Behavior Of Mesenchymal Stem Cells Research Articles

RGD Mutation of the Heparin Binding II Fragment of Fibronectin for Guiding Mesenchymal Stem Cell Behavior on Titanium Surfaces.

Installing bioactivity on metallic biomaterials by mimicking the extracellular matrix (ECM) is crucial for stimulating specific cellular responses to ultimately promote tissue regeneration. Fibronectin is an ECM protein commonly used for biomaterial functionalization. The use of fibronectin recombinant fragments is an attractive alternate to the use of full-length fibronectin because of the relatively low cost and facility of purification. However, it is necessary to combine more than one fragment, for example, the cell attachment site and the heparin binding II (HBII), either mixed or in one molecule, to obtain complete activity. In the present study, we proposed to install adhesion capacity to the HBII fragment by an RGD gain-of-function DNA mutation, retaining its cell differentiation capacity and thereby producing a small and very active protein fragment. The novel molecule, covalently immobilized onto titanium surfaces, maintained the growth factor-binding capacity and stimulated cell spreading, osteoblastic cell differentiation, and mineralization of human mesenchymal stem cells compared to the HBII native protein. These results highlight the potential capacity of gain-of-function DNA mutations in the design of novel molecules for the improvement of osseointegration properties of metallic implant surfaces.

Behavior of Mesenchymal Stem Cells Obtained From Dental Tissues: A Review of the Literature

The success of tissue engineering in combination with tissue regeneration depends on the behavior and cellular activity in the biological processes developed within a structure that functions as a support, better known as scaffolds, or directly at the site of the injury. The cell-cell and cell-biomaterial interaction are key factors for the induction of a specific cell behavior, together with the bioactive factors that allow the formation of the desired tissue. Mesenchymal Stem Cells (MSC) can be isolated from the umbilical cord and bone marrow; however, the behavior of Dental Pulp Stem Cells (DPSC) has been shown to have a high potential for the formation of bone tissue, and these cells have even been able to induce the process of angiogenesis. Advances in periodontal regeneration, dentin-pulp complex, and craniofacial bone defects through the induction of MSC obtained from tooth structures in in vitro-in vivo studies have permitted the obtaining of clinical evidence of the achievements obtained to date.

Topographical patterning: characteristics of current processing techniques, controllable effects on material properties and co-cultured cell fate, updated applications in tissue engineering, and improvement strategies.

Topographical patterning has recently attracted lots of attention in regulating cell fate, understanding the mechanism of cell-microenvironment interactions, and solving the great issues of regenerative medicine. The introduced patterns offer topographical cues that can affect the reconstruction of the cytoskeleton or stimulate cell membrane receptors. Numerous studies have focused on these effects on cell behavior including attachment, migration, proliferation, and differentiation. In this review, five aspects of topographical patterning are discussed: (1) the process of typical micro-/nanotechniques and their advantages and limitations; (2) the effects of patterning on the mechanical properties and surface properties of substrates; (3) the influences of micro-/nanopatterns on the behavior of mesenchymal stem cells, as well as the underlying mechanisms; (4) the application of patterns to solve the issues of targeted organs (e.g., skin, nerves, blood vessels, bones, and heart). In the end, future perspectives that would help promote the efficiency of topographical patterning are proposed.

Mesenchymal stem cells and the neuronal microenvironment in the area of spinal cord injury.

Cell-based technologies are used as a therapeutic strategy in spinal cord injury (SCI). Mesenchymal stem cells (MSCs), which secrete various neurotrophic factors and cytokines, have immunomodulatory, anti-apoptotic and anti-inflammatory effects, modulate reactivity/phenotype of astrocytes and the microglia, thereby promoting neuroregeneration seem to be the most promising. The therapeutic effect of MSCs is due to a paracrine mechanism of their action, therefore the survival of MSCs and their secretory phenotype is of particular importance. Nevertheless, these data are not always reported in efficacy studies of MSC therapy in SCI. Here, we provide a review with summaries of preclinical trials data evaluating the efficacy of MSCs in animal models of SCI. Based on the data collected, we have tried (1) to establish the behavior of MSCs after transplantation in SCI with an evaluation of cell survival, migration potential, distribution in the area of injured and intact tissue and possible differentiation; (2) to determine the effects MSCs on neuronal microenvironment and correlate them with the efficacy of functional recovery in SCI; (3) to ascertain the conditions under which MSCs demonstrate their best survival and greatest efficacy.

