Porous Bone Research Articles

Early Bone Healing on Hydroxyapatite-Coated and Chemically-Modified Hydrophilic Implant Surfaces in an Ovine Model.

Implant topography affects early peri-implant bone healing by changing the osteoconduction rate in the surrounding biological environment. Implant surfaces have been designed to promote faster and stronger bone formation for rapid and stable prosthesis loading. Early peri-implant bone healing has been observed with a sandblasted, acid-etched implant that was chemically modified to be hydrophilic (cmSLA). The present study investigates whether early peri-implant bone healing extends to a rough surface implant with a high crystalline hydroxyapatite surface (TSV MP-1 HA). Three implants were randomly placed in porous trabecular bone within both medial femoral condyles of 10 sheep. Early peri-implant bone stability was measured at 3- and 6-weeks healing time following implant insertion. Results indicated a similar implant stability quotient between the implants at insertion and over time. The significant increase over time of reverse torque values with respect to insertion torque (p < 0.001) did not differ between the implants. However, the bone-to-implant contact of TSV MP-1 HA was significantly higher than that of cmSLA implants at 6 weeks (p < 0.01). These data validate previous findings of a hydrophilic implant surface and extend the observation of early osseointegration to a rough surface implant in porous trabecular bone.

Isotropic porous structure design methods based on triply periodic minimal surfaces

Recently, triply periodic minimal surface (TPMS) is emerging as an ideal tool to generate porous structures. Yet, most of the current work only focuses on controlling the elastic modulus by the relative density. For special engineering applications, such as porous bone implants or energy absorbers, the generated porous TPMS may still be broken due to anisotropy. In this work, two strategies are proposed to design isotropic TPMS structures. The numerical homogenization theory and finite element analysis methods are utilized to study the relationship between TPMS parameters and the elastic modulus or anisotropy properties. Based on that, a Curvature-Wall thickness (CW) adjustment method is proposed for sheet TPMS structures whose performances are close to isotropy properties. In virtue of the constructed design map, both elastic modulus and anisotropy properties can be controlled. For sheet TPMS structures whose performances are far from the isotropy properties, the TPMS units can be combined to generate composite TPMS, which can be further designed by the proposed Curvature-Wall thickness adjustment method. Experimental results verify the effectiveness and accuracy of the proposed approaches. Appropriate elastic modulus and ideal isotropy properties can be acquired at the same time.

Treatment of pre-collapse non-traumatic osteonecrosis of the femoral head through Orthopdische Chirurgie M\xfcnchen approach combined with autologous bone mixed with \u03b2-tricalcium phosphate porous bioceramic bone graft: a retrospective study of mid-term results

BackgroundThis study aimed to evaluate the clinical efficacy of femoral head and neck fenestration combined with autologous bone mixed with β-tricalcium phosphate porous bioceramic bone (light bulb procedure) through Orthopdische Chirurgie München approach (OCM approach) for pre-collapse non-traumatic osteonecrosis of the femoral head(ONFH).MethodsThe clinical data of 47 patients (47 hips) with ONFH were retrospectively reviewed. The Harris hip score (HHS) was used to evaluate the clinical outcomes. Imaging was assessed by X-ray. Clinical failure was defined as postoperative total hip arthroplasty (THA) or the HHS was poor (< 70). The Kaplan–Meier survival curve was used to conduct a univariate analysis of risk factors. The analysis factors included gender, age, International Association Research Circulation Osseous (ARCO) stage, etiology, body mass index (BMI), 25-hydroxyvitamin D (25(OH)D), and type I collagen carboxy-terminal peptide (CTX). The COX multivariate risk model was used to analyze the risk factors.ResultsAll the 47 hips were followed up for 24–58 months, with an average of 45 months. The Harris score (76.29 ± 10.38) at the last follow-up was significantly higher than the preoperative HHS (64.45 ± 2.93) (P < 0.05). The postoperative HHS was excellent with a success rate of 36.17%. Postoperative imaging evaluation showed that 9 hips improved, 28 hips stabilized, and 10 hips progressed. Moreover, 17 out of 47 hips were defined as a postoperative clinical failure and the success rate was 63.83%. 25(OH)D and preoperative ARCO stage were risk factors for postoperative clinical failure (P < 0.05). The COX multivariate risk model analysis showed that IIIA stage was an independent risk factor for postoperative clinical failure (P < 0.05).ConclusionsThe head and neck fenestration and bone grafting via the OCM approach in the treatment of non-traumatic ONFH in the pre-collapse stage can achieve good clinical outcomes. 25(OH)D deficient patients and ARCO IIIA patients had a higher failure rate of bone graft using this approach.

