The odontogenic differentiation of human dental pulp stem cells on hydroxyapatite-coated biodegradable nanofibrous scaffolds

The authors aimed to design nanofibrous (NF) scaffolds that facilitate odontogenic and osteogenic differentiation of human dental pulp-derived mesenchymal stem cells (DPSCs) in vitro. For this purpose, hydroxyapatite (HA)–loaded poly (L-lactic acid)/poly (?-caprolactone) (PLLA:PCL 2;1) blend NFs were prepared using the electrospinning method. Alizarin red activity and cell viability were evaluated by MTT assay, and SEM revealed the proliferation properties of NF scaffolds. QRT-PCR results demonstrated that HA-loaded PLLA/PCL can lead to osteoblast/odontoblast differentiation in DPSCs through the up-regulation of related genes, thus indicating that electrospun biodegradable PCL/PLA/HA has remarkable prospects as scaffolds for bone and tooth tissue engineering.     

Development of poly (L-lactide-co-caprolactone) multichannel nerve conduit with aligned electrospun nanofibers for Schwann cell proliferation

Biomaterials are playing a significant role in understanding and promoting the plasticity and repair of the nervous system. Biomimetic nanofibrous scaffolds mimicking important features of the native extracellular matrix provide a promising strategy to restore functions or achieve favorable responses for tissue regeneration and autograft nerve conduit is one of the most promising nerve regeneration strategies. The present study is based on novel fabrication method by using a special collector for 3D multichannel nerve conduit, longitudinally oriented with aligned electrospun nanofibers. The conduit contained a high number of channels (varying from 7 to 19) and each channel showed a separate morphology. Nerve channels were fabricated with the varying length ranging from 4 to 9 cm and total diameter ranging from 2200 ± 40 µm to 3951 ± 196 µm, while the channel diameter ranging from 350 ± 86 µm to 780 ± 20 µm. It has been clearly shown that the average porosity of nerve conduits has reached almost 89%. In this study, we optimized the parameters to control the structural stability, including the size and the number of channels in the nerve conduit. We also checked in vitro cell biocompatibility of multichannel nerve conduit and demonstrated that Schwann cells have the tendency to grow along the direction of nanofibers and high cell growth was observed in high number of channels compared to low number of channels. These results elaborated the potential use of this biocompatible multichannel nerve conduit for further in vivo testing.     

Fabrication and evaluation of nanofibrous polyhydroxybutyrate valerate scaffolds containing hydroxyapatite particles for bone tissue engineering

Tissue engineering is a new approach for regeneration of damaged tissues. The current clinical methods such as autograft and allograft transplantation are not effective for repairing bone damages, mainly due to the limited available sources and the donor-site side effects. In this research, the nanocomposite poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV)/nano hydroxyapatite (nHA) scaffolds with different nHA ratios for bone regeneration were utilized. The diameter and porosity of scaffolds were approximately 200?nm and 74%, respectively. The degradability test of the scaffolds suggests a low degradation rate with total degradation of 30% after 3 months. Cytotoxicity result showed that cultured osteoblast cells (MC3T3) on nanocomposite scaffolds had superiority in terms of higher proliferation and attachment in comparison with PHBV scaffold. The protein expression of alkaline phosphatase illustrated that nanofibrous scaffold containing hydroxyapatite had the highest alkaline phosphatase activities as a result of better proliferation. These results recommend that PHBV/nHA scaffolds are suitable candidates for bone tissue engineering.     

