The effect of light compressive and tensile mechanical forces on SOST and POSTN expressions in human periodontal ligament cells: an in vitro study.

Periodontal ligament (PDL) cells play an important role in mechanosensing and secretion of signaling molecules during bone remodeling. However, the regulatory mechanism is unknown. The aim of the present study is to investigate the expression pattern of periostin and sclerostin in response to orthodontic forces in periodontal ligament cells in vitro. PDL cells were isolated from extracted teeth and treated with compressive forces of 25 gr/cm2 or equiaxial tension forces at frequency 1 Hz for 0, 24, 48, and 72 h. qRT-PCR was applied to evaluate the gene expressions. The secretion of sclerostin and periostin was assessed using ELISA. DAPI staining was used to evaluate apoptosis. The expression of sclerostin elevated significantly at protein and gene levels under compression forces after 24 h, while the application of tensile forces induced the expression of periostin and its upstream regulator RUNX2 (p < 0.05). Gene expression up-regulation was significant for POSTN and RUNX2 after 48 and 72 h tensile forces. Also, the gene expression of sclerostin reduced in a time-dependent manner after application of tensile force. The compression forces enhanced apoptosis to 7.5 ± 3.5% and induced gene expression of apoptotic markers of CASP9, and BCL2 within 72 h of exposure. Periostin and sclerostin play an important role in orthodontic loads and their expressions are affected oppositely by compressive and tensile forces that might be suggested as a biomarker for assessment of bone remodeling during orthodontic treatment.

Biomimetic vascular tissue engineering by decellularized scaffold and concurrent cyclic tensile and shear stresses.

Decellularization by chemical approaches has harmful effects on extracellular matrix (ECM) proteins, and damages lots of functional peptides and biomolecules present in the ultrastructure. In this study, we employed a combination of chemical and physical decellularization methods to overcome these disadvantages. The induced osmotic pressure by hypertonic/hypotonic solutions dissociated and removed most of cellular membranes significantly without any detergent or chemical agent. In total, 0.025% trypsin solution was found adequate to remove the remaining debrides, and ultimately 1% Triton X-100 was utilized for final cleansing. In addition, conducting all the decellularization processes at 4 °C yielded an ECM with least damages in the ultrastructure which could be inferred by close mechanical strength and swelling ratio to the native vessel, and high quality and quantity of cell attachment, migration and proliferation which were examined by optical microscopy and scanning electron microscopy (SEM) of the histology samples. Moreover, the obtained biological scaffold (BS) had no cytotoxicity according to the MTT assay, and this scaffold is storable at -20 °C. Employing bioreactor for concurrent cyclic tensile and shear stresses improved the cell migration into pores of the BS and made the cells and the scaffold compact in analogous to native tissue. As opening angle test showed by decellularizing of the blood vessel, the residual stress dropped significantly which revealed the role of cells in the amount of induced stress in the structure. However, intact and healthy ECM explicitly recovered upon recellularization and beat the initial residual stress of the native tissue. The tensile test of the blood vessels in longitudinal and radial directions revealed orthotropic behavior which can be explained by collagen fibers direction in the ECM. Furthermore, by the three regions of the stress-strain curve can be elucidated the roles of cells, elastin and collagen fibers in mechanical behavior of the vascular tissues.

Introduction of an efficient method for placenta decellularization with high potential to preserve ultrastructure and support cell attachment.

