Establishment of human cerebral organoid systems to model early neural development and assess the central neurotoxicity of environmental toxins

Human brain development is a complex process,and animal models often have significant limitations.To address this,researchers have developed pluripotent stem cell-derived three-dimensional structures,known as brain-like organoids,to more accurately model early human brain development and disease.To enable more consistent and intuitive reproduction of early brain development,in this study,we incorporated forebrain organoid culture technology into the traditional unguided method of brain organoid culture.This involved embedding organoids in matrigel for only 7 days during the rapid expansion phase of the neural epithelium and then removing them from the matrigel for further cultivation,resulting in a new type of human brain organoid system.This cerebral organoid system replicated the temporospatial characteristics of early human brain development,including neuroepithelium derivation,neural progenitor cell production and maintenance,neuron differentiation and migration,and cortical layer patterning and formation,providing more consistent and reproducible organoids for developmental modeling and toxicology testing.As a proof of concept,we applied the heavy metal cadmium to this newly improved organoid system to test whether it could be used to evaluate the neurotoxicity of environmental toxins.Brain organoids exposed to cadmium for 7 or 14 days manifested severe damage and abnormalities in their neurodevelopmental patterns,including bursts of cortical cell death and premature differentiation.Cadmium exposure caused progressive depletion of neural progenitor cells and loss of organoid integrity,accompanied by compensatory cell proliferation at ectopic locations.The convenience,flexibility,and controllability of this newly developed organoid platform make it a powerful and affordable alternative to animal models for use in neurodevelopmental,neurological,and neurotoxicological studies.

The Protective Effects of Flavonoids from Scutellaria Baicalensis Georgi Stems and Leaves on Oligodendrocyte Damage Induced by Aβ1-42

Aim: This study aimed to investigate the protective effects of flavonoids from the stem and leaves of Scutellaria baicalensis Georgi (SSFs) against Aβ1-42-induced oligodendrocytes (OL) damage. Methods: Immunofluorescence was used for the detection of myelin-associated glycoprotein (MAG), a characteristic protein of rat oligodendrocytes (OLN-93 cells). To evaluate the potential protective effects of SSFs on OLN-93 cells injured by Aβ1-42, an injury model was established by subjecting OLN-93 cells to Aβ1-42 exposed. Cell morphology was examined using an inverted microscope, while cell viability was assessed using the colorimetric method of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Additionally, lactate dehydrogenase (LDH) was measured using the pyruvic acid reduction assay. The Ginkgo biloba leaf extract (GBE) injection was used as a positive control. Results: A total of >95% of the MAG immunofluorescence-positive cells were identified as oligodendrocytes. Gradually increasing concentrations of SSFs impaired the cells, and the maximum nondetrimental dose for OLN-93 cells was 75 mg/L. This study assessed the effects of SSFs on OLN-93 cells damaged by Aβ1-42. The results indicated that SSFs significantly improved OLN-93 cell morphological abnormal changes, increased the OLN-93 cell survival rate, and reduced LDH release. Conclusion: SSFs can alleviate Aβ1-42-induced damage of OL.

Mesenchymal stem cells-based drug delivery systems for diabetic foot ulcer:A review

The complication of diabetes,which is known as diabetic foot ulcer(DFU),is a significant concern due to its association with high rates of disability and mortality.It not only severely affects patients’quality of life,but also imposes a substantial burden on the healthcare system.In spite of efforts made in clinical practice,treating DFU remains a challenging task.While mesenchymal stem cell(MSC)therapy has been extensively studied in treating DFU,the current efficacy of DFU healing using this method is still inadequate.However,in recent years,several MSCs-based drug delivery systems have emerged,which have shown to increase the efficacy of MSC therapy,especially in treating DFU.This review summarized the application of diverse MSCs-based drug delivery systems in treating DFU and suggested potential prospects for the future research.