Immunoporosis: Role of Innate Immune Cells in Osteoporosis.

Osteoporosis or porous bone disorder is the result of an imbalance in an otherwise highly balanced physiological process known as ‘bone remodeling’. The immune system is intricately involved in bone physiology as well as pathologies. Inflammatory diseases are often correlated with osteoporosis. Inflammatory mediators such as reactive oxygen species (ROS), and pro-inflammatory cytokines and chemokines directly or indirectly act on the bone cells and play a role in the pathogenesis of osteoporosis. Recently, Srivastava et al. (Srivastava RK, Dar HY, Mishra PK. Immunoporosis: Immunology of Osteoporosis-Role of T Cells. Frontiers in immunology. 2018;9:657) have coined the term “immunoporosis” to emphasize the role of immune cells in the pathology of osteoporosis. Accumulated pieces of evidence suggest both innate and adaptive immune cells contribute to osteoporosis. However, innate cells are the major effectors of inflammation. They sense various triggers to inflammation such as pathogen-associated molecular patterns (PAMPs), damage-associated molecular patterns (DAMPs), cellular stress, etc., thus producing pro-inflammatory mediators that play a critical role in the pathogenesis of osteoporosis. In this review, we have discussed the role of the innate immune cells in great detail and divided these cells into different sections in a systemic manner. In the beginning, we talked about cells of the myeloid lineage, including macrophages, monocytes, and dendritic cells. This group of cells explicitly influences the skeletal system by the action of production of pro-inflammatory cytokines and can transdifferentiate into osteoclast. Other cells of the myeloid lineage, such as neutrophils, eosinophils, and mast cells, largely impact osteoporosis via the production of pro-inflammatory cytokines. Further, we talked about the cells of the lymphoid lineage, including natural killer cells and innate lymphoid cells, which share innate-like properties and play a role in osteoporosis. In addition to various innate immune cells, we also discussed the impact of classical pro-inflammatory cytokines on osteoporosis. We also highlighted the studies regarding the impact of physiological and metabolic changes in the body, which results in chronic inflammatory conditions such as ageing, ultimately triggering osteoporosis.

Characterization of the Functional Properties of Polycaprolactone Bone Scaffolds Fabricated Using Pneumatic Micro-Extrusion

Abstract Pneumatic micro-extrusion (PME) is a direct-write additive manufacturing process, which has emerged as a robust, high-resolution method for the fabrication of a broad spectrum of biological tissues and organs. PME allows for noncontact multimaterial deposition of functional inks for tissue engineering applications. In spite of the advantages and engendered potential applications, the PME process is inherently complex, governed not only by complex physical phenomena but also by material–process interactions. Consequently, investigation of the influence of PME process parameters as well as the underlying physical phenomena behind material transport and deposition in PME would be inevitably a need. The overarching goal of this research work is to fabricate biocompatible, porous bone tissue scaffolds for the treatment of osseous fractures, defects, and diseases. In pursuit of this goal, the objectives of the work are: (i) to investigate the influence of seven consequential scaffold design factors and PME process parameters on the mechanical properties of fabricated bone tissue scaffolds and (ii) to explore the underlying dynamics behind material transport in the PME process, using a three-dimensional computational fluid dynamics (CFD) model. To investigate the effects of the design and process parameters, a series of experiments were designed and conducted. Layer height was identified as the most significant factor in this study. An increase in the layer height led to less overlap between subsequent layers, which allowed for more shrinkage and ultimately a reduction in scaffold diameter. In addition, print speed appeared as an influential factor in this study. An increase in the print speed resulted in a decline in linear mass density and thus in the extent of fusion between subsequent deposited layers. Besides, it was observed that there was a strong correlation between deposition mass and compression modulus. Overall, the results of this study pave the way for future investigation of PME-deposited polycaprolactone (PCL) scaffolds with optimal functional and medical properties for incorporation of stem cells toward the treatment of osseous fractures and defects.

