Earlier studies have shown that the influence of fixation stability on bone healing diminishes wi... more Earlier studies have shown that the influence of fixation stability on bone healing diminishes with advanced age. The goal of this study was to unravel the relationship between mechanical stimulus and age on callus competence at a tissue level. Using 3D in vitro micro-computed tomography derived metrics, 2D in vivo radiography, and histology, we investigated the influences of age and varying fixation stability on callus size, geometry, microstructure, composition, remodeling, and vascularity. Compared were four groups with a 1.5-mm osteotomy gap in the femora of Sprague-Dawley rats: Young rigid (YR), Young semirigid (YSR), Old rigid (OR), Old semirigid (OSR). Hypothesis was that calcified callus microstructure and composition is impaired due to the influence of advanced age, and these individuals would show a reduced response to fixation stabilities. Semirigid fixations resulted in a larger DeltaCSA (Callus cross-sectional area) compared to rigid groups. In vitro microCT analysis at 6 weeks postmortem showed callus bridging scores in younger animals to be superior than their older counterparts (p<0.01). Younger animals showed (i) larger callus strut thickness (p<0.001), (ii) lower perforation in struts (p<0.01), and (iii) higher mineralization of callus struts (p<0.001). Callus mineralization was reduced in young animals with semirigid fracture fixation but remained unaffected in the aged group. While stability had an influence, age showed none on callus size and geometry of callus. With no differences observed in relative osteoid areas in the callus ROI, old as well as semirigid fixated animals showed a higher osteoclast count (p<0.05). Blood vessel density was reduced in animals with semirigid fixation (p<0.05). In conclusion, in vivo monitoring indicated delayed callus maturation in aged individuals. Callus bridging and callus competence (microstructure and mineralization) were impaired in individuals with an advanced age. This matched with increased bone resorption due to higher osteoclast numbers. Varying fixator configurations in older individuals did not alter the dominant effect of advanced age on callus tissue mineralization, unlike in their younger counterparts. Age-associated influences appeared independent from stability. This study illustrates the dominating role of osteoclastic activity in age-related impaired healing, while demonstrating the optimization of fixation parameters such as stiffness appeared to be less effective in influencing healing in aged individuals.Copyright 2010 Elsevier Inc. All rights reserved.
Ample evidence supports that prostate tumor metastasis originates from a rare population of cance... more Ample evidence supports that prostate tumor metastasis originates from a rare population of cancer cells, known as cancer stem cells (CSCs). Unfortunately, little is known about the identity of these cells, making it difficult to target the metastatic prostate tumor. Here, for the first time, we report the identification of a rare population of prostate cancer cells that express the Tie-2 protein. We found that this Tie-2High population exists mainly in prostate cancer cell lines that are capable of metastasizing to the bone. These cells not only express a higher level of CSC markers but also demonstrate enhanced resistance to the chemotherapeutic drug Cabazitaxel. In addition, knockdown of the expression of the Tie-2 ligand angiopoietin (Ang-1) led to suppression of CSC markers, suggesting that the Ang-1/Tie-2 signaling pathway functions as an autocrine loop for the maintenance of prostate CSCs. More importantly, we found that Tie-2High prostate cancer cells are more adhesive than ...
The International journal of oral & maxillofacial implants
This paper reports on a 2-phase study of a novel membrane-scaffold graft construct, its ability t... more This paper reports on a 2-phase study of a novel membrane-scaffold graft construct, its ability to support periodontal ligament fibroblast (PDLF) and alveolar osteoblast (AO) growth in vitro, and its use for tissue engineering a PDL-AO interface in vivo. Human PDLFs were seeded onto perforated poly(epsilon-caprolactone) membranes (n=30) at 78,000 cells/cm2; human AOs were seeded on poly(epsilon-caprolactone) scaffolds (n=30) with fibrin glue at 625,000 cells/cm3. Cell attachment, morphology, viability, and metabolic activity were monitored for 3 weeks in vitro. Subsequently, cell-seeded membrane-scaffold constructs (experimental group, n=9) and nonseeded constructs (control group, n=4) assembled with fibrin glue were implanted subcutaneously into 7 athymic mice for 4 weeks. PDLFs formed confluent layers on membranes, whereas AOs produced mineralized matrices within scaffolds upon osteoinduction in vitro. Well-vascularized tissue formation was observed after implantation. Integration...