The placenta, as a large discarded tissue and rich in extracellular matrix (ECM), is an excellent candidate for biological scaffolds in reconstructive medicine. Considering the importance of ECM structure in cell fate, the aim of this study was to achieve human placenta decellularization protocol that preserve the structure of scaffolds. Thus, human placenta was decellularized by four protocols and decellularization efficacy was compared by hematoxylin and eosin (H&E), 4',6-diamidino-2-phenylindole (DAPI) staining, and DNA measurement. Decellularized placenta structure preservation was assessed by Masson's trichrome staining, scanning electron microscopy (SEM), and immunofluorescence (IF) for collagen I, IV, and fibronectin. Finally, liquid displacement measured scaffolds' porosity. After culturing menstrual blood-derived stem cells (MenSCs) on placenta scaffolds, cell adhesion was investigated by SEM imaging, and cell viability and proliferation were assessed by MTT assay. According to H&E and DAPI staining, only protocols 1 and 3 could completely remove cells from the scaffolds. DNA measurements confirmed a significant reduction in the genetic material of decellularized scaffolds compared to native placenta. According to Masson's trichrome, IF, and SEM imaging, scaffold structure is better preserved in P3 than P1 protocol. Liquid displacement showed higher porosity of P3 scaffold than P1. SEM imaging confirmed cells adhesion to the decellularized placenta, and the attached cells showed good viability and maintained their proliferative capacity, indicating the suitability of the scaffolds for cell growth. Results introduced an optimized protocol for placenta decellularization that preserves the scaffold structure and supports cell adhesion and proliferation.

Evaluation of alginate modification effect on cell-matrix interaction, mechanotransduction and chondrogenesis of encapsulated MSCs.

Mesenchymal stem cells (MSCs) are promising cell candidates for cartilage regeneration. Furthermore, it is important to control the cell-matrix interactions that have a direct influence on cell functions. Providing an appropriate microenvironment for cell differentiation in response to exogenous stimuli is a critical step towards the clinical utilization of MSCs. In this study, hydrogels consisted of different proportions of alginates that were modified using gelatin, collagen type I and arginine-glycine-aspartic acid (RGD) and were evaluated regarding their effects on mesenchymal stem cells. The effect of applying hydrostatic pressure on MSCs encapsulated in collagen-modified alginate with and without chondrogenic medium was evaluated 7, 14 and 21 days after culture, which is a comprehensive evaluation of chondrogenesis in 3D hydrogels with mechanical and chemical stimulants. Alcian blue, safranin O and dimethyl methylene blue (DMMB) staining showed the chondrogenic phenotype of cells seeded in the collagen- and RGD-modified alginate hydrogels with the highest intensity after 21 days of culture. The results of real-time PCR for cartilage-specific extracellular matrix genes indicated the chondrogenic differentiation of MSCs in all hydrogels. Also, the synergic effects of chemical and mechanical stimuli are indicated. The highest expression levels of the studied genes were observed in the cells embedded in collagen-modified alginate by loading after 14 days of exposure to the chondrogenic medium. The effect of using IHP on encapsulated MSCs in modified alginate with collagen type I is equal or even higher than using TGF-beta on encapsulated cells. The results of immunohistochemical assessments also confirmed the real-time PCR data.

A conductive cell-imprinted substrate based on CNT-PDMS composite.

Cell function regulation is influenced by continuous biochemical and biophysical signal exchange within the body. Substrates with nano/micro-scaled topographies that mimic the physiological niche are widely applied for tissue engineering applications. As the cartilage niche is composed of several stimulating factors, a multifunctional substrate providing topographical features while having the capability of electrical stimulation is presented. Herein, we demonstrate a biocompatible and conductive chondrocyte cell-imprinted substrate using polydimethylsiloxane (PDMS) and carbon nanotubes (CNTs) as conductive fillers. Unlike the conventional silicon wafers or structural photoresist masters used for molding, cell surface topographical replication is challenging as biological cells showed extremely sensitive to chemical solvent residues during molding. The composite showed no significant difference compared with PDMS with regard to cytotoxicity, whereas an enhanced cell adhesion was observed on the conductive composite's surface. Integration of nanomaterials into the cell seeding scaffolds can make tissue regeneration process more efficient.

Graphene oxide incorporated polycaprolactone/chitosan/collagen electrospun scaffold: Enhanced osteogenic properties for bone tissue engineering.

