Fourier transform infrared microscopic imaging: effects of estrogen and estrogen deficiency on fracture healing in rat femurs

Infrared spectroscopic imaging with 6-10 microm spatial resolution was used to characterize the changes in fracture callus mineral content, carbonate content, mineral crystallinity, and collagen maturity in femurs of 3-month-old ovariectomized rats treated with estrogen (estrogen sufficiency) or vehicle (estrogen deficiency). Comparisons were also made in these animals to cortical bone at a distance from the callus. Analyses at 4, 8, and 12 weeks post fracture demonstrated that healing was accelerated in the estrogen-sufficient animals as demonstrated by increasing mineral content and collagen maturity and decreasing carbonate incorporation.     

Microstructural evolution in external callus of human long bone

Accurate knowledge of bone fracture healing process is of clinical and theoretical importance in bone repair and regeneration, and biomineralization. It is well known that the histological healing occurs via the formation of hematoma, fibrocartilage, bony callus and bone modeling/remodeling. However, the detailed process from fracture to healing at the microstructural level remains unclear. In the present study, an evolutionary model of external callus is proposed, in which five representative stages are presented in terms of the organization of collagen and minerals during the formation of bony callus. The first stage is the formation of loose, disordered collagen fibrils, which is followed by mineralization on some of these individual microfibrils. Then the matrix is characterized by the fusion of mineralized individual fibrils into bundles. In the third stage, the absorption of disordered matrix occurs. This is gradually replaced by ordered collagen in stage four. Finally, completely ordered mineralized tissue is formed. The proper sequence of the process plays an important role in deciding the success of healing.In addition to the common mineral phase of hydroxyapatite (HA), dicalcium phosphate dihydrate (DCPD) phase was also found in early stage of healing, especially in rapid healing (children's callus). It vanished in the following process of healing. The deposition of DCPD is supposed to be brought about by some non-collagenous protein.     

Intramedullary bone formation after polylactic acid wire implantation

Semi-crystalline poly-L-lactic acid (PLLA) wire was implanted intramedullary in rat tibiae to evaluate the fracture healing processes and the tissue reaction on the PLLA in a fractured bone. The fracture healing after PLLA implantation was compared to a sham-operated group of animals. In all animals with intramedullary PLLA wire implantation newly formed bone was seen immediately against the implant 2 and 6 months after implantation. None of the shamoperated animals showed newly formed intramedullary bone formation other than fracture healing callus and normal trabecular bone.     

Effect of mendable polymer stitch density on the toughening and healing of delamination cracks in carbon–epoxy laminates

This paper presents an investigation into the effect of stitch density on the delamination toughening and self-healing properties of carbon–epoxy laminates. The stitches provide the laminate with the synergistic combination of high mode I interlaminar fracture toughness to resist delamination cracking and healing properties to repair delamination damage. The results show that the fracture toughness of the laminate increased with stitch density, due to higher traction (crack closure) loads exerted by the stitches bridging the delamination. During the healing process these bridging stitches first melt and then flow into the delamination, leading to self-healing with full restoration of the mode I fracture toughness. Furthermore, the stitches were capable of repairing delamination cracks many times larger than the original size of the stitches. The effect of stitch density on the healing process of delamination cracks and restoration of fracture toughness was found to remain approximately the same under multiple repair operations.     

The finite element analysis of a fractured tibia applied by composite bone plates considering contact conditions and time-varying properties of curing tissues

This paper addresses the use of composite bone plates in healing long-bone fractures such as transverse fractures of the tibia using finite element analysis, that takes into consideration contact conditions and material property variations of calluses in relation to the healing period. For the time-varying properties of calluses in relation to the healing period, stepwise material properties were imposed on the callus part based on the time elapsed, and the loading conditions were coupled with the callus properties based on the length of the healing period. The strain distributions at the fracture site were calculated according to the stacking sequence of the bone plate and healing time. The analysis results showed that composite bone plates with stacking sequences of [0]_12T for the Kevlar/BCP composites generated the most appropriate strain distributions at the fracture site during the early healing process.     

