Hemin Nie

Optimization of release pattern of FGF-2 and BMP-2 for osteogenic differentiation of low-population density hMSCs

In the modern design, most delivery systems for bone regeneration focus on a single growth factor (GF) or a simple mixture of multiple GFs, overlooking the coordination of proliferation and osteogenesis induced by various factors. In this study, core-shell microspheres with poly-l-lactide core-poly(lactic-co-glycolic acid) shell were fabricated, and two GFs, basic fibroblast growth factor 2 (FGF-2) and bone morphogenetic protein 2 (BMP-2) were encapsulated into the core or/and shell. The effects of different release patterns (parallel or sequential manners) of FGF-2 and BMP-2 from these core-shell microspheres on the osteogenic differentiation of low-population density human mesenchymal stem cells (hMSCs) were investigated and the temporal organization of GF release was optimized. In vitro experiments suggested that induction of osteogenic differentiation of low-population density hMSCs by the sequential delivery of FGF-2 followed by BMP-2 from the core-shell microspheres (group S2) was much more efficient than that by the parallel release of the two factors from uniform microspheres (group U). The osteogenic induction by the sequential delivery of BMP-2 followed by FGF-2 from core-shell microspheres (group S1) was even worse than that from microspheres loaded with BMP-2 in both core and shell (group B), although comparable to the cases of parallel delivery of dual GFs (group P). This study showed the advantages of group S2 microspheres in inducing osteogenic differentiation of low-population density hMSCs and the necessity of time sequence studies in tissue engineering while multiple GFs are involved.

Core–shell microspheres delivering FGF-2 and BMP-2 in different release patterns for bone regeneration

Bone regeneration by applying fibroblast growth factor-2 (FGF-2) plus bone morphogenetic protein-2 (BMP-2) is considered advantageous over a single growth factor (GF) treatment because the spontaneous repair of bone defects is essentially regulated by a series of GFs, including FGF-2 and BMP-2. However, the temporal interactions between FGF-2 and BMP-2 remain elusive and how to take full advantage of the interactions is a bottleneck in translating this dual-GF strategy into clinical applications. To compare the long-term effects of different temporal patterns of FGF-2 and BMP-2 on bone regeneration, a novel delivery system is needed. Herein, we report a type of PLLA core-PLGA shell double-walled microsphere. Different release patterns of FGF-2 and BMP-2 in the core and shell, respectively, were achieved due to different distributions of these two GFs. In vivo evaluations of different release patterns of dual GFs using a rat bone graft model suggested that a sequential delivery of FGF-2 and BMP-2 helped bridge and remodel the critical-sized bone grafts more efficiently than the other release patterns. More importantly, core-shell microspheres simultaneously and continuously releasing FGF-2 and BMP-2 resulted in the obvious resorption of grafted bone and bone non-union at 4 weeks. Consistently, the in vitro treatment of bovine bone sections by FGF-2 plus BMP-2 led to enhanced osteoclastogenesis compared to single GF treatments. The temporal organization of FGF-2 and BMP-2 by taking advantage of the core-shell microspheres will enable the development of more efficient devices for bone defects than existing delivery systems.

Effects of fiber alignment on stem cells–fibrous scaffold interactions

Considerable research has been focused on the utilization of electrospun ultrafine fibers to construct biomimetic scaffolds for tissue engineering applications. However, there is still a limited understanding of the effects of fiber alignment on stem cells-fibrous scaffold interactions and their role in deciding the fate of stem cells and scaffolds. Here, we tracked the real-time interactions between mesenchymal stem cells (MSCs) and aligned/non-aligned electrospun PCL-gelatin ultrafine fibers using time-lapse microscopy, and investigated the effects of fiber alignment on the behaviour of cell spreading, migration and differentiation, and scaffold remodeling. Cells were found to spread and migrate with increased velocity in the direction of aligned fibers as compared to non-aligned ones. Moreover, enhanced interactions between cells and aligned fibers were found to facilitate the infiltration of cells into the interior of fibrous constructs, which further led to enhanced micro-integration and scaffold remodeling as well as increased gene expression of late osteogenic markers as compared to non-aligned fibrous constructs. This study advances our understanding of the effects of fiber alignment on stem cells-fibrous scaffold interactions, and will facilitate the design of fibrous scaffolds with enhanced cell-matrix interactions for tissue engineering applications.

