Why 3D cell culture?

Today, there is increasing awareness of the drawbacks of 2D cell culture to predict the complex behavior of a biological system of many different interacting cell types and to reproduce the anatomy or physiology of a tissue for informative or useful study.

3D cell culture models are a more accurate representation of the natural environment experienced by the cells in the living organism, which allows for intercellular interactions with more realistic biochemical and physiological responses. Cells behave and respond more like they would in vivo to internal and external stimuli, such as changes in temperature, pH, nutrient absorption, transport, and differentiation. Therefore, scientists are shifting their focus from 2D to 3D cell culture in the fields of drug screening, tissue engineering, preclinical study, cell therapy and basic cell biological study.

Examples publications of 3D Cell Culture in areas of:
Differentiation  |  Morphology  |  Drug Metabolism  |  Proliferation |  Gene Expression  |  General Cell Function  | Viability

3D Cell Culture vs 2D cell Culture

Examples of publications showing the effects of 3D cell culture vs 2D cell culture on cell behaviors

  • Increased initial osteogenesis of adult rat mesenchymal stem cells as evidenced by collagen type I expression
  • Decreased later osteogenesis of adult rat mesenchymal stem cells as evidenced by collagen type I expression
  • Increased relative expression of osteogenic marker CBFA-1 of human adipose-derived stem cells
  • Increased relative expression of osteogenic marker CBFA-1 of human adipose-derived stem cells
  • Increased absolute expression of osteogenic markers alkaline phosphatase, osteonectin, osteopontin, Collagen 1, and JNK2 of human adipose-derived stem cells
  • Increased osteogenic differentiation of rat bone marrow stem cells
  • Increased differentiation of mouse embryonic stem cells into hematopoietic precursor cells
  • Increased differentiation activities after proliferation equilibrium of human trophoblast ED27 cells
  • Increased expression of contractile markers indicative of differentiated phenotype of human smooth muscle cells
  • Decreased cellular attachment, enabling phenotypic changes of primary cow chondrocytes
  • Increased osteogenic differentiation of cell line H1 human embryonic stem cells
  • Enhanced chondrogenesis as assessed by metachromasia and expression of chondrocyte-specific genes AGGRECAN, COL II, and COL X of human marrow stromal cells
  • Increased differentiation efficiency based on differentiation rate and leptin secretion of human adipose derived stem cells
  • Increased duration of undifferentiated state of mouse embryonic stem cells
  • Increased differentiation of primary human osteoblasts into osteocytes
  • Induced differentiated polarized phenotype of MDCK cells
  • Enabled formation of numerous bile canniculi and other in vivo similar morphological characteristics of human HepG2 liver cells
  • Maintained structural integrity on human HepG2 liver cells
  • Induced clear differences in morphology, arrangement and polarization of human bladder carcinoma cell line RT112 cells
  • Regenerated multicellular spheroids which maintained a tissue-like, geometrically well-ordered and highly prismatic organization in human bladder carcinoma cell line RT112 cells
  • Caused formation of aggregates of tumor epithelial (TEC) and human umbilical vein endothelial cells (HUVEC)
  • Increased tissue coherence and cell-cell contact of differentiated mouse 3T3-L1 preadipocytes
  • Altered morphology of human adipose-derived stem cells
  • Altered morphology of mouse osteoblast cell line MC3T3-E1 and primary mouse calvarial osteoblasts
  • Increased neurite extension length of human SH-SY5Y retinal cells
  • Decreased stress fibers and focal adhesions of vascular smooth muscle cells
  • Enabled integration with in situ generated extracellular matrix of mouse embryonic stem cells
  • Decreased cell size of hybridoma cells
  • Increased inter-cell connectivity / junction protein connections of human arachnoidal cells
  • Enabled formation of calcium and magnesium dependent tube-like structures mimicking angiogenesis of human endothelial cells
  • Decreased focal adhesion assemblies of human smooth muscle cells
  • Maintained original spindle-shape morphology of rat olfactory ensheathing cells
  • Increased cellular organization and cell-cell adhesions of human hepatoma cell line HepG2
