Three-dimensional cell culture systems as an in vitro platform for cancer and stem cell modeling

Three-dimensional(3D)culture systems are becoming increasingly popular due to their ability to mimic tissue-like structures more effectively than the monolayer cultures.In cancer and stem cell research,the natural cell characteristics and architectures are closely mimicked by the 3D cell models.Thus,the 3D cell cultures are promising and suitable systems for various proposes,ranging from disease modeling to drug target identification as well as potential therapeutic substances that may transform our lives.This review provides a comprehensive compendium of recent advancements in culturing cells,in particular cancer and stem cells,using 3D culture techniques.The major approaches highlighted here include cell spheroids,hydrogel embedding,bioreactors,scaffolds,and bioprinting.In addition,the progress of employing 3D cell culture systems as a platform for cancer and stem cell research was addressed,and the prominent studies of 3D cell culture systems were discussed.

The Interrelationship between Culture and Capital Structure:Evidence from International Retailers

The present study investigates the influence of cultural factors in 2022 on the capital structure of global retailers.There are sixteen retailers from eight countries in the sample.In recent times,numerous academicians have taken an interest in examining the capital structure and business model of retailers,owing to their swift and consistent growth.However,the fact that the majority of research originates from the retailers’host country gives rise to debate regarding the applicability of the capital structure of said retailers to countries with distinct cultural environments.Consequently,academics have begun to investigate whether the capital structure of multinational retailers is impacted by the diversity of national cultures.

Study of the geomagnetic field's regional gradients in Chinese continent using three-dimensional surface Spline model

We combined domestic ground-based and satellite magnetic measurements to create a regional three-dimensional surface Spline(3DSS)gradient model of the main geomagnetic field over the Chinese continent.To improve the precision of the model,we considered the data gap between the ground and satellite data.We compared and analyzed the results of the Taylor polynomial,surface Spline,and CHAOS-6(the CHAMP,?rsted and SAC-C model of Earth’s magnetic field)gradient models.Results showed that the gradients in the south-north and east-west directions of the four models were consistent.The 3DSS model was able to express not only gradients at different altitudes,but also average gradients inside the research area.The two Spline models were able to capture more information on gradient anomalies than were the fitted models.Strong local anomalies were observed in northern Xinjiang,Beijing,and the junction area between Jiangsu and Zhejiang,and the total intensity F decreased whereas the altitude increased.The gradient decreased by 21.69%in the south-north direction and increased by 11.78%in the east-west direction.In addition,the altitude gradient turned from negative to positive while the altitude increased.The Spline model and the two fitted models differed mainly in the field sources they expressed and the modeling theory.

Can preoperative planning using IRIS™three-dimensional anatomical virtual models predict operative findings during robot-assisted partial nephrectomy?

Objective To evaluate the predictive validity of IRIS™(Intuitive Surgical®,Sunnyvale,CA,USA)as a planning tool for robot-assisted partial nephrectomy(RAPN)by assessing the degree of overlap with intraoperative execution.Methods Thirty-one patients scheduled for RAPN by four experienced urologists were enrolled in a prospective study.Prior to surgery,urologists reviewed the IRIS™three-dimensional model on an iphone Operating System(iOS)app and completed a questionnaire outlining their surgical plan including surgical approach,and ischemia technique as well as confidence in executing this plan.Postoperatively,questionnaires assessing the procedural approach,clinical utility,efficiency,and effectiveness of IRIS™were completed.The degree of overlap between the preoperative and intraoperative questionnaires and between the planned approach and actual execution of the procedure was analyzed.Questionnaires were answered on a 5-point Likert scale and scores of 4 or greater were considered positive.Results Mean age was 65.1 years with a mean tumor size of 27.7 mm(interquartile range 17.5-44.0 mm).Hilar tumors consisted of 32.3%;48.4%of patients had R.E.N.A.L.nephrometry scores of 7-9.On preoperative questionnaires,the surgeons reported that in 67.7%cases they were confident that they can perform the procedure successfully,and on intraoperative questionnaires,the surgeons reported that in 96.8%cases IRIS™helped achieve good spatial sensation of the anatomy.There was a high degree of overlap between preoperative and intraoperative questionnaires for the surgical approach,interpreting anatomical details and clinical utility.When comparing plans for selective or off-clamp,the preoperative plan was executed in 90.0%of cases intraoperatively.Conclusion A high degree of overlap between the preoperative surgical approach and intraoperative RAPN execution was found using IRIS™.This is the first study to evaluate the predictive accuracy of IRIS™during RAPN by comparing preoperative plan and intraoperative execution.