Modification of nanofibrous scaffolds to mimic extracellular matrix in physical and chemical structuring

Regulating the surface chemistry and topography of the scaffolds is an effective way to improve the scaffolds' biocompatibility. A nanofibrous scaffold with topographical and chemical biomimicry to ECM was developed in this study. In which, PCL nanofibrous scaffold was employed as a substrate which was shish-kebab structured first to provide the topographical similarity with ECM and then coated with polydopamine and arginylglycylaspartic acid (RGD) to endow the chemical biomimicry. Fourier transform infrared (FTIR) spectroscopy and X-ray photoelectron spectroscopy (XPS) results confirmed the successful grafting of RGD on the scaffolds. It was proved that the introduced shish-kebab structure and grafted RGD not only enhanced the surface roughness but also improved the surface wettability of the nanofibrous scaffolds. Human umbilical vein endothelial cells (HUVECs) exhibited a more flourishing living state onto the modified scaffolds. Moreover, the present modification method was further employed to graft other bioactive molecules (VEGF) onto the scaffold to further investigate its applicability. HEF1 fibroblast cells displayed great affinity on the modified scaffolds, indicating that the green modification approach was highly feasible to enhance the synthesized scaffolds' biocompatibility, which not only provides topographical similarity but also enhances the chemical biomimicry simultaneously.     

Study on the properties of the electrospun silk fibroin/gelatin blend nanofibers for scaffolds

Silk fibroin (SF)/gelatin blend nanofibers membranes as scaffolds were fabricated successfully via electrospinning with different composition ratios in formic acid. The formation of intermolecular hydrogen bonds and the conformational transition of SF provided scaffolds with excellent mechanical properties. FTIR and DTA analysis showed the SF/gelatin nanofibers had more β‐sheet structures than the pure SF nanofibers. The former's breaking tenacity increased from 0.95 up to 1.60 MPa, strain at break was 7.6%, average fiber diameter was 89.2 nm, porosity was 87%, and pore diameter was 142 nm. MTT, H&E stain, and SEM results showed that the adhesion, spreading, and proliferation of human umbilic vein endothelium cells (HUVECs) and mouse fibroblasts on the SF/gelatin nanofibers scaffolds were definitely better than that on the SF nanofibers scaffolds. The scaffolds could replace the natural ECM proteins, support long‐term cell growth, form three‐dimensional networks of the nanofibrous structure, and grow in the direction of fiber orientation. Our results prove that the addition of gelatin improved the mechanical and biological properties of the pure SF nanofibers, these SF/gelatin blend nanofiber membranes are desirable for the scaffolds and may be a good candidate for blood vessel engineering scaffolds. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2009     

Fabrication and characterization of aligned nanofibrous PLGA/Collagen blends as bone tissue scaffolds

Aligned nanofibrous blends of poly (d, l-lactide-co-glycolide) (PLGA) and collagen with various PLGA/collagen compositions (80/20, 65/35 and 50/50) were fabricated by electrospinning and characterized for bone tissue engineering. Morphological characterization showed that the addition of collagen to PLGA resulted in narrowing of the diameter distribution and a reduction in average diameter. Differential scanning calorimetric (DSC) studies showed that the triple helix structure of the native collagen was not destroyed during the fabrication process. However, the blending had a marked effect on the overall enthalpy of the blends, whereby the total enthalpy decreased as the collagen content decreased. Thermogravimetric analysis showed the addition of collagen increased the hydrophilicity of the scaffolds. The crosslinking of collagen to increase the biostability was done using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) in ethanol and an overall ∼25% degree of crosslinking was achieved. The EDC crosslinking had little effect on the nanofibrous morphology of the 80/20 blend system; however, the nanofibrous features were compromised to some extent at higher collagen concentrations. The mechanical characterization under dry and wet conditions showed that increasing collagen content resulted in a tremendous decrease in the mechanical properties. However, crosslinking resulted in the increase in elastic modulus from 47 MPa to 83 MPa for the wet PLGA/Collagen 80/20 blend system, with little effect on the tensile strength. In conclusion, the aligned nanofibrous scaffold used in this study constitutes a promising material for bone tissue engineering.     

