Unraveling the influence of channel size and shape in 3D printed ceramic scaffolds on osteogenesis.

Bone has the capacity to regenerate itself for relatively small defects; however, this regenerative capacity is diminished in critical-size bone defects. The development of synthetic materials has risen as a distinct strategy to address this challenge. Effective synthetic materials to have emerged in recent years are bioceramic implants, which are biocompatible and highly bioactive. Yet nothing suitable for the repair of large bone defects has made the transition from laboratory to clinic. The clinical success of bioceramics has been shown to depend not only on the scaffold's intrinsic material properties but also on its internal porous geometry. This study aimed to systematically explore the implications of varying channel size, shape, and curvature in tissue scaffolds on in vivo bone regeneration outcomes. 3D printed bioceramic scaffolds with varying channel sizes (0.3 mm to 1.5 mm), shapes (circular vs rectangular), and curvatures (concave vs convex) were implanted in rabbit femoral defects for 8 weeks, followed by histological evaluation. We demonstrated that circular channel sizes of around 0.9 mm diameter significantly enhanced bone formation, compared to channel with diameters of 0.3 mm and 1.5 mm. Interestingly, varying channel shapes (rectangular vs circular) had no significant effect on the volume of newly formed bone. Furthermore, the present study systematically demonstrated the beneficial effect of concave surfaces on bone tissue growth in vivo, reinforcing previous in silico and in vitro findings. This study demonstrates that optimizing architectural configurations within ceramic scaffolds is crucial in enhancing bone regeneration outcomes. STATEMENT OF SIGNIFICANCE: Despite the explosion of work on developing synthetic scaffolds to repair bone defects, the amount of new bone formed by scaffolds in vivo remains suboptimal. Recent studies have illuminated the pivotal role of scaffolds' internal architecture in osteogenesis. However, these investigations have mostly remained confined to in silico and in vitro experiments. Among the in vivo studies conducted, there has been a lack of systematic analysis of individual architectural features. Herein, we utilized bioceramic 3D printing to conduct a systematic exploration of the effects of channel size, shape, and curvature on bone formation in vivo. Our results demonstrate the significant influence of channel size and curvature on in vivo outcomes. These findings provide invaluable insights into the design of more effective bone scaffolds.

Discovering an unknown territory using atom probe tomography: Elemental exchange at the bioceramic scaffold/bone tissue interface.

Here we report the first atom probe study to reveal the atomic-scale composition of in vivo bone formed in a bioceramic scaffold (strontium-hardystonite-gahnite) after 12-month implantation in a large bone defect in sheep tibia. The composition of the newly formed bone tissue differs to that of mature cortical bone tissue, and elements from the degrading bioceramic implant, particularly aluminium (Al), are present in both the newly formed bone and in the original mature cortical bone tissue at the perimeter of the bioceramic implant. Atom probe tomography confirmed that the trace elements are released from the bioceramic and are actively transported into the newly formed bone. NanoSIMS mapping, as a complementary technique, confirmed the distribution of the released ions from the bioceramic into the newly formed bone tissue within the scaffold. This study demonstrated the combined benefits of atom probe and nanoSIMS in assessing nanoscopic chemical composition changes at precise locations within the tissue/biomaterial interface. Such information can assist in understanding the interaction of scaffolds with surrounding tissue, hence permitting further iterative improvements to the design and performance of biomedical implants, and ultimately reducing the risk of complications or failure while increasing the rate of tissue formation. STATEMENT OF SIGNIFICANCE: The repair of critical-sized load-bearing bone defects is a challenge, and precisely engineered bioceramic scaffold implants is an emerging potential treatment strategy. However, we still do not understand the effect of the bioceramic scaffold implants on the composition of newly formed bone in vivo and surrounding existing mature bone. This article reports an innovative route to solve this problem, the combined power of atom probe tomography and nanoSIMS is used to spatially define elemental distributions across bioceramic implant sites. We determine the nanoscopic chemical composition changes at the Sr-HT Gahnite bioceramic/bone tissue interface, and importantly, provide the first report of in vivo bone tissue chemical composition formed in a bioceramic scaffold.

Design and evaluation of 3D-printed Sr-HT-Gahnite bioceramic for FDA regulatory submission: A Good Laboratory Practice sheep study.