  • Altered morphology of airway smooth muscle cells
  • Caused formation of complex, multilayered aggregates of Huh7 hepatoma cells
  • Induced cell alignment of human fibroblasts
  • Did not induce cell alignment of human endothelial cells
  • Significant differences in myoblast morphology and cytoskeletal organization
  • Increased cytochrome p-450 enzyme CYP1A2 ethoxyresorufin to resorufin conversion activity of C3A human hepatocyte cell line
  • Enabled response to CYP1A2 inducer beta-naphthoflavone and inhibitor furafylline of C3A human hepatocyte cell line
  • Increased cytochrome p-450 enzyme CYP3A4 testosterone to 6 beta-testosterone conversion activity of C3A human hepatocyte cell line
  • Increased cytochrome p-450 enzyme CYP3A4 phase II enzyme glucuronosyltransferases of C3A human hepatocyte cell line
  • Decreased the antiproliferative effect of doxorubicin, paclitaxel and tamoxifen on human MCF-7 cells
  • Did not affect chemosensitivity of PC-3 cell line cells
  • Decreased chemosensitivity of Lovo cell line cells
  • Decreased chemosensitivity of MCF-7 cell line cells
  • Increased chemosensitivity with anti-E-cadherin antibody treatment of MCF-7 cell line cells
  • Increased resistance to drug-induced apoptosis of human hepatoma cell line HepG2
  • Better preserved multidrug resistance associated protein 2 (Mrp2) of rat hepatocytes
  • Increased drug resistance of rat hepatocytes to methotrexate (MTX)
  • Linear metabolism of EFC to HFC in HepG2 cells producing liver-specific enzymes CYP1A1 and CYP3A4 with stable cell proliferation
  • Increased albumin and urea production in hepatocytes
  • Increased survival rates after exposure to the anti-cancer drug paclitaxel were observed in cell spheroids
  • Antiinvasive effect of doxorubicin on cell migration properties, beta1 intigrin expression nor the state of phosphorylation of FAK and PhoA in tumor cells
  • Change in cytotoxicity of Paclitaxel, KU174, Alimta, Zacitma, Doxorubicin, Vinorelbine, KU363, and 17AAg in H358 cells
  • Improved hepatotoxicity prediction efficacy
  • Increased growth rate and attachment of mesenchymal stem cells
  • Increased cell growth and metabolic rates of human osteosarcoma cells
  • Increased proliferation of tumor epithelial (TEC) and human umbilical vein endothelial cells (HUVEC)
  • Did not alter proliferation of SH-SY5Y human neuroblastoma cells on collagen constructs
  • Decreased proliferation of rat anterior tibialis muscle cells
  • Decreased proliferation of Lovo cell line cells
  • Decreased proliferation of MCF-7 cell line cells
  • Did not affect proliferation of PC-3 cell line cells
  • Increased progenitor cell number of primary human cord blood mononuclear cells
  • Supported cell growth under conditions of calcium and nutrients deprivation of rat hippocampal neurons and astrocytes
  • Decreased proliferation of sheep bone marrow stromal cells
  • Decreased proliferation rate of human breast cancer (MCF-7) cells
  • Increased proliferation rates of rat olfactory ensheathing cells
  • Enhanced inhibition of the proliferation of human epidermal growth factor receptor (HER2) over-expressing cancer cells with trastuzumab (Herceptin)
  • Decreased basal proliferation of airway smooth muscle cells
  • Dependent on cell-integrin engagement and epithelial ovarian cancer cell proteolytically remodeling abilities
  • Exponential cell proliferation trend in human glioblastoma cells
  • High-seeding density of HAC is critical for generation of cartilaginous constructs
  • Increase in CHO and fibroblast cell numbers on 3D fibrous matrices
  • Caused differences in gene expression of cow articular cartilage cells
  • Altered differential expression of 1,766 genes, including those relevant to cytoskeleton, extracellular matrix, and neurite outgrowth of human SH-SY5Y neuroblastoma cells
  • Doubled expression of 77 genes; halved the expression of 22 genes of vascular smooth muscle cells
  • Increased expression of cyclin-dependent kinase inhibitor 1 (p21) and collagen I of vascular smooth muscle cells
  • Increased monoclonal antibody productivity of hybridoma cells
  • Did not increase type IV collagen, fibronectin, and laminin accumulation of TGF-beta 1 treated human endothelial cells
  • Increased synthesis and secretion of type V collagen of both TGF-beta 1 treated and untreated human endothelial cells