HPLC fingerprint of Nauclea officinalis stems in different harvesting period

Objective:To establish HPLC fingerprint of Nauclea officinalis stems in different harvesting periods,and analyze the effect of harvest time on the quality of the medicinal materials by combining with chemical pattern recognition.Methods:The analysis was performed on Sun Fire-C18(150 mm×4.6 mm,5μm)column.The mobile phase consisted of acetonitrile-0.1%phosphoric acid solution(gradient elution)at a flow rate of 1.0 mL/min.Results:The HPLC fingerprint of Nauclea officinalis stems was established and 12 common peaks were determined,and 3 chromatographic peaks were identified by comparison with the mixed references.There were some differences in the quality of Nauclea officinalis in different harvesting periods.The OPLS-DA analysis successfully predicted four main markers of quality difference.Conclusion:The established HPLC fingerprint could reflect the composition characteristics of Nauclea officinalis stems in different harvesting period,and the main markers that influence the composition difference of the stems could be used as key indicators for the quality control.

Study on the Action Mechanism of Isodon suzhouensis Stems in the Treatment of Prostate Cancer Based on UPLC-Q-TOF-MS/MS Combined with Network Pharmacology and Molecular Docking

[Objectives]This study was conducted to explore the action mechanism of chemical components from stems of Isodon suzhouensis on prostate cancer based on UPLC-Q-TOF-MS/MS technique,network pharmacology,and molecular docking validation.[Methods]UPLC-Q-TOF-MS/MS was applied to search active components in stems of I.suzhouensis,and SwissTargetPrediction and PharmaMapper databases were used to predict their potential targets.The GeneCards database was adopted to screen the targets of prostate cancer,and the targets of the active components and the targets of prostate cancer were intersected to obtain common targets.A protein-protein interaction network was constructed,and also,an"active component-target-disease-pathway"network was constructed using Metascape for gene ontology function and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis.Moreover,AutoDock was adopted to perform molecular docking verification on key active components and core targets.[Results]Sixty three active components were screened from stems of I.suzhouensis,and they had 233 common targets with prostate cancer,mainly involving biological processes such as protein phosphorylation,response to hormones,active regulation of cell movement,cell response to lipids,and response to oxidative stress,as well as pathways in cancer,prostate cancer,micro ribonucleic acids in cancer,p53 signaling pathway,NF-κB signaling pathway and other signaling pathways,exerting anti-tumor effects.Molecular docking showed that the top 5 key active components had good binding ability with the top 5 core targets.[Conclusions]Active components from stems of I.suzhouensis can exert therapeutic effects on prostate cancer through multiple targets and pathways.

Cemented versus uncemented stems for revision total hip replacement:A systematic review and meta-analysis

BACKGROUND The popularity of uncemented stems in revision total hip arthroplasty(THA)has increased in the last decade.AIM To assess the outcomes of both cemented and uncemented stems after mid-term follow up.METHODS This study was performed following both the Preferred Reporting Items for Systematic Reviews and Meta-analyses Statement and the Cochrane Handbook for systematic reviews and meta-analysis guidelines.Articles were chosen irrespective of country of origin or language utilized for the article full texts.This paper included studies that reviewed revision THA for both cemented or uncemented long stems.RESULTS Three eligible studies were included in the meta-analysis.Analysis was conducted by using Review Manager version 5.3.We computed the risk ratio as a measure of the treatment effect,taking into account heterogeneity.We used random-effect models.There were no significant differences found for intraoperative periprosthetic fractures[risk ratio(RR)=1.25;95%confidence interval(CI):0.29-5.32;P=0.76],aseptic loosening(RR=2.15,95%CI:0.81-5.70;P=0.13),dislocation rate(RR=0.50;95%CI:0.10-2.47;P=0.39),or infection rate(RR=0.99,95%CI:0.82-1.19;P=0.89),between the uncemented and the cemented long stems for revision THA after mid-term follow-up.CONCLUSION This study has evaluated the mid-term outcomes of both cemented and uncemented stems at first-time revision THA.In summary,there were no significant differences in the dislocation rate,aseptic loosening,intraoperative periprosthetic fracture and infection rate between the two cohorts.

