Progress in microsphere-based scaffolds in bone/cartilage tissue engineering.

Bone/cartilage repair and regeneration have been popular and difficult issues in medical research. Tissue engineering is rapidly evolving to provide new solutions to this problem, and the key point is to design the appropriate scaffold biomaterial. In recent years, microsphere-based scaffolds have been considered suitable scaffold materials for bone/cartilage injury repair because microporous structures can form more internal space for better cell proliferation and other cellular activities, and these composite scaffolds can provide physical/chemical signals for neotissue formation with higher efficiency. This paper reviews the research progress of microsphere-based scaffolds in bone/chondral tissue engineering, briefly introduces types of microspheres made from polymer, inorganic and composite materials, discusses the preparation methods of microspheres and the exploration of suitable microsphere pore size in bone and cartilage tissue engineering, and finally details the application of microsphere-based scaffolds in biomimetic scaffolds, cell proliferation and drug delivery systems.

Progress of biomaterials for bone tumor therapy.

Bone tumors are currently a major clinical challenge. In recent decades, strategies using well-designed versatile biomaterials for the treatment of bone tumors have emerged and attracted extensive research interest. Suitable biomaterials not only facilitate repair for bone defects aroused by surgical intervention but also help deliver antineoplastic drugs to the target site or provide photothermal/magnetothermal therapy to kill bone tumor cells. Thus, the development of biomaterials exhibits a great perspective for future bone tumor treatment.We summarize the recent progress of versatile biomaterials for bone tumor therapy, with an emphasis on photothermal/magnetothermal therapy and drug delivery.With the further understanding and development of biomaterials, multifunctional biomaterials have been proposed for bone tumor treatment. Through the interdisciplinary cooperation from the fields of biomedicine, clinical medicine and engineering, multifunctional biomaterials will perfectly match individual bone defects in the clinic with low cost in the future.

Prospects of cell chemotactic factors in bone and cartilage tissue engineering.

INTRODUCTION: Tissue engineering has brought hope for the repair of bone and cartilage injury. As potential therapeutic molecules for use in tissue engineering, chemokines promote the development of cell-free tissue engineering, avoiding dilemmas faced by cell-based tissue engineering. The main role of chemokines in tissue engineering is to recruit progenitor/stem cells to the site of damaged tissue in vivo and induce differentiation into the corresponding tissue, thus remodeling tissue function. In recent years, many studies have demonstrated the great potential of chemokines in the regeneration and repair of various tissues, such as heart, bone and cartilage tissue. AREAS COVERED: The classification, structure, and function of chemokines and the application of several common chemokines in diseases, especially in bone/cartilage tissue regeneration are discussed. EXPERT OPINION: Many studies have demonstrated that the combinatory use of cell chemotactic factors (CCFs) and growth factors can exert synergistic effects on chondrogenesis and osteogenesis. With further understanding of biomaterials and the development of powerful bio-fabrication techniques, intelligent biomaterials will be created to meet the requirements for controlled bioactive factor release and biomimetic architecture. Also, a better understanding of the biological cascade reactions and pathways of CCFs is beneficial to guide the design of innovative biomaterials.

The Role of Mechanical Regulation in Cartilage Tissue Engineering.

Due to the lack of vascular distribution and the slow metabolism, cartilage tissue cannot repair itself, which remains a huge challenge in cartilage regeneration. Tissue engineering using stem cells appears to be a promising method for cartilage repair. Tissue engineers demonstrated that mechanical stimulation can enhance the quality of engineered cartilage, making it more similar to natural cartilage in structure and function. In this review, we summarize recent studies on the role of mechanical stimuli in chondrogenesis, focusing on the applications of extrinsic mechanical loading and the studies on mechanical properties of biomaterials in cartilage tissue engineering. This review will provide fresh insights into the potential use of mechanical stimuli for clinical use.

Gene-modified BMSCs encapsulated with carboxymethyl cellulose facilitate osteogenesis in vitro and in vivo.

Critical size bone defects are one of the most serious complications in orthopedics due to the lack of effective osteogenesis treatment. We fabricated carboxymethyl cellulose with phenol moieties (CMC-ph) microcapsules loaded with gene-modified rat bone mesenchymal stem cells (rBMSCs) that secrete hBMP2 following doxycycline (DOX) induction. The results showed that the morphology of microcapsules was spherical, and their diameters have equally distributed in the range of 100-150 µm; the viability of rBMSCs was unchanged over time. Through real-time PCR and Western blot analyses, the rBMSCs in microcapsules were found to secrete hBMP2 and to have upregulated mRNA and protein expression of osteogenesis-related genes in vitro and in vivo. Furthermore, the in vivo results suggested that the group with the middle concentration of cells expressed the highest amount of osteogenic protein over time. In this study, we showed that gene-modified rBMSCs in CMC-ph microcapsules had good morphology and viability. The BMP2-BMSCs/CMC-Ph microcapsule system could upregulate osteogenic mRNA and protein in vitro and in vivo. Further analysis demonstrated that the medium concentration of cells had a suitable density for transplantation in nude mice. Therefore, BMP2-BMSCs/CMC-Ph microcapsule constructs have potential for bone regeneration in vivo.