  • Induced expression of morphogenesis-associated distal epithelial gene fibroblast growth factor receptor 2 (FGFr2) of C3A human hepatocyte cell line
  • Increased expression of E-cadherin, catenin, and p27 of MCF-7 cell line cells
  • Decreased vinculin expression of human smooth muscle cells
  • Increased expression of contractile markers indicative of differentiated phenotype of human smooth muscle cells
  • Enabled synthesis of osteonectin and osteocalcin of mouse bone marrow stem cells
  • Increased osteocalcin secretion and alkaline phosphatase (AP) activities of cell line H1 human embryonic stem cells
  • Maintained P75NTR gene activity of rat olfactory ensheathing cells
  • Upregulated NGF, BDNF, and PLP of rat olfactory ensheathing cells
  • Reduced alpha-SM actin mRNA, protein and promoter activity of rat primary cultured mesangial cells
  • Increased post-transfection gene expression level and expression time of NIH 3T3 fibroblasts
  • Did not affect expression of MGMT and glutathione-S-transferase (GST) detoxifying enzymes of human glioblastoma cells
  • Increased expression of metabolic (transketolase, triosephosphate isomerase, pyruvate kinase M1/M2, alpha enolase, and phosphoglycerate mutase-1) polypeptides of human neuroblastoma cells
  • Increased expression of cell stress response (heat shock proteins (HSP) 90, 70, and 60; antioxidant, thioredoxin) polypeptides of human neuroblastoma cells
  • Increased expression of cell structure (septin 2, adenyl cyclase-associated protein-1), tubulin beta-2 chain, actin, translationally controlled tumor protein and cofilin) polypeptides of human neuroblastoma cells
  • Increased expression of signal transduction (peptidyl prolyl cis/trans isomerase A), biosynthetic (phosphoserine aminotransferase) polypeptides of human neuroblastoma cells
  • Increased expression of transport (cellular retinoic acid binding protein 1) polypeptides of human neuroblastoma cells
  • Increased E-cadherin, CD44v6, VEGF, KDR, endostatin, and cytochrome-c levels of human hepatoma cell line HepG2
  • Enabled expression of chondrocyte-associated antigen of rat chondrocytes
  • Increased mRNA levels of collagen type II and aggrecan of rat chondrocytes
  • Did not change collagen type I mRNA of rat chondrocytes
  • Decreased versican mRNA level of rat chondrocytes
  • Decreased alpha-SMA levels and pFAKY(397)/totalFAK ratios of normal and tumor ovarian fibroblasts
  • Upregulated gene expression of collagen Type I and III, and tropoelastin and integrins alpha 1, beta 1 and beta 3 of rat vascular smooth muscle cells
  • Decreased alpha-smooth muscle actin and vimentin expression of airway smooth muscle cells
  • Increased active matrix metalloproteinase (MMP)-2 of airway smooth muscle cells
  • Upregulated Phase I and Phase II xenobiotic drug metabolism genes, as well as hepatocyte-specific transcripts HNF4alpha, Albumin, TTR and alpha1AT of Huh7 hepatoma cells
  • Increased expression of HCV receptors, cell adhesion and tight junction markers (CD81, scavenger receptor class B member 1, claudin-1, occludin, ZO-1, beta-Catenin and E-Cadherin) of Huh7 hepatoma cells
  • Increased and quickened upregulated mRNA expression levels of integrin beta 1 and ILK of rat skeletal myoblasts
  • Increased production of adipogenic markers leptin, adiponectin, glycerol of human preadipocytes
  • Enhanced secretion of leptin and caused differences in laminin expression (mRNA and protein level) of differentiated mouse 3T3-L1 preadipocytes
  • Did not change mRNA expression of PPARgamma and Glut-4 of differentiated mouse 3T3-L1 preadipocytes
  • Increased relative expression of osteogenic marker CBFA-1 of human adipose-derived stem cells
  • Increased absolute expression of osteogenic markers alkaline phosphatase, osteonectin, osteopontin, Collagen 1, and JNK2 of human adipose-derived stem cells
  • Increased collagen synthesis on human MCF-7 cells
  • Increased level of BMP-2 expression and osteocalcin content of rat mesenchymal stem cells
  • Increased osteocalcin content of rat bone marrow stem cells
  • Decreased growth in mRNA expression of type I collagen over time of rabbit bone marrow stem cells
  • Did not change absolute expression of type I collagen or chordin of rabbit bone marrow stem cells
  • Increased mRNA expression of vascular endothelial growth factor (VEGF) and osteopontin of rabbit bone marrow stem cells
  • Increased leptin secretion of mouse preadipocyte 3T3-L1 cells