Three-Dimensional Analytical Modeling of Axial-Flux Permanent Magnet Drivers

In this paper, the axial-flux permanent magnet driver is modeledand analyzed in a simple and novel way under three-dimensional cylindricalcoordinates. The inherent three-dimensional characteristics of the deviceare comprehensively considered, and the governing equations are solved bysimplifying the boundary conditions. The axial magnetization of the sectorshapedpermanent magnets is accurately described in an algebraic form bythe parameters, which makes the physical meaning more explicit than thepurely mathematical expression in general series forms. The parameters of theBessel function are determined simply and the magnetic field distribution ofpermanent magnets and the air-gap is solved. Furthermore, the field solutionsare completely analytical, which provides convenience and satisfactoryaccuracy for modeling a series of electromagnetic performance parameters,such as the axial electromagnetic force density, axial electromagnetic force,and electromagnetic torque. The correctness and accuracy of the analyticalmodels are fully verified by three-dimensional finite element simulations and a15 kW prototype and the results of calculations, simulations, and experimentsunder three methods are highly consistent. The influence of several designparameters on magnetic field distribution and performance is studied and discussed.The results indicate that the modeling method proposed in this papercan calculate the magnetic field distribution and performance accurately andrapidly, which affords an important reference for the design and optimizationof axial-flux permanent magnet drivers.

Isolation and Identification of Cancer Stem Cells from Human Osteosarcom by Serum-free Three-dimensional Culture Combined with Anticancer Drugs

The cancer stem cells(CSCs)from human osteosarcoma by serum-free three-dimensional culture combined with anticancer drugs were isolated and identified.The primary cells derived from human osteosarcoma were digested by trypsin to prepare a single-cell suspension,and mixed homogeneously into 1.2% alginate gel.Single-cell alginate gel was cultured with serum-free DMEM/F12 medium.Epirubicin(0.8μg/mL)was added to the medium to enrich CSCs.After cultured conventionally for 7 to 10 days,most of cells suspended in ...

Simplified three-dimensional culture system for long-term expansion of embryonic stem cells

