ABSTRACT
Objective: Three-dimensional [3D] biomimetic nanofiber scaffolds have widespread applications in biomedical tissue engineering. They provide a suitable environment for cellular adhesion, survival, proliferation and differentiation, guide new tissue formation and development, and are one of the outstanding goals of tissue engineering. Electrospinning has recently emerged as a leading technique for producing biomimetic scaffolds with micro to nanoscale topography and a high porosity similar to the natural extracellular matrix [ECM]. These scaffolds are comprised of synthetic and natural polymers for tissue engineering applications. Several kinds of cells such as human embryonic stem cells [hESCs] and mouse ESCs [mESCs] have been cultured and differentiated on nanofiber scaffolds. mESCs can be induced to differentiate into a particular cell lineage when cultured as embryoid bodies [EBs] on nano-sized scaffolds
Materials and Methods: We cultured mESCs [2500 cells/100 Mul] in 96-well plates with knockout Dulbecco's modified eagle medium [DMEM-KO] and Roswell Park Memorial Institute-1640 [RPMI-1640], both supplemented with 20% ESC grade fetal bovine serum [FBS] and essential factors in the presence of leukemia inhibitory factor [LIF]. mESCs were seeded at a density of 2500 cells/100 Mul onto electrospun polycaprolactone [PCL] nanofibers in 96-well plates. The control group comprised mESCs grown on tissue culture plates [TCP] at a density of 2500 cells/100 Mul. Differentiation of mESCs into mouse hematopoietic stem cells [mHSCs] was performed by stem cell factor [SCF], interleukin-3 [IL-3], IL-6 and Fms-related tyrosine kinase ligand [Flt3-L] cytokines for both the PCL and TCP groups. We performed an experimental study of mESCs differentiation
Results: PCL was compared to conventional TCP for survival and differentiation of mESCs to mHSCs. There were significantly more mESCs in the PCL group. Flowcytometric analysis revealed differences in hematopoietic differentiation between the PCL and TCP culture systems. There were more CD34+ [Sca1+] and CD133+ cells subpopulations in the PCL group compared to the conventional TCP culture system
Conclusion: The nanofiber scaffold, as an effective surface, improves survival and differentiation of mESCs into mHSCs compared to gelatin coated TCP. More studies are necessary to understand how the topographical features of electrospun fibers affect cell growth and behavior. This can be achieved by designing biomimetic scaffolds for tissue engineering
Materials and Methods: We cultured mESCs [2500 cells/100 Mul] in 96-well plates with knockout Dulbecco's modified eagle medium [DMEM-KO] and Roswell Park Memorial Institute-1640 [RPMI-1640], both supplemented with 20% ESC grade fetal bovine serum [FBS] and essential factors in the presence of leukemia inhibitory factor [LIF]. mESCs were seeded at a density of 2500 cells/100 Mul onto electrospun polycaprolactone [PCL] nanofibers in 96-well plates. The control group comprised mESCs grown on tissue culture plates [TCP] at a density of 2500 cells/100 Mul. Differentiation of mESCs into mouse hematopoietic stem cells [mHSCs] was performed by stem cell factor [SCF], interleukin-3 [IL-3], IL-6 and Fms-related tyrosine kinase ligand [Flt3-L] cytokines for both the PCL and TCP groups. We performed an experimental study of mESCs differentiation
Results: PCL was compared to conventional TCP for survival and differentiation of mESCs to mHSCs. There were significantly more mESCs in the PCL group. Flowcytometric analysis revealed differences in hematopoietic differentiation between the PCL and TCP culture systems. There were more CD34+ [Sca1+] and CD133+ cells subpopulations in the PCL group compared to the conventional TCP culture system
Conclusion: The nanofiber scaffold, as an effective surface, improves survival and differentiation of mESCs into mHSCs compared to gelatin coated TCP. More studies are necessary to understand how the topographical features of electrospun fibers affect cell growth and behavior. This can be achieved by designing biomimetic scaffolds for tissue engineering
ABSTRACT
Superparamagnetic iron oxide nanoparticles [SPIONs] have been used to label mammalian cells and to monitor their fate in vivo using magnetic resonance imaging [MRI]. However, the effectiveness of phenotype of labeled cells by SPIONs is still a matter of question. The aim of this study was to investigate the efficiency and biological effects of labeled mouse embryonic stem cells [mESCs] using ferumoxide- protamine sulfate complex. In an experimental study, undifferentiated mESCs, C571 line, a generous gift of Stem Cell Technology Company, were cultured on gelatin-coated flasks. The proliferation and viability of SPION-labeled cells were compared with control. ESCs and embryoid bodies [EBs] derived from differentiated hematopoietic stem cells [HSCs] were analyzed for stage-specific cell surface markers using fluorescence-activated cell sorting [FACS]. Our observations showed that SPIONs have no effect on the self-renewal ability of mESCs. Reverse microscopic observations and prussian blue staining revealed 100% of cells were labeled with iron particles. SPION-labeled mESCs did not significantly alter cell viability and proliferation activity. Furthermore, labeling did not alter expression of representative surface phenotypic markers such as stage-specific embryonic antigen 1 [SSEA1] and cluster of differentiation 117 [CD117] on undifferentiated ESC and CD34, CD38 on HSCs, as measured by flowcytometry. According to the results of the present study, SPIONs-labeling method as MRI agents in mESCs has no negative effects on growth, morphology, viability, proliferation and differentiation that can be monitored in vivo, noninvasively. Non-invasive cell tracking methods are considered as new perspectives in cell therapy for clinical use and as an easy method for evaluating the placement of stem cells after transplantation