This in vitro study aimed to evaluate the physicochemical and biological activity of the polycaprolactone/chitosan/collagen scaffolds incorporated with 0, 0.5, 3, and 6 wt% of graphene oxide (GO). Using standard tests and MG-63 cells, the characteristics of scaffolds were evaluated, and the behavior of osteoblasts were simulated, respectively. A non-significant decrease in nanofibers diameter was noted in scaffolds with a higher ratio of GO. The hydrophilicity and bioactivity of the scaffold surface, as well as cell attachment and proliferation, increased in correspondence to an increase in GO. The higher ratio of GO also improved the osteogenesis activity. GO increased the degradation rate, but it was negligible and seemed not enough to endanger stability. Modifying the scaffolds with GO did not make a significant change to the antibacterial effect.

Mechanical and Chemical Predifferentiation of Mesenchymal Stem Cells Into Cardiomyocytes and Their Effectiveness on Acute Myocardial Infarction.

Myocardial infarction is one of the leading causes of death all over the world. Mesenchymal stem cells (MSCs) transplantation has shown a promising potential to recovery of ischemic heart disease due to their capability in differentiating into cardiac cells. However, various investigations have been performed to optimize the efficacy of cardiac cell therapy in recent years. Here, we sought to interrogate the effect of autologous transplantation of undifferentiated and predifferentiated adipose and bone marrow-derived MSCs in a rabbit model of myocardial infarction and also to investigate whether cardiac function could be improved by mechanically induced MSCs via equiaxial cyclic strain. The two sources of MSCs were induced toward cardiomyocyte phenotype using mechanical loading and chemical factors and thereafter injected into the infarcted myocardium of 35 rabbits. Echocardiography and histopathology studies were used to evaluate cardiac function after 2 months. The results demonstrated significant scar size reduction and greater recovery of left ventricle ejection fraction after transplantation of predifferentiated cells, though the differences were not significant when comparing mechanically with chemically predifferentiated MSCs. Thus, although there was no significant improvement in infarcted myocardium between chemically and mechanically predifferentiated MSCs, mechanically induced cells are more preferred due to lack of any chemical intervention and cost reasonableness in their preparation method. Outcomes of this study may be useful for developing future therapeutic strategies, however long-term assessments are still required to further examine their effectiveness.

Delivery of molecular cargoes in normal and cancer cell lines using non-viral delivery systems.

OBJECTIVE: In this study, transfection efficiency of human papillomavirus (HPV) E7 DNA and protein constructs into HEK-293T normal cell line, and A549 and TC-1 tumor cell lines was evaluated by four delivery systems including supercharge GFP, hPP10 cell penetrating peptide, TurboFect and Lipofectamine using fluorescence microscopy and flow cytometry. RESULTS: The results indicated that Lipofectamine 2000 and TurboFect produced more effective transfection for GFP and E7-GFP DNA constructs in HEK-293T cells compared to in A549 and TC-1 cells (p < 0.05). In contrast, the supercharge GFP was efficient for E7 DNA and E7 protein delivery in both normal cell (~ 83.94 and ~ 77.01% for HEK-293T), and cancer cells (~ 71.69 and ~ 67.19% for TC-1, and ~ 73.86 and ~ 67.49% for A549), respectively. Indeed, in these cell lines, transfection efficiency by +36 GFP reached ~ 60-80%. Moreover, the hPP10 produced the best transfection result for E7-GFP protein in HEK-293T cells (~ 63.66%) compared to TurboFect (~ 32.95%); however, the efficiency level of hPP10 was only ~ 17.51 and ~ 16.36% in TC-1 and A549 cells. CONCLUSIONS: Our data suggested that the supercharge GFP is the most suitable transfection vehicle for DNA and protein delivery into TC-1 and A549 tumor cell lines compared to other carriers.

Extremely Low Frequency Electromagnetic Field in Mesenchymal Stem Cells Gene Regulation: Chondrogenic Markers Evaluation.