PHB和PHBV注塑和挤出成型工艺研究

对聚3-羟基丁酸酯(PHB)和聚3-羟基丁酸3-羟基戊酸酯(PHBV)的热性能和流变性能进行了测试和比较。结果表明:两者的加工温度范围均较窄,剪切变稀严重;PHBV比PHB的熔融温度和分解温度稍低。以此为依据,通过注塑和挤出成型实验,分析了PHB和PHBV的加工特点,确定了其注塑和挤出成型工艺条件。     

PHBV降解菌株的选育及其降解性能研究

从炼油厂污水处理车间活性污泥中筛选出一株具有降解3-羟基丁酸-co-3-羟基戊酸共聚酯(PHBV)能力的菌株。经菌体形态特征、菌落培养特征、生理生化反应考察以及16S rDNA序列比对,确定菌株属于食酸菌属。菌株经24 h培养,发酵液PHBV降解酶活力可达13.3 U/ml,实验条件下菌株可使PHBV薄膜在32 h内完全降解质,谱检验显示降解产物中仅见β-羟基丁酸单体未,见其他寡聚体。     

Inhibiting the vascular smooth muscle cells proliferation by EPC and DPPC liposomes encapsulated magnolol

Percutaneous transluminal coronary angioplasty (PTCA) is a non-surgical modality for treating stennosis. However, the recurrence of restenosis in 30-50% patients within 6 months is the major drawback of PTCA. The major reason of restenosis is the proliferation of the vascular smooth muscle cells (VSMCs). Magnolol, a pure compound extracted from Magnolia officinalis, encapsulated by liposome was investigated for inhibiting the VSMCs proliferation leading to restenosis by PTCA. 1,2-Diacyl-Sn-glycero-3-phosphocholine (EPC) and 1,2-dipalmitoyl-Sn-glycero-3-phosphocholine (DPPC) liposomes were utilized to encapsulate the magnolol. EPC liposome obtained the higher encapsulation efficiency than DPPC lipsomes from UV-vis spectroscopy study. The inhibiting efficiency of EPC and DPPC liposomes encapsulated magnolol was higher than pure magnonol. Magnolol encapsulated by EPC liposomes had better efficiency on inhibiting VSMCs than DPPC liposome. Addition of cholesterol in liposomes could slightly enhance the encapsulation efficiency. The particles sizer analysis revealed the average particles size of EPC and DPPC liposomes encapsulated magnolol became larger than pure EPC or DPPC liposomes. From the transmission electron microscopy (TEM) analysis, the magnolol seems to interfere with EPC and DPPC liposomes to form a homogeneous lipid bilayer.     

Effects of high glucose on vascular endothelial growth factor synthesis and secretion in aortic vascular smooth muscle cells from obese and lean zucker rats

Type 1 diabetes is characterized by insulin deficiency, type 2 by both insulin deficiency and insulin resistance: in both conditions, hyperglycaemia is accompanied by an increased cardiovascular risk, due to increased atherosclerotic plaque formation/instabilization and impaired collateral vessel formation. An important factor in these phenomena is the Vascular Endothelial Growth Factor (VEGF), a molecule produced also by Vascular Smooth Muscle Cells (VSMC). We aimed at evaluating the role of high glucose on VEGF-A(164) synthesis and secretion in VSMC from lean insulin-sensitive and obese insulin-resistant Zucker rats (LZR and OZR). In cultured aortic VSMC from LZR and OZR incubated for 24 h with d-glucose (5.5, 15 and 25 mM) or with the osmotic controls l-glucose and mannitol, we measured VEGF-A(164) synthesis (western, blotting) and secretion (western blotting and ELISA). We observed that: (i) d-glucose dose-dependently increases VEGF-A(164) synthesis and secretion in VSMC from LZR and OZR (n = 6, ANOVA p = 0.002-0.0001); (ii) all the effects of 15 and 25 mM d-glucose are attenuated in VSMC from OZR vs. LZR (p = 0.0001); (iii) l-glucose and mannitol reproduce the VEGF-A(164) modulation induced by d-glucose in VSMC from both LZR and OZR. Thus, glucose increases via an osmotic mechanism VEGF synthesis and secretion in VSMC, an effect attenuated in the presence of insulin resistance.     