Sonochemical nanostructuring of titanium for regulation of human mesenchymal stem cells behavior for implant development

The influence of surface nanotopography of sonochemically generated mesoporous titania coatings (TMS) on the adhesion, proliferation, and osteogenic differentiation of human mesenchymal stem cells (hMSCs) have been investigated in vitro for the first time. It has been revealed that adhesion and proliferation of hMSCs is higher on disordered TMS surfaces compared to smooth polished titania surface after five days of incubation. Surprisingly, the sonochemically generated disordered nanotopography induces the differentiation of hMSCs into osteogenic direction in the absence of osteogenic medium in 14 days of incubation. Thus sonochemical nanostructuring of titanium based implants stimulates the regenerative process of bone tissue.

Effect of Poly(sophorolipid) Functionalization on Human Mesenchymal Stem Cell Osteogenesis and Immunomodulation.

Sophorolipids are a class of glycolipids that can be polymerized via ring-opening metathesis polymerization giving rise to bioresorbable biomaterials. The surface chemistry of the resulting poly(sophorolipids) (pLSLs) can be modified using a combination of enzymatic and "click" chemistries to insert bioactive groups that influence cellular behavior. Mesenchymal stem cells (MSCs) are being actively investigated for engineered bone grafts for fracture repair due to their osteogenic potential, and more recently, due to their immunomodulatory capacity. The long-term goal of this work is to utilize functionalized pLSL foams loaded with MSCs as bioresorbable scaffolds for bone fracture healing. Toward this goal, the present study evaluated the effect of various pLSL chemistries on the osteogenic and immunomodulatory behavior of MSCs. pLSLs functionalized with PO4, NH2, or COOH small functional groups were fabricated into open porous foams and then cultured with MSCs in the presence of osteogenic medium for 72 h. Protein level assessments demonstrated that the PO4-functionalized pLSL foams supported the highest degree of MSC osteogenesis as well as the highest levels of immunomodulatory factors pertinent to improve bone fracture healing. Cumulatively, these results suggest that further investigation of the long-term osteogenic commitment of MSCs in PO4-functionalized pLSL foams is warranted.

Laser Surface Microstructuring of a Bio-Resorbable Polymer to Anchor Stem Cells, Control Adipocyte Morphology, and Promote Osteogenesis.

New strategies in regenerative medicine include the implantation of stem cells cultured in bio-resorbable polymeric scaffolds to restore the tissue function and be absorbed by the body after wound healing. This requires the development of appropriate micro-technologies for manufacturing of functional scaffolds with controlled surface properties to induce a specific cell behavior. The present report focuses on the effect of substrate topography on the behavior of human mesenchymal stem cells (MSCs) before and after co-differentiation into adipocytes and osteoblasts. Picosecond laser micromachining technology (PLM) was applied on poly (L-lactide) (PLLA), to generate different microstructures (microgrooves and microcavities) for investigating cell shape, orientation, and MSCs co-differentiation. Under certain surface topographical conditions, MSCs modify their shape to anchor at specific groove locations. Upon MSCs differentiation, adipocytes respond to changes in substrate height and depth by adapting the intracellular distribution of their lipid vacuoles to the imposed physical constraints. In addition, topography alone seems to produce a modest, but significant, increase of stem cell differentiation to osteoblasts. These findings show that PLM can be applied as a high-efficient technology to directly and precisely manufacture 3D microstructures that guide cell shape, control adipocyte morphology, and induce osteogenesis without the need of specific biochemical functionalization.