Reporter gene systems for the identification and characterization of cancer stem cells

Cancer stem cells(CSCs)are tumor cells that share functional characteristics with normal and embryonic stem cells.CSCs have increased tumor-initiating capacity and metastatic potential and lower sensitivity to chemo-and radiotherapy,with important roles in tumor progression and the response to therapy.Thus,a current goal of cancer research is to eliminate CSCs,necessitating an adequate phenotypic and functional characterization of CSCs.Strategies have been developed to identify,enrich,and track CSCs,many of which distinguish CSCs by evaluating the expression of surface markers,the initiation of specific signaling pathways,and the activation of master transcription factors that control stemness in normal cells.We review and discuss the use of reporter gene systems for identifying CSCs.Reporters that are under the control of aldehyde dehydrogenase 1A1,CD133,Notch,Nanog homeobox,Sex-determining region Y-box 2,and POU class 5 homeobox can be used to identify CSCs in many tumor types,track cells in real time,and screen for drugs.Thus,reporter gene systems,in combination with in vitro and in vivo functional assays,can assess changes in the CSCs pool.We present relevant examples of these systems in the evaluation of experimental CSCs-targeting therapeutics,demonstrating their value in CSCs research.

Pharmacological Activity and Application of Roots, Stems and Leaves of Scutellaria baicalensis Georgi

The root of Scutellaria baicalensis Georgi is traditionally used as medicine,and it has been confirmed that S.baicalensis Georgi has flavonoid chemical constituents,pharmacological activity and cosmetic efficacy.With the extensive application of S.baicalensis Georgi roots,the resource of S.baicalensis Georgi has been increasingly short.The above-ground part of stems and leaves of S.baicalensis Georgi has also been gradually recognized and developed.Studies have found that the chemical constituents from stems and leaves of S.baicalensis Georgi are also a group of flavonoids with a lot of pharmacological activity and have a great application value.Based on this,the present review will be reported on the chemical constituents and application of the roots,stems and leaves of S.baicalensis Georgi.

Preparation and Adsorption Properties of Biomass Activated Carbon from Ginger Stems

Biomass activated carbon(BAC)was produced from ginger stems by carbonization and activation presented high specific surface areas and mesoporous structures.The carbonization temperature of the ginger stems were controlled within 500~900℃.The optimal carbonization condition is as follows:carbonization temperature of 700℃,carbonization time of 6 h.The determined optimum activation condition is:temperature of 800℃,activator of KOH and carbonized product/alkali ratio of 1:4(w/w).The carbonization yield,BAC yield and Brunauer-Emmett-Teller(BET)surface area were measured and the adsorption performance of BAC to nitrogen was investigated.The results showed that the nitrogen adsorption isotherm curve was as type I isotherm.It was finally determined that the BET surface area was 660 m2/g under the abovementioned optimal conditions of carbonization and activation.The FESEM analysis indicates that the obtained BAC is of micropore structure.

Pharmacognostic standardization of stems of Thespesia lampas(Cav.)Dalz & Gibs

Objective:To establish the standardization parameters for complete pharmacoguoslic evaluation of stems of Tlespesia lampas(T.lampas)(Cav.) Dalz & Gibs(Malvaceae),an important planl in the Indian system of medicine.Methods:Morphological,microscopical,physico-chemical evaluations,florescence analysis of T.lampas stems were investigated and preliminary phytochemical analysis,GO-MS analysis and HPTLC fingerprinting were carried out for qualitative phytochemical evaluation of various extracts of stems of T.lampas.Results:Chemomieroscopy revealed the presence of lignin.starch grains and calcium oxalate crystals.Physicochemical evaluation used to detennine numerical slandards showed a result with total ash(9.03± 0.05)%w/w,acid insoluble ash(1.50±0.01)%w/w,water soluble ash(2.51±0.02)%w/w,sulphated ash(7.50+0.01)%w/w,ethanol soluble extractive(0.24±0.02)%w/w,water soluble extractive(0.08 ±0.01)<7,w/w,moisture content(6.03±0.05)%w/w and total crude fibre content of stem powder(47.36±0.32)%w/w.Behavior characteristics of the stem powder showed presence of steroids, starch,alkaloid,flavonoids and proteins.Preliminary phytochemical analysis revealed presence of glycosides,phenolic compounds,tannins,steroids,saponins,flavonoids,carbohydrates and proteins.GC-MS analysis showed the presence of fatty acids such as dodecanoic acid, lelraclecanoic acid,n-hexadecanoic acid,9-lelradecenal and HPTLC fingerprinting revealed the presence of β-sitosterol and quercetin in stems of T.lampas.Conclusions:The pharmacognostic standardization of T.lampas is useful towards establishing standards for quality, purity and sample identification.

