Saeid Vakilian

The synergistic effect of surface topography and sustained release of TGF‐β1 on myogenic differentiation of human mesenchymal stem cells

A combination of topographical cues and controlled release of biochemical factors is a potential platform in controlling stem cells differentiation. In this study the synergistic effect of nanotopography and sustained release of biofunctional transforming growth factor beta 1 (TGF-β1) on differentiation of human Whartons Jelly-derived mesenchymal stem cell (hWJ-derived UC-MSCs) toward myogenic lineage was investigated. In order to achieve a sustained release of TGF-β1, this factor was encapsulated within chitosan nanoparticles. Afterwards the aligned composite mats were fabricated using poly-ɛ-caprolacton (PCL) containing TGF-β1-loaded chitosan nanoparticles and poly-L-lactic acid (PLLA). The nanofiber topography notably up-regulated the expressions of calponin1 and SM22α compared with tissue culture polystyrene (TCP). Moreover, the combination of nanofiber topography and sustained TGF-β1release resulted in more significant enhancement of SMC marker, in particular smooth muscle α-actin (ASMA) expression, compared with bolus delivery despite lower amounts of TGF-β1 (>10 times lower). Additionally, immunofluorescence staining showed that ASMA and desmin were expressed at higher intensity in cells exposed to controlled TGF-β1 delivery rather than bolus delivery. These results demonstrated the importance of combined effect of topography and drug delivery in directing stem cell fate and the potential of such biofunctional scaffolds for cell transplantation applications in bladder tissue engineering.

Structural stability and sustained release of protein from a multilayer nanofiber/nanoparticle composite

The cellular microenvironment can be engineered through the utilization of various nano-patterns and matrix-loaded bioactive molecules. In this study, a multilayer system of electrospun scaffold containing chitosan nanoparticles was introduced to overcome the common problems of instability and burst release of proteins from nanofibrous scaffolds. Bovine serum albumin (BSA)-loaded chitosan nanoparticles was fabricated based on ionic gelation interaction between chitosan and sodium tripolyphosphate. Suspension electrospinning was employed to fabricate poly-ɛ-caprolacton (PCL) containing protein-loaded chitosan nanoparticles with a core-shell structure. To obtain the desired scaffold mechanical properties with enough elasticity for expansion and contraction, a hybrid mono and multilayer electrospun scaffold was fabricated using PCL containing protein-loaded chitosan nanoparticles and poly-L-lactic acid (PLLA). According to the BSA release profile, the multi-layered structure of nanofibers with two barrier layers provided a programmable release pattern of the loaded protein. Moreover, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and circular dichroism spectra results showed that the electrospinning process had no significant effect on the primary and secondary structure of the protein. The results indicated a desirable biocompatibility and mechanical cues of the multilayer nanofibrous scaffolds supporting structural stability and controlled release of the protein, which can offer diverse applications in hollow organ tissue engineering.

Improved proliferation and osteogenic differentiation of mesenchymal stem cells on polyaniline composited by polyethersulfone nanofibers

Tissue engineering is a promising emerged method trying to reconstruct lost tissues that using synthetic and biomaterials and their combination with cells. The purpose of this study is increase osteoinductivity of polyethersulfone (PES) by using polyaniline (PANi). In this study, after fabrication of PES and composited PES-PANi scaffolds by electrospinning, scaffolds were characterized morphologically and mechanically. Then osteoinductivity of scaffolds was investigated by osteogenic differentiation of human mesenchymal stem cells (MSCs) cultured on the PES and PES-PANI in comparison to the tissue culture polystyrene as a control. The Osteogenic potential of MSCs was evaluated by Alizarin Red staining, ALP activity, calcium content assay and bone related gene expression assay. Scaffolds were smooth, bead free and in the scale of nanometers and PES mechanical stability was decreased significantly after composite with PANi. Highest growth, ALP activity and deposited calcium of cells were observed in the PES-PANi groups compared to the PES and TCPS. Significantly higher expression of collagen type one and osteonectin was also detected in the PES-PANi group. It can be concluded that PES-PANi construct has potential to be a good candidate as bone grafting substitute and using in tissue engineering applications.