Mesenchymal Stem Cells from Cervix and Age: New Insights into CIN Regression Rate.

Cervical intraepithelial neoplasia (CIN) is a precancerous lesion of the uterine cervix that can regress or progress to cervical cancer; interestingly, it has been noted that young women generally seem to have higher rates of spontaneous regression and remission, suggesting a correlation between the patient's age and regression/progression rates of CIN. Even if the underlying mechanisms are still unclear, inflammation seems to play a pivotal role in CIN fate and inflammatory processes are often driven by mesenchymal stem cells (MSCs). This study was aimed at evaluating if age affects the behavior of MSCs from the cervix (C-MSCs) that in turn may modulate inflammation and, finally, regression rate. Fourteen samples of the human cervix were recovered from two groups of patients, “young” (mean age 28 ± 2) and “old” (mean age 45 ± 3), during treatment using the loop electrosurgical excision procedure (LEEP) technique. Progenitor cells were isolated, deeply characterized, and divided into young (yC-MSCs) and old cervixes (oC-MSCs); the senescence, expression/secretion of selected cytokines related to inflammation, and the effects of indirect cocultures with HeLa cells were analyzed. Our results show that isolated cells satisfy the fixed criteria for stemness and display age-related properties; yC-MSCs express a higher level of cytokines related to acute inflammation than oC-MSCs. Finally, in the crosstalk with HeLa cells, MSCs derived from the cervixes of young patients play a stronger antitumoral role than oC-MSCs. In conclusion, the immunobiology of MSCs derived from the cervix is affected by the age of donors and this can correlate with the regression rate of CIN by influencing their paracrine effect. In addition, MSCs from a young cervix drives an antitumoral effect by sustaining an acute inflammatory environment.

Culture of Mesenchymal Stem Cells in a Hydrogel Model of Vocal Fold Lamina Propria.

Stem cell injection has been proposed as an alternative approach for the restoration of vocal fold (VF) function in patients with VF scarring. To assess the therapeutic efficacy of this treatment strategy, we evaluated the behaviors of human mesenchymal stem cells (hMSCs) in hydrogels derived from thiolated hyaluronic acid (HA-SH) and poly(ethylene glycol) diacrylate (PEG-DA) entrapping assembled collagen fibrils (abbreviated as HPC gels). Three hydrogel formulations with varying amounts of collagen (0, 1 and 2 mg/mL) but a fixed HA-SH (5 mg/mL) and PEG-DA (2 mg/mL) concentration, designated as HPC0, HPC1 and HPC2, were investigated. The HPC gels exhibit similar pore sizes (35-50 nm) and AFM indentation moduli (~175 Pa), although the elastic shear modulus for HPC1 (~32 Pa) is lower than HPC0 and HPC2 (~55 Pa). Although HPC1 and HPC2 gels both promoted the development of an elongated cell morphology, greater cell spreading was observed in HPC2 than in HPC1 by day 7. At the transcript level, cells cultured in HPC1 and HPC2 gels had an increased expression of fibronectin and integrin β1, but a decreased expression of tissue inhibitor of metalloproteinase-1, collagen types I/III and HA synthase-1 when compared to cells cultured in HPC0 gels. Cellular expression of connective tissue growth factor was also elevated in HPC1 and HPC2 cultures. Importantly, the HPC2 hydrogels promoted a signficant up-regulation of matrix metalloproteinase 1, transforming growth factor β1, and epithelial growth factor receptor, indicating an increased tissue turnover. Overall, hMSCs cultured in HPC2 gels adopt a phenotype reminiscent of cells involved in the wound healing process, providing a platform to study the effectiveness of therapeutic stem cell treatments for VF scarring.