Progress of co-culture systems in cartilage regeneration.

INTRODUCTION: Cartilage tissue engineering has rapidly developed in recent decades, exhibiting promising potential to regenerate and repair cartilage. However, the origin of a large amount of a suitable seed cell source is the major bottleneck for the further clinical application of cartilage tissue engineering. The use of a monoculture of passaged chondrocytes or mesenchymal stem cells results in undesired outcomes, such as fibrocartilage formation and hypertrophy. In the last two decades, co-cultures of chondrocytes and a variety of mesenchymal stem cells have been intensively investigated in vitro and in vivo, shedding light on the perspective of co-culture in cartilage tissue engineering. AREAS COVERED: We summarize the recent literature on the application of heterologous cell co-culture systems in cartilage tissue engineering and compare the differences between direct and indirect co-culture systems as well as discuss the underlying mechanisms. EXPERT OPINION: Co-culture system is proven to address many issues encountered by monocultures in cartilage tissue engineering, including reducing the number of chondrocytes needed and alleviating the dedifferentiation of chondrocytes. With the further development and knowledge of biomaterials, cartilage tissue engineering that combines the co-culture system and advanced biomaterials is expected to solve the difficult problem regarding the regeneration of functional cartilage.

Function of sustained released resveratrol on IL-1ß-induced hBMSC MMP13 secretion inhibition and chondrogenic differentiation promotion.

Metalloproteinase-13 is the major type II collagenase that directly implicates cartilage matrix destruction. Metalloproteinase-13 is inducted and activated by interleukin-1ß, which is a commonly observed proinflammatory cytokine in the joint cavity of arthritic patients. Depression of interleukin-1ß function can inhibit metalloproteinase-13 expression and protect the cartilage extracellular matrix. In this study, resveratrol release microspheres were developed, and the direct function of the released resveratrol on the interleukin-1ß was discussed. The resveratrol-loaded microspheres were fabricated using oil-in-water emulsion and solution-evaporation methods. The particle size and the encapsulation efficiency for the techniques, which used different fabrication conditions, were within 8.3-63.9 µm and 37%-82%, respectively. The effect of drug release lasted for more than 650 h in a PBS solution at 37℃. Human bone mesenchymal stem cells were chosen for cell experiments. Interleukin-1ß was used to induce an inflammatory condition. The effect of sustained resveratrol release from the microspheres on the cells' gene expression was observed using the transwell co-culturing method. The results indicated that metalloproteinase-13 mRNA expression was upregulated after interleukin-1ß induction. The released resveratrol directly inhibited the function of interleukin-1ß and thus downregulated metalloproteinase-13 mRNA expression. Moreover, the upregulation of Col2, aggrecan and Sox9 mRNA expressions, which are major chondrocyte markers, was observed after resveratrol was released into the culture medium. Resveratrol was observed to maintain the cells' chondrogenic gene expression when subject to the inflammation condition. The sustained released resveratrol inhibited interleukin-1ß-inducted metalloproteinase-13 activation and promoted chondrocyte differentiation. This drug-loading microsphere is a promising candidate for arthritis therapy.

Redifferentiation of dedifferentiated chondrocytes in a novel three-dimensional microcavitary hydrogel.

Although chondrocytes exist in native cartilage all over the body, it is still a challenge to use them as therapeutic cells for cartilage tissue engineering (TE) because of their easy dedifferentiation in in vitro culture. An improved culture system to maintain the characteristics of chondrocytes or recover their chondrocytic phenotype should be developed. In this study, we have set up an innovative microcavitary alginate hydrogel in an easy way. We compared this culture system with the conventional hydrogel and found that the microcavitary hydrogel exhibited outstanding superiorities in helping the dedifferentiated chondrocytes recover the capability for synthesizing cartilaginous extracellular matrix. In addition, we explored the correlation between chondrocyte redifferentiation in microcavitary hydrogels and changes in p38 and Erk1/2 activity. Our findings indicated that this microcavitary hydrogel would be a promising culture system to provide sufficient competent cells for cartilage regeneration and TE.

Effect of microcavitary alginate hydrogel with different pore sizes on chondrocyte culture for cartilage tissue engineering.

