The design of biomimetic nanomaterials that can directly influence the behavior of cells and facilitate the regeneration of tissues and organs has become an active area of research. Here, the production of materials based on nano‐hydroxyapatite composites in scaffolds with nanofibrous and nanoporous topographies, designed to mimic the native bone matrix for applications in bone tissue engineering, is reported. Human mesenchymal stem cells grown on these nanocomposites are stimulated to rapidly produce bone minerals in situ, even in the absence of osteogenic supplements in the cell‐culture medium. Nanocomposites comprising type I collagen and nano‐hydroxyapatite are found to be especially efficient at inducing mineralization. When subcutaneously implanted into nude mice, this biomimetic nanocomposite is able to form a new bone matrix within only two weeks. Furthermore, when the nanocomposite is enriched with human mesenchymal stem cells before implantation, development of the bone matrix is accelerated to within one week. To the best of the authors' knowledge, this study provides the first clear in vitro and in vivo demonstration of osteoinduction controlled by the material characteristics of a biomimetic nanocomposite. This approach can potentially facilitate the translation of de novo bone‐formation technologies to the clinic.

Nanostructured materials are gaining new impetus owing to the advancements in material fabrication techniques and their unique properties (their nanosize, high surface area-to-volume ratio, and high porosity). Such nanostructured materials mimic the subtleties of extracellular matrix (ECM) proteins, creating artifi cial microenvironments which resemble the native niches in the body. On the other hand, the isolation of mesenchymal stem cells (MSCs) from various tissue sources has resulted in the interest to study the multiple differentiation lineages for various therapeutic treatments. In this review, our focus is tailored towards the potential of biomimetic nanostructured materials as osteoinductive scaffolds for bone regeneration to differentiate MSCs towards osteoblastic cell types without the presence of soluble factors. In addition to mimicking the nanostructure of native bone, the supplement of collagen and hydroxyapatite which mimic the main components of the ECM also brings signifi cant advantages to these materials.

Mechanical stimulation is one of the factors that regulating bone regeneration and healing. In this study, the biological responses of bone marrow derived mesenchymal stem cells (MSCs) to mechanical stimuli on aligned nanofibers and cast films were investigated. The uniaxial cyclic strain (1% strain and 1 Hz) was applied continuously to the cell substrates and osteoblastic activities were assessed at weeks 1, 2, and 4. The MSCs morphology on the aligned nanofibers was more elongated and spindle-like than MSCs on the cast films. Strain stimulation significantly attenuated the proliferation at week one but was significantly enhanced at week 4 for both types of substrates. Only the MSCs on strained nanofibers had greater alkaline phosphatase (ALP) levels at week one, while the ALP hindered the MSCs on both substrates at week 4. Strain application played a greater influence on osteocalcin expression for the cast films than the nanofibers at week 4. Clearly, the cellular response to strain induction was highly dependent on the surface-cell adhesion, which itself was greatly influenced by the surface texture of the substrate.

This chapter contains sections titled: Introduction Bone Grafts Types of Materials for Bone Applications Sources of Stem Cells Used for Bone Tissue Engineering to Increase Osteogenic Differentiation Current Perspective Conclusion References

Mineralized type I collagen (collagen I) nanofibers and their nanofibril bundles make up the microstructure of natural bone tissue, which range from nanometers to micrometers. However, attempts to achieve this hierarchically assembled structure in vitro have been unsuccessful. In this study, we added osteonectin into the collagen I solution, either at a high or low weight ratio (osteonectin: collagen I = 1:30 or 1:90) before co-precipitation. Results indicated that spindle-like nano-hydroxyapatite (nano-HA) was deposited on collagen/osteonectin and pure osteonectin (control) groups. Furthermore, transmission electron microscope (TEM) and scanning electron microscope (SEM) results showed that the assembled mineralized fiber bundles were formed randomly at different levels from 50 nm, 250 nm to 1100 nm. However, when we replaced collagen I with collagen II, osteonectin addition did not induce the formation of mineralized fiber bundles. The participation of osteonectin in the assembly of the mineralized fibers could provide new insights into the novel mineralization function of osteonectin for bone development in vivo and advancing new biomimetic methods for bone graft applications.

Bone is a nanocomposite consisting of two main components, nano-hydroxyapatite (n-HA) and Type I collagen (Col). The aim is to exploit the nano-scale functional and material characteristics of natural bone in order to modulate cellular functions for optimal bone repair in bone graft systems. Here, we present an effective and novel technique in obtaining n-HA in cognate with native apatite on electrospun nanofibers within minutes without any pre-treatment. Using an alternate calcium and phosphate (Ca–P) solution dipping method, n-HA was formed on poly(lactide-co-glycolide) acid (PLGA) and blended PLGA/Col nanofibers. The presence of the functional groups of collagen significantly hastened n-HA deposition closed to nine-fold. The quantity of n-HA impinged upon the specific surface area, whereby mineralized PLGA/Col had a greater surface area than non-mineralized PLGA/Col, whereas n-HA did not significantly improve the specific surface area of mineralized PLGA compared to pure PLGA. The novelty of the process was that n-HA on PLGA had a positive modulation on early osteoblast capture (within minutes) compared to pure PLGA. Contrary, cell capture on mineralized PLGA/Col was comparable to pure PLGA/Col. Interestingly, although n-HA impeded proliferation during the culture period (days 1, 4 and 7), the cell functionality such as alkaline phosphatase (ALP) and protein expressions were ameliorated on mineralized nanofibers. The amount of n-HA appeared to have a greater effect on the early stages of osteoblast behavior (cell attachment and proliferation) rather than the immediate/late stages (proliferation and differentiation).