Modification of electrospun poly(L-lactic acid)/polyethylenimine nanofibrous scaffolds for biomedical application

In this study, modification of poly(L-lactic acid) (PLLA) electrospun nanofibrous scaffolds blending with polyethylenimine (PEI) in different blend ratios was performed. The sample with 85:15 blend ratio revealed most promising results, and was selected for further modification with gelatin. It was found that the presence of PEI could enhanced porosity, mechanical properties, surface/bulk hydrophilicity and also gelatin grafting density about five times with positive effect on cell behavior. The results indicated that the limitations of PLLA electrospun nanofibers for potential application as a functional tissue engineering scaffold (i.e., poor cell adhesion and necrosis of host tissues as a result of providing acidic environment while degradation) could be overcome through blending with PEI and grafting with gelatin.     

Green fabrication route of robust,biodegradable silk sericin and poly(vinyl alcohol) nanofibrous scaffolds

Silk sericin (SS) has been extensively used to fabricate scaffolds for tissue engineering. However, due to its inferior mechanical properties, it has been found to be a poor choice of material when being electrospun into nanofibrous scaffolds. Here, SS has been combined with poly(vinyl alcohol) (PVA) and electrospun to create scaffolds with enhanced physical properties. Crucially, these SS/PVA nanofibrous scaffolds were created using only distilled water as a solvent with no added crosslinker in an environmentally friendly process. Temperature has been shown to have a marked effect on the formation of the SS sol–gel transition and thus influence the final formation of fibers. Heating the spinning solutions to 70 °C delivered nanofibers with enhanced morphology, water stability and mechanical properties. This is due to the transition of SS from β‐sheets into random coils that enables enhanced molecular interactions between SS and PVA. The most applicable SS/PVA weight ratios for the formation of nanofibers with the desired properties were found to be 7.5/1.5 and 10.0/1.5. The fibers had diameters ranging from 60 to 500 nm, where higher PVA and SS concentrations promoted larger diameters. The crystallinity within the fibers could be controlled by manipulation of the balance between PVA and SS loadings. In vitro degradation (in phosphate buffer solution, pH 7.4 at 37 °C) was 30–50% within 42 days and fibers were shown to be nontoxic to skin fibroblast cells. This work demonstrates a new green route for incorporating SS into nanofibrous fabrics, with potential use in biomedical applications. © 2019 Society of Chemical Industry     

Fabrication of gelatin/chitosan nanofibrous scaffold: process optimization and empirical modeling

Electrospinning is a very useful technique for producing polymeric nanofibers by applying electrostatic forces. This study reports on the modeling and optimization of the electrospinning process of gelatin/chitosan, using response surface methodology. The individual and the interaction effects of the gelatin/chitosan blend ratio (50/50, 60/40 and 70/30), applied voltage (20, 25 and 30 kV) and feeding rate (0.2, 0.4 and 0.6 mL h^?1) on the mean fiber diameter and standard deviation of the fiber diameter were investigated on optimization section, using scanning electron microscopy. To fabricate the nanofibrous gelatin/chitosan blend, trifluoroacetic acid/dichloromethane was selected as the solvent system. The model obtained for the mean fiber diameter has a quadratic relationship with applied voltage and feeding rate. The interaction between applied voltage and flow rate were found significant but the interactions of blend ratio and flow rate and also blend ratio and applied voltage were negligible. A quadratic relationship was obtained for applied voltage and flow rate with standard deviation of the fiber diameter and there was no interaction between the parameters in the model. The optimum condition for electrospinning of gelatin/chitosan was also introduced using the model obtained in this study. Scanning electron micrographs of human dermal fibroblast cells on the nanofibrous structures show good attachment and proliferation on the fabricated scaffold surface. Electrospun gelatin/chitosan nanofibrous mats have great potential for use as a scaffold for skin tissue engineering. © 2014 Society of Chemical Industry     

Influence of the nanofibrous morphology on the catalytic activity of NiO nanostructures: an effective impact toward methanol electrooxidation