AIM: To devise a simplified and efficient method for long-term culture and maintenance of embryonic stem cells requiring less frequent passaging. METHODS: Mouse embryonic stem cells(ESCs) labeled with enhanced yellow fluorescent protein were cultured in three-dimensional(3-D) self-assembling scaffolds and compared with traditional two-dimentional(2-D) culture techniques requiring mouse embryonic fibroblast feeder layers or leukemia inhibitory factor. 3-D scaffolds encapsulating ESCs were prepared by mixing ESCs with polyethylene glycol tetra-acrylate(PEG-4-Acr) and thiolfunctionalized dextran(Dex-SH). Distribution of ESCs in 3-D was monitored by confocal microscopy. Viability and proliferation of encapsulated cells during long-term culture were determined by propidium iodide as well as direct cell counts and PrestoB lue(PB) assays. Genetic expression of pluripotency markers(Oct4, Nanog, Klf4, and Sox2) in ESCs grown under 2-D and 3-D cultureconditions was examined by quantitative real-time polymerase chain reaction. Protein expression of selected stemness markers was determined by two different methods, immunofluorescence staining(Oct4 and Nanog) and western blot analysis(Oct4, Nanog, and Klf4). Pluripotency of 3-D scaffold grown ESCs was analyzed by in vivo teratoma assay and in vitro differentiation via embryoid bodies into cells of all three germ layers. RESULTS: Self-assembling scaffolds encapsulating ESCs for 3-D culture without the loss of cell viability were prepared by mixing PEG-4-Acr and Dex-SH(1:1 v/v) to a final concentration of 5%(w/v). Scaffold integrity was dependent on the degree of thiol substitution of Dex-SH and cell concentration. Scaffolds prepared using Dex-SH with 7.5% and 33% thiol substitution and incubated in culture medium maintained their integrity for 11 and 13 d without cells and 22 ± 5 d and 37 ± 5 d with cells, respectively. ESCs formed compact colonies, which progressively increased in size over time due to cell proliferation as determined by confocal microscopy and PB staining. 3-D scaffold cultured ESCs expressed significantly higher levels(P < 0.01) of Oct4, Nanog, and Kl4, showing a 2.8, 3.0 and 1.8 fold increase, respectively, in comparison to 2-D grown cells. A similar increase in the protein expression levels of Oct4, Nanog, and Klf4 was observed in 3-D grown ESCs. However, when 3-D cultured ESCs were subsequently passaged in 2-D culture conditions, the level of these pluripotent markers was reduced to normal levels. 3-D grown ESCs produced teratomas and yielded cells of all three germ layers, expressing brachyury(mesoderm), NCAM(ectoderm), and GATA4(endoderm) markers. Furthermore, these cells differentiated into osteogenic, chondrogenic, myogenic, and neural lineages expressing Col1, Col2, Myog, and Nestin, respectively. CONCLUSION: This novel 3-D culture system demonstrated long-term maintenance of mouse ESCs without the routine passaging and manipulation necessary for traditional 2-D cell propagation.

Microfluidic three-dimensional cell culture of stem cells for high-throughput analysis

Although the recent advances in stem cell engineering have gained a great deal of attention due to their high potential in clinical research,the applicability of stem cells for preclinical screening in the drug discovery process is still challenging due to difficulties in controlling the stem cell microenvironment and the limited availability of high-throughput systems.Recently,researchers have been actively developing and evaluating three-dimensional(3D)cell culture-based platforms using microfluidic technologies,such as organ-on-a-chip and organoid-on-a-chip platforms,and they have achieved promising breakthroughs in stem cell engineering.In this review,we start with a comprehensive discussion on the importance of microfluidic 3D cell culture techniques in stem cell research and their technical strategies in the field of drug discovery.In a subsequent section,we discuss microfluidic 3D cell culture techniques for high-throughput analysis for use in stem cell research.In addition,some potential and practical applications of organ-on-a-chip or organoid-on-a-chip platforms using stem cells as drug screening and disease models are highlighted.

Optimization of Three-Dimensional Culture Conditions of HepG2 Cells with Response Surface Methodology Based on the VitroGel System

Objective This study optimizes three-dimensional(3D) culture conditions of HepG2 using response surface methodology(RSM) based on the VitroGel system to facilitate the cell model in vitro for liver tissues.Method HepG2 cell was 3D cultured on the VitroGel system.Cell viability was detected using Cell Counting Kit-8(CCK-8) assay of HepG2 lived cell numbers.The proliferation of HepG2 cell and clustering performance was measured via fluorescence staining test.Albumin concentration in the culture medium supernatant as an index of HepG2 cell biological function was measured with ELISA kit.Independent factor tests were conducted with three key factors:inoculated cell concentration,cultured time,and dilution degree of the hydrogel.The preliminary results of independent factor tests were used to determine the levels of factors for RSM.Result The selected optimal culture conditions are as follows:concentration of inoculated cells was4.44 × 10^(5)/mL,culture time was 4.86 days,and hydrogel dilution degree was 1:2.23.The result shows that under optimal conditions,the predicted optical density(OD) value of cell viability was 3.10 and measured 2.978 with a relative error of 3.94%.Conclusion This study serves as a reference for the 3D HepG2 culture and constructs liver tissues in vitro.Additionally,it provides the foundation for repeated dose high-throughput toxicity studies and other scientific research work.