There is little evidence demonstrating the effects of electromagnetic fields (EMFs) generated within the biological entity and the effect of extrinsic fields on cellular programing. Taking the path of the more studied stimuli into attention, mechanical forces, it could be understood that nonchemical factors play a consequential role in transcriptional regulatory networks. Cartilaginous tissue consists of collagen protein that is considered as a piezoelectric substrate and is influenced by electric fields making chondrogenic specific genes an exciting candidate for bioelectromagnetic studies. As electromagnetic properties highly depend on the frequencies applied, this study delves into the ability of two EMFs with the frequency of 25 Hz and 50 Hz in inducing SOX9 and COL2 gene expressions in a three-dimensional (3D) mesenchymal stem cell (MSC)-alginate construct. Cell-alginate beads were divided into six groups and treated for a time period of 21 days. To determine the results, qualitative and quantitative data were both reviewed. On observation of real-time polymerase chain reaction (PCR) data, it was apparent that TGF-ß1 treatment had a greater COL2 and SOX9 gene expression impact on MSCs compared to pulsed electromagnetic field (PEMF) treatments alone. COL2 was shown to have a greater transcriptional tendency to PEMF, whereas under defined electromagnetic parameters applied in this study, no significant difference was detected in SOX9 gene expressions compared to the control group. PEMF co-treatments enhanced the deposition of extracellular matrix molecules, as the matrix-rich beads were positively stained by Alcian blue. This genre of study is the venue for the control and healing of connective tissue defects.

Fluid-Structure Interactions Analysis of Shear-Induced Modulation of a Mesenchymal Stem Cell: An Image-Based Study.

Although effects of biochemical modulation of stem cells have been widely investigated, only recent advances have been made in the identification of mechanical conditioning on cell signaling pathways. Experimental investigations quantifying the micromechanical environment of mesenchymal stem cells (MSCs) are challenging while computational approaches can predict their behavior due to in vitro stimulations. This study introduces a 3D cell-specific finite element model simulating large deformations of MSCs. Here emphasizing cell mechanical modulation which represents the most challenging multiphysics phenomena in sub-cellular level, we focused on an approach attempting to elicit unique responses of a cell under fluid flow. Fluorescent staining of MSCs was performed in order to visualize the MSC morphology and develop a geometrically accurate model of it based on a confocal 3D image. We developed a 3D model of a cell fixed in a microchannel under fluid flow and then solved the numerical model by fluid-structure interactions method. By imposing flow characteristics representative of vigorous in vitro conditions, the model predicts that the employed external flow induces significant localized effective stress in the nucleo-cytoplasmic interface and average cell deformation of about 40%. Moreover, it can be concluded that a lower strain level is made in the cell by the oscillatory flow as compared with steady flow, while same ranges of effective stress are recorded inside the cell in both conditions. The deeper understanding provided by this study is beneficial for better design of single cell in vitro studies.

Comparative analysis of effects of cyclic uniaxial and equiaxial stretches on gene expression of human umbilical vein endothelial cells.

It has been well established that biomechanical environment can influence functionality of biological cells. There are evidences that show mechanical cyclic stretch can promote smooth muscle cell (SMC) markers in endothelial cells (ECs). The objective of this study was to determine whether mechanical stimuli in the forms of uniaxial and equiaxial cyclic stretches (UNCS and EQCS) can affect endothelial and smooth muscle gene expressions in mRNA level of human umbilical vein endothelial cells (HUVECs). For this purpose, 10% uniaxial UNCS and EQCS (60 cycles/min for 24 h) were applied on HUVECs, and using real-time PCR expressions of three EC specific markers, vascular endothelial growth factor receptor-2 (VEGFR-2, also known as FLK-1), von Willebrand Factor (vWF) and vascular endothelial-cadherin (VE-cadherin) and two SMC specific genes, α-smooth muscle actin (α-SMA) and smooth muscle myosin heavy chain (SMMHC) were quantified. Moreover, alterations in cell height were analyzed by atomic force microscopy (AFM). Results showed that cyclic UNCS for 24 h downregulated the expression of all EC markers and upregulated the expression of all SMC markers while low effects on HUVECs height were observed. Cyclic EQCS in the same conditions resulted in minor effect on SMC gene expression in HUVECs, while led to strong reduction in vWF with no significant change in other two endothelial genes. Cyclic EQCS considerably elevated cell height. Results proposed that ECs can transdifferentiate to SMC phenotype under specific microenvironmental conditions.

Alteration of human umbilical vein endothelial cell gene expression in different biomechanical environments.