Biodegradable materials are widely used in the biomedical field because there is no postoperative surgery after implantation. Widely used synthetic biodegradable materials are polyesters, especially those used in tissue engineering. Advances in the tissue engineering field have brought much attention in terms of scaffold fabrication, such as with biodegradable polyester nanofibers. The rationale for using nanofibers for tissue engineering is that the nonwoven polymeric meshwork is a close representation of the nanoscale protein fiber meshwork in native extracellular matrix (ECM). Electrospinning technique is a promising way to fabricate controllable continuous nanofiber scaffold mimicking the ECM structure. Electrospun nanofibers provide high surface-to-volume ratio and high porosity as a promising scaffold for tissue engineering. Because the degradation behaviors of scaffolds significantly affect new tissue regeneration, the degradation of the material becomes one of the crucial factors when considering using polyester nanofibers as scaffolds in tissue engineering. In this review paper, we focus on the degradation studies of several bioresorbable polyester nanofibrous scaffolds used in tissue engineering. The degradable properties of nanofibers were compared with the corresponding degradable materials in macroscale. The factors that might affect the degradation behaviors were analyzed.

Chemical guiding cues are being exploited to stimulate neuron adhesion and neurite outgrowth. In this study, an amino-functioned PLLA, lysine-capped PLLA [K-(CH2)n-PLLA (n=2, 5, 8)], was synthesized with different length of linking spaces between lysine molecule and PLLA backbone. Drop-cast films were fabricated from K-(CH2)n-PLLA/PLLA blends (10/90, w/w) and amino groups were detected on the surfaces of the resultant films. More amine groups were detected on the surface and the hydrophilicity of the films was obviously improved by annealing the films in water. The representative atomic force microscopy (AFM) images indicated that incorporation of lysine-capped PLLA into PLLA matrix increased the roughness of the films and resulted in a phase separation with distinct two nano-domains which may correspond to the hydrophilic and hydrophobic domains. Furthermore, the laminin-derived peptides, CYIGSR (Cys-Tyr-Ile-Gly-Ser-Arg) and CSIKVAV (Cys-Ser-Ile-Lys-Val-Ala-Val), were jointly tethered to the amine groups of lysine-capped PLLA by a linking reagent sulfo-succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (Sulfo-SMCC). The neonatal mouse cerebellum C17.2 stem cells were seeded on the peptides-grafted K-(CH2)n-PLLA/PLLA (n=2, 5, 8) films and pure PLLA films were used as controls. Improved viability and longer neurites were obtained on the peptide-grafted films than PLLA film over the cultivation period, especially for K-(CH2)5-PLLA/PLLA, which had the highest peptide density of 0.28±0.03μg/cm2. This study highlights the potential of using the lysine-cappeded PLLA with laminin-derived peptides for promoting nerve regeneration.

By the method of injection molding combined with thermally induced phase separation (TIPS), a novel nerve conduit with a plurality of channels and macro-/microporous architecture was fabricated using poly (lactide-co-glycolide) (PLGA, 75:25; Mn=1.22x10(5)). The diameter of the conduits and the number of channels could be regulated by changing the parameters of the mold, and the porosity of the conduit was as high as 95.4%. Meanwhile, the hierarchical pore architecture of the walls could be controlled through varying the solution concentration and the contents of porogen. The degradation study in vitro showed that 7-channel conduit could hold its apparent geometry for about 12 weeks in phosphate buffer solution (PBS) at 37degreesC, and the pH values of the degradation solution were detected in the range 4.1-4.5. The influences of the conduit architecture on the cell attachment, spreading, and proliferation were evaluated by culturing rat mesenchymal stem cells alone or together with Schwann cells in vitro. The implantation of the PLGA conduit in the spinal cord showed that it had good biocompatibility, and no obvious inflammatory response was detected. Therefore, the results implied that these PLGA multiple-channel nerve conduits have the potential use for spinal cord injury.