There is an unmet clinical need for a spinal fusion implant material that recapitulates the biological and mechanical performance of natural bone. We have developed a bioceramic, Sr-HT-Gahnite, which has been identified as a potential fusion device material. This material has the capacity to transform the future of the global interbody devices market, with follow on social, economic, and environmental benefits, rooted in its remarkable combination of mechanical properties and bioactivity. In this study, and in line with FDA requirements, the in vivo preclinical systemic biological safety of a Sr-HT-Gahnite interbody fusion device is assessed over 26 weeks in sheep under good laboratory practice (GLP). Following the in-life phase, animals are assessed for systemic biological effects via blood haematology and clinical biochemistry, strontium dosage analysis in the blood and wool, and histopathology examination of the distant organs including adrenals, brain, heart, kidneys, liver, lungs and bronchi, skeletal muscle, spinal nerves close to the implanted sites, ovaries, and draining lymph nodes. Our results show that no major changes in blood haematology or biochemistry parameters are observed, no systemic distribution of strontium to the blood and wool, and no macroscopic or histopathological abnormalities in the distant organs when Sr-HT-Gahnite was implanted, compared to baseline and control values. Together, these results indicate the systemic safety of the Sr-HT-Gahnite interbody fusion device. The results of this study extend to the systemic safety of other Sr-HT-Gahnite implanted medical devices in contact with bone or tissue, of similar size and manufactured using the described processes. STATEMENT OF SIGNIFICANCE: This paper is considered original and innovative as it is the first that thoroughly reports the systemic biological safety of previously undescribed bioceramic material, Sr-HT-Gahnite. The study has been performed under good laboratory practice, in line with FDA requirements for assessment of a new interbody fusion device, making the results broadly applicable to the translation of sheep models to the human cervical spine; and also the translation of Sr-HT-Gahnite as a biomaterial for use in additional applications. We expect this study to be of broad interest to the readership of Acta Biomaterilia. Its findings are directly applicable to researchers and clinicians working in bone repair and the development of synthetic biomaterials.

Inclusion of the in-chain sulfur in 3-thiaCTU increases the efficiency of mitochondrial targeting and cell killing by anticancer aryl-urea fatty acids.

Mitochondria in tumor cells are functionally different from those in normal cells and could be targeted to develop new anticancer agents. We showed recently that the aryl-ureido fatty acid CTU is the prototype of a new class of mitochondrion-targeted agents that kill cancer cells by increasing the production of reactive oxygen species (ROS), activating endoplasmic reticulum (ER)-stress and promoting apoptosis. However, prolonged treatment with high doses of CTU were required for in vivo anti-tumor activity. Thus, new strategies are now required to produce agents that have enhanced anticancer activity over CTU. In the present study we prepared a novel aryl-urea termed 3-thiaCTU, that contained an in-chain sulfur heteroatom, for evaluation in tumor cell lines and in mice carrying tumor xenografts. The principal finding to emerge was that 3-thiaCTU was several-fold more active than CTU in the activation of aryl-urea mechanisms that promoted cancer cell killing. Thus, in in vitro studies 3-thiaCTU disrupted the mitochondrial membrane potential, increased ROS production, activated ER-stress and promoted tumor cell apoptosis more effectively than CTU. 3-ThiaCTU was also significantly more active than CTUin vivo in mice that carried MDA-MB-231 cell xenografts. Compared to CTU, 3-thiaCTU prevented tumor growth more effectively and at much lower doses. These findings indicate that, in comparison to CTU, 3-thiaCTU is an aryl-urea with markedly enhanced activity that could now be suitable for development as a novel anticancer agent.

Nicotinamide Mononucleotide Alleviates Osteoblast Senescence Induction and Promotes Bone Healing in Osteoporotic Mice.

Combating the accumulated senescent cells and the healing of osteoporotic bone fractures in the older remains a significant challenge. Nicotinamide mononucleotide (NMN), a precursor of NAD+, is an excellent candidate for mitigating aging-related disorders. However, it is unknown if NMN can alleviate senescent cell induction and enhance osteoporotic bone fracture healing. Here we show that NMN treatment partially reverses the effects of tumor necrosis factor-alpha (TNF-α) on human primary osteoblasts (HOBs): senescent cell induction, diminished osteogenic differentiation ability, and intracellular NAD+ and NADH levels. Mechanistically, NMN restores the mitochondrial dysfunction in HOBs induced by TNF-α evidenced by increased mitochondrial membrane potential and reduced reactive oxidative species and mitochondrial mass. NMN also increases mitophagy activity by down-regulating P62 expression and up-regulating light chain 3B-II protein expression. In addition, the cell senescence protective effects of NMN on HOBs are mitigated by a mitophagy inhibitor (Bafilomycin A1). In vivo, NMN supplementation attenuates senescent cell induction in growth plates, partially prevents osteoporosis in an ovariectomized mouse model, and accelerates bone healing in osteoporotic mice. We conclude that NMN can be a novel and promising therapeutic candidate to enhance bone fracture healing capacity in the older.