Biomechanical environments affect the function of cells. In this study we analysed the effects of five mechanical stimuli on the gene expression of human umbilical vein endothelial cells (HUVECs) in mRNA level using real-time PCR. The following loading regimes were applied on HUVECs for 48 h: intermittent (0-5 dyn/cm(2) , 1 Hz) and uniform (5 dyn/cm(2) ) shear stresses concomitant by 10% intermittent equiaxial stretch (1 Hz), uniform shear stress alone (5 dyn/cm(2) ), and intermittent uniaxial and equiaxial stretches (10%, 1 Hz). A new bioreactor was made to apply uniform/cyclic shear and tensile loadings. Three endothelial suggestive specific genes (vascular endothelial growth factor receptor-2 (VEGFR-2, also known as FLK-1), von Willebrand Factor (vWF) and vascular endothelial-cadherin (VE-cadherin)), and two smooth muscle genes (α-smooth muscle actin (α-SMA) and smooth muscle myosin heavy chain (SMMHC)) were chosen for assessment of alteration in gene expression of endothelial cells and transdifferentiation toward smooth cells following load applications. Shear stress alone enhanced the endothelial gene expression significantly, while stretching alone was identified as a transdifferentiating factor. Cyclic equiaxial stretch contributed less to elevation of smooth muscle genes compared to uniaxial stretch. Cyclic shear stress in comparison to uniform shear stress concurrent with cyclic stretch was more influential on promotion of endothelial genes expression. Influence of different mechanical stimuli on gene expression may open a wider horizon to regulate functions of cell for tissue engineering purposes.

Comparing the effect of equiaxial cyclic mechanical stimulation on GATA4 expression in adipose-derived and bone marrow-derived mesenchymal stem cells.

Myocardium is prone to mechanical stimuli among which pulsatile blood flow exerts both radial and longitudinal strains on the heart. Recent studies have shown that mechanical stimulation can notably influence regeneration of cardiac muscle cells. GATA4 is a cardiac-specific transcription factor that plays an important role in late embryonic heart development. Our study aimed at investigating the effect of equiaxial cyclic strain on GATA4 expression in adipose-derived (ASCs) and bone marrow-derived (BMSCs) mesenchymal stem cells. For this reason, both ASCs and BMSCs were studied in four distinct groups of chemical, mechanical, mechano-chemical and negative control. According to this categorisation, the cells were exposed to cyclic mechanical loading and/or 5-azacytidine as the chemical factor. The level of GATA4 expression was then quantified using real-time PCR method on the first, fourth and seventh days. The results show that: (1) equiaxial cyclic stimulation of mesenchymal stem cells could promote GATA4 expression from the early days of induction and as it went on, its combination with chemical factor elevated expression; (2) cyclic strain could accelerate GATA4 expression compared to the chemical factor; (3) in this regard, these results indicate a higher capacity of ASCs than BMSCs to express GATA4.

Applying shear stress to endothelial cells in a new perfusion chamber: hydrodynamic analysis.

Perfusion bioreactors have been proved to be an impartible part of vascular tissue engineering due to its broad range of applications as a means to distribute nutrients within porous scaffold along with providing appropriate physical and mechanical stimuli. To better understand the mechanical phenomena inside a bioreactor, computational fluid dynamics (CFD) was adopted followed by a validation technique. The fluid dynamics of the media inside the bioreactor was modeled using the Navier-Stokes equation for incompressible fluids while convection through the scaffold was described by Brinkman's extension of Darcy's law for porous media. Flow within the reactor determined the orientation of endothelial cells on the scaffold. To validate flow patterns, streamlines and shear stresses, colorimetry technique was used following attained results from CFD. Our bioreactor was modeled to simulate the optimum condition and flow patterns over scaffold to culture ECs for in vitro experimentation. In such experiments, cells were attached firmly without significant detachment and more noticeably elongation process was triggered even shortly after start up.

Evaluation of rabbit adipose derived stem cells fate in perfused multilayered silk fibroin composite scaffold for Osteochondral repair.