Irradiation Response of Adipose-derived Stem Cells under Three-dimensional Culture Condition

Objective Adipose tissue distributes widely in human body. The irradiation response of the adipose cells in vivo remains to be investigated. In this study we investigated irradiation response of adipose-derived stem cells (ASCs) under three-dimensional culture condition. Methods ASCs were isolated and cultured in low attachment dishes to form three-dimensional (3D) spheres in vitro. The neuronal differentiation potential and stem-liked characteristics was monitored by using immunofluoresence staining and flow cytometry in monolayer and 3D culture. To investigate the irradiation sensitivity of 3D sphere culture, the fraction of colony survival and micronucleus were detected in monolayer and 3D culture. Soft agar assays were performed for measuring malignant transformation for the irradiated monolayer and 3D culture. Results The 3D cultured ASCs had higher differentiation potential and an higher stem-like cell percentage. The 3D cultures were more radioresistant after either high linear energy transfer (LET) carbon ion beam or low LET X-ray irradiation compared with the monolayer cell. The ASCs’ potential of cellular transformation was lower after irradiation by soft agar assay. Conclusion These findings suggest that adipose tissue cell are relatively genomic stable and resistant to genotoxic stress.

Inhibition of mammalian target of rapamycin induces phenotypic reversion in three-dimensional cultures of malignant breast epithelial cells

Inhibition of mammalian target of rapamycin (m- TOR) is a potential method for cancer treatment. Effects of rapamycin (RAP) on the reversion of malignant breast epithelial cells were investigated on three-dimensional (3D) basement membrane extract (BME) cultures. Through continuous exposure to 20 nM of RAP, cell colony size was significantly reduced in 3D BME cultures of malignant breast epithelial cells, while normal cell colony size appeared unaffected. In unfixed 3D BME cultures of normal and RAP-treated malignant breast epithelial cells, the presence of luminal cell death was confirmed by ethidium bromide and propidium iodide labeling. Increased structural organization was observed by im- munofluorescence staining of F-actin and β-catenin in RAP-treated malignant breast epithelial cells. In monolayer cultures of normal and malignant breast epithelial cells, continuous exposure to 20 nM of RAP increased caspase 3/7 activity and decreased proliferation. Reverse transcriptase polymerase ch- ain reaction (RT-PCR) array analysis indicated a fold increase in the expression of a number of proteins related to polarity, cell-cell adhesion, and cell-matrix adhesion in the presence of RAP. Our data showed that phenotypic reversion of malignancy can be ach- ieved through RAP exposure on 3D BME cultures. This 3D BME culture system will provide correct microenvironments for observing the effects of other mTOR inhibitors on phenotypic reversion of malignant breast epithelial cells.

Construction of Intelligent Recommendation Retrieval Model of FuJian Intangible Cultural Heritage Digital Archives Resources

In order to improve the consistency between the recommended retrieval results and user needs,improve the recommendation efficiency,and reduce the average absolute deviation of resource retrieval,a design method of intelligent recommendation retrieval model for Fujian intangible cultural heritage digital archive resources based on knowledge atlas is proposed.The TG-LDA(Tag-granularity LDA)model is proposed on the basis of the standard LDA(Linear Discriminant Analysis)model.The model is used to mine archive resource topics.The Pearson correlation coefficient is used to measure the relevance between topics.Based on the measurement results,the FastText deep learning model is used to achieve archive resource classification.According to the classification results,TF-IDF(term frequency–inverse document frequency)algorithm is used to calculate the weight of resource retrieval keywords to achieve resource retrieval,and a recommendation model of intangible cultural heritage digital archives resources is built through the knowledge map to achieve comprehensive and personalized recommendation of resources.The experimental results show that the recommendation and retrieval results of the proposed method are more in line with users’needs,can provide users with personalized digital archive resources,and the average absolute deviation of resource retrieval is low,the recommendation efficiency is high,and the utilization effect of archive resources is effectively improved.