Functional Ultra-High Molecular Weight Polyethylene Composites for Ligament Reconstructions and Their Targeted Applications in the Restoration of the Anterior Cruciate Ligament.

The selection of biomaterials as biomedical implants is a significant challenge. Ultra-high molecular weight polyethylene (UHMWPE) and composites of such kind have been extensively used in medical implants, notably in the bearings of the hip, knee, and other joint prostheses, owing to its biocompatibility and high wear resistance. For the Anterior Cruciate Ligament (ACL) graft, synthetic UHMWPE is an ideal candidate due to its biocompatibility and extremely high tensile strength. However, significant problems are observed in UHMWPE based implants, such as wear debris and oxidative degradation. To resolve the issue of wear and to enhance the life of UHMWPE as an implant, in recent years, this field has witnessed numerous innovative methodologies such as biofunctionalization or high temperature melting of UHMWPE to enhance its toughness and strength. The surface functionalization/modification/treatment of UHMWPE is very challenging as it requires optimizing many variables, such as surface tension and wettability, active functional groups on the surface, irradiation, and protein immobilization to successfully improve the mechanical properties of UHMWPE and reduce or eliminate the wear or osteolysis of the UHMWPE implant. Despite these difficulties, several surface roughening, functionalization, and irradiation processing technologies have been developed and applied in the recent past. The basic research and direct industrial applications of such material improvement technology are very significant, as evidenced by the significant number of published papers and patents. However, the available literature on research methodology and techniques related to material property enhancement and protection from wear of UHMWPE is disseminated, and there is a lack of a comprehensive source for the research community to access information on the subject matter. Here we provide an overview of recent developments and core challenges in the surface modification/functionalization/irradiation of UHMWPE and apply these findings to the case study of UHMWPE for ACL repair.

The aryl-ureido fatty acid CTU activates endoplasmic reticulum stress and PERK/NOXA-mediated apoptosis in tumor cells by a dual mitochondrial-targeting mechanism.

The cancer cell mitochondrion is functionally different from that in normal cells and could be targeted to develop novel experimental therapeutics. The aryl-ureido fatty acid CTU (16({[4-chloro-3-(trifluoromethyl)phenyl]-carbamoyl}amino)hexadecanoic acid) is the prototype of a new class of mitochondrion-targeted agents that kill cancer cells. Here we show that CTU rapidly depolarized the inner mitochondrial membrane, selectively inhibited complex III of the electron transport chain and increased reactive oxygen species (ROS) production. From RNA-seq analysis, endoplasmic reticulum (ER)-stress was a major activated pathway in CTU-treated cells and in MDA-MB-231 tumor xenografts from CTU-treated nu/nu mice. Mitochondrion-derived ROS activated the PERK-linked ER-stress pathway and induced the BH3-only protein NOXA leading to outer mitochondrial membrane (OMM) disruption. The lipid peroxyl scavenger α-tocopherol attenuated CTU-dependent ER-stress and apoptosis which confirmed the critical role of ROS. Oleic acid protected against CTU-mediated apoptosis by activating Mcl-1 expression, which increased NOXA sequestration and prevented OMM disruption. Taken together, CTU both uncouples mitochondrial electron transport and activates ROS production which promotes ER-stress-dependent OMM disruption and tumor cell death. Dual-mitochondrial targeting agents like CTU offer a novel approach for development of new anti-cancer therapeutics.

PTU, a novel ureido-fatty acid, inhibits MDA-MB-231 cell invasion and dissemination by modulating Wnt5a secretion and cytoskeletal signaling.