Understanding cell homing-based tissue regeneration from the perspective of materials

Homing of cells to their target organs for tissue defect repair poses a significant challenge to biomaterials scientists and tissue engineers, due to the low efficiency of homing of effective cells to defect sites as well as the difficulties in coordinating cell migration, adhesion, spreading and differentiations. Recent advances in biomaterials have successfully improved the efficiency of homing of mesenchymal stem cells (MSCs) and cell homing-based tissue regeneration. In this review, the process of cell-homing based tissue regeneration was discussed from three different perspectives, including cell surface engineering, scaffold optimization and signaling molecule interactions. Cell surface modification by using polymeric materials offers a simple way to administrate cell migration. Besides, the ordered or anisotropic structures are proved to be more efficient for cell adhesion, spreading and infiltration than relatively random or isotropy structures. Moreover, the coordinated release of different growth factors (GFs), e.g. achieved via core-shell microspheres, can orchestrate the biological processes, including cell growth and differentiations, and significantly enhance the osteogenic differentiation of low population density of MSCs. These developments in biomaterials are not only important for the fundamental understanding of material-cell interactions, but also help understand cell homing-based tissue regeneration from the perspective of materials, which is crucial for the design and fabrication of a new generation of highly functional biomaterials for tissue regeneration.

Bioinspired Engineering of Poly(ethylene glycol) Hydrogels and Natural Protein Fibers for Layered Heart Valve Constructs

Layered constructs from poly(ethylene glycol) (PEG) hydrogels and chicken eggshell membranes (ESMs) are fabricated, which can be further cross-linked by glutaraldehyde (GA) to form GA-PEG-ESM composites. Our results indicate that ESMs composed of protein fibrous networks show elastic moduli ∼3.3-5.0 MPa and elongation percentages ∼47-56%, close to human heart valve leaflets. Finite element simulations reveal obvious stress concentration on a partial number of fibers in the GA-cross-linked ESM (GA-ESM) samples, which can be alleviated by efficient stress distribution among multiple layers of ESMs embedded in PEG hydrogels. Moreover, the polymeric networks of PEG hydrogels can prevent mineral deposition and enzyme degradation of protein fibers from incorporated ESMs. The fibrous structures of ESMs retain in the GA-PEG-ESM samples after subcutaneous implantation for 4 weeks, while those from ESM and GA-ESM samples show early degradation to certain extent, suggesting the prevention of enzymatic degradation of protein fibers by the polymeric network of PEG hydrogels in vivo. Thus, these GA-PEG-ESM layered constructs show heterogenic structures and mechanical properties comparable to heart valve leaflets, as well as improved functions to prevent progressive calcification and enzymatic degeneration, which are likely used for artificial heart valves.

Fabrication of graphene oxide-β-cyclodextrin nanoparticle releasing doxorubicin and topotecan for combination chemotherapy

Controlled delivery of dual chemical drugs may increase the efficiency of chemotherapy. In this work, doxorubicin (DOX) was bound to adamantanecarboxylic acid (AD-COOH) through covalent linkage to form AD-DOX, while graphene oxide (GO) was modified by ethylenediamino-β-cyclodextrin (EDA-CD) to form graphene oxide-β-cyclodextrin (GO-CD), then GO-CD was assembled with as-produced AD-DOX via host–guest interaction. In contrast to the conjugation of DOX via covalent linkage, topotecan (TPT) was loaded onto GO through π–π stacking interaction. It was found that TPT/GO-CD/AD-DOX nanoparticles synchronised the delivery of DOX and TPT, and pH value played a critical role in determining the release profiles of dual drugs. Moreover, in vitro cellular investigations proved that TPT/GO-CD/AD-DOX nanoparticles were superior to free drugs and single-drug loaded nanoparticles in inhibiting the growth of HeLa cells. As a kind of injectable biomaterials, this controlled drug delivery device offers a solution to simultaneous delivery of dual chemical drugs in cancer lesion positions. Authors expect that this platform can be exploited for controlled delivery of various drug combinations for combination chemotherapy.

Inhibition of HeLa cell growth by doxorubicin-loaded and tuftsin-conjugated arginate-PEG microparticles

In order to improve the release pattern of chemotherapy drug and reduce the possibility of drug resistance, poly(ethylene glycol amine) (PEG)-modified alginate microparticles (ALG-PEG MPs) were developed then two different mechanisms were employed to load doxorubicin (Dox): 1) forming Dox/ALG-PEG complex by electrostatic attractions between unsaturated functional groups in Dox and ALG-PEG; 2) forming Dox-ALG-PEG complex through EDC-reaction between the amino and carboxyl groups in Dox and ALG, respectively. Additionally, tuftsin (TFT), a natural immunomodulation peptide, was conjugated to MPs in order to enhance the efficiency of cellular uptake. It was found that the Dox-ALG-PEG-TFT MPs exhibited a significantly slower release of Dox than Dox/ALG-PEG-TFT MPs in neutral medium, suggesting the role of covalent bonding in prolonging Dox retention. Besides, the release of Dox from these MPs was pH-sensitive, and the release rate was observably increased at pH 6.5 compared to the case at pH 7.4. Compared with Dox/ALG-PEG MPs and Dox-ALG-PEG MPs, their counterparts further conjugated with TFT more efficiently inhibited the growth of HeLa cells over a period of 48 h, implying the effectiveness of TFT in enhancing cellular uptake of MPs. Over a period of 48 h, Dox-ALG-PEG-TFT MPs inhibited the growth of HeLa cells less efficiently than Dox/ALG-PEG-TFT MPs but the difference was not significant (p > 0.05). In consideration of the prolonged and sustained release of Dox, Dox-ALG-PEG-TFT MPs possess the advantages for long-term treatment.

