Guorui Jin

Guided orientation of cardiomyocytes on electrospun aligned nanofibers for cardiac tissue engineering.

Cardiac tissue engineering (TE) is one of the most promising strategies to reconstruct the infarct myocardium and the major challenge involves producing a bioactive scaffold with anisotropic properties that assist in cell guidance to mimic the heart tissue. In this study, random and aligned poly(ε-caprolactone)/gelatin (PG) composite nanofibrous scaffolds were electrospun to structurally mimic the oriented extracellular matrix (ECM). Morphological, chemical and mechanical properties of the electrospun PG nanofibers were evaluated by scanning electron microscopy (SEM), water contact angle, attenuated total reflectance Fourier transform infrared spectroscopy and tensile measurements. Results indicated that PG nanofibrous scaffolds possessed smaller fiber diameters (239 ± 37 nm for random fibers and 269 ± 33 nm for aligned fibers), increased hydrophilicity, and lower stiffness compared to electrospun PCL nanofibers. The aligned PG nanofibers showed anisotropic wetting characteristics and mechanical properties, which closely match the requirements of native cardiac anisotropy. Rabbit cardiomyocytes were cultured on electrospun random and aligned nanofibers to assess the biocompatibility of scaffolds, together with its potential for cell guidance. The SEM and immunocytochemical analysis showed that the aligned PG scaffold greatly promoted cell attachment and alignment because of the biological components and ordered topography of the scaffolds. Moreover, we concluded that the aligned PG nanofibrous scaffolds could be more promising substrates suitable for the regeneration of infarct myocardium and other cardiac defects.

Stem cell differentiation to epidermal lineages on electrospun nanofibrous substrates for skin tissue engineering.

Bone marrow (BM) mesenchymal stem cells (MSC) capable of differentiating along the epidermal lineage on engineered nanofibrous scaffolds have great potential for bionanomaterial-cell transplantation therapy of skin wounds. MSC have been the focus of many tissue engineering studies, mainly because of their multipotential properties. We investigated the potential of human BM-derived MSC for epidermal cell differentiation in vitro on electrospun collagen/poly(l-lactic acid)-co-poly(3-caprolactone) (Coll/PLLCL) nanofibrous scaffolds. PLLCL and Coll/PLLCL nanofibrous scaffolds were fabricated by an electrospinning process and their chemical and mechanical characterization carried out by scanning electron microscopy (SEM), water contact angle determination, Fourier transform infrared spectroscopy, and tensile testing. The differentiation of MSC was carried out using epidermis inducing factors, including epidermal growth factor (EGF) and 1,25-dihydroxyvitamin D(3), in culture medium. The proliferation of MSC evaluated by cell proliferation assay showed that the number of cells grown on Coll/PLLCL nanofibrous scaffolds was significantly higher than those on PLLCL scaffolds. The SEM results showed that MSC differentiated on Coll/PLLCL nanofibrous scaffolds showed a round keratinocyte morphology and expressed keratin 10, filaggrin and partial involucrin protein by immunofluorescent microscopic studies. The interaction of MSC and nanofibers was studied and we concluded that the electrospun Coll/PLLCL nanofibers could mimic the native skin extracellular matrix environment and are promising substrates for advanced skin tissue engineering. Our studies on the differentiation of MSC along the epidermal lineage on nanofibrous scaffolds suggest their potential application in skin regeneration without regional differentiation.

Electrospun conducting polymer nanofibers and electrical stimulation of nerve stem cells