  • Increased activated paxillin, FAK, AKT, C-Src, and ERK1/2 antibodies of mouse MC3T3-E1 preosteoblasts
  • Src induced changes in expression of over 100 genes in MDCK cells
  • Upregulation of OPN and FGF-2 and downregulation of collagen 1 gene expression in 3D osteoblast cultures under static conditions
  • Downregulation of OPN and FGF-2 gene expression in 3D osteoblast cultures under shear stress conditions
  • Induced a more tumor-related expression profile in Hodgkin lymphoma cell lines
  • Upregulated genes involved in immune response and apoptosis and downregulated genes involved in cell division in Hodgkin lymphoma cell lines
  • Correlating gene expression profiles and breast cancer tumor cell morphology
  • Increased expression of mesenchymal genes including vimentin, n-cadherin, zeb1, snail and slug as well as pro-metastatic genes RhoC, Tenascin C and MTA1 in 293T cells
  • Silenced GFP expression through shRNA plasmid delivered by PEI-coated SPMNs
  • Suppressed genes associated with cell adhesion and cell cycling in MC3T3 E1 osteoblast-like cells
  • Differential protein expression relevant to tumor cell proliferation, survival, and chemoresistance and thus may reveal novel targets for cancer therapy
  • Regulation of MT1-MMP during matrix invasion and lumen formation of endothelial cells
  • Co-association of Cdc42, Par6b, Par3, Jam-B and Jam-C, MT1-MMP, and integrin {alpha}2{beta}1 in the endothelial cell (EC) lumen signaling complex to control human EC tubulogenesis
  • Reduced expression of SM-MHC, smoothelin, and calponin in vSMCs through HSPG-mediated ERK1/2 activation in response to fluid flow
  • Increased expression of a-SMA and SM22 in vSMCs through HSPG-mediated ERK1/2 activation in response to fluid flow
  • More closely recapitulates in vivo protein expression of N-cadherin in human glioblastoma cells
  • Regain ability to assemble fibronectin matrix correlating with decreased Raf-1 protein expression
  • Upregulation of genes characteristic of retinal neurons following growth factor stimulation
  • Upregulation of antioxidant proteins and proteins involved in dynamic remodeling of the actin cyctoskeleton in MDCK cells
  • Decresed expression of several mitochondrial proteins
  • Increased expression of proteins in anaerobic glycolysis
  • Upregulation of creatine kinase in hypoxic conditions
  • Increased expression of adhesion protein transcripts in C2C12 cells
  • Resembled gene expression profiles from spontaneous human cancers
  • Decreased responsiveness to intracellular Ca(2+) of SH-SY5Y human neuroblastoma cells on collagen constructs
  • Eliminated anti-proliferative effect of TGF-beta 1 of human endothelial cells
  • Eliminated requirement for supporting substrates poly-D-lysine and laminin of rat hippocampal neurons and astrocytes
  • Slowed response to K(+) depolarization of SH-SY5Y human neuroblastoma cells on Cytodex 3 microbead array
  • Did not affect dose response to radiation of human MCF-10A cells
  • Assembly of tissue-like aggregates with high mass transport in a low shear microgravity model using a Rotating Wall Vessel (RWV)
  • Enhanced cell proliferation and cell morphology of osteoblasts exposed to fluid shear stress
  • Reduced expression of SM-MHC, smoothelin, and calponin in vSMCs through HSPG-mediated ERK1/2 activation in response to fluid flow
  • Increased expression of a-SMA and SM22 in vSMCs through HSPG-mediated ERK1/2 activation in response to fluid flow
  • Relationship between cell morphology and cellular radiosensitivity based on chromatin organization
  • Upregulation of stem cell survival signaling including beta-catenin, Notch1 and Survivin in response to radiation in 293T cells
  • Significantly enhanced function of human HepG2 liver cells
  • Enhanced performance and functional activity of human HepG2 liver cells
  • Induced well developed cell-cell contacts, a distinct endoplasmatic reticulum, and marked Golgi apparatus in human bladder carcinoma cell line RT112 cells
  • Enabled colocalization of alpha-6 integrin and alpha-V integrin of mesenchymal stem cells
  • Did not alter resting membrane potential of SH-SY5Y human neuroblastoma cells on collagen constructs
  • Did not change triglyceride content of differentiated mouse 3T3-L1 preadipocytes
  • Increased alkaline phosphatase activity of rat mesenchymal stem cells
  • Increased alkaline phosphatase (ALP) activity of rat bone marrow stem cells