Experimental measurement and simulation of mechanical strength and biological behavior of porous bony scaffold coated with alginate-hydroxyapatite for femoral applications

In the present study, the 3D printer method for fabricating porous bone scaffolds with Polylactic Acid (PLA) printed was used by Fused Deposition Modelling (FDM). Then, the prepared 3D scaffold coated with alginate composed with various amounts of Hydroxyapatite (HA) using the freeze-drying technique. After the fabrication of novel functional graded materials scaffold, the mechanical strength and biological behavior were investigated. The Scanning Electron Microscopy (SEM) and X-ray Diffraction (XRD) analysis used to characterize the morphology and phase characterization. The structure of the porous scaffold was simulated using the ABAQUS analysis method and their mechanical and physical properties were extracted. Also, strong antibacterial properties were observed for all samples with increasing HA nanoparticles against gram-positive and gram-negative bacterial suspensions. These bio-nano composites are compatible with the pH of the blood. In general, it was found that the most suitable bio-nano composite is the sample with 10 wt % HA nanoparticles. The elastic modulus increased from 350 MPa to 394 MPa with the addition of HA nanoparticles, while the porosity percentages decreased from 44 % to 36 %. The compressive strength increases from 25.2 MPa to 32.7 MPa with the addition of 30 wt% HA nanoparticles. The obtained results showed that the bio-nano composites prepared in this study were suitable for further development in bone substitutes with desirable mechanical properties. The alginate/HA material properties reported from the numerical approach, have an acceptable agreement with experimental results. Furthermore, the numerical study indicated that the scaffold compressive strength increases from 22.8 MPa (S1) to 31.2 MPa (S4) with the addition of 30 wt % HA nanoparticles. Nanoindentation test explains that the indenter penetration decreases from 82.163 nm to 69.338 nm with the addition of 30 wt% HA nanoparticles. As a consequence, PLA-alginate/20 wt % HA (S3) and, PLA-alginate/30 wt% HA (S4) have the best mechanical properties. Although due to lower biological advantages of S4 concerning S3, PLA-alginate/20 wt % HA (S3) chose as an appropriate bio-nanocomposite for orthopedic application.

Proliferation and osteogenic differentiation of mesenchymal stem cells on three-dimensional scaffolds made by thermal sintering method

This article presents a thermal sintering method to fabricate porous bone tissue engineering scaffolds based on polycaprolactone (PCL), polylactic acid (PLA), and their composites. The mechanical properties, porous structure, biodegradability, and biocompatibility of sintered scaffolds were evaluated. The scaffolds showed a porosity in the range of 86–91% with a pore size of 75 m $$\mu$$ to 400 m $$\mu$$ . PCL/PLA composite scaffolds showed a Young’s modulus of around 49 MPa, which was between the modulus values of PCL (24 MPa) and PLA (63 MPa) scaffolds. Fibroblast cells (SNL) exhibited spreading and adhesion on the scaffolds, and scaffolds demonstrated a significant difference in the osteogenic differentiation of human mesenchymal stem cells (hMSCs) after 7 and 14 days of culture in comparison with the control (tissue culture polystyrene). Our results demonstrated that the thermally sintered PCL/PLA composite scaffold could be a promising candidate for bone tissue regeneration.

Delamination-fretting wear failure evaluation at HAp-Ti-6Al–4V interface of uncemented artificial hip implant

Present research aims to develop a finite element computational model to examine delamination-fretting wear behaviour that can suitably mimic actual loading conditions at HAp-Ti-6Al–4V interface of uncemented hip implant femoral stem component. A simple finite element contact configuration model based on fretting fatigue experimental arrangement subjected to different mechanical and tribological properties consist of contact pad (bone), HAp coating and Ti–6Al–4V substrate are developed using adaptive wear modelling approach adopting modified Archard wear equation to be examined under static simulation. The developed finite element model is validated and verified with reported literatures. The findings revealed that significant delamination-fretting wear is recorded at contact edge (leading edge) as a result of substantial contact pressure and contact slip driven by stress singularity effect. The delamination-fretting wear behaviour is promoted under higher delamination length, lower normal loading with higher fatigue loading, increased porous (cancellous) and cortical bone elastic modulus with higher cycle number due to significant relative slip amplitude as the result of reduced interface rigidity. Tensile-compressive condition (R=−1) experiences most significant delamination-fretting wear behaviour (8 times higher) compared to stress ratio R=0.1 and R=10.