Tumour microenvironment greatly influences the development and metastasis of cancer progression. ... more Tumour microenvironment greatly influences the development and metastasis of cancer progression. The development of three dimensional (3D) culture models which mimic that displayed in vivo can improve cancer biology studies and accelerate novel anticancer drug screening. Inspired by a systems biology approach, we have formed 3D in vitro bioengineered tumour angiogenesis microenvironments within a glycosaminoglycan-based hydrogel culture system. This microenvironment model can routinely recreate breast and prostate tumour vascularisation. The multiple cell types cultured within this model were less sensitive to chemotherapy when compared with two dimensional (2D) cultures, and displayed comparative tumour regression to that displayed in vivo. These features highlight the use of our in vitro culture model as a complementary testing platform in conjunction with animal models, addressing key reduction and replacement goals of the future. We anticipate that this biomimetic model will provide a platform for the in-depth analysis of cancer development and the discovery of novel therapeutic targets.
SummarySince the isolation of fetal stem cell populations from perinatal tissues, such as umbilic... more SummarySince the isolation of fetal stem cell populations from perinatal tissues, such as umbilical cord blood and placenta, interest has been growing in understanding their greater plasticity compared with adult stem cells and exploring their potential in regenerative medicine. The phenomenon of fetal microchimerism (FMC) naturally occurring during pregnancy through the transfer of fetal stem/progenitor cells to maternal blood and tissues has been integral in developing this dogma. Specifically, microchimeric mesenchymal stem cells and endothelial progenitors of fetal origin have now demonstrated a capacity for tissue repair in the maternal host. However, the use of similar fetal stem cells in therapy has been significantly hampered by the availability of clinically relevant cell numbers and/or contamination with cells of maternal origin, particularly when using the chorionic and decidual placenta. In the present prospective review, we highlight the importance of FMC to the field o...
Bone metastasis is a frequent and life-threatening complication of breast cancer. The molecular m... more Bone metastasis is a frequent and life-threatening complication of breast cancer. The molecular mechanisms supporting the establishment of breast cancer cells in the skeleton are still not fully understood, which may be attributed to the lack of suitable models that interrogate interactions between human breast cancer cells and the bone microenvironment. Although it is well-known that integrins mediate adhesion of malignant cells to bone extracellular matrix, their role during bone colonization remains unclear. Here, the role of β1 integrins in bone colonization was investigated using tissue-engineered humanized in vitro and in vivo bone models. In vitro, bone-metastatic breast cancer cells with suppressed integrin β1 expression showed reduced attachment, spreading, and migration within human bone matrix compared to control cells. Cell proliferation in vitro was not affected by β1 integrin knockdown, yet tumor growth in vivo within humanized bone microenvironments was significantly ...
On 31st March 2003 Advanced Tissue Sciences (ATS) was liquidated, with the effect that in excess ... more On 31st March 2003 Advanced Tissue Sciences (ATS) was liquidated, with the effect that in excess of US$300 million of stakeholder financing was destroyed. Although successful in the development of breakthrough technologies in the regenerative medicine arena and the building of a substantial portfolio of patents, the company never made a profit. In this case study, ATS’ business strategy, market and competitive environment will be discussed in the context of the company’s historical development. A number of important lessons from this case are discussed. From a management perspective the most critical lesson is the importance of effective financial planning and management of costs, and in particular R&D costs, including the significant costs associated with clinical trials. In addition, a clear strategic focus is extremely important due to the significant resources required in the development of a new therapy. From an investor’s perspective the lessons to be gath...
Materials science & engineering. C, Materials for biological applications, 2013
We have compared the effects of different sterilization techniques on the properties of Bombyx mo... more We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the var...