Poly-L-lactic acid (PLLA) and PLLA/collagen (50% PLLA+50% collagen; PLLA/Col) nanofibers were fabricated using electrospinning. Mineralization of these nanofibers was processed using a modified alternating soaking method. The structural properties and morphologies of mineralized PLLA and PLLA/Col nanofibers were investigated using X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and contact angle measurements. Human bone-derived osteoblasts were cultured on the materials for up to 1 week to assess the biological properties of the nanofibrous composites. Cell attachment on these nanocomposites was also tested within 1 h of culture at room temperature. The mechanical properties of the cell-nanocomposite constructs were determined using tensile testing. From our results, the bone-like nano-hydroxyapatite (n-HA) was successfully deposited on the PLLA and PLLA/Col nanofibers. We observed that the formation of n-HA on PLLA/Col nanofibers was faster and significantly more uniform than on pure PLLA nanofibers. The n-HA significantly improved the hydrophilicity of PLLA/Col nanofibers. From the results of cell attachment studies, n-HA deposition enhanced the cell capture efficacy at the 20-minute time point for PLLA nanofibers. The E-modulus values for PLLA+n-HA with cells (day 1 and day 4) were significantly higher than for PLLA+n-HA without cells. Based on these observations, we have demonstrated that n-HA deposition on nanofibers is a promising strategy for early cell capture.

A novel biomimetic self-assembly method was designed to create nano-carbonated hydroxyapatite/collagen (nCHAC) composites by means of incorporating various collagen and carbonate concentrations using solutions such as CaCl(2), H(3)PO(4), and Na(2)CO(3). At a given range of collagen and carbonate content, the nanosized inorganic phase of the newly synthesized material has a low degree of crystallinity which resembles that of natural bone. By manipulating the concentrations of collagen and carbonates, various morphologies of the nCHAC can be obtained. The crystal size of nCHAC is dependent on the concentration of carbonate and collagen present in the composites. For instance, higher collagen concentration results in smaller crystal nCHAC crystal size. Conversely, the higher the carbonate content, the smaller are the crystal size and the collagen fibril assembly. As the carbonate content increased, the plate-like crystals first became needle-like structures, subsequently short needle-like crystals and eventually became spherical particles. From this study, our method showcased the flexibility of fabricating various types of nCHAC composites which can be designed for different bone applications.

Despite the large number of publications in peer review literature in the field of nanofibers, there is still uncertainty as to what aspects of these research results have commercial applications. In an effort to better understand the technological progress made in the field of nanofibers, we surveyed the patents issued in the United States from 1976 up to end 2006. The present review will provide an overall view of the current patent landscape including trends and key applications. Key assignees and key inventors were identified and their contributions were discussed. Patents were identified using keywords such as nanofibers, ultrafine, and electrospinning. After patents were downloaded, we reviewed each patent for relevancy and identified 100 patents to be related to nanofibers. 75% of the current issued patents on nanofibers are directed at either fabrication methods or the use of nanofibers in filtration systems. The patent data indicates that medical applications and medical products using nanofibers appear to be the emerging application for nanofibers. We anticipate a growing number of patents on novel applications for nanofiber would originate from academic centers in the future.

We prepared nano-carbonated hydroxyapatite (CHA) with different additions of carbonated contents at room temperature. The resulting materials were tested by X-ray diffraction (XRD), Fourier transform Infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) respectively. Results were shown that the materials were platelet-like CHA polycrystals that were similar to natural bone minerals, except for the material sample with the highest carbonated content. The presence of carbonated content in hydroxyapatite (HA) significantly decreased the crystallinity of synthesized CHA. Moreover, the morphological change of CHA was observed with the increase of carbonated content. The sequence of morphological change was from flat-like platelet, needle-arrayed platelet, particle-consisted platelet, smaller platelet, and finally spherical particle. This tendency is accordant to controllable solubility of biological HA with certain carbonated content.

Allograft bone, dematerialized bone matrix and calcium-based synthetic materials have long been used as bone graft substitutes. First-generation bone graft substitutes as stand-alone graft substitutes have not developed as hoped. It remains a great challenge to design an ideal bone graft that emulates nature's own structures or functions. To further improve the performance of such bone graft substitutes, scientists are investigating biomimetic processes to incorporate the desirable nano-features into the next generation of biomaterials. In this regard, nanostructured biomaterials less than 100 nm in at least one dimension, in particular nanocomposites, are perceived to be beneficial and potentially ideal for bone applications, owing to their nanoscale functional characteristics that facilitate bone cell growth and subsequent tissue formation. In fact, bone itself is a nanocomposite system with a complex hierarchical structure. This review reports the impact of biomimetically derived nanocomposite biomaterials for use in bone applications and provides possible suggestions for future research and development.

Touted as the technological revolution of the 21st century, nanotechnology has already been the topic of numerous publications, patents and news stories. While the commercial applications of nanotechnology are still relatively few, they are beginning to attract the attention of the commercial sector. For successful commercialization of this emerging technology, patent protection is imperative. Patents are publicly available documents forming a rich repository of current knowledge. By a systematic method of collecting, organizing and analyzing these publicly available documents, valuable information can be distilled about the activities and opportunities within this emerging field. As this field is rapidly progressing, this paper provides a summary of current patent issues in nanotechnology, an overview of nanotechnology patent analyses and the latest update of the current landscape of nanotechnology patents.