Canine ACL reconstruction with an injectable hydroxyapatite/collagen paste for accelerated healing of tendon-bone interface.

Healing of an anterior cruciate ligament (ACL) autologous graft in a bone tunnel occurs through the formation of fibrovascular scar tissue, which is structurally and compositionally inferior to normal fibrocartilaginous insertion and thus may increase the reconstruction failure and the rate of failure recurrence. In this study, an injectable hydroxyapatite/type I collagen (HAp/Col Ⅰ) paste was developed to construct a suitable local microenvironment to accelerate the healing of bone-tendon interface. Physicochemical characterization demonstrated that the HAp/Col Ⅰ paste was injectable, uniform and stable. The in vitro cell culture illustrated that the paste could promote MC3T3-E1 cells proliferation and osteogenic expression. The results of a canine ACL reconstruction study showed that the reconstructive ACL had similar texture and color as the native ACL. The average width of the tunnel, total bone volume, bone volume/tissue volume and trabecular number acquired from micro-CT analysis suggested that the healing of tendon-bone interface in experimental group was better than that in control group. The biomechanical test showed the maximal loads in experimental group achieved approximately half of native ACL's maximal load at 24 weeks. According to histological examination, Sharpey fibers could be observed as early as 12 weeks postoperatively while a typical four-layer transitional structure of insertion site was regenerated at 48 weeks in the experimental group. The injectable HAp/Col Ⅰ paste provided a biomimetic scaffold and microenvironment for early cell attachment and proliferation, further osteogenic expression and extracellular matrix deposition, and in vivo structural and functional regeneration of the tendon-bone interface.

Hydroxyapatite-collagen nanoparticles reinforced polyanhydride based injectable paste for bone substitution: effect of dopant addition in vitro.

The present study focuses on the synthesis and characterization of hydroxyapatite-collagen nanoparticles incorporated polyanhydride paste and investigating its bone regeneration capacity in vitro. Photocrosslinkable polyanhydride paste was prepared after synthesizing methacrylate derivatives of 1,6-bis(p-carboxyphenoxy)hexane (MCPH) and sebacic acid dimethacrylate (MSA). These multifunctional monomers, namely 45 wt% MSA, 45 wt% MCPH in addition to 10 wt% poly(ethylene glycol)diacrylate (PEGDA) were photopolymerized under ultraviolet light (365 nm) to produce highly crosslinked polyanhydride networks using camphroquinone (CQ)/ethyl 4-(dimethylamino)benzoate [4-EDMAB] for light initiated crosslinking and benzoyl peroxide (BPO)/dimethyl toludine (DMT) for chemically initiated crosslinking. Separately, using the co-precipitation process, (1 wt%) Si, (1 wt%) Sr, and (0.5 + 0.5) wt% Si/Sr was doped into hydroxyapatite-collagen nanoparticles in size range between 50 and 70 nm. Si, Sr, and both Si/Sr doped hydroxyapatite-collagen nanoparticles to the extent 10 wt% were added to polyanhydride monomer mixture containing 40 wt% MSA, 40 wt% MCPH and 10 wt% PEGDA and subsequently photopolymerized as previously mentioned. Incorporation of hydroxyapatite-collagen nanoparticles to the extent of 10 wt% into polyanhydride matrix enhanced compressive strength of the hardened paste from 30 to 49 MPa. Mesenchymal stem cells obtained from the human umbilical cord were cultured onto pure polyanhydride and hydroxyapatite-collagen added scaffold to assess their cellular proliferation and differentiation capacity to bone cell. MTT assay showed that mesenchymal stem cell proliferation was significantly higher in Si/Sr binary doped hydroxyapatite-collagen-polyanhydride sample as compared to other samples. Again from immunocytochemistry study using confocal images suggested that expression of osteocalcin, a marker indicating differentiation into osteoblast, was the highest in Si/Sr binary doped hydroxyapatite-collagen-polyanhydride sample against the other samples studied in this case. This study thus summarizes the development of photocurable biocomposites containing polyanhydride and Si, Sr doped hydroxyapatite-collagen nanoparticles that exhibited tremendous promise to regenerate bone tissues in complex-shaped musculoskeletal defect sites.

Formulating injectable pastes of porous calcium phosphate glass microspheres for bone regeneration applications.