An implicit constitutive relation for describing the small strain response of porous elastic solids whose material moduli are dependent on the density

In this short note, we develop a constitutive relation that is linear in both the Cauchy stress and the linearized strain, by linearizing implicit constitutive relations between the stress and the deformation gradient that have been put into place to describe the response of elastic bodies (Rajagopal, KR. On implicit constitutive theories. Applications of Mathematics 2003; 28: 279–319), by assuming that the displacement gradient is small. These implicit equations include the classical linearized elastic constitutive approximation as well as some classes of constitutive relations that imply limiting strain in tension, as special subclasses. Moreover, the constitutive relations that are developed allow the material moduli to depend on the density; thus, they can be used to describe the response of porous materials, such as porous metals, bone, rocks, and concrete undergoing small deformations.

Effect of Skull Porous Trabecular Structure on Transcranial Ultrasound Imaging in the Presence of Elastic Wave Mode Conversion at Varying Incidence Angle

With the advancement of aberration correction techniques, transcranial ultrasound imaging has exhibited great potential in applications such as imaging neurological function and guiding therapeutic ultrasound. However, the feasibility of transcranial imaging varies among individuals because of the differences in skull acoustic properties. To better understand the fundamental mechanisms underlying the variation in imaging performance, the effect of the structure of the porous trabecular bone on transcranial imaging performance (i.e., target localization errors and resolution) was investigated for the first time through the use of elastic wave simulations and experiments. Simulation studies using high-resolution computed tomography data from ex vivo skull samples revealed that imaging at large incidence angles reduced the target localization error for skulls having low porosity; however, as skull porosity increased, large angles of incidence resulted in degradation of resolution and increased target localization errors. Experimental results indicate that imaging at normal incidence introduced a localization error of 1.85 ± 0.10 mm, while imaging at a large incidence angle (40°) resulted in an increased localization error of 6.54 ± 1.33 mm and caused a single point target to no longer appear as a single, coherent target in the resulting image, which is consistent with simulation results. This first investigation of the effects of skull microstructure on transcranial ultrasound imaging indicates that imaging performance is highly dependent on the porosity of the skull, particularly at non-normal angles of incidence.

Local and systemic inflammation after implantation of a novel iron based porous degradable bone replacement material in sheep model

Despite the high potential of healthy bone to regenerate, the reconstruction of large bone defects remains a challenge. Due to the lack of mechanical stability of existing bone substitutes, recently developed degradable metallic alloys are an interesting alternative providing higher load-bearing capabilities. Degradable iron-based alloys therefore might be an attractive innovation. To test the suitability of a newly-designed iron-based alloy for such applications, an animal experiment was performed. Porous iron-based degradable implants with two different densities and a control group were tested. The implants were positioned in the proximal tibia of Merino sheep. Over a period of 6 and 12 months, blood and histological parameters were monitored for signs of inflammation and degradation. In the histological evaluation of the implants` environment we found degraded alloy particles, but no inflammatory reaction. Iron particles were also found within the popliteal lymph nodes on both sides. The serum blood levels of phosphorus, iron and ferritin in the long term groups were elevated. Other parameters did not show any changes. Iron-based degradable porous bone replacement implants showed a good biocompatibility in this experiment. For a clinical application, however, the rate of degradation would have to be significantly increased. Biocompatibility would then have to be re-evaluated.