Migration and invasion promote tumor cell metastasis, which is the leading cause of cancer death. At present there are no effective treatments. Epidemiological studies have suggested that ω-3 polyunsaturated fatty acids (PUFA) may decrease cancer aggressiveness. In recent studies epoxide metabolites of ω-3 PUFA exhibited anti-cancer activity, although increased in vivo stability is required to develop useful drugs. Here we synthesized novel stabilized ureido-fatty acid ω-3 epoxide isosteres and found that one analogue - p-tolyl-ureidopalmitic acid (PTU) - inhibited migration and invasion by MDA-MB-231 breast cancer cells in vitro and in vivo in xenografted nu/nu mice. From proteomics analysis of PTU-treated cells major regulated pathways were linked to the actin cytoskeleton and actin-based motility. The principal finding was that PTU impaired the formation of actin protrusions by decreasing the secretion of Wnt5a, which dysregulated the Wnt/planar cell polarity (PCP) pathway and actin cytoskeletal dynamics. Exogenous Wnt5a restored invasion and Wnt/PCP signalling in PTU-treated cells. PTU is the prototype of a novel class of agents that selectively dysregulate the Wnt/PCP pathway by inhibiting Wnt5a secretion and actin dynamics to impair MDA-MB-231 cell migration and invasion.

Personalized Baghdadite scaffolds: stereolithography, mechanics and in vivo testing.

An ongoing challenge in the field of orthopedics is to produce a clinically relevant synthetic ceramic scaffold for the treatment of 'critical-sized' bone defects, which cannot heal without intervention. We had developed a bioactive ceramic (baghdadite, Ca3ZrSi2O9) and demonstrated its outstanding bioactivity using traditional manufacturing techniques. Here, we report on the development of a versatile stereolithography printing technology that enabled fabrication of anatomically-shaped and -sized Baghdadite scaffolds. We assessed the in vivo bioactivity of these scaffolds in co-delivering of bone morphogenetic protein-2 (BMP2) and zoledronic acid (ZA) through bioresorbable coatings to induce bone formation and increase retention in a rat model of heterotopic ossification. Micro-computed tomography, histology, mechanical tests pre- and post-implantation, and mechanical modelling were used to assess bone ingrowth and its effects on the mechanics of the scaffolds. Bone ingrowth and the consequent mechanical properties of the scaffolds improved with increasing BMP2 dose. Co-delivery of ZA with BMP2 further improved this outcome. The significant bone formation within the scaffolds functionalized with 10 µg BMP2 and 2 µg ZA made them 2.3 × stiffer and 2.7 × stronger post-implantation and turned these inherently brittle scaffolds into a tough and deformable material. The effects of bone ingrowth on the mechanical properties of scaffolds were captured in a mechanical model that can be used in future clinical studies for non-destructive evaluation of scaffold's stiffness and strength as new bone forms. These results support the practical utilization of our versatile stereolithographic printing methods and BMP2/ZA functionalization to create fit-for-purpose personalized implants for clinical trials. STATEMENT OF SIGNIFICANCE: In this study, we addressed a long-standing challenge of developing a ceramic printing technology that enables fabrication of customizable anatomically-shaped and -sized bioceramic scaffolds with precise internal architectures using an inexpensive desktop printer. We also addressed another challenge related to delivery of pharmaceuticals. BMP2, currently available as a bone-inducing bioactive protein, is clinically administered in a collagen scaffold that has limited moldability and poor mechanical properties. The comparably stiffer and stronger 3D printed personalized Baghdadite scaffolds developed here can be readily functionalized with bioresorbable coatings containing BMP2 ± ZA. These innovations considerably improve on the prior art and are scalable for use in human surgery.

Baghdadite Ceramics Prevent Senescence in Human Osteoblasts and Promote Bone Regeneration in Aged Rats.

Bone fractures and critical-sized bone defects present significant health threats for the elderly who have limited capacity for regeneration due to the presence of functionally compromised senescent cells. A wide range of synthetic materials has been developed to promote the regeneration of critical-sized bone defects, but it is largely unknown if a synthetic biomaterial (scaffold) can modulate cellular senescence and improve bone regeneration in aged scenarios. The current study investigates the interaction of Baghdadite (Ca3ZrSi2O9) ceramic scaffolds with senescent human primary osteoblast-like cells (HOBs) and its bone regeneration capacity in aged rats. A senescent HOB model was established by repeatedly passaging HOBs till passage 7 (P7). Compared to the clinically used hydroxyapatite/tricalcium phosphate (HA/TCP), Baghdadite prevented senescence induction in P7 HOBs and markedly negated the paracrine effect of P7 HOB secretomes that mediated the up-regulations of cellular senescence-associated gene expression levels in P2 HOBs. We further demonstrated that conditioned media extracted from Baghdadite corrected the dysfunctional mitochondria in P7 HOBs. In vivo, the bone regeneration capacity was enhanced when 3D printed Baghdadite scaffolds were implanted in a calvaria critical-sized bone defect model in both young and aged rats compared to HA/TCP scaffolds, but a better effect was observed in aged rats than in young rats. This study suggests that Baghdadite ceramic represents a novel and promising biomaterial approach to promote bone regeneration capacity in the elderly by providing an anti-senescent microenvironment.