Shear behavior of intact granite under thermo-mechanical coupling and three-dimensional morphology of shear-formed fractures

The shear failure of intact rock under thermo-mechanical(TM)coupling conditions is common,such as in enhanced geothermal mining and deep mine construction.Under the effect of a continuous engineering disturbance,shear-formed fractures are prone to secondary instability,posing a severe threat to deep engineering.Although numerous studies regarding three-dimensional(3D)morphologies of fracture surfaces have been conducted,the understanding of shear-formed fractures under TM coupling conditions is limited.In this study,direct shear tests of intact granite under various TM coupling conditions were conducted,followed by 3D laser scanning tests of shear-formed fractures.Test results demonstrated that the peak shear strength of intact granite is positively correlated with the normal stress,whereas it is negatively correlated with the temperature.The internal friction angle and cohesion of intact granite significantly decrease with an increase in the temperature.The anisotropy,roughness value,and height of the asperities on the fracture surfaces are reduced as the normal stress increases,whereas their variation trends are the opposite as the temperature increases.The macroscopic failure mode of intact granite under TM coupling conditions is dominated by mixed tensileeshear and shear failures.As the normal stress increases,intragranular fractures are developed ranging from a local to a global distribution,and the macroscopic failure mode of intact granite changes from mixed tensileeshear to shear failure.Finally,3D morphological characteristics of the asperities on the shear-formed fracture surfaces were analyzed,and a quadrangular pyramid conceptual model representing these asperities was proposed and sufficiently verified.

A three-dimensional semi-implicit unstructured grid finite volume ocean model

A new three-dimensional semi-implicit finite-volume ocean model has been developed for simulating the coastal ocean circulation, which is based on the staggered C-unstructured non-orthogonal grid in the hor- izontal direction and z-level grid in the vertical direction. The three-dimensional model is discretized by the semi-implicit finite-volume method, in that the free-surface and the vertical diffusion are semi-implicit, thereby removing stability limitations associated with the surface gravity wave and vertical diffusion terms. The remaining terms in the momentum equations are discretized explicitly by an integral method. The partial cell method is used for resolving topography, which enables the model to better represent irregular topography. The model has been tested against analytical cases for wind and tidal oscillation circulation, and is applied to simulating the tidal flow in the Bohal Sea. The results are in good agreement both with the analytical solutions and measurement results.

Three-Dimensional Tidal Model and Its Application to Numerical Simulation of Water Quality in Coastal Waters

The turbulence mechanism plays an important part in the mixing process and momentum transfer of turbulence. A three-dimensional Prandtl mixing length tidal model has been developed to simulate tidal flows and water quality. The eddy viscosities and diffusivities are computed from the Prandtl mixing length model. In order to model the water quality of an estuary or coastal area many interdependent processes need to be simulated. These may be conveniently separated into three main groups: transport and mixing processes, biochemical interaction of water quality variables and the utilization and re-cycling of nutrients by living matter. The model simulates full oxygen and nutrient balance, primary productivity and the transport, reaction mechanism and fate of pollutants over tidal time-scales. The model is applied to numerical simulation of tidal flows and water quality in Dalian Bay. The model has been calibrated against a limited data set of historical water quality observations and in general demonstrates excellent agreement with all available data.

The study on three-dimensional numerical model and fronts of the Jiulong Estuary and the Xiamen Bay

Applying the methods of on-site observation and dynamic model, the research on the fronts at the Jiulong Estuary has been carried out, during which spatial and temporal distribution, dynamic characteristics and formation mechanism of salinity fronts are analyzed and discussed. The research shows that the estuarine fronts mainly lie in the area from the Jiyu Islet to the Haimen Island, outside of Yuweizai to Hulishan cross-section, the near coast of Yuweizai and the south of the Songyu-Gulangyu Channel. The fronts in the former two regions are formed directly by plume, while the one near the coast of Yuweizai is a tidal intrusion front caused by flood current and the one at the south of the Songyu-Gulangyu Channel is the result of current shear transformation. Under normal circumstances, fresh water of the Jiulong River mainly influences the inside of the Xiamen Bay, and when it is in typhoon seasons, plume front can affect the Taiwan Strait and has an effect on the biogeochemical Drocesses in the strait.