Tissue engineering of nerve grafts requires synergistic combination of scaffolds and techniques to promote and direct neurite outgrowth across the lesion for effective nerve regeneration. In this study, we fabricated a composite polymeric scaffold which is conductive in nature by electrospinning and further performed electrical stimulation of nerve stem cells seeded on the electrospun nanofibers. Poly-L-lactide (PLLA) was blended with polyaniline (PANi) at a ratio of 85:15 and electrospun to obtain PLLA/PANi nanofibers with fiber diameters of 195 ± 30 nm. The morphology, chemical and mechanical properties of the electrospun PLLA and PLLA/PANi scaffolds were carried out by scanning electron microscopy (SEM), X-ray photo electron spectroscopy (XPS) and tensile instrument. The electrospun PLLA/PANi fibers showed a conductance of 3 × 10⁻⁹ S by two-point probe measurement. In vitro electrical stimulation of the nerve stem cells cultured on PLLA/PANi scaffolds applied with an electric field of 100 mV/mm for a period of 60 min resulted in extended neurite outgrowth compared to the cells grown on non-stimulated scaffolds. Our studies further strengthen the implication of electrical stimulation of nerve stem cells on conducting polymeric scaffolds towards neurite elongation that could be effective for nerve tissue regeneration.

Tissue engineered plant extracts as nanofibrous wound dressing

Use of plant extracts for treatment of burns and wound is a common practice followed over the decades and it is an important aspect of health management. Many medicinal plants have a long history of curative properties in wound healing. Electrospun nanofibers provide high porosity with large surface area-to-volume ratio and are more appropriate for cell accommodation, nutrition infiltration, gas exchange and waste excretion. Electrospinning makes it possible to combine the advantages of utilizing these plant extracts in the form of nanofibrous mats to serve as skin graft substitutes. In this study, we investigated the potential of electrospinning four different plant extracts, namely Indigofera aspalathoides, Azadirachta indica, Memecylon edule (ME) and Myristica andamanica along with a biodegradable polymer, polycaprolactone (PCL) for skin tissue engineering. The ability of human dermal fibroblasts (HDF) to proliferate on the electrospun nanofibrous scaffolds was evaluated via cell proliferation assay. HDF proliferation on PCL/ME nanofibers was found the highest among all the other electrospun nanofibrous scaffolds and it was 31% higher than the proliferation on PCL nanofibers after 9 days of cell culture. The interaction of HDF with the electrospun scaffold was studied by F-actin and collagen staining studies. The results confirmed that PCL/ME had the least cytotoxicity among the different plant extract containing scaffolds studied here. Therefore we performed the epidermal differentiation of adipose derived stem cells on PCL/ME scaffolds and obtained early and intermediate stages of epidermal differentiation. Our studies demonstrate the potential of electrospun PCL/ME nanofibers as substrates for skin tissue engineering.

Polypyrrole-contained electrospun conductive nanofibrous membranes for cardiac tissue engineering.

Cardiac tissue engineering (TE) is one of the most promising strategies to reconstruct infarct myocardium and the major challenge is to generate a bioactive substrate with suitable chemical, biological, and conductive properties, thus mimicking the extracellular matrix (ECM) both structurally and functionally. In this study, polypyrrole/poly(ε-caprolactone)/gelatin nanofibrous scaffolds were electrospun by incorporating different concentrations of polypyrrole (PPy) to PCL/gelatin (PG) solution. Morphological, chemical, mechanical, and biodegradation properties of the electrospun nanofibers were evaluated. Our data indicated that by increasing the concentration of PPy (0-30%) in the composite, the average fiber diameters reduced from 239 ± 37 nm to 191 ± 45 nm, and the tensile modulus increased from 7.9 ± 1.6 MPa to 50.3 ± 3.3 MPa. Conductive nanofibers containing 15% PPy (PPG15) exhibited the most balanced properties of conductivity, mechanical properties, and biodegradability, matching the requirements for regeneration of cardiac tissue. The cell proliferation assay, SEM, and immunostaining analysis showed that the PPG15 scaffold promote cell attachment, proliferation, interaction, and expression of cardiac-specific proteins better than PPG30. Electrospun PPG15 conductive nanofibrous scaffold could be desirable and promising substrates suitable for the regeneration of infarct myocardium and cardiac defects.