  • Created dose dependent effect of MMP inhibitor (GM6001) of mouse osteoblast cell line MC3T3-E1 and primary mouse calvarial osteoblasts
  • Decreased tyrosine phosphorylation of focal adhesion kinase (FAK) of vascular smooth muscle cells
  • Decreased metabolic rates of human trophoblast cells
  • Altered cell cycle progression of human trophoblast ED27 cells
  • Decreased metabolic rate of human arachnoidal cells
  • Did not affect Lucifer Yellow permeability or hydraulic conductivity of human arachnoidal cells
  • Did not affect plating efficiency of rat anterior tibialis muscle cells
  • Altered the synergistic effects of extracellular matrix mechanics and RhoA activity of human smooth muscle cells
  • Enabled calcification of mouse bone marrow stem cells
  • Enabled in unison contracting structurally organized cell-matrix cardiac constructs of neonatal rat heart ventricular cells
  • Altered resting membrane potentials of SH-SY5Y human neuroblastoma cells on Cytodex 3 microbead array
  • Slowed development of VGCC function of SH-SY5Y human neuroblastoma cells on Cytodex 3 microbead array
  • Did not affect methylation of O-6-methylguanine-DNA-methyltransferase (MGMT) promoter of human glioblastoma cells
  • Increased fraction of cells in G I phase of human hepatoma cell line HepG2
  • Caused formation of HER2 homodimers, as opposed to heterodimers of human epidermal growth factor receptor (HER2) over-expressing cancer cells
  • Increased polarization of Huh7 hepatoma cells
  • Enabled Elucidation of Prostate Cancer Biomarkers of human prostatic epithelial cells
  • Use of human platelet lysate (PL) as a serum substitute for stromal vascular fraction (SVF) cells
  • Regulation of cell speed and persistence by affecting protrusion activity and matrix deformation through focal adhesion proteins in matrix-embedded cells
  • Possessed greater viability and were less susceptible to cell death at higher levels of methotrexate on human HepG2 liver cells
  • Decreased apoptosis and cell death even under suboptimal conditions (e.g., nutrient depletion) of human osteosarcoma cells
  • Increased cell death in response to loading / mechanical insult of primary rat cortical neurons
  • Increased attachment and proliferation of mesenchymal stem cells
  • Increased cell spreading and proliferation of mouse MC3T3-E1 preosteoblasts
  • Caused differences in growth kinetics of cow articular cartilage cells
  • Increased long term cell growth of mouse MC3T3-E1 cells
  • Increased proliferation of total cells and progenitors without exogenous cytokines of human cord blood cells
  • Decreased apoptotic rate of hybridoma cells
  • Decreased proliferation activities of human trophoblast cells
  • Decreased growth rate of human arachnoidal cells
  • Decreased mortality rate of rat hippocampal neurons and astrocytes
  • Supported cell survival under conditions of calcium and nutrients deprivation of rat hippocampal neurons and astrocytes
  • Decreased cell viability and degree of spreading of human smooth muscle cells
  • Decreased apoptosis of rat olfactory ensheathing cells
  • Decreased apoptosis of human hepatoma cell line HepG2

Ref:  http://3dcellculture.com/Just_The_Facts_3D_vs_2D_Cell_Culture

According to PubMed, the published number about 3D cell culture have grown dramatically in past 25 years.

As 3D culture systems become more mature and relevant to human and animal physiology, the ability to design and develop co-cultures becomes possible as does the ability to integrate stem cells. The primary objectives for developing 3D cell culture systems vary widely and range from engineering tissues for clinical delivery through to the development of models for drug screening.

How does VitroGel bridge in vitro and in vivo studies?

Mixing VitroGel solution and the cell culture media at proper ratio (We recommend VitroGel: cell media=4:1 v/v), the final hydrogel is injectable through a syringe. This injectable property give a further advantage of 3D cell culture system by creating the possibility for in vivo study with the same platform system. Cells or molecular compounds which have been successfully tested in vitro 3D cell culture system can be injected into animals using the same system for in vivo study. In combine with existing/novel analysis technology, we hope our technology can shorten the R&D time of drug discovery, cell therapy and tissue engineering studies and benefit pharmaceutical and clinical industries.