Elevated extracellular calcium ions promote proliferation and migration of mesenchymal stem cells via increasing osteopontin expression

Supplementation of mesenchymal stem cells (MSCs) at sites of bone resorption is required for bone homeostasis because of the non-proliferation and short lifespan properties of the osteoblasts. Calcium ions (Ca2+) are released from the bone surfaces during osteoclast-mediated bone resorption. However, how elevated extracellular Ca2+ concentrations would alter MSCs behavior in the proximal sites of bone resorption is largely unknown. In this study, we investigated the effect of extracellular Ca2+ on MSCs phenotype depending on Ca2+ concentrations. We found that the elevated extracellular Ca2+ promoted cell proliferation and matrix mineralization of MSCs. In addition, MSCs induced the expression and secretion of osteopontin (OPN), which enhanced MSCs migration under the elevated extracellular Ca2+ conditions. We developed in vitro osteoclast-mediated bone resorption conditions using mouse calvaria bone slices and demonstrated Ca2+ is released from bone resorption surfaces. We also showed that the MSCs phenotype, including cell proliferation and migration, changed when the cells were treated with a bone resorption-conditioned medium. These findings suggest that the dynamic changes in Ca2+ concentrations in the microenvironments of bone remodeling surfaces modulate MSCs phenotype and thereby contribute to bone regeneration.

Prolonged incubation with Metformin decreased angiogenic potential in human bone marrow mesenchymal stem cells.

Metformin is commonly prescribed as a hypoglycemic agent following the onset of type 2 diabetes mellitus. This study aimed to investigate pro- and/or anti-angiogenic effects of Metformin on human bone marrow mesenchymal stem cells. Cells were incubated with different doses of Metformin including 0.5, 1, 10, 50, 100, 200 and 500 μM for 14 days. Cell viability and total fatty acids profile were examined by MTT and gas chromatography methods. Differentiation of cells to endothelial lineage was studied by monitoring the expression of VEGFR-2 and Tie-2 receptors and VE-cadherin via real-time PCR and western blotting. Angiogenic potential and migration of cells were assessed by tubulogenesis and Transwell migration assays. PCR array was performed to analyze mTOR signaling. CD133+ and VEGFR-2+ cells were detected in blood samples of non-diabetic control, diabetic subjects and diabetics received Metformin. Metformin dose-dependently reduced cell survival. Decreased content of palmitate and oleate coincided increased level of stearate, palmitoleate, and linoleate (p < 0.05). Metformin decreased the angiogenic potential of cells by decreasing VEGFR-2 and Tie-2 expression (p < 0.05). The protein level of VE-cadherin decreased in cells received Metformin. Compared to the control, Metformin blunted the expression of VEGF subtypes and directed cells to energy status by induction of PRKAA1, PRKAB2, and PRKAG1 genes (p < 0.05). Non-significant differences were observed regarding the number of CD133 and VEGFR-2 cells in blood samples (p > 0.05). These data support a notion that Metformin could blunt the angiogenic behavior of human mesenchymal stem cells by modulating mTOR signaling pathway.

Substrate topography interacts with substrate stiffness and culture time to regulate mechanical properties and smooth muscle differentiation of mesenchymal stem cells.

Substrate stiffness and topography are two powerful means by which mesenchymal stem cells (MSCs) activities can be modulated. The effects of substrate stiffness on the MSCs mechanical properties were investigated previously, however, the role of substrate topography in this regard is not yet well understood. Moreover, in vessel wall, these two physical cues act simultaneously to regulate cellular function, hence it is important to investigate their cooperative effects on cellular activity. Herein, we investigated the combined effects of substrate stiffness, substrate topography and culture time on the mechanical behavior of MSCs. The MSCs were cultured on the stiff and soft substrates with or without micro-grooved topography for 10 days and their viscoelastic properties and smooth muscle (SM) gene expression were investigated on days 2, 6 and 10. In general, substrate topography significantly interacted with substrate stiffness as well as culture time in the modulation of cell viscoelastic behavior and SM gene expression. The micro-grooved, stiff substrates resulted in the maximum cell stiffness and gene expression of α-actin and h1-calponin, and these values were detected to be minimum in the smooth, soft substrates. The findings can be helpful in the mechano-regulation of MSCs for vascular tissue engineering applications.