Reprogramming of human fibroblasts into osteoblasts by insulin-like growth factor-binding protein 7.

The induced pluripotent stem cell (iPSC) is a promising cell source for tissue regeneration. However, the therapeutic value of iPSC technology is limited due to the complexity of induction protocols and potential risks of teratoma formation. A trans-differentiation approach employing natural factors may allow better control over reprogramming and improved safety. We report here a novel approach to drive trans-differentiation of human fibroblasts into functional osteoblasts using insulin-like growth factor binding protein 7 (IGFBP7). We initially determined that media conditioned by human osteoblasts can induce reprogramming of human fibroblasts to functional osteoblasts. Proteomic analysis identified IGFBP7 as being significantly elevated in media conditioned with osteoblasts compared with those with fibroblasts. Recombinant IGFBP7 induced a phenotypic switch from fibroblasts to osteoblasts. The switch was associated with senescence and dependent on autocrine IL-6 signaling. Our study supports a novel strategy for regenerating bone by using IGFBP7 to trans-differentiate fibroblasts to osteoblasts.

High-Strength Fiber-Reinforced Composite Hydrogel Scaffolds as Biosynthetic Tendon Graft Material.

The development of suitable synthetic scaffolds for use as human tendon grafts to repair tendon ruptures remains a significant engineering challenge. Previous synthetic tendon grafts have demonstrated suboptimal tissue ingrowth and synovitis due to wear particles from fiber-to-fiber abrasion. In this study, we present a novel fiber-reinforced hydrogel (FRH) that mimics the hierarchical structure of the native human tendon for synthetic tendon graft material. Ultrahigh molecular weight polyethylene (UHMWPE) fibers were impregnated with either biosynthetic polyvinyl alcohol/gelatin hydrogel (FRH-PG) or with polyvinyl alcohol/gelatin + strontium-hardystonite (Sr-Ca2ZnSi2O7, Sr-HT) composite hydrogel (FRH-PGS). The scaffolds were fabricated and assessed to evaluate their suitability for tendon graft applications. The microstructure of both FRH-PG and FRH-PGS showed successful impregnation of the hydrogel component, and the tendon scaffolds exhibited equilibrium water content of ∼70 wt %, similar to the values reported for native human tendon, compared to ∼50 wt % water content retained in unmodified UHMWPE fibers. The tensile strength of FRH-PG and FRH-PGS (77.0-81.8 MPa) matched the range of human Achilles' tendon tensile strengths reported in the literature. In vitro culture of rat tendon stem cells showed cell and tissue infiltration into both FRH-PG and FRH-PGS after 2 weeks, and the presence of Sr-HT ceramic particles influenced the expression of tenogenic markers. On the other hand, FRH-PG supported the proliferation of murine C2C12 myoblasts, whereas FRH-PGS seemingly did not support it under static culture conditions. In vivo implantation of FRH-PG and FRH-PGS scaffolds into full-thickness rat patellar tendon defects showed good collagenous tissue ingrowth into these scaffolds after 6 weeks. This study demonstrates the potential viability for our FRH-PG and FRH-PGS scaffolds to be used for off-the-shelf biosynthetic tendon graft material.

A Novel Bone Substitute with High Bioactivity, Strength, and Porosity for Repairing Large and Load-Bearing Bone Defects.