Self-healing of delamination fatigue cracks in carbon fibre–epoxy laminate using mendable thermoplastic

This article examines the self-healing repair of delamination damage in mendable carbon fibre–epoxy laminates under static or fatigue interlaminar loading. The healing of delamination cracks in laminates containing particles or fibres of the mendable thermoplastic poly[ethylene-co-(methacrylic acid)] (EMAA) was investigated. The results showed that the formation of large-scale bridging zone of EMAA ligaments along the crack upon healing yielded a large increase (~300%) in the static mode I interlaminar fracture toughness, exceeding the requirement of full restoration. The mendable laminates retained high healing efficiency with multiple repair cycles because of the capability of EMAA to reform the bridging zone under static delamination crack growth conditions. Under fatigue loading, healing by the EMAA was found to restore the mode I fatigue crack growth resistance, with the rates of growth being slightly less than that pertinent to the unmodified laminate. The EMAA bridging zone, which generated high toughness under static loading conditions, does not develop under fatigue loading because of rapid fatigue failure of the crack bridging ligaments. Similar to the multiple healing capability of EMAA under static loading, multiple healing of delamination fatigue cracks is confirmed, with the fatigue crack growth rates remaining approximately unchanged. This study shows that EMAA was capable of full recovery of fatigue crack growth resistance and superior healing efficiency for static loading.     

Application of bridging-law concepts to short-fibre composites 4. FEM analysis of notched tensile specimens

This is the fourth paper in a series of four where notch sensitivities, fracture energies and bridging laws in short-fibre polymer composites are investigated. In this paper finite-element modelling (FEM) of centre-hole-notched tensile specimens is performed, with different bridging laws governing crack growth. Crack lengths, crack profiles and stress distributions are predicted. The results are compared with experimentally determined crack shapes from an earlier investigation. Only with softening bridging laws can the experimental results be matched. The predicted crack lengths are sensitive to bridging-law parameters. When bridging laws determined by the double cantilever beam (DCB) method are applied, the predicted crack lengths and profiles show good correlation with the experimental results. The results support the validity of the DCB method to determine bridging laws in short-fibre composites.     

Nanofibers prepared by needleless electrospinning technology as scaffolds for wound healing

Electrospun gelatin and poly-ε-caprolactone (PCL) nanofibers were prepared using needleless technology and their biocompatibility and therapeutic efficacy have been characterized in vitro in cell cultures and in an experimental model of a skin wound. Human dermal fibroblasts, keratinocytes and mesenchymal stem cells seeded on the nanofibers revealed that both nanofibers promoted cell adhesion and proliferation. The effect of nanofibers on wound healing was examined using a full thickness wound model in rats and compared with a standard control treatment with gauze. Significantly faster wound closure was found with gelatin after 5 and 10 days of treatment, but no enhancement with PCL nanofibers was observed. Histological analysis revealed enhanced epithelialisation, increased depth of granulation tissue and increased density of myofibroblasts in the wound area with gelatin nanofibers. The results show that gelatin nanofibers produced by needleless technology accelerate wound healing and may be suitable as a scaffold for cell transfer and skin regeneration.     

Controlling Mechanical Properties of Cell‐Laden Hydrogels by Covalent Incorporation of Graphene Oxide

Graphene‐based materials are useful reinforcing agents to modify the mechanical properties of hydrogels. Here, an approach is presented to covalently incorporate graphene oxide (GO) into hydrogels via radical copolymerization to enhance the dispersion and conjugation of GO sheets within the hydrogels. GO is chemically modified to present surface‐grafted methacrylate groups (MeGO). In comparison to GO, higher concentrations of MeGO can be stably dispersed in a pre‐gel solution containing methacrylated gelatin (GelMA) without aggregation or significant increase in viscosity. In addition, the resulting MeGO‐GelMA hydrogels demonstrate a significant increase in fracture strength with increasing MeGO concentration. Interestingly, the rigidity of the hydrogels is not significantly affected by the covalently incorporated GO. Therefore, this approach can be used to enhance the structural integrity and resistance to fracture of the hydrogels without inadvertently affecting their rigidity, which is known to affect the behavior of encapsulated cells. The biocompatibility of MeGO‐GelMA hydrogels is confirmed by measuring the viability and proliferation of the encapsulated fibroblasts. Overall, this study highlights the advantage of covalently incorporating GO into a hydrogel system, and improves the quality of cell‐laden hydrogels.     

