and stem cell-like spheroids by modulation of Notch signaling pathway and induction of autophagic cell death. PLoS ONE, 2015, 10:e0137614

[39] Page H, Flood P, Reynaud EG. Three-dimensional tissue cultures: current trends and beyond. Cell. Tissue Res., 2013, 352:123-131

[40] Kim JB. Three-dimensional tissue culture models in cancer biology. Semin. Cancer Biol., 2005, 15:365-377

[41] Becker JL, Souza GR. Using space-based investigations to inform cancer research on Earth. Nat. Rev. Cancer, 2013, 13:315-327

[42] Sutherland RM, MacDonald HR, Howell RL. Multicellular spheroids: a new model target for in vitro studies of immunity to solid tumor allografts. J. Natl. Cancer Inst., 1977, 58:1849-1853

[43] Hoffmann TK et al. A novel mechanism for anti-EGFR antibody action involves chemokine-mediated leukocyte infiltration. Int. J. Cancer, 2009, 124:2589-2596

[44] Holmes TD et al. A human NK cell activation/inhibition threshold allows small changes in the target cell surface phenotype to dramatically alter susceptibility to NK cells. J. Immunol., 2011, 186:1538-1545

[45] van Kasteren SI et al. Chemical biology of antigen presentation by MHC molecules. Curr. Opin. Immunol., 2014, 26:21-31

[46] Dietl K et al. Lactic acid and acidification inhibit TNF secretion and glycolysis of human monocytes. J. Immunol., 2010, 184:1200-1209

[47] Hauptmann S et al. Macrophages and multicellular tumor spheroids in co-culture: a three-dimensional model to study tumor-host interactions. Evidence for macrophage-mediated tumor cell proliferation and migration. Am. J. Pathol., 1993, 143:1406-1415

[48] Gottfried E et al. Tumor-derived lactic acid modulates dendritic cell activation and antigen expression. Blood, 2006, 107:2013-2021

[49] Kim TE et al. Three-dimensional culture and interaction of cancer cells and dendritic cells in an electrospun nano-submicron hybrid fibrous scaffold. Int. J. Nanomed., 2016, 11:823-835

[50] Sung SY et al. Coevolution of prostate cancer and bone stroma in three-dimensional coculture: implications for cancer growth and metastasis. Cancer Res., 2008, 68:9996-10003

[51] Hammond TG, Hammond JM. Optimized suspension culture: the rotating-wall vessel. Am. J. Physiol., 2001, 281:F12-F25

[52] Kaur P et al. Human breast cancer histoid: an in vitro 3-dimensional co-culture model that mimics breast cancer tissue. J. Histochem., 2011, 59:1087-1100

[53] Krishnan V et al. Dynamic interaction between breast cancer cells and osteoblastic tissue: comparison of two- and three-dimensional cultures. J. Cell. Physiol., 2011, 226:2150-2158

[54] Dhurjati R et al. Metastatic breast cancer cells colonize and degrade three-dimensional osteoblastic tissue in vitro. Clin. Exp. Metastas-., 2008, 25:741-752

[55] Mastro AM, Vogler EA. A three-dimensional osteogenic tissue model for the study of metastatic tumor cell interactions with bone. Cancer Res., 2009, 69:4097-4100

[56] Krishnan V et al. In vitro mimics of bone remodeling and the vicious cycle of cancer in bone. J. Cell. Physiol., 2014, 229:453-462

[57] Krishnan V, Vogler EA, Mastro AM. Three-dimensional in vitro model to study osteobiology and ssteopathology. J. Cell. Biochem., 2015, 116:2715-2723

[58] Liu M et al. Injectable hydrogels for cartilage and bone tissue engineering. Bone Res., 2017, 5:17014

[59] Yi H et al. Recent advances in nano scaffolds for bone repair. Bone Res., 2016, 4:16050

[60] Velazquez OC et al. Fibroblast-dependent differentiation of human microvascular endothelial cells into capillary-like three-dimensional networks. Faseb. J., 2002, 16:1316-1318

[61] Feng X et al. Fibrin and collagen differentially but synergistically regulate sprout angiogenesis of human dermal microvascular endothelial cells in 3-dimensional matrix. Int. J. Cell Biol., 2013, 2013:231279

[62] Benton G et al. Matrigel: from discovery and ECM mimicry to assays and models for cancer research. Adv. Drug Deliv. Rev., 2014, 79-80:3-18

[63] Correa de Sampaio P et al. A heterogeneous in vitro three dimensional model of tumour-stroma interactions regulating sprouting angiogenesis. PLoS ONE, 2012, 7:e30753