The treatment of challenging fractures and large osseous defects presents a formidable problem fo... more The treatment of challenging fractures and large osseous defects presents a formidable problem for orthopaedic surgeons. Tissue engineering/regenerative medicine approaches seek to solve this problem by delivering osteogenic signals within scaffolding biomaterials. In this study, we introduce a hybrid growth factor delivery system that consists of an electrospun nanofiber mesh tube for guiding bone regeneration combined with peptide-modified alginate hydrogel injected inside the tube for sustained growth factor release. We tested the ability of this system to deliver recombinant bone morphogenetic protein-2 (rhBMP-2) for the repair of critically-sized segmental bone defects in a rat model. Longitudinal μ-CT analysis and torsional testing provided quantitative assessment of bone regeneration. Our results indicate that the hybrid delivery system resulted in consistent bony bridging of the challenging bone defects. However, in the absence of rhBMP-2, the use of nanofiber mesh tube and ...
Journal of investigative and clinical dentistry, Jan 22, 2014
The aim of the present study was to compare bacterial load using preoperative rinsing and swabbin... more The aim of the present study was to compare bacterial load using preoperative rinsing and swabbing techniques for oral surgery with 0.2% chlorhexidine (CHX). Participants were healthy volunteers undergoing a general anesthetic for the removal of teeth. Participants were randomly allocated to receive 15 mL of 0.2% CHX for 60 s as either a rinse or have their mouths swabbed. Plaque samples were aseptically collected pre- and post-rinsing from the same sites in all patients (the distal surface of all second molar teeth). Patients in the swab group had similar bacteria counts before and after the application of CHX (143.4 vs 138.5 colonies, P = 1.000). After rinsing with CHX, there was an eightfold reduction of bacterial load (71 vs 8.8 colonies, P < 0.001). The present study demonstrates that the use of CHX as an antimicrobial agent is effective in reducing the overall number of bacterial colonies in the oral cavity. Rinsing is a more effective method of doing this.
Mesenchymal stem cells derived from human bone marrow (hBMSCs) and human adipose tissue (hAMSCs) ... more Mesenchymal stem cells derived from human bone marrow (hBMSCs) and human adipose tissue (hAMSCs) represent a useful source of progenitor cells for cell therapy and tissue engineering. However, it is not clear what the similarities and differences between them are. Like hBMSCs, hAMSCs can differentiate into osteogenic, adipogenic, and chondrogenic cells. Whether MSCs derived from different tissue sources represent fundamentally similar or different cell types is not clear. Given the possible different sources of MSCs for cell therapy, a comprehensive comparison of the different MSCs would be very useful. Here, we compared the transcriptome profile of hAMCS and hBMSCs during directed differentiation into bone, cartilage, and fat. Our data revealed considerable similarities between bone marrow-derived MSCs (BMSCs) and adipose tissue-derived MSCs (AMSCs). We uncovered an interesting bifurcation of pathways in both BMSCs and AMSCs, in which osteogenesis and adipogenesis appear to be link...
High renewal and maintenance of multipotency of human adult stem cells (hSCs), are a prerequisite... more High renewal and maintenance of multipotency of human adult stem cells (hSCs), are a prerequisite for experimental analysis as well as for potential clinical usages. The most widely used strategy for hSC culture and proliferation is using serum. However, serum is poorly defined and has a considerable degree of inter-batch variation, which makes it difficult for large-scale mesenchymal stem cells (MSCs) expansion in homogeneous culture conditions. Moreover, it is often observed that cells grown in serum-containing media spontaneously differentiate into unknown and/or undesired phenotypes. Another way of maintaining hSC development is using cytokines and/or tissue-specific growth factors; this is a very expensive approach and can lead to early unwanted differentiation. In order to circumvent these issues, we investigated the role of sphingosine-1-phosphate (S1P), in the growth and multipotency maintenance of human bone marrow and adipose tissue-derived MSCs. We show that S1P induces g...