In this study, the influence of the morphology on the electrocatalytic activity of nickel oxide nanostructures toward methanol oxidation is investigated. Two nanostructures were utilized: nanoparticles and nanofibers. NiO nanofibers have been synthesized by using the electrospinning technique. Briefly, electrospun nanofiber mats composed of polyvinylpyrolidine and nickel acetate were calcined at 700°C for 1 h. Interestingly, compared to nanoparticles, the nanofibrous morphology strongly enhanced the electrocatalytic performance. The corresponding current densities for the NiO nanofibers and nanoparticles were 25 and 6 mA/cm², respectively. Moreover, the optimum methanol concentration increased to 1 M in case of the nanofibrous morphology while it was 0.1 M for the NiO nanoparticles. Actually, the one-dimensional feature of the nanofibrous morphology facilitates electrons'' motion which enhances the electrocatalytic activity. Overall, this study emphasizes the distinct positive impact of the nanofibrous morphology on the electrocatalytic activity which will open a new avenue for modification of the electrocatalysts.     

Cellular Response of Limbal Stem Cells on PHBV/Gelatin Nanofibrous Scaffold for Ocular Epithelial Regeneration

The aim of this study was to develop a blend of nanofibrous poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV)/gelatin substrate for limbal stem cell (LSC) expansion that can serve as a potential alternative substrate to replace human amniotic membrane. The human Limbus stem cell was used to evaluate the biocompatibility of substrates (nanofibrous scaffold, and human amniotic membrane) based on their phenotypic profile, viability, proliferation, and attachment ability. Biocompatibility results indicated that the all substrates were highly biocompatible, as LSCs could favorably attach and proliferate on the nanofibrous surface. Microscopic figures showed that the human LSCs were firmly anchored to the substrates and were able to retain a normal corneal stem cell phenotype. Microscopic analyses illustrated that cells infiltrated the nanofibers and successfully formed a three-dimensional corneal epithelium, which was viable for two weeks. Immunocytochemistry (ICC) and real time–PCR results revealed no change in the expression profile of LSCs grown on nanofibrous substrate when compared to those grown on human amniotic membrane. In addition, electrospun nanofibrous PHBV substrate provides not only a milieu supporting LSCs expansion, but also serves as a useful alternative carrier for ocular surface tissue engineering and could be used as an alternative substrate to amniotic membrane.     

Evaluation of the chondrogenic differentiation of mesenchymal stem cells on hybrid biomimetic scaffolds

Hybrid materials are widely and promisingly used as scaffolds in cartilage tissue remodeling. In this study, hybrid scaffolds consist of polycaprolactone (PCL), poly(vinyl alcohol) (PVA) with/without gelatin (GEL) to mimic natural cartilage extracellular matrix (ECM) were investigated. Scaffolds were prepared by freeze drying and characterized by scanning electron microscopy and compressive mechanical testing. Biological assays of mesenchymal stem cell (MSC) cultures, 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide, and dimethyl methylene blue were performed, and real‐time polymerization chain reaction analysis of the cartilage‐specific ECM gene marker expression was done. The results show an open interconnected porous structure with a compression modulus of 1.27 ± 0.04 MPa. The surface of the scaffolds showed an excellent efficiency in the adhesion and proliferation of MSCs. A significant increase in the proteoglycan content from 3.70 ± 0.96 to 5.4 ± 1.13 μg/mL was observed after 14 days in the PCL–PVA–GEL scaffolds. The expression amount of the sex‐determining region Y–Box 9 (SOX9) and collagen II (COL2) mRNA levels of the MSCs showed significant increases in SOX9 and COL2, respectively in comparison with PCL–PVA scaffold. The study revealed that the aforementioned scaffold as a blend of natural and synthetic polymers may be a promising substrate in tissue engineering for cartilage repair with MSC transplantation. © 2014 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2014 , 131, 40635.     