[64] Fischbach C et al. Engineering tumors with 3D scaffolds. Nat. Methods, 2007, 4:855-860

[65] Kimlin LC, Casagrande G, Virador VM. In vitro three-dimensional (3D) models in cancer research: an update. Mol. Carcinog., 2013, 52:167-182

[66] Tsurkan MV et al. Defined polymer-peptide conjugates to form cell-instructive starPEG-heparin matrices in situ. Adv. Mater., 2013, 25:2606-2610

[67] Taubenberger AV et al. 3D extracellular matrix interactions modulate tumour cell growth, invasion and angiogenesis in engineered tumour microenvironments. Acta Biomater., 2016, 36:73-85

[68] Roudsari LC et al. A 3D Poly(ethylene glycol)-based tumor angiogenesis model to study the influence of vascular cells on lung tumor cell behavior. Sci. Rep., 2016, 6

[69] Wartenberg M et al. Inhibition of tumor-induced angiogenesis and matrix-metalloproteinase expression in confrontation cultures of embryoid bodies and tumor spheroids by plant ingredients used in traditional chinese medicine. Lab. Invest., 2003, 83:87-98

[70] Timmins NE, Dietmair S, Nielsen LK. Hanging-drop multicellular spheroids as a model of tumour angiogenesis. Angiogenesis, 2004, 7:97-103

[71] Seano G et al. Modeling human tumor angiogenesis in a three-dimensional culture system. Blood, 2013, 121:e129-e137

[72] Oyanagi J et al. Epithelial-mesenchymal transition stimulates human cancer cells to extend microtubule-based invasive protrusions and suppresses cell growth in collagen gel. PLoS ONE, 2012, 7:e53209

[73] Chen L et al. The enhancement of cancer stem cell properties of MCF-7 cells in 3D collagen scaffolds for modeling of cancer and anti-cancer drugs. Biomaterials, 2012, 33:1437-1444

[74] Serebriiskii I et al. Fibroblast-derived 3D matrix differentially regulates the growth and drug-responsiveness of human cancer cells. Matrix Biol., 2008, 27:573-585

[75] Xu X, Farach-Carson MC, Jia X. Three-dimensional in vitro tumor models for cancer research and drug evaluation. Biotechnol. Adv., 2014, 32:1256-1268

[76] Fong EL et al. Modeling Ewing sarcoma tumors in vitro with 3D scaffolds. Proc. Natl Acad. Sci. U.S.A., 2013, 110:6500-6505

[77] Kim JW, Ho WJ, Wu BM. The role of the 3D environment in hypoxia-induced drug and apoptosis resistance. Anticancer Res., 2011, 31:3237-3245

[78] Huang YJ, Hsu SH. Acquisition of epithelial-mesenchymal transition and cancer stem-like phenotypes within chitosan-hyaluronan membrane-derived 3D tumor spheroids. Biomaterials, 2014, 35:10070-10079

[79] Tevis KM et al. Mimicking the tumor microenvironment to regulate macrophage phenotype and assessing chemotherapeutic efficacy in embedded cancer cell/macrophage spheroid models. Acta Biomater., 2017, 50:271-279

[80] Ramgolam K et al. Melanoma spheroids grown under neural crest cell conditions are highly plastic migratory/invasive tumor cells endowed with immunomodulator function. PLoS ONE, 2011, 6:e18784

[81] Herter S et al. A novel three-dimensional heterotypic spheroid model for the assessment of the activity of cancer immunotherapy agents. Cancer Immunol. Immunother., 2017, 66:129-140

[82] Phan-Lai V et al. CCL21 and IFN gamma recruit and activate tumor specific T cells in 3D scaffold model of breast cancer. Anticancer Agents Med. Chem., 2014, 14:204-210

[83] Campbell JJ et al. Development of three-dimensional collagen scaffolds with controlled architecture for cell migration studies using breast cancer cell lines. Biomaterials, 2017, 114:34-43

[84] Provenzano PP et al. Collagen density promotes mammary tumor initiation and progression. BMC Med., 2008, 6

[85] Mi K, Xing Z. CD44(+)/CD24(-) breast cancer cells exhibit phenotypic reversion in three-dimensional self-assembling peptide RADA16 nanofiber scaffold. Int. J. Nanomed., 2015, 10:3043-3053

[86] Da Sie Y et al. Fabrication of three-dimensional multi-protein microstructures for cell migration and adhesion enhancement. Biomed. Opt. Express, 2015, 6:480-490