Achieving adequate healing in large or load-bearing bone defects is highly challenging even with surgical intervention. The clinical standard of repairing bone defects using autografts or allografts has many drawbacks. A bioactive ceramic scaffold, strontium-hardystonite-gahnite or "Sr-HT-Gahnite" (a multi-component, calcium silicate-based ceramic) is developed, which when 3D-printed combines high strength with outstanding bone regeneration ability. In this study, the performance of purely synthetic, 3D-printed Sr-HT-Gahnite scaffolds is assessed in repairing large and load-bearing bone defects. The scaffolds are implanted into critical-sized segmental defects in sheep tibia for 3 and 12 months, with bone autografts used for comparison. The scaffolds induce substantial bone formation and defect bridging after 12 months, as indicated by X-ray, micro-computed tomography, and histological and biomechanical analyses. Detailed analysis of the bone-scaffold interface using focused ion beam scanning electron microscopy and multiphoton microscopy shows scaffold degradation and maturation of the newly formed bone. In silico modeling of strain energy distribution in the scaffolds reveal the importance of surgical fixation and mechanical loading on long-term bone regeneration. The clinical application of 3D-printed Sr-HT-Gahnite scaffolds as a synthetic bone substitute can potentially improve the repair of challenging bone defects and overcome the limitations of bone graft transplantation.

A comparative histomorphological and micro computed tomography study of the primary stability and the osseointegration of The Sydney Mini Screw; a qualitative pilot animal study in New Zealand rabbits.

OBJECTIVE: The aim of this study was to assess the potential of improving orthodontic miniscrews' (MSs) primary stability in vivo by evaluating the dispersion capacity of an injectable bone graft substitute (iBGS) through a newly designed hollow MS [The Sydney Mini Screw (SMS)] and its integration with the cortical and trabecular bone by using the femur and tibia in a New Zealand rabbit animal model. METHODS: In total, 24 MSs were randomly placed in each proximal tibia and femur of 6 New Zealand rabbits with an open surgery process. Aarhus MSs were used as controls and the effect of injection of iBGS was studied by implanting SMSs with and without iBGS injection. The dispersion of iBGS and the integration of the SMS were studied by using micro Computed Tomography (µCT) and histochemical analysis at two time points, 0 day and 8 weeks post-implantation. RESULTS: iBGS was successfully injected through the SMS and hardened in situ. After 8 weeks, µCT results revealed that the iBGS particles were resorbed and bone tissue was formed around the SMS and within its lateral exit holes. CONCLUSIONS: This pilot animal study showed the high potential of the combined use of iBGS and SMS as a newly developed technique to promote the primary stability of MSs.

Architectural Design of 3D Printed Scaffolds Controls the Volume and Functionality of Newly Formed Bone.

The successful regeneration of functional bone tissue in critical-size defects remains a significant clinical challenge. To address this challenge, synthetic bone scaffolds are widely developed, but remarkably few are translated to the clinic due to poor performance in vivo. Here, it is demonstrated how architectural design of 3D printed scaffolds can improve in vivo outcomes. Ceramic scaffolds with different pore sizes and permeabilities, but with similar porosity and interconnectivity, are implanted in rabbit calvaria for 12 weeks, and then the explants are harvested for microcomputed tomography evaluation of the volume and functionality of newly formed bone. The results indicate that scaffold pores should be larger than 390 µm with an upper limit of 590 µm to enhance bone formation. It is also demonstrated that a bimodal pore topology-alternating large and small pores-enhances the volume and functionality of new bone substantially. Moreover, bone formation results indicate that stiffness of new bone is highly influenced by the scaffold's permeability in the direction concerned. This study demonstrates that manipulating pore size and permeability in a 3D printed scaffold architecture provides a useful strategy for enhancing bone regeneration outcomes.

Nanoemulsion-Enabled Oral Delivery of Novel Anticancer ω-3 Fatty Acid Derivatives.

Lipid-based drugs are emerging as an interesting class of novel anticancer drugs with the potential to target specific cancer cell metabolic pathways linked to their proliferation and invasiveness. In particular, ω-3 polyunsaturated fatty acids (PUFA) derivatives such as epoxides and their bioisosteres have demonstrated the potential to suppress growth and promote apoptosis in triple-negative human breast cancer cells MDA-MB-231. In this study, 16-(4'-chloro-3'-trifluorophenyl)carbamoylamino]hexadecanoic acid (ClFPh-CHA), an anticancer lipid derived from ω-3,17,18-epoxyeicosanoic acid, was formulated as a stable nanoemulsion with size around 150 nm and narrow droplet size distribution (PDI < 0.200) through phase-inversion emulsification process followed by high pressure homogenization in view of an oral administration. The ClFPh-CHA-loaded nanoemulsions were able to significantly decrease the relative tumor volume in mice bearing an intramammary tumor xenograft at all doses tested (2.5, 10 and 40 mg/kg) after 32 days of daily oral administration. Furthermore, absolute tumor weight was decreased to 50% of untreated control at 10 and 40 mg/kg, while intraperitoneal administration could achieve a significant reduction only at the highest dose of 40 mg/kg. Results suggest that oral administration of ClFPh-CHA formulated as a nanoemulsion has a sufficient bioavailability to provide an anticancer effect in mice and that the activity is at least equal if not superior to that obtained by a conventional parenteral administration of equivalent doses of the same drug.