Alginate microcapsules co-embedded with MSCs and anti-EGF mAb for the induction of hair cell-like cells in guinea pigs by taking advantage of host EGF

As the irreversibility of hair cell loss in mammals is among the main reasons giving rise to permanent hearing loss, studies have been focused on the development of biological technologies to generate new hair cells as a means of replacing lost hair cells. Bone marrow-derived mesenchymal stem cells (BMSCs) possess the capacity to differentiate into hair cell-like cells, and may find applications in the regeneration of mammalian cochlear hair cells. In order to efficiently induce BMSCs into hair cells, alginate microcapsules co-delivering rat bone marrow-derived mesenchymal stem cells (rBMSCs) and anti-EGF monoclonal antibody (mAb) were developed to examine the feasibility of differentiating rBMSC into hair cell-likes cells in vitro and in vivo by taking advantage of epidermal growth factor (EGF) ligands. In vitro analysis showed that anti-EGF mAbs bonded with exogenous EGF ligands activated the EGF receptors on rBMSCs, enhancing the expression of myosin 7a (a hair cell marker) and Notch1 (supporting cell marker) via the EGFR/Ras/Raf/ERK1/2 signal pathway. In in vivo experiments, alginate microcapsules loaded with rBMSCs (2 × 106 cells per microcapsule) together with Iso mAbs or anti-EGF mAbs were grafted into guinea pig cochlea. After 6 weeks of treatment, immunofluorescence analyses indicated that the rBMSCs embedded into anti-EGF microcapsules were more efficiently transformed into hair cells compared with the group with Iso mAb microcapsules and displayed an ordered arrangement in Reissners membranes. The results highlighted the significance of engineering the microenvironment of stem cells for hair cell differentiation, and particularly the advantage of hair cell differentiation of rBMSCs by recruiting host EGF ligands via tethered mAbs. In conclusion, this strategy of co-embedding rBMSCs and anti-EGF mAb in alginate microcapsules is a promising modality for the regeneration of hair cell-like cells.

The Design and Improvement of Aptamer-based Fluorescent Probes

Fluorescent probes exhibit the characteristics of high sensitivity, diversified design, and strong quantitative analysis ability. Therefore, they have become one of the research hotspots in analytical chemistry and biological analysis. Aptamers are single-stranded DNAs or RNAs that are obtained in vitro and can selectively bind to the targets, making them ideal candidates as identifying units in fluorescent probes. However, most of the current aptamer probes are not regulatable because their fluorescence signals can’t be changed after the fluorescent probes are bound to the target, resulting in a high signal background and limited contrast. Actually, the structure of aptamers is flexible and can be combined with a variety of nanomaterials to achieve high sensitivity fluorescence for quantitative analysis, and can be used for a variety of target determinations, such as metal ions, small molecules, proteins and cells. In addition, with the development of research on DNA nanotechnology, researchers have designed many aptamer-based fluorescent probes with special properties, which have good application prospects in complex sample analyses and even in vivo cell imaging. In this review, the development of research in the field of aptamer-based fluorescent probes is surveyed and four mechanisms of aptamer fluorescent probes, including dye intercalation, conversion of DNA conformation, signal amplification, and mediation of nanomaterials, are comprehensively discussed. A R T I C L E H I S T O R Y Received: April 14, 2018 Revised: July 11, 2018 Accepted: September 14, 2018 DOI: 10.2174/2405465803666180919114755

Zero-dimensional, one-dimensional, two-dimensional and three-dimensional biomaterials for cell fate regulation

&NA; The interaction of biological cells with artificial biomaterials is one of the most important issues in tissue engineering and regenerative medicine. The interaction is strongly governed by physical and chemical properties of the materials and displayed with differentiated cellular behaviors, including cell self‐renewal, differentiation, reprogramming, dedifferentiation, or transdifferentiation as a result. A number of engineered biomaterials with micro‐ or nano‐structures have been developed to mimic structural components of cell niche and specific function of extra cellular matrix (ECM) over past two decades. In this review article, we briefly introduce the fabrication of biomaterials and their classification into zero‐dimensional (0D), one‐dimensional (1D), two‐dimensional (2D) and three‐dimensional (3D) ones. More importantly, the influence of different biomaterials on inducing cell self‐renewal, differentiation, reprogramming, dedifferentiation, and transdifferentiation was discussed based on the progress at 0D, 1D, 2D and 3D levels, following which the current research limitations and research perspectives were provided.