Earlier studies have shown that the influence of fixation stability on bone healing diminishes wi... more Earlier studies have shown that the influence of fixation stability on bone healing diminishes with advanced age. The goal of this study was to unravel the relationship between mechanical stimulus and age on callus competence at a tissue level. Using 3D in vitro micro-computed tomography derived metrics, 2D in vivo radiography, and histology, we investigated the influences of age and varying fixation stability on callus size, geometry, microstructure, composition, remodeling, and vascularity. Compared were four groups with a 1.5-mm osteotomy gap in the femora of Sprague-Dawley rats: Young rigid (YR), Young semirigid (YSR), Old rigid (OR), Old semirigid (OSR). Hypothesis was that calcified callus microstructure and composition is impaired due to the influence of advanced age, and these individuals would show a reduced response to fixation stabilities. Semirigid fixations resulted in a larger DeltaCSA (Callus cross-sectional area) compared to rigid groups. In vitro microCT analysis at 6 weeks postmortem showed callus bridging scores in younger animals to be superior than their older counterparts (p<0.01). Younger animals showed (i) larger callus strut thickness (p<0.001), (ii) lower perforation in struts (p<0.01), and (iii) higher mineralization of callus struts (p<0.001). Callus mineralization was reduced in young animals with semirigid fracture fixation but remained unaffected in the aged group. While stability had an influence, age showed none on callus size and geometry of callus. With no differences observed in relative osteoid areas in the callus ROI, old as well as semirigid fixated animals showed a higher osteoclast count (p<0.05). Blood vessel density was reduced in animals with semirigid fixation (p<0.05). In conclusion, in vivo monitoring indicated delayed callus maturation in aged individuals. Callus bridging and callus competence (microstructure and mineralization) were impaired in individuals with an advanced age. This matched with increased bone resorption due to higher osteoclast numbers. Varying fixator configurations in older individuals did not alter the dominant effect of advanced age on callus tissue mineralization, unlike in their younger counterparts. Age-associated influences appeared independent from stability. This study illustrates the dominating role of osteoclastic activity in age-related impaired healing, while demonstrating the optimization of fixation parameters such as stiffness appeared to be less effective in influencing healing in aged individuals.Copyright 2010 Elsevier Inc. All rights reserved.
Ample evidence supports that prostate tumor metastasis originates from a rare population of cance... more Ample evidence supports that prostate tumor metastasis originates from a rare population of cancer cells, known as cancer stem cells (CSCs). Unfortunately, little is known about the identity of these cells, making it difficult to target the metastatic prostate tumor. Here, for the first time, we report the identification of a rare population of prostate cancer cells that express the Tie-2 protein. We found that this Tie-2High population exists mainly in prostate cancer cell lines that are capable of metastasizing to the bone. These cells not only express a higher level of CSC markers but also demonstrate enhanced resistance to the chemotherapeutic drug Cabazitaxel. In addition, knockdown of the expression of the Tie-2 ligand angiopoietin (Ang-1) led to suppression of CSC markers, suggesting that the Ang-1/Tie-2 signaling pathway functions as an autocrine loop for the maintenance of prostate CSCs. More importantly, we found that Tie-2High prostate cancer cells are more adhesive than ...
The International journal of oral & maxillofacial implants
This paper reports on a 2-phase study of a novel membrane-scaffold graft construct, its ability t... more This paper reports on a 2-phase study of a novel membrane-scaffold graft construct, its ability to support periodontal ligament fibroblast (PDLF) and alveolar osteoblast (AO) growth in vitro, and its use for tissue engineering a PDL-AO interface in vivo. Human PDLFs were seeded onto perforated poly(epsilon-caprolactone) membranes (n=30) at 78,000 cells/cm2; human AOs were seeded on poly(epsilon-caprolactone) scaffolds (n=30) with fibrin glue at 625,000 cells/cm3. Cell attachment, morphology, viability, and metabolic activity were monitored for 3 weeks in vitro. Subsequently, cell-seeded membrane-scaffold constructs (experimental group, n=9) and nonseeded constructs (control group, n=4) assembled with fibrin glue were implanted subcutaneously into 7 athymic mice for 4 weeks. PDLFs formed confluent layers on membranes, whereas AOs produced mineralized matrices within scaffolds upon osteoinduction in vitro. Well-vascularized tissue formation was observed after implantation. Integration...