Application of bridging-law concepts to short-fibre composites Part 3: Bridging law derivation from experimental crack profiles

This is the third paper in a series of four where notch sensitivity, fracture energy and bridging laws are studied in short-fibre polymer composites. Here, bridging laws are derived from experimental crack-opening profiles in centre-hole notched tensile specimens. The materials studied are three types of commercial glass–mat composites with different reinforcement structures and matrices. The materials have softening bridging laws and the calculated fracture energies from bridging laws are in good agreement with values determined directly by experiment. The calculated maximum local bridging stress is found to be higher than the uniaxial tensile strength. An outline of a failure criterion for notched specimens based on the crack-bridging approach is presented.     

Poroelastic analysis of bone tissue differentiation by using the boundary element method

Fracture healing is initiated and tightly regulated mainly by growth factors and by mechanical environment around the callus site. Biomechanics of fracture healing have been previously studied. Most computational models are based on finite elements and some of them study the level of strain or stress in the different tissues. These strain/stress fields are the main mechanical stimuli affecting cell differentiation and ossification pathway. In this work, we incorporated that hypothesis into a poroelastic axi-symmetric boundary element callus model, where the pore pressure was included as a part of the stimuli function. This analysis allowed us to extend the observations made by other authors and a new poroelastic correlation between mechanical conditions and local tissue formation is proposed. This work shows the capability of the boundary element method to characterize the tissue phenotypes during a progressive healing process. The results were in good agreement with those reported in previous works.     

Simulation of the bone healing process of fractured long bones applied with a composite bone plate with consideration of the blood vessel growth

The healing process of long bones such as the tibia was simulated on the basis of a mechanoregulation theory by taking blood vessel growth into consideration. The tissue differentiation process of calluses by taking into consideration blood vessel growth was simulated by a user subroutine program based on the mechanoregulation model and a diffusion equation. Composite bone plates made of a plain weave carbon/epoxy composite (WSN3k) and a plain weave glass/polypropylene composite (Twintex) were applied to the fracture site to investigate the effect of plate modulus on the healing performance. The simulation results revealed that the flexible composite bone plate made of Twintex [0]₁₈, which had a slightly higher Young’s modulus than a cortical bone, provided the highest healing performance. Moreover, it was found that the effect of the plate modulus on the healing performance reduced when the blood vessel growth at the fracture site was considered, which reflected a more realistic bone healing process.     

Influence of high-frequency cyclical stimulation on the bone fracture-healing process: mathematical and experimental models

Mechanical stimulation affects the evolution of healthy and fractured bone. However, the effect of applying cyclical mechanical stimuli on bone healing has not yet been fully clarified. The aim of the present study was to determine the influence of a high-frequency and low-magnitude cyclical displacement of the fractured fragments on the bone-healing process. This subject is studied experimentally and computationally for a sheep long bone. On the one hand, the mathematical computational study indicates that mechanical stimulation at high frequencies can stimulate and accelerate the process of chondrogenesis and endochondral ossification and consequently the bony union of the fracture. This is probably achieved by the interstitial fluid flow, which can move nutrients and waste from one place to another in the callus. This movement of fluid modifies the mechanical stimulus on the cells attached to the extracellular matrix. On the other hand, the experimental study was carried out using two sheep groups. In the first group, static fixators were implanted, while, in the second one, identical devices were used, but with an additional vibrator. This vibrator allowed a cyclic displacement with low magnitude and high frequency (LMHF) to be applied to the fractured zone every day; the frequency of stimulation was chosen from mechano-biological model predictions. Analysing the results obtained for the control and stimulated groups, we observed improvements in the bone-healing process in the stimulated group. Therefore, in this study, we show the potential of computer mechano-biological models to guide and define better mechanical conditions for experiments in order to improve bone fracture healing. In fact, both experimental and computational studies indicated improvements in the healing process in the LMHF mechanically stimulated fractures. In both studies, these improvements could be associated with the promotion of endochondral ossification and an increase in the rate of cell proliferation and tissue synthesis.     