Prodrug suicide gene therapy for cancer targeted intracellular by mesenchymal stem cell exosomes.

The natural behavior of mesenchymal stem cells (MSCs) and their exosomes in targeting tumors is a promising approach for curative therapy. Human tumor tropic mesenchymal stem cells (MSCs) isolated from various tissues and MSCs engineered to express the yeast cytosine deaminase::uracil phosphoribosyl transferase suicide fusion gene (yCD::UPRT-MSCs) released exosomes in conditional medium (CM). Exosomes from all tissue specific yCD::UPRT-MSCs contained mRNA of the suicide gene in the exosome's cargo. When the CM was applied to tumor cells, the exosomes were internalized by recipient tumor cells and in the presence of the prodrug 5-fluorocytosine (5-FC) effectively triggered dose-dependent tumor cell death by endocytosed exosomes via an intracellular conversion of the prodrug 5-FC to 5-fluorouracil. Exosomes were found to be responsible for the tumor inhibitory activity. The presence of microRNAs in exosomes produced from naive MSCs and from suicide gene transduced MSCs did not differ significantly. MicroRNAs from yCD::UPRT-MSCs were not associated with therapeutic effect. MSC suicide gene exosomes represent a new class of tumor cell targeting drug acting intracellular with curative potential.

In vitro response of dental pulp stem cells in 3D scaffolds: A regenerative bone material

Three-dimensional-porous scaffolds of bone graft substitutes play a critical role in both cell targeting and transplantation strategies. These scaffolds provide surfaces that facilitate the response of stem cells related to attachment, survival, migration, proliferation, and differentiation. ObjectiveThe aim of this study was to evaluate the in vitro behavior of human dental pulp mesenchymal stem cells cultured on scaffolds of polylactic/polyglycolic acid with and without hydroxyapatite. MethodWe performed an in vitro experimental study using dental pulp stem cells obtained from samples of premolars, molars. The cells were cultured on scaffolds with osteogenic differentiation medium. Cell proliferation, adhesion and cell differentiation to an osteoblastic linage in the biomaterial were evaluated at three different time points: 7, 15 and 30 days. Each experiment was performed in triplicate. Analysis of the data was performed with the Split Plot block and MANOVA model. ResultsThe differentiation capability of hDPSCs towards the osteoblast lineage was better in the scaffold of PLGA/HA at 7, 15 and 30 days, as indicated by the high expression of osteogenic markers RUNX2, ALP, OPN and COL-I, compared with differentiation in the PLGA scaffold. No statistically significant differences were found in cell adhesion between the two types of scaffolds. ConclusionThe PLGA/HA scaffold provided better physical and chemical signals, as judged by the ability of dental pulp stem cells to adhere, proliferate and differentiate toward the osteogenic lineage.

Abstract 339: A Novel Strategy to Harness Stem Cell-Intrinsic Mechanobiological Properties for Advanced Tissue Repair: Implication in Cardiac Tissue Injury