Enhanced Cardiac Differentiation of Human Cardiovascular Disease Patient-Specific Induced Pluripotent Stem Cells by Applying Unidirectional Electrical Pulses Using Aligned Electroactive Nanofibrous Scaffolds

In the embryonic heart, electrical impulses propagate in a unidirectional manner from the sinus venosus and appear to be involved in cardiogenesis. In this work, aligned and random polyaniline/polyetersulfone (PANI/PES) nanofibrous scaffolds doped by Camphor-10-sulfonic acid (β) (CPSA) were fabricated via electrospinning and used to conduct electrical impulses in a unidirectional and multidirectional fashion, respectively. A bioreactor was subsequently engineered to apply electrical impulses to cells cultured on PANI/PES scaffolds. We established cardiovascular disease-specific induced pluripotent stem cells (CVD-iPSCs) from the fibroblasts of patients undergoing cardiothoracic surgeries. The CVD-iPSCs were seeded onto the scaffolds, cultured in cardiomyocyte-inducing factors, and exposed to electrical impulses for 1 h/day, over a 15-day time period in the bioreactor. The application of the unidirectional electrical stimulation to the cells significantly increased the number of cardiac Troponin T (cTnT+) cells in comparison to multidirectional electrical stimulation using random fibrous scaffolds. This was confirmed by real-time polymerase chain reaction for cardiac-related transcription factors (NKX2.5, GATA4, and NPPA) and a cardiac-specific structural gene (TNNT2). Here we report for the first time that applying electrical pulses in a unidirectional manner mimicking the unidirectional wave of electrical stimulation in the heart, could increase the derivation of cardiomyocytes from CVD-iPSCs.

Adapted dexamethasone delivery polyethylene oxide and poly(ɛ-caprolactone) construct promote mesenchymal stem cells chondrogenesis

Abstract Issues associated with tissue transplantation and shortage of donors has always been a concern, whereas tissue engineering has provided the hopeful opportunities. The aim of this study was to investigate the chondrogenic differentiation potential of mesenchymal stem cells (MSCs) in the presence of embedded Dexamethasone into electrospun Poly(ethyleneoxide) nanofibers composited with Poly(ɛ-caprolactone) nanofibers. Electrospun-fabricated scaffolds were characterized by SEM, tensile, contact angle, release profile, MTT assay, and chonderogenic differentiation of stem cells. Results demonstrated Poly(ɛ-caprolactone) properties improved by composite with a Poly(ethyleneoxide). Overall, stem cells seeded on nanofibers content drug showed highest chondrogenic potential in comparison to the other groups.

G-CSF loaded nanofiber/nanoparticle composite coated with collagen promotes wound healing in vivo

Sustained release of functional growth factors can be considered as a beneficial methodology for wound healing. In this study, recombinant human granulocyte colony-stimulating factor (G-CSF)-loaded chitosan nanoparticles were incorporated in Poly(ε-caprolactone) (PCL) nanofibers, followed by surface coating with collagen type I. Physical and mechanical properties of the PCL nanofibers containing G-CSF loaded chitosan nanoparticles PCL/NP(G-CSF) and in vivo performance for wound healing were investigated. G-CSF structural stability was evaluated through SDS_PAGE, reversed phase (RP) HPLC and size-exclusion chromatography, as well as circular dichroism. Nanofiber/nanoparticle composite scaffold was demonstrated to have appropriate mechanical properties as a wound dresser and a sustained release of functional G-CSF. The PCL/NP(G-CSF) scaffold showed a suitable proliferation and well-adherent morphology of stem cells. In vivo study and histopathological evaluation outcome revealed that skin regeneration was dramatically accelerated under PCL/NP(G-CSF) as compared with control groups. Superior fibroblast maturation, enhanced collagen deposition and minimum inflammatory cells were also the beneficial properties of PCL/NP(G-CSF) over the commercial dressing. The synergistic effect of extracellular matrix-mimicking nanofibrous membrane and G-CSF could develop a suitable supportive substrate in order to extensive utilization for the healing of skin wounds.

The role of miR-17-92 cluster in the expression of tumor suppressor genes in unrestricted somatic stem cells

The miR-17-92 cluster consisted of seven miRNAs (mir-17-5p, -17-3p, -18a, -19a, -20a, -19b-1, and -92a-1). Previous studies have shown this cluster has been over-expressed in several cancers. The aim of this study was to evaluate the over-expression impacts of miR-17-92 on stem cells. In the current work, the effect of miR-17-92 cluster which was cloned in Lentiviral vector has been investigated on unrestricted somatic stem cells (USSCs). Tumor suppressor genes (p53, p15, RBL1, SMAD2, SMAD4, and MAPK-1) expression, especially p53, was considerably reduced. These data show the potential of miR-17-92 for oncogenesis regulation in stem cells. In conclusion, the role of miR-17-92 in USSCs may provide a better understanding of its function in tumorigenesis and for the possible use in cell therapy of the anti-mir-17-92 cluster.