Interlayer self-healing and toughening of carbon fibre/epoxy composites using copolymer films

This paper presents an investigation of the combined self-healing and toughening performance of two copolymers: thermoplastic poly(ethylene-co-methyl acrylate) (EMA) and poly(ethylene-co-methacrylic acid) (EMAA). Carbon fibre composites were manufactured from unidirectional prepregs with rectangular-shaped patches being placed between composite plies. Results from double-cantilever-beam and short-beam-shear testing show that the incorporation of mendable polymers improves interlaminar fracture toughness but causes a reduction in interlaminar shear strength. The healing efficiency in terms of restoration of the interlaminate fracture energy scales linearly with the areal percentage of self-healing material. Microstructure study revealed distinct difference in the fracture surfaces of composites with EMA and EMAA, with EMA displaying extensive nano-scale porous structures in contrast to the more homogenous single phase structure from EMAA.     

Materials evolution of bone plates for internal fixation of bone fractures:A review

Bone plates play a vital role in bone fracture healing by providing the necessary mechanical fixation for fracture fragments through modulating biomechanical microenvironment adjacent to the fracture site.Good treatment effect has been achieved for fixation of bone fracture with conventional bone plates,which are made of stainless steel or titanium alloy.However,several limitations still exist with traditional bone plates including loosening and stress shielding due to significant difference in modulus between metal material and bone tissue that impairs optimal fracture healing.Additionally,due to demographic changes and non-physiological loading,the population suffering from refractory fractures,such as osteoporosis fractures and comminuted fractures,is increasing,which imposes a big challenge to traditional bone plates developed for normal bone fracture repair.Therefore,optimal fracture treatment with adequate fixation implants in terms of materials and design relevant to special conditions is desirable.In this review,the complex physiological process of bone healing is introduced,followed by reviewing the development of implant design and biomaterials for bone plates.Finally,we discuss recent development of hybrid bone plates that contains bioactive elements or factors for fracture healing enhancement as a promising direction.This includes biodegradable Mg-based alloy used for designing bone screw-plates that has been proven to be beneficial for fracture healing,an innovative development that attracts more and more attention.This paper also indicates that the tantalum bone plates with porous structure are also emerging as a new fracture internal fixation implants.The reduction of the stress shielding is verified to be useful to accelerate bone fracture healing.Potential application of biodegradable metals may also avoid a second operation for implant removal.Further developments in biometals and their design for orthopedic bone plates are expected to improve the treatment of bone fracture,especially the refractory fractures.     

Enhancement of delamination fatigue resistance in carbon nanotube reinforced glass fiber/polymer composites

We show that the addition of small volume fractions of multi-walled carbon nanotubes (CNTs) to the matrix of glass–fiber composites reduces cyclic delamination crack propagation rates significantly. In addition, both critical and sub-critical inter-laminar fracture toughness values are increased. These results corroborate recent experimental evidence that the incorporation of CNTs improve fatigue life by a factor of two to three in in-plane cyclic loading. We show that in both the critical and sub-critical cases, the degree of delamination suppression is most pronounced at lower levels of applied cyclic strain energy release rate, ΔG. High-resolution scanning electron microscopy of the fracture surfaces suggests that the presence of the CNTs at the delamination crack front slows the propagation of the crack due to crack bridging, nanotube fracture, and nanotube pull-out. Further examination of the sub-critical fracture surfaces shows that the relative proportion of CNT pull-out to CNT fracture is dependent on the applied cyclic strain energy, with pull-out dominating as ΔG is reduced. The conditions for crack propagation via matrix cracking and nanotube pull-out and fracture are studied analytically using fracture mechanics theory and the results compared with data from the experiments. It is believed that the shift in the fracture behavior of the CNTs is responsible for the associated increase in the inter-laminar fracture resistance that is observed at lower levels of ΔG relative to composites not containing CNTs.     