Development of osteochondral tissue engineering approaches using scaffolds seeded with stem cells in association with mechanical stimulations has been recently considered as a promising technique for the repair of this tissue. In this study, an integrated and biomimetic trilayered silk fibroin (SF) scaffold containing SF nanofibers in each layer was fabricated. The osteogenesis and chondrogenesis of stem cells seeded on the fabricated scaffolds were investigated under a perfusion flow. 3-Dimethylthiazol-2,5-diphenyltetrazolium bromide assay showed that the perfusion flow significantly enhanced cell viability and proliferation. Analysis of gene expression by stem cells revealed that perfusion flow had significantly upregulated the expression of osteogenic and chondrogenic genes in the bone and cartilage layers and downregulated the hypertrophic gene expression in the intermediate layer of the scaffold. In conclusion, applying flow perfusion on the prepared integrated trilayered SF-based scaffold can support osteogenic and chondrogenic differentiation for repairing osteochondral defects.

Differential effects of cyclic uniaxial stretch on human mesenchymal stem cell into skeletal muscle cell.

Both fetal and adult skeletal muscle cells are continually being subjected to biomechanical forces. Biomechanical stimulation during cell growth affects proliferation, differentiation and maturation of skeletal muscle cells. Bone marrow-derived hMSCs [human MSCs (mesenchymal stem cells)] can differentiate into a variety of cell types, including skeletal muscle cells that are potentially a source for muscle regeneration. Our investigations involved a 10% cyclic uniaxial strain at 1 Hz being applied to hMSCs grown on collagen-coated silicon membranes with or without IGF-I (insulin-like growth factor-I) for 24 h. Results obtained from morphological studies confirmed the rearrangement of cells after loading. Comparison of MyoD and MyoG mRNA levels between test groups showed that mechanical loading alone can initiate myogenic differentiation. Furthermore, comparison of Myf5, MyoD, MyoG and Myf6 mRNA levels between test groups showed that a combination of mechanical loading and growth factor results in the highest expression of myogenic genes. These results indicate that cyclic strain may be useful in myogenic differentiation of stem cells, and can accelerate the differentiation of hMSCs into MSCs in the presence of growth factor.

Intermittent hydrostatic pressure enhances growth factor-induced chondroinduction of human adipose-derived mesenchymal stem cells.

Hydrostatic pressure (HP) plays an essential role in regulating function of chondrocytes and chondrogenic differentiation. The objective of this study was to examine effects of intermittent HP on chondrogenic differentiation of human adipose-derived mesenchymal stem cells (hASCs) in the presence or absence of chemical chondrogenic medium. Cells were isolated from abdominal fat tissue and confirmed for expression of ASC surface proteins and differentiation potential. Passage 3 pellets were treated with chemical (growth factor), mechanical (HP of 5 MPa and 0.5 Hz with duration of 4 h/day for 7 consecutive days), and combined chemical-mechanical stimuli. Using real-time polymerase chain reaction, the expression of Sox9, collagen II, and aggrecan as three major chondrogenic markers were quantified among three experimental groups and compared to those of stem cells and human cartilage tissue. In comparison to the chemical and mechanical groups, the chemical-mechanical group showed the highest expression for all three chondrogenic genes close to that of cartilage tissue. Results show the beneficial role of intermittent HP on chondrogenic differentiation of hASCs, and that this loading regime in combination with chondrogenic medium can be used in cartilage tissue engineering.

Replacement of Trypsin by Proteases for Medical Applications.