[87] Wu M, Chen G, Li YP. TGF-beta and BMP signaling in osteoblast, skeletal development, and bone formation, homeostasis and disease. Bone Res., 2016, 4:16009

[88] Taubenberger AV et al. Delineating breast cancer cell interactions with engineered bone microenvironments. J. Bone Miner. Res., 2013, 28:1399-1411

[89] Reichert JC et al. Mineralized human primary osteoblast matrices as a model system to analyse interactions of prostate cancer cells with the bone microenvironment. Biomaterials, 2010, 31:7928-7936

[90] Sieh S et al. Interactions between human osteoblasts and prostate cancer cells in a novel 3D in vitro model. Organogenesis, 2010, 6:181-188

[91] Sieh S et al. Paracrine interactions between LNCaP prostate cancer cells and bioengineered bone in 3D in vitro culture reflect molecular changes during bone metastasis. Bone, 2014, 63:121-131

[92] Herroon MK, Diedrich JD, Podgorski I. New 3D-Culture approaches to study interactions of bone marrow adipocytes with metastatic prostate cancer cells. Front. Endocrinol., 2016, 7:84

[93] Marlow R et al. A novel model of dormancy for bone metastatic breast cancer cells. Cancer Res., 2013, 73:6886-6899

[94] Fitzgerald KA et al. The use of collagen-based scaffolds to simulate prostate cancer bone metastases with potential for evaluating delivery of nanoparticulate gene therapeutics. Biomaterials, 2015, 66:53-66

[95] Moreau JE et al. Tissue-engineered bone serves as a target for metastasis of human breast cancer in a mouse model. Cancer Res., 2007, 67:10304-10308

[96] Subia B et al. Target specific delivery of anticancer drug in silk fibroin based 3D distribution model of bone-breast cancer cells. ACS Appl. Mater. Interfaces, 2015, 7:2269-2279

[97] Cox RF et al. Osteomimicry of mammary adenocarcinoma cells in vitro; increased expression of bone matrix proteins and proliferation within a 3D collagen environment. PLoS ONE, 2012, 7:e41679

[98] de la Puente P et al. 3D tissue-engineered bone marrow as a novel model to study pathophysiology and drug resistance in multiple myeloma. Biomaterials, 2015, 73:70-84

[99] Holzapfel BM et al. Species-specific homing mechanisms of human prostate cancer metastasis in tissue engineered bone. Biomaterials, 2014, 35:4108-4115

[100] Han Q et al. Kinsenoside screening with a microfluidic chip attenuates gouty arthritis through inactivating NF-κB signaling in macrophages and protecting endothelial cells. Cell Death Dis., 2016, 7

[101] Hsiao AY et al. Microfluidic system for formation of PC-3 prostate cancer co-culture spheroids. Biomaterials, 2009, 30:3020-3027

[102] Buchanan CF et al. Three-dimensional microfluidic collagen hydrogels for investigating flow-mediated tumor-endothelial signaling and vascular organization. Tissue Eng., Part C., 2014, 20:64-75

[103] Ching-Te Kuo CLC et al. Configurable 2D and 3D spheroid tissue cultures on bioengineered surfaces with acquisition of epithelial–mesenchymal transition characteristics. NPG Asia Mater., 2012, 4

[104] Arai K et al. A novel high-throughput 3D screening system for EMT inhibitors: a pilot screening discovered the EMT inhibitory cctivity of CDK2 inhibitor SU9516. PLoS ONE, 2016, 11:e0162394

[105] Mantovani A et al. The chemokine system in cancer biology and therapy. Cytokine Growth Factor. Rev., 2010, 21:27-39

[106] Roussos ET, Condeelis JS, Patsialou A. Chemotaxis in cancer. Nat. Rev. Cancer, 2011, 11:573-587

[107] Huang CP et al. Engineering microscale cellular niches for three-dimensional multicellular co-cultures. Lab. Chip., 2009, 9:1740-1748

[108] Vickerman V et al. Design, fabrication and implementation of a novel multi-parameter control microfluidic platform for three-dimensional cell culture and real-time imaging. Lab. Chip., 2008, 8:1468-1477

[109] Polacheck WJ, Charest JL, Kamm RD. Interstitial flow influences direction of tumor cell migration through competing mechanisms. Proc. Natl Acad. Sci. U.S.A., 2011, 108:11115-11120

[110] Kim BJ et al. Cooperative roles of SDF-1alpha and EGF gradients on tumor cell migration revealed by a robust 3D microfluidic model. PLoS ONE, 2013, 8:e68422