Intralaminar fracture mechanism in unidirectional CFRP composites — part II: analysis

Three kinds of representative carbon fiber reinforced unidirectional composite materials are used, and their intralaminar fracture behavior is investigated by using the double-cantilever beam (DCB) specimen with a simultaneous acoustic emission measuring. In Part I, the experimental results on the crack propagation, the bridging fibers, the intralaminar fracture toughness acoustic emission characteristics and microscope observations were obtained. Here, we use a bridging fiber model to analyze the debonding force acting on a bridging fiber and try to estimate the number of bridging fibers during the crack propagating process. At the same time, the intralaminar fracture toughness is calculated by both the adhesive force model and the finite element analysis. As a result, it is found that the intralaminar fracture toughness without the bridging fibers will have a constant value during the crack propagation, but it increases greatly when bridging fibers exist. It is clear that the bridging fibers play an important role in the intralaminar fracture toughness. The debonding forces acting on the bridging fibers and the number of bridging fibers are obtained. Furthermore, the quantitative estimation of the increment of the intralaminar fracture toughness contributed by bridging fibers is made according to the adhesive force model and it is comparable with the results obtained by the finite element analysis.     

Mixed-mode quasi-static failure criteria for adhesively-bonded pultruded GFRP joints

The strain energy release rates of adhesively-bonded pultruded GFRP joints were determined experimentally. The crack propagated in the adherend along paths outside the symmetry plane accompanied by fiber bridging. A new method, designated the “extended global method”, was introduced to facilitate mode partitioning in the mixed-mode experiments. Non-linear finite element models were developed in order to quantify the effect of the observed fiber bridging on crack propagation. An exponential traction-separation cohesive law was used to model the fiber bridging zone and calculate the energy release rate due to the fiber bridging, while the virtual crack closure technique was used for calculation of the fracture components at the crack tip. Experimental, analytical and numerical analyses were used to establish quasi-static mixed-mode failure criteria for crack initiation and propagation. The derived mixed-mode failure criteria can be used for simulating progressive crack propagation in other joint configurations comprising the same adhesive and adherends.     

Small extracellular vesicles secreted by urine-derived stem cells enhanced wound healing in aged mice by ameliorating cellular senescence

Regeneration of chronic skin wounds in tissue is still a key challenge in regenerative medicine because of the accumulation of senescent cells and increasing secretion of s¨enescence-associated secretory phenotype(SASP)in the wound site.Recently,some studies have reported that small extracellular vesicles(sEVs)derived from stem cells can alleviate cellular senescence with very low risk of tumorigenesis and immune responses.As our previous studies have shown that urine-derived stem cells(USCs)can be obtained easily and noninvasively and sEVs derived from USCs(USC-sEVs)have capabilities of regenerating tissue injuries,using USC-sEVs to enhance chronic skin wound healing in aged tissue might be a feasible and efficient strategy.Therefore,in this study,the USC-sEVs were collected and firstly loaded in a human acellular amniotic membrane(HAAM)for controlled releasing and locating the USC-sEVs in the wound site before they were implanted into a chronic skin wound in aged mice.In vivo results showed that the USC-sEVs in HAAM could effectively accelerate the wound healing by ameliorating cellular senescence and reducing the secretion of SASP in the aged skin wounds.To elucidate the mechanism,USC-sEVs were used to in vitro culture human dermal fibroblasts(HDFs)and results showed that USC-sEVs could rejuvenate senescent fibroblasts by reversing the aging phenotypes of senescent HDFs and efficiently reducing the secretion of SASP after they activated the Sirt1 pathway.Therefore,USC-sEVs are efficient for enhancing wound healing in aged mice by ameliorating cellular senescence.