Background: Cell culture has a crucial role in many applications in biotechnology. The production of vaccines, recombinant proteins, tissue engineering, and stem cell therapy all need cell culture. Most of these activities needed adherent cells to move, which should be trypsinized several times until received on a large scale. Although trypsin is manufactured from the bovine or porcine pancreas, the problem of contamination by unwanted animal proteins, unwanted immune reactions, or contamination to pathogen reagents is the main problem. Objectives: This study investigated microbial proteases as a safe alternative for trypsin replacement in cell culture experiments for the detachment of adherent cells. Methods: The bacteria were isolated from the leather industry effluent based on their protease enzymes. After sequencing their 16S ribosomal deoxyribonucleic acid, their protease enzymes were purified, and their enzyme activities were assayed. The alteration of enzymatic activities using different substrates and the effect of substrate concentrations on enzyme activities were determined. The purified proteases were evaluated for cell detachment in the L929 fibroblast cells compared to trypsin. The separated cells were cultured again, and cell proliferation was determined by the MTT assay. Results: The results showed that the isolated bacteria were Bacillus pumilus, Stenotrophomonas sp., Klebsiella aerogenes, Stenotrophomonas maltophilia, and Bacillus licheniformis. Among the isolated bacteria, the highest and the lowest protease activity belonged to Stenotrophomonas sp. and K. aerogenes, with 60.34 and 11.09 U/mL protease activity, respectively. All the isolated microbial proteases successfully affected L929 fibroblast cells' surface proteins and detached the cells. A significant induction in cell proliferation was observed in the cells treated with K. aerogenes protease and B. pumilus protease, respectively (P < 0.05). Conclusions: The obtained results suggested that microbial proteases can be used as safe and efficient alternatives to trypsin in cell culture in biopharmaceutical applications.

The Role of Low-Frequency Electromagnetic Fields on Mesenchymal Stem Cells Differentiation: A Systematic Review.

BACKGROUND: Low-frequency electromagnetic fields (EMFs) influence biological processes. This present study was aimed at the scientific literature on the use of EMFs in the mesenchymal stem cell differentiation process. MATERIALS AND METHODS: The electronic search was carried out in PubMed and Web of Science, a database with a combination of the sinusoidal and pulsed low- and extremely low-frequency electromagnetic fields stimulation and mesenchymal stem cells differentiation, considering the period of publication until December 2021. The literature search identified 118 references in PubMed and Web of Science of which 46 articles were selected, respectively, according to the eligibility requirements. CONCLUSION: The analysis of research indicated that EMFs are an easy-to-apply and practical way in cell therapy and tissue engineering when regulation of stem cells is required. Studies have shown that EMFs have positive effects on stem cell differentiation, accelerating its process regardless of the parameters and type of stem cells. However, the exact amplitude, frequency, duration of the electrical field, and application method remain elusive and need more study in future work.

In vitrostatic and dynamic cell culture study of novel bone scaffolds based on 3D-printed PLA and cell-laden alginate hydrogel.

The aim of this paper was to design and fabricate a novel composite scaffold based on the combination of 3D-printed polylactic acid-based triply periodic minimal surfaces (TPMSs) and cell-laden alginate hydrogel. This novel scaffold improves the low mechanical properties of alginate hydrogel and can also provide a scaffold with a suitable pore size, which can be used in bone regeneration applications. In this regard, an implicit function was used to generate some gyroid TPMS scaffolds. Then the fused deposition modeling process was employed to print the scaffolds. Moreover, the micro computed tomography technique was employed to assess the microstructure of 3D-printed TPMS scaffolds and obtain the real geometries of printed scaffolds. The mechanical properties of composite scaffolds were investigated under compression tests experimentally. It was shown that different mechanical behaviors could be obtained for different implicit function parameters. In this research, to assess the mechanical behavior of printed scaffolds in terms of the strain-stress curves on, two approaches were presented: equivalent volume and finite element-based volume. Results of strain-stress curves showed that the finite-element based approach predicts a higher level of stress. Moreover, the biological response of composite scaffolds in terms of cell viability, cell proliferation, and cell attachment was investigated. In this vein, a dynamic cell culture system was designed and fabricated, which improves mass transport through the composite scaffolds and applies mechanical loading to the cells, which helps cell proliferation. Moreover, the results of the novel composite scaffolds were compared to those without alginate, and it was shown that the composite scaffold could create more viability and cell proliferation in both dynamic and static cultures. Also, it was shown that scaffolds in dynamic cell culture have a better biological response than in static culture. In addition, scanning electron microscopy was employed to study the cell adhesion on the composite scaffolds, which showed excellent attachment between the scaffolds and cells.