Controlled release of multiple epidermal induction factors through core–shell nanofibers for skin regeneration

With advances in the field of tissue engineering, it is increasingly recognized that biodegradable and biocompatible scaffolds incorporated with multiple wound healing mediators might serve as the most promising medical devices for skin tissue regeneration. Through controlled drug delivery, these medical devices can reduce the toxicity effects and optimize clinical efficiency. In this study, we first encapsulated multiple epidermal induction factors (EIF) such as the epidermal growth factor (EGF), insulin, hydrocortisone, and retinoic acid (RA) with gelatin and poly(L-lactic acid)-co-poly-(ε-caprolactone) (PLLCL) solutions and performed electrospinning by two different approaches: blend spinning and core-shell spinning. No burst release was detected from EIF encapsulated core-shell nanofibers; however, an initial 44.9% burst release from EIF blended nanofibers was observed over a period of 15 days. The epidermal differentiation potential of adipose-derived stem cells (ADSCs) was evaluated for EIF-containing scaffolds prepared either by core-shell spinning or by blend spinning. After 15 days of cell culture, the proliferation of ADSCs on EIF encapsulated core-shell nanofibers was the highest. Moreover, a higher percentage of ADSCs got differentiated to epidermal lineages on EIF encapsulated core-shell nanofibers compared to the cell differentiation on EIF blended nanofibers, which can be attributed to the sustained release of EIF from the core-shell nanofibers. Our study demonstrated that the EIF encapsulated core-shell nanofibers might serve as a promising tissue engineered graft for skin regeneration.

Development of nanofibrous cellulose acetate/gelatin skin substitutes for variety wound treatment applications:

The major component of fibrous extracellular matrix of dermis is composed of a complex combination of proteins and polysaccharides. Electrospun cellulose acetate/gelatin might be an effective simulator of the structure and composition of native skin and during this study, we electrospun cellulose acetate/gelatin membranes in various compositions and their performance as a scaffold for either skin tissue engineering or as a wound dressing was evaluated. Skin treatment products, whether tissue-engineered scaffolds or wound dressings, should be sufficiently hydrophilic to allow for gas and fluid exchange and absorb excess exudates while controlling the fluid loss. However, a wound dressing should be easily removable without causing tissue damage and a tissue-engineered scaffold should be able to adhere to the wound, and support cell proliferation during skin regeneration. We showed that these distinct adherency features are feasible just by changing the composition of cellulose acetate and gelatin in composite cellulose acetate/gelatin scaffolds. High proliferation of human dermal fibroblasts on electrospun cellulose acetate/gelatin 25:75 confirmed the capability of cellulose acetate/gelatin 25:75 nanofibers as a tissue-engineered scaffold, while the electrospun cellulose acetate/gelatin 75:25 can be a potential low-adherent wound dressing.

Electrospun synthetic and natural nanofibers for regenerative medicine and stem cells

Nanofibers are attractive substrates for tissue regeneration applications because they structurally mimic the native extracellular matrix. Electrospinning has been recognized as one of the most efficient techniques to fabricate polymer nanofibers. Recent research has demonstrated that cellular responses, for example attachment, proliferation and differentiation, can be modulated by tuning nanofiber properties. In combination with other processing techniques, such as particulate leaching or three‐dimensional printing, nanofibrous scaffolds incorporating macroporous networks could be developed to enhance infiltration of cells. Three dimensional nanofiber‐based constructs offer an opportunity to achieve advanced functional tissue regeneration. This review explores the advantageous effects of nanofibers on cell behaviors compared to traditional scaffolds.

Facile Layer-by-Layer Self-Assembly toward Enantiomeric Poly(lactide) Stereocomplex Coated Magnetite Nanocarrier for Highly Tunable Drug Deliveries