Rationale: Stem cells are constantly exposed to biomechanical and biochemical signals when transplanted in a continuously beating cardiac environment for treatment. Here, we postulated that an in vitro miniaturized beating cardiac microenvironment can induce unique biomechanical stimuli to co-cultured MSCs that can influence therapeutic potential of the later. This project aimed (i) to pre-condition mesenchymal stem cells (MSCs) with continuous mechanobiological stimuli using a 3D cardiac tissue environment (ii) to intramyocardially inject pre-conditioned MSCs into mice heart with myocardial infarction (MI) for tissue repair. Methods: We fabricated an in vitro cardiac platform as shown in Fig 1 , using an electro-conductive hydrogel and human iPSC-derived cardiac cells. Adipose MSCs were then injected to the beating hydrogel platform. After 7 days, activated (MB+) and non-activated MSCs (MB-) were isolated for in vitro and in vivo analysis. Results: The force generated by the constant beating of cardiomyocytes induced profound effects in on MSC behavior in the MB+ group. Several mechano-transduction factors (YAP1, TAZ), actin polymerization genes involved in cardiac muscle contractions ( ACTA2 , CDC42, TNNT2 ) and angiogenic genes were upregulated in MB+. RNA-seq analysis further confirmed these findings. 3 week study in mice with MI demonstrated significant increase in capillary density and reduction in infarct sizes in MB+ (n=6) compared to MB- group. Furthermore, the MB+ group showed significantly higher cardiac performance in echocardiography. We can conclude MB-activated MSCs can have significant impact in future treatment strategies for patients with MI.

Adhesion and differentiation behaviors of mesenchymal stem cells on titanium with micrometer and nanometer-scale grid patterns produced by femtosecond laser irradiation.

To clarify the effects of grid topographies with different scales on cell morphology and functionalization, we investigated the adhesion and differentiation of human mesenchymal stem cells (hMSCs) to titanium surfaces with micron, nano, and micron/nano (hybrid) grid topographies created by femtosecond laser irradiation. The results showed that cellular adhesion and differentiation strongly depended on the scales of the grid topography. hMSCs cultured on micron and hybrid grid topographies showed regulation of cellular adhesion plaques following the surface topography and were vinculin-positive, whereas filamentous vinculin was evident at the filopodia of hMSCs cultured on nanogrids. The findings indicate that the micron grid topography was beneficial for cell colonization by anchoring the cells to the substrate surface, whereas the nanogrid topography was beneficial for cell locomotion. With the superposition effect of the micron and nanogrids, micro/nanohybrid grid topography strongly promoted cell adhesion. This differential adhesion induced differences cell differentiation. Nanogrids promoted differentiation of hMSCs, particularly osteogenic differentiation. These findings provide a basis for the design of novel biomaterial surfaces that can regulate specific cellular functions. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 2735-2743, 2018.

Development of a Novel Orthogonal Double Gradient for High‐Throughput Screening of Mesenchymal Stem Cells–Materials Interaction

AbstractSurface gradients provide a powerful platform to accelerate multiscale design by efficiently studying of material–cell interactions to ultimately enhance function of synthetic clinical biomaterials. Herein, a novel orthogonal double gradient is reported in which surface stiffness and wettability vary independently and continuously in perpendicular directions, providing unique combinations of stiffness and wettability over a broad range (stiffness: 6–89 MPa; water contact angle: 29°–90°). It is found that mesenchymal stem cell behavior is nonlinearly regulated by surface stiffness and wettability. These combined stiffness and wettability properties of a material significantly affect stem cell adhesion, spreading, nucleus size, and vinculin expression. This high‐throughput screening system enables elucidation of the relationships between biointerface properties and biological behavior, and thereby serves a potential tool for accelerating the development of high‐performance biomaterials.

Poly(amidoamine)-alginate hydrogels: directing the behavior of mesenchymal stem cells with charged hydrogel surfaces

The surface charge of a biomaterial represents a promising tool to direct cellular behavior, which is crucial for therapeutic approaches in regenerative medicine. To expand the understanding of how the material surface charge affects protein adsorption and mesenchymal stem cell behavior, differently charged surfaces with zeta potentials spanning from −25 mV to +15 mV were fabricated by the conjugation of poly(amidoamine) to alginate-based hydrogels. We showed that the increase of the biomaterials surface charge resulted in enhanced quantities of biologically available, surface-attached proteins. Since different surface charges were equalized after protein adsorption, mesenchymal stem cells interacted rather with diverse protein compositions instead of different surface features. Besides an enhanced cell attachment to increasingly positively charged surfaces, the cell spreading area and the expression of adhesion-related genes integrin α5 and tensin 1 were found to be increased after adhesion. Moreover, first results indicate a potential impact of the surface charge on mesenchymal stem cell differentiation towards bone and fat cells. The improved understanding of surface charge-related cell behavior has significant impact on the design of biomedical devices and artificial organs.