Current trends in regenerative medicine treatments for bone repair applications focus on cell-based therapies. These aim to deliver the treatment via a minimally invasive injection to reduce patient trauma and to improve efficacy. This paper describes the injectability of porous calcium phosphate glass microspheres to be used for bone repair based on their formulation, rheology and flow behavior. The use of excipients (xanthan gum, methyl cellulose and carboxyl methyl cellulose) were investigated to improve flow performance. Based on our results, the flow characteristics of the glass microsphere pastes vary according to particle size, surface area, and solid to liquid ratio, as well as the concentration of viscosity modifiers used. The optimal flow characteristics of calcium phosphate glass microsphere pastes was found to contain 40 mg/mL of xanthan gum which increased viscosity whilst providing elastic properties (∼29,000 Pa) at shear rates that mirror the injection process and the resting period post injection, preventing the glass microspheres from both damage and dispersion. It was established that a base formulation must contain 1 g of glass microspheres (60-125 µm in size) per 1 mL of cell culture media, or 0.48 g of glass microspheres of sizes between 125 and 200 µm. Furthermore, the glass microsphere formulations with xanthan gum were readily injectable via a syringe-needle system (3-20 mL, 18G and 14G needles), and have the potential to be utilized as a cell (or other biologics) delivery vehicle for bone regeneration applications.

Study on an injectable biomedical paste using cross-linked sodium hyaluronate as a carrier of hydroxyapatite particles.

Exploring the long-term filler for minimally invasive plastic surgery has been widely concerned. In the present study, a series of injectable paste composed of hydroxyapatite (HAp) spherical particles and cross-linked sodium hyaluronate (cHA) solution were prepared. The physicochemical properties of cHA as a carrier of high content HAp microspheres (>50%) and as-obtained injectable HAp/cHA paste were studied. The cross-linking degree (DC), viscosity and molecular weight (Mw and Mn) of cHA increased with the increasing of the cross-linker dosage from 7.5 to 17.5 wt% under the certain conditions. HAp/cHA pastes were fabricated by homogeneously blending different sizes of HAp microspheres with cHA solution. The stability, rheological performance and push-out force of the pastes were studied, and the influence factors were discussed. The results indicated that moderate crosslinked cHA with 60% middle size HAp (HAp-M60/cHA-15.0) had appropriate comprehensive property. Finally, the in vitro cell culture approved the paste had no cytotoxicity. Although the biological performance of the pastes still need to be investigated, this preliminary study demonstrates that it is possible to carry high content HAp in cHA, expecting the better volumetric maintenance after long term implantation.

Engineering bone regeneration with novel cell-laden hydrogel microfiber-injectable calcium phosphate scaffold.

Cell-based tissue engineering is promising to create living functional tissues for bone regeneration. The implanted cells should be evenly distributed in the scaffold, be fast-released to the defect and maintain high viability in order to actively participate in the regenerative process. Herein, we report an injectable calcium phosphate cement (CPC) scaffold containing cell-encapsulating hydrogel microfibers with desirable degradability that could deliver cells in a timely manner and maintain cell viability. Microfibers were synthesized using partially-oxidized alginate with various concentrations (0-0.8%) of fibrinogen to optimize the degradation rate of the alginate-fibrin microfibers (Alg-Fb MF). A fibrin concentration of 0.4% in Alg-Fb MF resulted in the greatest enhancement of cell migration, release and proliferation. Interestingly, a significant amount of cell-cell contact along the long-axis of the microfibers was established in Alg-0.4%Fb MF as early as day 2. The injectable tissue engineered construct for bone reconstruct was fabricated by mixing the fast-degradable Alg-0.4%Fb MF with CPC paste at 1:1 volume ratio. In vitro study showed that cells re-collected from the construct maintained good viability and osteogenic potentials. In vivo study demonstrated that the hBMSC-encapsulated CPC-MF tissue engineered construct displayed a robust capacity for bone regeneration. At 12weeks after implantation, osseous bridge in the rat mandibular defect was observed in CPC-MF-hBMSCs group with a new bone area fraction of (42.1±7.8) % in the defects, which was >3-fold that of the control group. The novel tissue-engineered construct presents an excellent prospect for a wide range of dental, craniofacial and orthopedic applications.