Generation and Characterization of Human Mesenchymal Stem Cell-Derived Smooth Muscle Cells

Cardiovascular diseases are the leading cause of death worldwide. A completely autologous treatment can be achieved by using elastogenic mesenchymal stem cell (MSC)-derived smooth muscle cells (SMC) at the affected tissue site of vascular diseases such as abdominal aortic aneurysms (AAA). Thus, our work focused on evaluating the efficacy of (a) the combination of various growth factors, (b) different time periods and (c) different MSC lines to determine the treatment combination that generated SMCs that exhibited the greatest elastogenicity among the tested groups using Western blotting and flow cytometry. Additionally, total RNA sequencing was used to confirm that post-differentiation cells were upregulating SMC-specific gene markers. Results indicated that MSCs cultured for four days in PDGF + TGFβ1 (PT)-infused differentiation medium showed significant increases in SMC markers and decreases in MSC markers compared to MSCs cultured without differentiation factors. RNA Seq analysis confirmed the presence of vascular smooth muscle formation in MSCs differentiated in PT medium over a seven-day period. Overall, our results indicated that origin, growth factor treatment and culture period played a major role in influencing MSC differentiation to SMCs.     

Leptin in Atherosclerosis: Focus on Macrophages,Endothelial and Smooth Muscle Cells

Increasing adipose tissue mass in obesity directly correlates with elevated circulating leptin levels. Leptin is an adipokine known to play a role in numerous biological processes including regulation of energy homeostasis, inflammation, vascular function and angiogenesis. While physiological concentrations of leptin may exhibit multiple beneficial effects, chronically elevated pathophysiological levels or hyperleptinemia, characteristic of obesity and diabetes, is a major risk factor for development of atherosclerosis. Hyperleptinemia results in a state of selective leptin resistance such that while beneficial metabolic effects of leptin are dampened, deleterious vascular effects of leptin are conserved attributing to vascular dysfunction. Leptin exerts potent proatherogenic effects on multiple vascular cell types including macrophages, endothelial cells and smooth muscle cells; these effects are mediated via an interaction of leptin with the long form of leptin receptor, abundantly expressed in atherosclerotic plaques. This review provides a summary of recent in vivo and in vitro studies that highlight a role of leptin in the pathogenesis of atherosclerotic complications associated with obesity and diabetes.     

Engagement of the CXCL12–CXCR4 Axis in the Interaction of Endothelial Progenitor Cell and Smooth Muscle Cell to Promote Phenotype Control and Guard Vascular Homeostasis

Endothelial progenitor cells (EPCs) are involved in vascular repair and modulate properties of smooth muscle cells (SMCs) relevant for their contribution to neointima formation following injury. Considering the relevant role of the CXCL12–CXCR4 axis in vascular homeostasis and the potential of EPCs and SMCs to release CXCL12 and express CXCR4, we analyzed the engagement of the CXCL12–CXCR4 axis in various modes of EPC–SMC interaction relevant for injury- and lipid-induced atherosclerosis. We now demonstrate that the expression and release of CXCL12 is synergistically increased in a CXCR4-dependent mechanism following EPC–SMC interaction during co-cultivation or in response to recombinant CXCL12, thus establishing an amplifying feedback loop Additionally, mechanical injury of SMCs induces increased release of CXCL12, resulting in enhanced CXCR4-dependent recruitment of EPCs to SMCs. The CXCL12–CXCR4 axis is crucially engaged in the EPC-triggered augmentation of SMC migration and the attenuation of SMC apoptosis but not in the EPC-mediated increase in SMC proliferation. Compared to EPCs alone, the alliance of EPC–SMC is superior in promoting the CXCR4-dependent proliferation and migration of endothelial cells. When direct cell–cell contact is established, EPCs protect the contractile phenotype of SMCs via CXCL12–CXCR4 and reverse cholesterol-induced transdifferentiation toward a synthetic, macrophage-like phenotype. In conclusion we show that the interaction of EPCs and SMCs unleashes a CXCL12–CXCR4-based autoregulatory feedback loop promoting regenerative processes and mediating SMC phenotype control to potentially guard vascular homeostasis.