Nanoscale Electrical Potential and Roughness of a Calcium Phosphate Surface Promotes the Osteogenic Phenotype of Stromal Cells.

Mesenchymal stem cells (MSCs) and osteoblasts respond to the surface electrical charge and topography of biomaterials. This work focuses on the connection between the roughness of calcium phosphate (CP) surfaces and their electrical potential (EP) at the micro- and nanoscales and the possible role of these parameters in jointly affecting human MSC osteogenic differentiation and maturation in vitro. A microarc CP coating was deposited on titanium substrates and characterized at the micro- and nanoscale. Human adult adipose-derived MSCs (hAMSCs) or prenatal stromal cells from the human lung (HLPSCs) were cultured on the CP surface to estimate MSC behavior. The roughness, nonuniform charge polarity, and EP of CP microarc coatings on a titanium substrate were shown to affect the osteogenic differentiation and maturation of hAMSCs and HLPSCs in vitro. The surface EP induced by the negative charge increased with increasing surface roughness at the microscale. The surface relief at the nanoscale had an impact on the sign of the EP. Negative electrical charges were mainly located within the micro- and nanosockets of the coating surface, whereas positive charges were detected predominantly at the nanorelief peaks. HLPSCs located in the sockets of the CP surface expressed the osteoblastic markers osteocalcin and alkaline phosphatase. The CP multilevel topography induced charge polarity and an EP and overall promoted the osteoblast phenotype of HLPSCs. The negative sign of the EP and its magnitude at the micro- and nanosockets might be sensitive factors that can trigger osteoblastic differentiation and maturation of human stromal cells.

Carbon nanotube loaded electrospun scaffolds based on thermoplastic urethane (TPU) with enhanced proliferation and neural differentiation of rat mesenchymal stem cells: The role of state of electrical conductivity

Electromagnetic stimulation has been shown as an effective strategy to enhance neural cells regeneration. The use of electrically conductive materials including polymer nanocomposites for the construction of scaffolds with potential to repair the injured nervous system has attracted great attentions. In the present study, attempts have been made to fabricate nanostructured fibrous scaffolds based on flexible thermoplastic polyurethane (TPU) and surface functionalized multiwalled carbon nanotubes (MWNT) via electrospinning process in order to examine and highlight the influence of state of electrical conductivity of scaffolds upon neural cells proliferation and differentiation. For this purpose, various scaffolds comprising different wt.% of MWNTs were prepared and their electrical conductivity, microstructural and mechanical characteristics were studied using two probe electrometer, scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM) and tensile measurements. Cellular behavior, proliferation and neural differentiation of rat mesenchymal stem cells (RMSC) seeded on the fabricated scaffolds with different state of conductivity were investigated both in the presence and absence of an external electromagnetic stimulation. Results revealed a linear correlation between electrical conductivity and cell signaling as well as neural gene expression. It is demonstrated that MWNT particles are encapsulated by the TPU and mostly oriented along the nanofibers axis by the electric field imposed on the jetted fibers during electrospinning process. Reinforcement of both bulk and surface modulus was exhibited by MWNT loaded scaffolds, indicating intensification of the interface between TPU and surface modified MWNT partices. Electrical conductivity of scaffolds versus MWNT wt% showed to follow percolation model with a threshold around 2.5 wt% of MWNTs, implying the involvement of both tunneling and conduction mechanisms for the passage of the electron current throughout the scaffolds upon stimulation. RMSCs cultured on MWNT loaded nanofibrous scaffolds with conductivity above threshold exhibited enhanced neural differentiation, suggesting the crucial role of state of electrical conductivity of the scaffolds for directing the proliferation and differentiation of stem cells towards the neural tissues.