Modeling and optimization of gelatin-chitosan micro-carriers preparation for soft tissue engineering: Using Response Surface Methodology

Electrospray ionization is a wide spread technique for producing polymeric microcarriers (MCs) by applying electrostatic force and ionic cross-linker, simultaneously. In this study, fabrication process of gelatin-chitosan MCs and its optimization using the Response Surface Methodology (RSM) is reported. Gelatin/chitosan (G/C) blend ratio, applied voltage and feeding flow rate, their individual and interaction effects on the diameter and mechanical strength of the MCs were investigated. The obtained models for diameter and mechanical strength of MCs have a quadratic relationship with G/C blend ratio, applied voltage and feeding flow rate. Using the desirability curve, optimized G/C blend ratios that are introduced, include the desirable quantities for MCs diameter and mechanical strength. MCs of the same desirable diameter (350μm) and different G/C blend ratio (1, 2, and 3) were fabricated and their elasticity was investigated via Atomic Force Microscopy (AFM). The biocompatibility of the MCs was evaluated using MTT assay. The results showed that human Umbilical Cord Mesenchymal Stem Cells (hUCMSCs) could attach and proliferate on fabricated MCs during 7days of culturing especially on those prepared with G/C blend ratios of 1 and 2. Such gelatin-chitosan MCs may be considered as a promising candidate for injectable tissue engineering scaffolds, supporting attachment and proliferation of hUCMSCs.

Renal Differentiation of Mesenchymal Stem Cells Seeded on Nanofibrous Scaffolds Improved by Human Renal Tubular Cell Lines-conditioned Medium

Kidney injuries and renal dysfunctions are one of the most important clinical problems, and tissue engineering could be a valuable method for solving it. The objective of this study was to investigate the synergistic effect of renal cell line-conditioned medium and Polycaprolactone (PCL) nanofibers on renal differentiation of human mesenchymal stem cells (MSCs). In the current study, after stem cells isolation and characterization, PCL nanofibrous scaffold was fabricated using electrospinning methods and characterized morphologically, mechanically, and for biocompatibility. The renal differentiation of seeded MSCs on the surface of PCL nanofibers with and without human renal tubular cell lines-conditioned medium was investigated by evaluation of eight important renal-related genes expression by real-time reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry. Fabricated nanofibrous scaffolds were good in all characterized items. Almost highest expression of all genes was detected in stem cells seeded on PCL under conditioned media in comparison with the stem cells seeded on PCL, tissue culture polystyrene (TCPS) under renal induction medium, and TCPS under conditioned medium. According to the results, PCL nanofibers in contribution with conditioned medium can provide the optimal conditions for renal differentiation of MSCs and could be a promising candidate for renal tissue engineering application.

Primordial germ cell differentiation of nuclear transfer embryonic stem cells using surface modified electroconductive scaffolds.

A combination of nanotopographical cues and surface modification of collagen and fibronectin is a potential platform in primordial germ cells (PGCs) differentiation. In the present study, the synergistic effect of nanotopography and surface modification on differentiation of nuclear transfer embryonic stem cells (nt-ESCs) toward PGC lineage was investigated. In order to achieve this goal, poly-anyline (PANi) was mix within poly-l-lactic acid (PLLA). Afterward, the random composite mats were fabricated using PLLA and PANi mix solution. The nanofiber topography notably upregulated the expressions of prdm14, mvh and c-kit compared with tissue culture polystyrene (TCP). Moreover, the combination of nanofiber topography and surface modification resulted in more enhancement of PGCs differentiation compared with non-modified nanofibrous scaffold. Additionally, gene expression results showed that mvh and c-kit were expressed at higher intensity in cells exposed to collagen and fibronectin rather than collagen or fibronectin solitary. These results demonstrated the importance of combined effect of collagen and fibronectin in order to develop a functional extracellular matrix (ECM) mimic in directing stem cell fate and the potential of such biofunctional scaffolds for treatment of infertility.