A Novel Three-Dimensional Mouse Embryonic Implantation Model In Vitro

To regenerate three-dimensional endometrium in vitro as a novel model for studying the mechanism of implantation of embryos, the luminal epithelial cells and stromal cells of the rabbit uterus were separated and cultured in vitro. The type Ⅰ mouse tail collagen was used as scaffolding material. The stromal cells were inoculated in the type I mouse tail collagen, and the luminal epithelial cells were inoculated on the type i mouse tall collagen to regenerate the endometrium in vitro. The regenerated endometrium was cultured in DMEM-F/12 media containing 100 nmol L^-1 progesterone, 10 nM β-estradiol, and 10% fetal bovine serum （FBS） for 3 d. The media were then replaced with CZB containing 100 nM progesterone, 10 nmol L-1 β-estradiol, and 10% FBS, and the mouse blastulas were co-cultured with it. The results of scanning electronic micrography showed that the epithelial cells on the surface of the reconstructed endometrium were covered with numerous slender microvilli and some epithelial cells protruded pinopodes. After culturing for 12 h with the mouse blastula, the shedding, attachment, and implantation of the blastula were observed. The blastula can escape from zona pellucida and attach to the three-dimensional endometrium and is then implanted into it. This study showed that the reconstructed three-dimensional endometrium can serve as a robust embryo implantation model in vitro.

A Three-Dimensional Water Quality Model and Its Application to Jiaozhou Bay, China

A three-dimensional coupled physical and water quality model was developed and applied to the Jiaozhou Bay to study water quality involving nutrients, biochemical oxygen demand, dissolved oxygen, and phytoplankton that are closely related to eutrophication process. The physical model is a modified ECOM-si version with inclusion of flooding/draining processes over the intertidal zone. The water quality model is based on WASP5 which quantifies processes governing internal nutrients cycling, dissolved oxygen balance and phytoplankton growth. The model was used to simulate the spatial distribution and the temporal variation of water quality in the Jiaozhou Bay for the period of May 2005 to May 2006. In addition, the effect of reduction of riverine nutrients load was simulated and evaluated. The simulated results show that under the influence of nutrients discharged from river, the concentrations of nutrients and phytoplankton were higher in the northwest and northeast of the bay, and decreased from the inner bay to the outer. Affected by strong tidal mixing, the concentrations of all state variables were vertically homogeneous except in the deeper regions where a small gradient was found. Obvious seasonal variation of phytoplankton biomass was found, which exhibited two peaks in March and July, respectively. The variation of riverine waste loads had remarkable impact on nutrients concentration in coastal areas, but slightly altered the distribution in the center of the bay.

Construction and Analysis of Three-dimensional Graphic Model of Single-chain Fv Derived from an Anti-human Placental Acidic Isoferritin Monoclonal Antibody by Computer

Summary: A three-dimensional (3D) graphic model of a single-chain Fv (scFv) which was derived from an anti-human placental acidic isoferritin (PAF) monoclonal antibody (MAb) was construct- ed by a homologous protein-predicting computer algorithm on Silicon graphic computer station. The structure, surface static electricity and hydrophobicity of scFv were investigated. Computer graphic modelling indicated that all regions of scFv including the linker, variable regions of the heavy (VH) and light (VL) chains were suitable. The VH region and the VL region were involved in composing the 'hydrophobic pocket'. The linker was drifted away VH and VL regions. The complementarity determining regions (CDRs) of VH and VL regions surrounded the 'hydrophobic pocket'. This study provides a theory basis for improving antibody affinity, investigating antibody structure and analyzing the functions of VH and VL regions in antibody activity.

Three-dimensional fuzzy logic system for process modeling and control

The traditional fuzzy logic system （FLS） can only model and control the process in two-dimensional nature. Many of real-world systems are of multidimensional features, such as, thermal and fluid processes with spatiotemporal dynamics, biological systems, or decision-making processes that contain stochastic and imprecise uncertainties. These types of systems are difficult for the traditional FLS to model and control because they require a third dimension for spatial or probabilistic information. The type-2 fuzzy set provides the possibility to develop a three-dimensional fuzzy logic system for modeling and controlling these processes in three-dimensional nature.