A highly tunable nanoparticle (NP) system with multifunctionalities was developed as drug nanocarrier via a facile layer-by-layer (LbL) stereocomplex (SC) self-assembly of enantiomeric poly(l-lactic acid) (PLLA) and poly(d-lactic acid) (PDLA) in solution using silica-coated magnetite (Fe3O4@SiO2) as template. The poly(lactide) (PLA) SC coated NPs (Fe3O4@SiO2@-SC) were further endowed with different stimuli-responsiveness by controlling the outermost layer coatings with respective pH-sensitive poly(lactic acid)-poly(2-dimethylaminoethyl methacrylate) (PLA-D) and temperature-sensitive poly(lactic acid)-poly(N-isopropylacrylamide) (PLA-N) diblock copolymers to yield Fe3O4@SiO2@SC-D and Fe3O4@SiO2@SC-N NPs, respectively, while the superparamagnetic properties of Fe3O4 were maintained. TEM images show a clearly resolved core-shell structure with a silica layer and sequential PLA SC co/polymer coating layers in the respective NPs. The well-designed NPs possess a size distribution in a range of 220-270 nm and high magnetization of 70.8-72.1 emu/g [Fe3O4]. More importantly, a drug release study from the as-constructed stimuli-responsive NPs exhibited sustained release profiles and the rates of release can be tuned by variation of external environments. Further cytotoxicity and cell culture studies revealed that PLA SC coated NPs possessed good cell biocompatibility and the doxorubicin (DOX)-loaded NPs showed enhanced drug delivery efficiency toward MCF-7 cancer cells. Together with the strong magnetic sensitivity, the developed hybrid NPs demonstrate a great potential of control over the drug release at a targeted site. The developed coating method can be further optimized to finely tune the nanocarrier size and operating range of pHs and temperatures for in vivo applications.

Design of polyhedral oligomeric silsesquioxane (POSS) based thermo-responsive amphiphilic hybrid copolymers for thermally denatured protein protection applications

A series of thermo-responsive amphiphilic hybrid copolymers with a random brush-like structure were synthesized by copolymerizing hydrophilic poly(ethylene glycol)methacrylate (PEGMA) and hydrophobic polyhedral oligomeric silsesquioxane methacrylate (POSSMA) together with temperature sensitive poly(propylene glycol)methacrylate (PPGMA) via atom transfer radical polymerization (ATRP). The resulting poly(PEGMA–PPGMA–POSSMA) (PEPS) hybrid copolymers show a lower critical solution temperature (LCST) in the range of 31–33 °C. Static and dynamic light scattering (SLS and DLS) studies show that micellar structures created by the PEPS copolymer in aqueous media were core-shell structures and possessed a thick hydration layer. The presence of a small amount of POSS (3.1 wt%) in the PEPS copolymers lowered the CMC of the micelles at room temperature by one order of magnitude compared to samples without POSSMA (PEP). Incorporation of POSSMA also enhanced the stability of the formed micelles, i.e. PEPS containing 6.7 wt% POSS exhibited a constant hydrodynamic radius, Rh (∼65 nm), and an aggregation number, Nagg (∼350), when the temperature was varied from 20 to 70 °C while PEP without POSS showed a large increase in both Rh and Nagg values. On the other hand, the change of Rg as the temperature increases could be attributed to the PPG brush adopting a more extended and compact conformation below and above LCST respectively. The thermo-responsiveness of the PPG brush in PEPS hybrid micelles was also exploited to mimic the natural GroEL–GroES chaperone functionalities for renaturation of thermally denatured proteins. Above LCST of PPG, the chaperone-like system comes into effect with hydrophobic PPG domains on the micelle surface, providing spontaneous capture and protection of the unfolded proteins, thus inhibiting the undesired protein aggregation at elevated temperatures. Upon cooling, PPG returns to its hydrophilic state, thereby inducing the release of the bound unfolded proteins. The renaturation process of the detached proteins is spontaneously accomplished by the presence of PEG and OH-groups in the micelle corona. The working mechanism and thermal denaturation protection effect were also investigated by DLS, SLS and circular dichroism (CD) spectroscopy. In the presence of PEPS hybrid micelles, the protection efficiencies for GFP, lipase and lysozyme that can be achieved during the heat-induced denaturation process are 81.4%, 89.3% and 88.7%, respectively. Cell culture and cytotoxicity studies revealed that the PEPS hybrid micelles could be effectively internalized by C6 glioma cells and possess good cell biocompatibility. These interesting findings open up new opportunities to exploit PEPS hybrid copolymers as artificial chaperones for protecting unfolded proteins from toxic aggregation at high temperatures.