Zn-substituted Mg2SiO4 nanoparticles-incorporated PCL-silk fibroin composite scaffold: A multifunctional platform towards bone tissue regeneration

Electrospun porous bone scaffolds are known to imitate the extracellular matrix very well and provide an environment through which the tissue formation is enhanced. Although polymeric scaffolds have a great potential in bone tissue regeneration, their weak bioactivity (bone bonding ability) and mechanical properties have left room for improvement. Therefore, the present study focused on the developing a ternary multifunctional platform composed of polycaprolactone (PCL)/silk fibroin (SF)/Zn-substituted Mg2SiO4 nanoparticles for bone tissue regeneration. This study is composed of two connected sections including synthesis and characterization of Mg(2-x)ZnxSiO4, x = 0, 0.5, 1, 1.5, 2 through surfactant-assisted sol-gel technique followed by incorporation of the nanoparticles into PCL/SF hybrid scaffold via electrospinning technique. The weight ratios of polymers and ceramic nanoparticles were optimized to reach desirable textural—porosity, pore size, and fiber diameter—and mechanical properties. Having optimized the ternary scaffold, it was then undergone different physical, chemical, and biological tests in vitro. A precise comparison study between the ternary (PCL/SF/ceramic nanoparticles), binary (PCL/SF), and pure PCL was made to shed light on the effect of each composition on the applicability of ternary scaffold. The overall results confirmed that the Mg1Zn1SiO4 nanoparticles-incorporated PCL/SF scaffold with fluorescence property was the one yielding the highest Young's modulus and desirable textural properties. The ternary scaffold showed improved biological properties making it a promising candidate for further studies towards bone tissue regeneration.

Design and Mechanical Properties Verification of Gradient Voronoi Scaffold for Bone Tissue Engineering.

In order to obtain scaffold that can meet the therapeutic effect, researchers have carried out research on irregular porous structures. However, there are deficiencies in the design method of accurately controlling the apparent elastic modulus of the structure at present. Natural bone has a gradient porous structure. However, there are few studies on the mechanical property advantages of gradient bionic bone scaffold. In this paper, an improved method based on Voronoi-tessellation is proposed. The method can get controllable gradient scaffolds to fit the modulus of natural bone, and accurately control the apparent elastic modulus of porous structure, which is conducive to improving the stress shielding. To verify the designed structure can be fabricated by additive manufacturing, several designed models are obtained by SLM and EBM. Through finite element analysis (FEA), it is verified that the irregular porous structure based on Voronoi-tessellation is more stable than the traditional regular porous structure of the same structure volume, the same pore number and the same material. Furthermore, it is verified that the gradient irregular structure has a better stability than the non-gradient structure. An experiment is conducted successfully to verify the stability performance got by FEA. In addition, a dynamic impact FEA is also performed to simulate impact resistance. The result shows that the impact resistance of the regular porous structure, the irregular porous structure and the gradient irregular porous structure becomes better in turn. The mechanical property verification provides a theoretical basis for the structural design of gradient irregular porous bone tissue engineering scaffolds.

Influence of layer thickness and build orientation on compressive strength of 3D printed scaffolds prototypes

The research on fabrication of customised porous bone scaffolds through additive manufacturing has been gaining momentum since last couple of years. This paper presents an analysis of compressive strength (CS) of two slightly different calcium sulfate based proprietary materials namely, Zp150 and VisiJet PXL Core. The comparison is performed on different layer thickness (LT) and builds orientation (BO) of ZPrinter ®450 using full factorial design approach. Initially, a computer aided design (CAD) model of the porous scaffold with designed porosity is prepared in a CAD modelling software and then additive manufacturing is done on four different LT (0.089 mm, 0.101 mm, 0.112 mm and 0.125 mm) and three BO (along x-axis, y-axis and z-axis) of ZPrinter ®450. Analysis of CS reveals that prototypes made of VisiJet PXL Core material have shown a better strength in comparison with their Zp150 counterparts for similar LT and BO. In general, prototypes of both the materials printed along x-axis posses better CS for all the LT. Moreover, the prototype of VisiJet PXL Core material fabricated along x-axis on layer thickness 0.089 mm possesses highest (1.17 MPa) CS among all the prototypes.

Radiation Characteristics of Cranial Leaky Lamb Waves.

We numerically and experimentally investigate the dispersion properties of leaky Lamb waves in the cranial bone. Cranial Lamb waves leak energy from the skull into the brain when propagating at speeds higher than the speed of sound in the surrounding fluid. The understanding of their radiation mechanism is significantly complicated by the geometric and mechanical characteristics of the cortical tables and the trabecular bone (diploë). Toward such understanding, we here analyze the sub-1.0 MHz radiation angle dispersion spectrum of porous bone phantoms and parietal bone geometries obtained from μ CT scans. Our numerical results show that, when diploic pores are physically modeled, leakage angles computed from time transient finite-element analyses correspond to those predicted by an equivalent three-layered fluid-loaded waveguide model. For the bone geometries analyzed, two main leaky branches are observed in the near-field dispersion spectrum: a fast wave radiated at small angles, which is related to the fastest fundamental Lamb mode supported by the cranial bone, and a slower wave radiated at larger angles. This observation is also confirmed by experimental tests carried out on an immersed parietal bone.