Tumour microenvironment greatly influences the development and metastasis of cancer progression. ... more Tumour microenvironment greatly influences the development and metastasis of cancer progression. The development of three dimensional (3D) culture models which mimic that displayed in vivo can improve cancer biology studies and accelerate novel anticancer drug screening. Inspired by a systems biology approach, we have formed 3D in vitro bioengineered tumour angiogenesis microenvironments within a glycosaminoglycan-based hydrogel culture system. This microenvironment model can routinely recreate breast and prostate tumour vascularisation. The multiple cell types cultured within this model were less sensitive to chemotherapy when compared with two dimensional (2D) cultures, and displayed comparative tumour regression to that displayed in vivo. These features highlight the use of our in vitro culture model as a complementary testing platform in conjunction with animal models, addressing key reduction and replacement goals of the future. We anticipate that this biomimetic model will provide a platform for the in-depth analysis of cancer development and the discovery of novel therapeutic targets.
SummarySince the isolation of fetal stem cell populations from perinatal tissues, such as umbilic... more SummarySince the isolation of fetal stem cell populations from perinatal tissues, such as umbilical cord blood and placenta, interest has been growing in understanding their greater plasticity compared with adult stem cells and exploring their potential in regenerative medicine. The phenomenon of fetal microchimerism (FMC) naturally occurring during pregnancy through the transfer of fetal stem/progenitor cells to maternal blood and tissues has been integral in developing this dogma. Specifically, microchimeric mesenchymal stem cells and endothelial progenitors of fetal origin have now demonstrated a capacity for tissue repair in the maternal host. However, the use of similar fetal stem cells in therapy has been significantly hampered by the availability of clinically relevant cell numbers and/or contamination with cells of maternal origin, particularly when using the chorionic and decidual placenta. In the present prospective review, we highlight the importance of FMC to the field o...
Bone metastasis is a frequent and life-threatening complication of breast cancer. The molecular m... more Bone metastasis is a frequent and life-threatening complication of breast cancer. The molecular mechanisms supporting the establishment of breast cancer cells in the skeleton are still not fully understood, which may be attributed to the lack of suitable models that interrogate interactions between human breast cancer cells and the bone microenvironment. Although it is well-known that integrins mediate adhesion of malignant cells to bone extracellular matrix, their role during bone colonization remains unclear. Here, the role of β1 integrins in bone colonization was investigated using tissue-engineered humanized in vitro and in vivo bone models. In vitro, bone-metastatic breast cancer cells with suppressed integrin β1 expression showed reduced attachment, spreading, and migration within human bone matrix compared to control cells. Cell proliferation in vitro was not affected by β1 integrin knockdown, yet tumor growth in vivo within humanized bone microenvironments was significantly ...
On 31st March 2003 Advanced Tissue Sciences (ATS) was liquidated, with the effect that in excess ... more On 31st March 2003 Advanced Tissue Sciences (ATS) was liquidated, with the effect that in excess of US$300 million of stakeholder financing was destroyed. Although successful in the development of breakthrough technologies in the regenerative medicine arena and the building of a substantial portfolio of patents, the company never made a profit. In this case study, ATS’ business strategy, market and competitive environment will be discussed in the context of the company’s historical development. A number of important lessons from this case are discussed. From a management perspective the most critical lesson is the importance of effective financial planning and management of costs, and in particular R&D costs, including the significant costs associated with clinical trials. In addition, a clear strategic focus is extremely important due to the significant resources required in the development of a new therapy. From an investor’s perspective the lessons to be gath...
Materials science & engineering. C, Materials for biological applications, 2013
We have compared the effects of different sterilization techniques on the properties of Bombyx mo... more We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the var...