[111] Sun YS et al. Electrotaxis of lung cancer cells in ordered three-dimensional scaffolds. Biomicrofluidics, 2012, 6:14102-1410214

[112] Shin MK, Kim SK, Jung H. Integration of intra- and extravasation in one cell-based microfluidic chip for the study of cancer metastasis. Lab. Chip., 2011, 11:3880-3887

[113] Jeon JS et al. Human 3D vascularized organotypic microfluidic assays to study breast cancer cell extravasation. Proc. Natl Acad. Sci. U.S.A., 2015, 112:214-219

[114] Bersini S et al. A microfluidic 3D in vitro model for specificity of breast cancer metastasis to bone. Biomaterials, 2014, 35:2454-2461

[115] Charbe N, McCarron PA, Tambuwala MM. Three-dimensional bio-printing: a new frontier in oncology research. World J. Clin. Oncol., 2017, 8:21-36

[116] Ma R et al. Bacterial inhibition potential of 3D rapid-prototyped magnesium-based porous composite scaffolds--an in vitro efficacy study. Sci. Rep., 2015, 5

[117] Yang Y et al. Anti-infective efficacy, cytocompatibility and biocompatibility of a 3D-printed osteoconductive composite scaffold functionalized with quaternized chitosan. Acta Biomater., 2016, 46:112-128

[118] Stanton MM, Samitier J, Sanchez S. Bioprinting of 3D hydrogels. Lab. Chip., 2015, 15:3111-3115

[119] Vanderburgh J, Sterling JA, Guelcher SA. 3D printing of tissue engineered constructs for in vitro modeling of disease progression and drug screening. Ann. Biomed. Eng., 2017, 45:164-179

[120] Zhang YS et al. Bioprinting the cancer microenvironment. ACS Biomater. Sci. Eng., 2016, 2:1710-1721

[121] Mou H et al. [Non-small cell lung cancer 95D cells co-cultured with 3D-bioprinted scaffold to construct a lung cancer model in vitro]. Zhonghua Zhongliu Zazhi, 2015, 37:736-740

[122] Xu F et al. A three-dimensional in vitro ovarian cancer coculture model using a high-throughput cell patterning platform. Biotechnol. J., 2011, 6:204-212

[123] Zhao Y et al. Three-dimensional printing of Hela cells for cervical tumor model in vitro. Biofabrication, 2014, 6:035001

[124] Ling KHG et al. Bioprinting-based high-throughput fabrication of three-dimensional MCF-7 human breast cancer cellular spheroids. Engineering, 2016, 1:269-274

[125] Grolman JM et al. Rapid 3D extrusion of synthetic tumor microenvironments. Adv. Mater., 2015, 27:5512-5517

[126] Huang TQ et al. 3D printing of biomimetic microstructures for cancer cell migration. Biomed. Micro., 2014, 16:127-132

[127] Soman P et al. Cancer cell migration within 3D layer-by-layer microfabricated photocrosslinked PEG scaffolds with tunable stiffness. Biomaterials, 2012, 33:7064-7070

[128] Guo R et al. Fabrication of 3D scaffolds with precisely controlled substrate modulus and pore size by templated-fused deposition modeling to direct osteogenic sifferentiation. Adv. Healthc. Mater., 2015, 4:1826-1832

[129] Zhou X et al. 3D bioprinting a cell-laden bone matrix for breast cancer metastasis study. ACS Appl. Mater. Interfaces, 2016, 8:30017-30026

[130] Zhu W et al. A 3D printed nano bone matrix for characterization of breast cancer cell and osteoblast interactions. Nanotechnology, 2016, 27:315103

[131] Zhu W et al. 3D printed nanocomposite matrix for the study of breast cancer bone metastasis. Nanomedicine, 2016, 12:69-79

[132] Sze WMRN, Sai F. From the printer: potential of three-dimensional printing for orthopaedic applications. J. Orthop. Transl., 2016, 6:42-49

[133] Qiao H et al. Structural simulation of adenosine phosphate via plumbagin and zoledronic acid competitively targets JNK/Erk to synergistically attenuate osteoclastogenesis in a breast cancer model. Cell Death Dis., 2016, 7

[134] Qiao H et al. Synergistic suppression of human breast cancer cells by combination of plumbagin and zoledronic acid In vitro. Acta Pharmacol. Sin., 2015, 36:1085-1098

[135] Amin R et al. 3D-printed microfluidic devices. Biofabrication, 2016, 8:022001