Long-Term Retarded Release for the Proteasome Inhibitor Bortezomib through Temperature-Sensitive Dendritic Glycopolymers as Drug Delivery System from Calcium Phosphate Bone Cement.

For the local treatment of bone defects, highly adaptable macromolecular architectures are still required as drug delivery system (DDS) in solid bone substitute materials. Novel DDS fabricated by host-guest interactions between β-cyclodextrin-modified dendritic glycopolymers and adamantane-modified temperature-sensitive polymers for the proteasome inhibitor bortezomib (BZM) is presented. These DDS induce a short- and long-term (up to two weeks) retarded release of BZM from calcium phosphate bone cement (CPC) in comparison to a burst release of the drug alone. Different release parameters of BZM/DDS/CPC are evaluated in phosphate buffer at 37°C to further improve the long-term retarded release of BZM. This is achieved by increasing the amount of drug (50-100µg) and/or DDS (100-400µg) versus CPC (1g), by adapting the complexes better to the porous bone cement environment, and by applying molar ratios of excess BZM toward DDS with 1:10, 1:25, and 1:100. The temperature-sensitive polymer shells of BZM/DDS complexes in CPC, which allow drug loading at room temperature but are collapsed at body temperature, support the retarding long-term release of BZM from DDS/CPC. Thus, the concept of temperature-sensitive DDS for BZM/DDS complexes in CPC works and matches key points for a local therapy of osteolytic bone lesions.

Mechanical performances of hip implant design and fabrication with PEEK composite

Artificial bone implant materials need porosity for nutrient distribution, moderate pore size to provide cell cultures and bone-like mechanical properties. The homogenisation of discrepancies between the microstructure of implants and bone is an important subject. This research aims to design microstructures with poly ether-ether-ketone (PEEK) and its composites to improve the compatibility of implants. Porous hip bone implants fabricated by fused deposition modelling (FDM) are proposed to mimic natural bone with various homogenisation lattice structures and excellent properties. Five isotropic lattice structures with homogenisation control strategies are printed with PEEK and composite PEEK with reduced graphene oxide (rGO) and calcium hydroxyapatite (cHAp). An examination is performed on a three-dimensional (3D) distribution of the effective module surface of the five composite porous unit lattice structures. The relationship between the modulus of elasticity, anisotropy and cell parameters are thoroughly investigated by finite element analysis (FEA). Analysis of the surface treatment used to create micropores in the scaffolding and the nanostructure yields a bioactive PEEK/hydroxyapatite (HAp) composite with various control configuration distributions and cell growths. The functionalised biocompatibility and degradability of rGO/HAp composite in various ratios to PEEK, and their nanostructure arrays, are studied by a surface functionalisation approach. The improved design eliminates slight imperfections, allowing for a more stable structure. The controlled homogenisation, porosity and particle size distribution helps to increase cellular infiltration and biological integration of the PEEK and hip implant composites.

Development and physicochemical characterization of novel porous phosphate glass bone graft substitute and in vitro comparison with xenograft

The process of bone regeneration in bone grafting procedures is greatly influenced by the physicochemical properties of the bone graft substitute. In this study, porous phosphate glass (PPG) morsels were developed and their physicochemical properties such as degradation, crystallinity, organic content, surface topography, particle size and porosity were evaluated using various analytical methods. The in vitro cytotoxicity of the PPG morsels was assessed and the interaction of the PPG morsels with Dental Pulp Stem Cells (DPSCs) was studied by measuring cell proliferation and cell penetration depth. The cell-material interactions between PPG morsels and a commercially available xenograft (XG) were compared. The PPG morsels were observed to be amorphous, biocompatible and highly porous (porosity = 58.45%). From in vitro experiments, PPG morsels were observed to be non-cytotoxic and showed better cell proliferation. The internal surface of PPG was easily accessible to the cells compared to XG.