The treatment of challenging fractures and large osseous defects presents a formidable problem fo... more The treatment of challenging fractures and large osseous defects presents a formidable problem for orthopaedic surgeons. Tissue engineering/regenerative medicine approaches seek to solve this problem by delivering osteogenic signals within scaffolding biomaterials. In this study, we introduce a hybrid growth factor delivery system that consists of an electrospun nanofiber mesh tube for guiding bone regeneration combined with peptide-modified alginate hydrogel injected inside the tube for sustained growth factor release. We tested the ability of this system to deliver recombinant bone morphogenetic protein-2 (rhBMP-2) for the repair of critically-sized segmental bone defects in a rat model. Longitudinal μ-CT analysis and torsional testing provided quantitative assessment of bone regeneration. Our results indicate that the hybrid delivery system resulted in consistent bony bridging of the challenging bone defects. However, in the absence of rhBMP-2, the use of nanofiber mesh tube and ...
Journal of investigative and clinical dentistry, Jan 22, 2014
The aim of the present study was to compare bacterial load using preoperative rinsing and swabbin... more The aim of the present study was to compare bacterial load using preoperative rinsing and swabbing techniques for oral surgery with 0.2% chlorhexidine (CHX). Participants were healthy volunteers undergoing a general anesthetic for the removal of teeth. Participants were randomly allocated to receive 15 mL of 0.2% CHX for 60 s as either a rinse or have their mouths swabbed. Plaque samples were aseptically collected pre- and post-rinsing from the same sites in all patients (the distal surface of all second molar teeth). Patients in the swab group had similar bacteria counts before and after the application of CHX (143.4 vs 138.5 colonies, P = 1.000). After rinsing with CHX, there was an eightfold reduction of bacterial load (71 vs 8.8 colonies, P < 0.001). The present study demonstrates that the use of CHX as an antimicrobial agent is effective in reducing the overall number of bacterial colonies in the oral cavity. Rinsing is a more effective method of doing this.
Mesenchymal stem cells derived from human bone marrow (hBMSCs) and human adipose tissue (hAMSCs) ... more Mesenchymal stem cells derived from human bone marrow (hBMSCs) and human adipose tissue (hAMSCs) represent a useful source of progenitor cells for cell therapy and tissue engineering. However, it is not clear what the similarities and differences between them are. Like hBMSCs, hAMSCs can differentiate into osteogenic, adipogenic, and chondrogenic cells. Whether MSCs derived from different tissue sources represent fundamentally similar or different cell types is not clear. Given the possible different sources of MSCs for cell therapy, a comprehensive comparison of the different MSCs would be very useful. Here, we compared the transcriptome profile of hAMCS and hBMSCs during directed differentiation into bone, cartilage, and fat. Our data revealed considerable similarities between bone marrow-derived MSCs (BMSCs) and adipose tissue-derived MSCs (AMSCs). We uncovered an interesting bifurcation of pathways in both BMSCs and AMSCs, in which osteogenesis and adipogenesis appear to be link...
High renewal and maintenance of multipotency of human adult stem cells (hSCs), are a prerequisite... more High renewal and maintenance of multipotency of human adult stem cells (hSCs), are a prerequisite for experimental analysis as well as for potential clinical usages. The most widely used strategy for hSC culture and proliferation is using serum. However, serum is poorly defined and has a considerable degree of inter-batch variation, which makes it difficult for large-scale mesenchymal stem cells (MSCs) expansion in homogeneous culture conditions. Moreover, it is often observed that cells grown in serum-containing media spontaneously differentiate into unknown and/or undesired phenotypes. Another way of maintaining hSC development is using cytokines and/or tissue-specific growth factors; this is a very expensive approach and can lead to early unwanted differentiation. In order to circumvent these issues, we investigated the role of sphingosine-1-phosphate (S1P), in the growth and multipotency maintenance of human bone marrow and adipose tissue-derived MSCs. We show that S1P induces g...
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