In our previous work, a novel microcavitary hydrogel was proven to be effective for proliferation of chondrocytes and maintenance of chondrocytic phenotype. In present work, we further investigated whether the size of microcavity would affect the growth and the function of chondrocytes. By changing the stirring rate, gelatin microspheres in different sizes including small size (80-120µm), middle size (150-200µm) and large size (250-300µm) were prepared. And then porcine chondrocytes were encapsulated into alginate hydrogel with various sizes of gelatin microspheres. Cell Counting Kit-8 (CCK-8), Live/dead staining and real-time PCR were used to analyze the effect of the pore size on cell proliferation and expression of specific chondrocytic genes. According to all the data, cells cultivated in microcavitary hydrogel, especially in small size, had preferable abilities of proliferation and higher expression of cartilaginous markers including type II collagen, aggrecan and cartilage oligomeric matrix protein (COMP). Furthermore, it was shown by western blot assay that the culture of chondrocytes in microcavitary hydrogel could improve the proliferation of cells potentially by inducing the Erk1/2-MAPK pathway. Taken together, this study demonstrated that chondrocytes favored microcavitary alginate hydrogel with pore size within the range of 80-120µm for better growth and ECM synthesis, in which Erk1/2 pathway was involved. This culture system would be promising for cartilage tissue engineering.

In vitro osteogenesis of synovium mesenchymal cells induced by controlled release of alendronate and dexamethasone from a sintered microspherical scaffold.

In vitro osteogenesis was successfully achieved with synovium-derived mesenchymal stem cells (SMSCs), which intrinsically have a strong chondrogenic tendency, by in situ release of alendronate (AL) and dexamethasone (Dex) from poly(lactic-co-glycolic acid) (PLGA)/hydroxyapatite (HA) sintered microspherical scaffold (PLGA/HA-SMS). Cumulative release profiles of AL and Dex from PLGA/HA-SMS and the influence on SMSCs osteogenic commitment were investigated. SMSCs seeded in Al-/Dex-loaded PLGA/HA-SMS (PLGA/HA-Com-SMS) exhibited significant osteogenic differentiation, as indicated by high yields of alkaline phosphatase (ALP) and bone calcification. In addition, mechanical properties (compressional) of PLGA/HA-Com-SMSs were also evaluated and approved. In conclusion, by promoting osteogenic commitment of SMSCs in vitro, this newly designed controlled-release system opens a new door to bone reparation and regeneration.

Poly(lactide-co-glycolide)/titania composite microsphere-sintered scaffolds for bone tissue engineering applications.

The objective of this study was to synthesize and characterize novel three-dimensional porous scaffolds made of poly(lactic-co-glycolic acid) (PLGA)/nano-TiO(2)-particle composite microspheres for potential bone repair applications. The introduction of TiO(2) component has been proven capable of largely enhancing mechanical properties of PLGA/TiO(2) microsphere-sintered scaffold ("PLGA/TiO(2)-SMS"). In addition, composite nano-TiO(2) additives are capable of inducing an increased arrest of adhesive proteins from the environment, which benefits cell attachment onto the scaffolds. Osteoblast proliferation and maturation were evaluated by MTT assay, alkaline phosphatase (ALP) activity, and bony calcification assay. The results indicate that osteoblasts cultured on the composite scaffolds with different TiO(2) content (0, 0.1, and 0.3 g/1 g PLGA) display increased cell proliferation compared with pure PLGA scaffold. When cultured on composite scaffolds, osteoblasts also exhibit significantly enhanced ALP activity and higher calcium secretion, with respect to those on the pure PLGA scaffolds. Taken together, PLGA/TiO(2)-SMSs deserve attention utilizing for potential bone-repairing therapeutics.

Concurrent extraction of proteins and RNA from cell-laden hydrogel scaffold free of polysaccharide interference.

Concurrent extractions of proteins and RNA from cell-laden scaffolds would be of great help to biomaterialists and tissue engineers. Here we describe a procedure to extract proteins from the discard solution generated during the RNA isolation from polysaccharide-rich hydrogel scaffolds. This approach allows to obtain proteins and RNA from same sample while eliminating the polysaccharide interference.

The control of anchorage-dependent cell behavior within a hydrogel/microcarrier system in an osteogenic model.

The use of injectable hydrogels for tissue engineering purposes such as bone regeneration has been hampered by the mass depletion of cells after encapsulation, due to the lack of a proper interface between hydrogel matrices and osteo-progenitor cells. Efforts to graft bioactive molecules as cell attachment moieties have achieved limited success. In this study, we devised a solution to promote cellular focal adhesion within hydrogels, and elicit the mechanism behind cellular survival/death therein. We found that the fulfillment of ligation between cellular integrins and extracellular ligands, instead of the expression of integrins per se, is essential to avoid apoptosis in gel-encapsulated anchorage-dependent cells (ADCs). Absence of such ligation brought about mass cell death in our osteogenic model with osteoblasts (as representative of ADCs) and failure of osteogenic commitment of mesenchymal stem cells (as representative of anchorage-dependent progenitors). We have designed a gel-based composite system that works as a suspension of injectable cell-laden microcarriers in hydrogel, as compared to the conventional cell-suspended hydrogels. Injectable microscopic anchors (microcarriers) not only provide platforms for cellular focal adhesion but also facilitate the cells to overcome gel enlacement and fully spread out into their natural morphology. Further in vitro and in vivo osteogenic investigations show the composites to be a competent potential injectable vehicle for the conveyance of ADCs and regenerations of bone and other tissues.