Podosome-Driven Defect Development in Lamellar Bone under the Conditions of Senile Osteoporosis Observed at the Nanometer Scale.

The degradation mechanism of human trabecular bone harvested from the central part of the femoral head of a patient with a fragility fracture of the femoral neck under conditions of senile osteoporosis was investigated by high-resolution electron microscopy. As evidenced by light microscopy, there is a disturbance of bone metabolism leading to severe and irreparable damages to the bone structure. These defects are evoked by osteoclasts and thus podosome activity. Podosomes create typical pit marks and holes of about 300–400 nm in diameter on the bone surface. Detailed analysis of the stress field caused by the podosomes in the extracellular bone matrix was performed. The calculations yielded maximum stress in the range of few megapascals resulting in formation of microcracks around the podosomes. Disintegration of hydroxyapatite and free lying collagen fibrils were observed at the edges of the plywood structure of the bone lamella. At the ultimate state, the disintegration of the mineralized collagen fibrils to a gelatinous matrix comes along with a delamination of the apatite nanoplatelets resulting in a brittle, porous bone structure. The nanoplatelets aggregate to big hydroxyapatite plates with a size of up to 10 x 20 μm2. The enhanced plate growth can be explained by the interaction of two mechanisms in the ruffled border zone: the accumulation of delaminated hydroxyapatite nanoplatelets near clusters of podosomes and the accelerated nucleation and random growth of HAP nanoplatelets due to a nonsufficient concentration of process-directing carboxylated osteocalcin cOC.

Biocomputational Analysis of ARAP1, a novel GTPase‐Activating‐Protein Involved in Osteoclast‐Mediated Bone Digestion

RhoGTPases are important regulators of many cellular activities including cell proliferation, trafficking of proteins, DNA transcription, and apoptosis. RhoGTPases are also involved in bone remodeling, the continuous process of bone digestion, and formation that occurs during one's lifespan. Any abnormalities in these Rho Proteins can lead to physiological effects such as osteoporosis, characterized by porous bone, or osteopetrosis, characterized by highly-dense bone tissue. RhoA-GTP is responsible for disrupting sealing zone dynamics and promoting cell motility in osteoclasts. On the other hand, Cdc42-GTP and Rac1-GTP maintain osteoclast attachment to the bone surface, thereby allowing it to continue enzymatic degradation of bone. To employ RhoGTPases as potential drug targets, we need to further understand the complex physiology and control mechanisms for these G-proteins. ARAP1 has been shown to localize to the osteoclast sealing zones upon the release of phosphoinositide (3,4,5) trisphosphate, which binds to its first and third PH domains. ARAP1 belongs to the RhoGAP family of proteins and has a RhoGAP domain responsible for catalyzing the intrinsic GTPase activity of G proteins. However, it is still unknown which GTPase (i.e. RhoA, Cdc42, Rac1) preferentially binds to the ARAP1 catalytic domain. Delineating the substrate specificity for the RhoGAP domain is crucial to its characterization as a pro-migration or pro-digestion protein, allowing us to specifically target its function in managing osteoporosis or osteopetrosis-related illnesses. In this computational study, we have modeled the ARAP1 RhoGAP domain using homology modeling and ab initio techniques, characterized its biophysical features, and used docking analysis to scrutinize its interactions with potential substrates. The docking results suggest that ARAP1 shows a preferential substrate specificity for RhoA. This indicates that ARAP1-RhoGAP may behave in a similar manner as ARAP3-RhoGAP, which has been shown to prefer RhoA-targeting in an experimental setup. Lastly, we report an in silico mutagenesis analysis that induces RhoA-like mutations in Cdc42/Rac1 and provides structural data that supports our hypothesis of an ARAP1-RhoA preferential attraction. Overall, this study reveals new insights into the structure-function relations of the RhoGAP domain, specifically pertaining to its role in osteoclast-mediated bone digestion, using an integrated and comprehensive strategy of sequence and structure analyses. The novel findings from our study regarding the molecular mechanism of GTPases in the osteoclast lays the groundwork towards developing therapies that improve the quality of life for those suffering from chronic bone pathologies.