Effects of Different Wall-breaking Methods on Extraction of Solanesol from Stems and Leaves of Potato

In order to extract solanesol from potato stems and leaves more effectively and improve the extraction rate of solanesol, the same batch of potato stems and leaves harvested from Guyuan, Ningxia was selected as a research object, cell wall of potato stems and leaves was broken by enzymolysis with cellulase and high- speed shearing, and then reflux-extracted with 95% ethanol. Solanesol content was determined by HPLC, and extract yield was calculated. Extract yield and solanesol extraction rate was used as an index for comparison of difference between the 2 wall-breaking methods, so as to select the optimal wall-breaking method. The results showed that enzymolysis with cellulase exhibited extraction rate and extract yield of solanesol of 91.38% and 8.02%, respectively, which were better than those under high-speed shear emulsification technique. The enzymolysis wall-breaking method has the advantages of simple operation and strong feasibility.

Effects of compatibility of Scutellaria baicalensis stems and Polygonum cuspidatum on TRPV1 expression and inflammatory cytokines in rats with acute lung injury

Objective:The protective effect of Scutellaria baicalensis Stems and Polygonum Cuspidatum compatibility on lipopolysaccharide(LPS)-induced acute lung injury(ALI)in rats was studied by observing the expression of TRPV1 and inflammatory cytokines.Methods:48 male SD rats were randomly divided into 6 groups:control group,model group,dexamethasone group(5mg/kg)and Scutellaria baicalensis Stems-Polygonum Cuspidatum(3.5,7 and 14g/kg).The administration group was gavaged for 7 days,and the control group and model group were given the same amount of 0.9%sodium chloride.On the 8th day,except the control group,rats in other groups were injected with 8mg/kg LPS through caudal vein to induce Ali model.Take the rat lung tissue 6 hours after modeling,and calculate the wet/dry weight ratio(W/D)of the rat lung tissue;HE staining to observe the pathological changes of lung tissue;Determine the content of tumor necrosis factor-α(TNF-α)and interleukin-1β(1L-1β)in alveolar lavage fluid(BALF)and the activity of superoxide dismutase(SOD)in serum;Detect the mRNA and protein expression levels of TRPV1 receptor in rat lung tissue.Results:Compared with the model group,Scutellaria baicalensis Stems-Polygonum Cuspidatum can significantly reduce the damage of lung tissue structure and bleeding state,W/D value and TNF-α、IL-1βThe content of TRPV1 decreased,the level of SOD increased,and the mRNA and protein expression of TRPV1 receptor decreased.Conclusion:The combination of Scutellaria baicalensis Stems-Polygonum has a protective effect on acute lung injury in rats,and its mechanism may be related to down-regulating the expression of TRPV1 and inhibiting the levels of TNF-αand IL-1βin inflammatory cells.

Inhibition of Flavonoids from Scutellaria baicalensis Stems and Leaves against Abnormal Formation of PHF and Regulation of PP in OA Induced Brain Injury in Rats