Effects of Material-Tissue Interactions on Bone Regeneration Outcomes Using Baghdadite Implants in a Large Animal Model.

Extensive bone loss due to trauma or disease leads to impaired healing. Current bone grafts and substitutes have major drawbacks that limit their effectiveness for treating large bone defects. A number of bone substitutes in development are undergoing preclinical testing, but few studies specifically investigate the in vivo material-tissue interactions that provide an important indicator to long-term implant safety and efficacy. This study is the first of its kind to specifically investigate in vivo material-tissue interactions at the bone-implant interface. Baghdadite scaffolds implanted in critical-sized segmental defects in sheep tibia for 26 weeks are analyzed by focused ion beam scanning electron microscopy, multiphoton microscopy, and histology. The scaffolds are seen to induce extensive bone formation that directly abut the implant surfaces with no evidence of chronic inflammation or fibrous capsule formation. Bone remodeling is influenced by slow in vivo degradation around and within the implant, causing portions of the implant to be incorporated into the newly formed bone. These findings have important implications for predicting the long-term effects of baghdadite ceramics in promoting defect healing, and support the translation of baghdadite scaffolds as a new generation of bone graft substitutes with improved properties for the repair of large bone defects.

A Novel Arylurea Fatty Acid That Targets the Mitochondrion and Depletes Cardiolipin To Promote Killing of Breast Cancer Cells.

Cancer cell mitochondria are promising anticancer drug targets because they control cell death and are structurally and functionally different from normal cell mitochondria. We synthesized arylurea fatty acids and found that the analogue 16-({[4-chloro-3-(trifluoromethyl)phenyl]carbamoyl}amino)hexadecanoic acid (13b) decreased proliferation and activated apoptosis in MDA-MB-231 breast cancer cells in vitro and in vivo. In mechanistic studies 13b emerged as the prototype of a novel class of mitochondrion-targeted agents that deplete cardiolipin and promote cancer cell death.

Loss of the vitamin D receptor in human breast and prostate cancers strongly induces cell apoptosis through downregulation of Wnt/ß-catenin signaling.

Vitamin D co-regulates cell proliferation, differentiation and apoptosis in numerous tissues, including cancers. The known anti-proliferative and pro-apoptotic actions of the active metabolite of vitamin D, 1,25-dihydroxy-vitamin D [1,25(OH)2D] are mediated through binding to the vitamin D receptor (VDR). Here, we report on the unexpected finding that stable knockdown of VDR expression in the human breast and prostate cancer cell lines, MDA-MB-231 and PC3, strongly induces cell apoptosis and inhibits cell proliferation in vitro. Implantation of these VDR knockdown cells into the mammary fat pad (MDA-MB-231), subcutaneously (PC3) or intra-tibially (both cell lines) in immune-incompetent nude mice resulted in reduced tumor growth associated with increased apoptosis and reduced cell proliferation compared with controls. These growth-retarding effects of VDR knockdown occur in the presence and absence of vitamin D and are independent of whether cells were grown in bone or soft tissues. Transcriptome analysis of VDR knockdown and non-target control cell lines demonstrated that loss of the VDR was associated with significant attenuation in the Wnt/ß-catenin signaling pathway. In particular, cytoplasmic and nuclear ß-catenin protein levels were reduced with a corresponding downregulation of downstream genes such as Axin2, Cyclin D1, interleukin-6 (IL-6), and IL-8. Stabilization of ß-catenin using the GSK-3ß inhibitor BIO partly reversed the growth-retarding effects of VDR knockdown. Our results indicate that the unliganded VDR possesses hitherto unknown functions to promote breast and prostate cancer growth, which appear to be operational not only within but also outside the bone environment. These novel functions contrast with the known anti-proliferative nuclear actions of the liganded VDR and may represent targets for new diagnostic and therapeutic approaches in breast and prostate cancer.