[Objectives] To explore the inhibitory effects of flavonoids from Scutellaria baicalensis Georgi stems and leaves against abnormal formation of PHF and regulation mechanism of protein phosphatase(PP) in okadaic acid(OA) induced brain injury in rats.[Methods]Male Sprague-Dawley rats were treated with OA(200 ng/kg) in the lateral ventricle to establish a memory impairment model,and Morris water maze was used to screen the memory impairment model.Rats were intragastrically administered with 25,50 and 100 mg/kg SSF daily for 36 d.The protein expressions of PHF,PP1,PP2A-Cα,PP2A-Cβ,PP2 CA and PP2 CB in rat cortex and hippocampus were determined by Western blot.Ginkgo biloba leaf flavonoids were used as positive control drugs.[Results]Compared with the sham-operated rats,the expression of PHF protein in the cortex and hippocampus of the model group was significantly increased(P < 0.01);protein expression of PP2A-Cα and PP2A-Cβin the cortex and hippocampus,and PP2 CB in the hippocampus were significantly decreased(P < 0.05);protein expression of PP2 CA and PP2 CB in the cortex was significantly increased(P < 0.01);protein expression of PP1 in the cortex was significantly decreased(P < 0.01).Compared with the model rats,25,50 and 100 mg/kg SSF 36 d reversed the protein expression of PHF,PP2A-Cα and PP2ACβ in rat cortex and hippocampus and expression of PP1 in rat cortex induced by OA,but had no significant effect on the protein expression of PP2 CA and PP2 CB,G.biloba flavonoids also showed similar results to SSF.[Conclusions] SSF can significantly inhibit the abnormal formation of PHF in rat brain induced by OA.The inhibitory effect may be related to the regulation of PP1,PP2A-Cα and PP2A-Cβ protein expression in cerebral cortex and hippocampus by SSF,but less related to the regulation of PP2 CA and PP2 CB.

A Neural Based Modeling Approach for Drying Kinetics Analysis of Mint Branches and Their Fractions (Leaves and Stems)

This work is aimed at investigating regular mint (Mentha × villosa) drying behavior and assessing how the heterogeneous composition of plants affects their drying kinetics. Drying kinetics and sorption isotherms were evaluated for whole branches and their fractions (leaves and stems). Stems and leaves were characterized by measurement of dimensions, apparent density and initial moisture content. The moisture sorption isotherms were obtained under temperatures of 30°C, 40°C and 50°C for branches, stems and leaves and the data were fitted to the GAB model. Mint branches and their fractions were oven dried at temperatures from 40°C to 70°C?and were obtained kinetic curves for each part. Water sorption patterns were similar for leaves and stems and the GAB model described well the sorption behavior of both materials. At a constant temperature, the drying rates were higher for leaves in comparison to stems and the differences increased as the temperature was raised. Therefore, depending on drying conditions, the moisture distribution in dried branches might be significantly different. Since the leaves constitute the major fraction in branches, the drying rates of branches were closer to those of leaves. The kinetic curves were fitted to a diffusion model based on an analytical solution of Fick’s second diffusion law and to an empirical model based on artificial neural network (ANN). The results showed that the model based on the ANN predicted the drying kinetics of the different parts better than the diffusive model. A single network was built to describe the kinetic behavior of branches and fractions in the whole range of temperatures investigated. The diffusive model based on fitting effective diffusivity did not provide good predictions of moisture content, probably because neither the dependence of effective diffusivity on the moisture content nor the heterogeneity and shrinking of static beds were considered.

The pharmacokinetics of aspirin in combination with total ginsenoside of ginseng stems and leaves in rats

Objective:To investigate the effect of total ginsenoside of ginseng stems and leaves (TGSL) on the pharmacokinetics of aspirin in rats.Methods:Sprague-Dawley rats were randomly divided into two groups (n =6),a combined group that received TGSL (625 mg/kg body weight) and aspirin (10 mg/kg body weight) by gavage,and an aspirin group that received aspirin (10 mg/kg body weight) by gavage.The concentration of salicylic acid,an important metabolite of aspirin,was determined by highperformance liquid chromatography in supernatant from blood obtained from the orbital sinus at various time points to examine the effect of TGSL on aspirin.Results:The results showed that the Tmax of salicylic acid was [0.92 (0.58)] hours in the aspirin group and [2.50 (1.22)] hours in the combined group,and was statistically significantly different between the groups (p <.05).Conclusions:TGSL can affect the pharmacokinetics of aspirin at Tmax in rats.

Pharmacognostical Study on Stems and Leaves of Campsis grandiflora(Thunb.) Schum.

[Objectives] To provide basis for identification,development and use of Campsis grandiflora( Thunb.) Schum. [Methods]Original plants,traits,microstructures and UV spectra were used for identification. [Results]For the first time,the pharmacognostical characteristics were elaborated about stems and leaves of Campsis grandiflora( Thunb.) Schum. [Conclusions] This study is expected to provide basis for pharmacognostical quality standard of stems and leaves of Campsis grandiflora( Thunb.) Schum.

STEMS教育与业余训练融合的探究

采用实证研究法、综合分析法、跨学科研究法,将STEM教育与业余训练内涵进行结合,构建“STEMS”教育模式。

Clinical translation of bioartificial liver support systems with human pluripotent stem cell-derived hepatic cells

There is currently a pressing need for alternative the-rapies to liver transplantation. The number of patients waiting for a liver transplant is substantially higher than the number of transplantable donor livers, resulting in a long waiting time and a high waiting list mortality. An extracorporeal liver support system is one possible approach to overcome this problem. However, the ideal cell source for developing bioartificial liver(BAL) support systems has yet to be determined. Recent advancements in stem cell technology allow researchers to generate highly functional hepatocyte-like cells from human pluripotent stem cells(h PSCs). In this mini-review, we summarize previous clinical trials with different BAL systems, and discuss advantages of and potential obstacles to utilizing h PSC-derived hepatic cells in clinical-scale BAL systems.

Stem cell autotomy and niche interaction in different systems

The best known cases of cell autotomy are the formation of erythrocytes and thrombocytes(platelets) from progenitor cells that reside in special niches. Recently, autotomy of stem cells and its enigmatic interaction with the niche has been reported from male germline stem cells(GSCs) in several insect species. First described in lepidopterans, the silkmoth, followed by the gipsy moth and consecutively in hemipterans, foremost the milkweed bug. In both, moths and the milkweed bug, GSCs form finger-like projections toward the niche, the apical cells(homologs of the hub cells in Drosophila). Whereas in the milkweed bug the projection terminals remain at the surface of the niche cells, in the gipsy moth they protrude deeply into the singular niche cell. In both cases, the projections undergo serial retrograde fragmentation with progressing signs of autophagy. In the gipsy moth, the autotomized vesicles are phagocytized and digested by the niche cell. In the milkweed bug the autotomized vesicles accumulate at the niche surface and disintegrate. Autotomy and sprouting of new projections appears to occur continuously. The significance of the GSC-niche interactions, however, remains enigmatic. Our concept on the signaling relationship between stem cell-niche in general and GSC and niche(hub cells and cyst stem cells) in particular has been greatly shaped by Drosophila melanogaster. In comparing the interactions of GSCs with their niche in Drosophila with those in species exhibiting GSC autotomy it is obvious that additional or alternative modes of stem cell-niche communication exist. Thus, essential signaling pathways, including niche-stem cell adhesion(E-cadherin) and the direction of asymmetrical GSC division- as they were found in Drosophila- can hardly be translated into the systems where GSC autotomywas reported. It is shown here that the serial autotomy of GSC projections shows remarkable similarities with Wallerian axonal destruction, developmental axon pruning and dying-back degeneration in neurodegenerative diseases. Especially the hypothesis of an existing evolutionary conserved "autodestruction program" in axons that might also be active in GSC projections appears attractive. Investigations on the underlying signaling pathways have to be carried out. There are two other well known cases of programmed cell autotomy: the enucleation of erythroblasts in the process of erythrocyte maturation and the segregation of thousands of thrombocytes(platelets) from one megakaryocyte. Both progenitor cell types- erythroblasts and megakaryocytes- are associated with a niche in the bone marrow, erythroblasts with a macrophage, which they surround, and the megakaryocytes with the endothelial cells of sinusoids and their extracellular matrix. Although the regulatory mechanisms may be specific in each case, there is one aspect that connects all described processes of programmed cell autotomy and neuronal autodestruction: apoptotic pathways play always a prominent role. Studies on the role of male GSC autotomy in stem cell-niche interaction have just started but are expected to reveal hitherto unknown ways of signal exchange. Spermatogenesis in mammals advance our understanding of insect spermatogenesis. Mammal and insect spermatogenesis share some broad principles, but a comparison of the signaling pathways is difficult. We have intimate knowledge from Drosophila, but of almost no other insect, and we have only limited knowledge from mammals. The discovery of stem cell autotomy as part of the interaction with the niche promises new general insights into the complicated stem cell-niche interdependence.

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.