Laurent Bozec

Collagen — Emerging collagen based therapies hit the patient

The choice of biomaterials available for regenerative medicine continues to grow rapidly, with new materials often claiming advantages over the short-comings of those already in existence. Going back to nature, collagen is one of the most abundant proteins in mammals and its role is essential to our way of life. It can therefore be obtained from many sources including porcine, bovine, equine or human and offer a great promise as a biomimetic scaffold for regenerative medicine. Using naturally derived collagen, extracellular matrices (ECMs), as surgical materials have become established practice for a number of years. For clinical use the goal has been to preserve as much of the composition and structure of the ECM as possible without adverse effects to the recipient. This review will therefore cover in-depth both naturally and synthetically produced collagen matrices. Furthermore the production of more sophisticated three dimensional collagen scaffolds that provide cues at nano-, micro- and meso-scale for molecules, cells, proteins and bulk fluids by inducing fibrils alignments, embossing and layered configuration through the application of plastic compression technology will be discussed in details. This review will also shed light on both naturally and synthetically derived collagen products that have been available in the market for several purposes including neural repair, as cosmetic for the treatment of dermatologic defects, haemostatic agents, mucosal wound dressing and guided bone regeneration membrane. There are other several potential applications of collagen still under investigations and they are also covered in this review.

Demineralization–remineralization dynamics in teeth and bone

Biomineralization is a dynamic, complex, lifelong process by which living organisms control precipitations of inorganic nanocrystals within organic matrices to form unique hybrid biological tissues, for example, enamel, dentin, cementum, and bone. Understanding the process of mineral deposition is important for the development of treatments for mineralization-related diseases and also for the innovation and development of scaffolds. This review provides a thorough overview of the up-to-date information on the theories describing the possible mechanisms and the factors implicated as agonists and antagonists of mineralization. Then, the role of calcium and phosphate ions in the maintenance of teeth and bone health is described. Throughout the life, teeth and bone are at risk of demineralization, with particular emphasis on teeth, due to their anatomical arrangement and location. Teeth are exposed to food, drink, and the microbiota of the mouth; therefore, they have developed a high resistance to localized demineralization that is unmatched by bone. The mechanisms by which demineralization–remineralization process occurs in both teeth and bone and the new therapies/technologies that reverse demineralization or boost remineralization are also scrupulously discussed. Technologies discussed include composites with nano- and micron-sized inorganic minerals that can mimic mechanical properties of the tooth and bone in addition to promoting more natural repair of surrounding tissues. Turning these new technologies to products and practices would improve health care worldwide.

Single-bacterium nanomechanics in biomedicine: unravelling the dynamics of bacterial cells

The use of the atomic force microscope (AFM) in microbiology has progressed significantly throughout the years since its first application as a high-resolution imaging instrument. Modern AFM setups are capable of characterizing the nanomechanical behaviour of bacterial cells at both the cellular and molecular levels, where elastic properties and adhesion forces of single bacterium cells can be examined under different experimental conditions. Considering that bacterial and biofilm-mediated infections continue to challenge the biomedical field, it is important to understand the biophysical events leading towards bacterial adhesion and colonization on both biological and non-biological substrates. The purpose of this review is to present the latest findings concerning the field of single-bacterium nanomechanics, and discuss future trends and applications of nanoindentation and single-cell force spectroscopy techniques in biomedicine.

Coherent X-ray diffraction from collagenous soft tissues

Coherent X-ray diffraction has been applied in the imaging of inorganic materials with great success. However, its application to biological specimens has been limited to some notable exceptions, due to the induced radiation damage and the extended nature of biological samples, the last limiting the application of most part of the phasing algorithms. X-ray ptychography, still under development, is a good candidate to overcome such difficulties and become a powerful imaging method for biology. We describe herein the feasibility of applying ptychography to the imaging of biological specimens, in particular collagen rich samples. We report here speckles in diffraction patterns from soft animal tissue, obtained with an optimized small angle X-ray setup that exploits the natural coherence of the beam. By phasing these patterns, dark field images of collagen within tendon, skin, bone, or cornea will eventually be obtained with a resolution of 60–70 nm. We present simulations of the contrast mechanism in collagen based on atomic force microscope images of the samples. Simulations confirmed the ‘speckled’ nature of the obtained diffraction patterns. Once inverted, the patterns will show the disposition and orientation of the fibers within the tissue, by enhancing the phase contrast between protein and no protein regions of the sample. Our work affords the application of the most innovative coherent X-ray diffraction tools to the study of biological specimens, and this approach will have a significant impact in biology and medicine because it overcomes many of the limits of current microscopy techniques.

Thermal analysis of model and historic tapestries

SummaryDynamic mechanical thermal analysis (DMTA), differential scanning calorimetry (DSC) and thermogravimetry (TG) have been used to characterize model tapestries, especially woven for the EC-funded project (MODHT) and to historic tapestries in royal palaces and museums. Modulus values of woollen threads from model tapestries are reported and the effects of traditional dyeing and mordanting processes quantified. TG, particularly of black woollen threads showed alterations in thermal stability. Tests of creep on immersion in water and subsequent heating to 90°C and on exposure to a controlled relative humidity programme were also used to rank these effects. Modulus values of historic woollen samples were also obtained and DSC studies of model and historic silk samples are reported together with preliminary atomic force microscopy (AFM) images of silk fibres.

Nanotechnology in dentistry: prevention, diagnosis, and therapy

Nanotechnology has rapidly expanded into all areas of science; it offers significant alternative ways to solve scientific and medical questions and problems. In dentistry, nanotechnology has been exploited in the development of restorative materials with some significant success. This review discusses nanointerfaces that could compromise the longevity of dental restorations, and how nanotechnolgy has been employed to modify them for providing long-term successful restorations. It also focuses on some challenging areas in dentistry, eg, oral biofilm and cancers, and how nanotechnology overcomes these challenges. The recent advances in nanodentistry and innovations in oral health-related diagnostic, preventive, and therapeutic methods required to maintain and obtain perfect oral health, have been discussed. The recent advances in nanotechnology could hold promise in bringing a paradigm shift in dental field. Although there are numerous complex therapies being developed to treat many diseases, their clinical use requires careful consideration of the expense of synthesis and implementation.

Regulation of post-Golgi LH3 trafficking is essential for collagen homeostasis

Post-translational modifications are necessary for collagen precursor molecules (procollagens) to acquire final shape and function. However, the mechanism and contribution of collagen modifications that occur outside the endoplasmic reticulum and Golgi are not understood. We discovered that VIPAR, with its partner proteins, regulate sorting of lysyl hydroxylase 3 (LH3, also known as PLOD3) into newly identified post-Golgi collagen IV carriers and that VIPAR-dependent sorting is essential for modification of lysines in multiple collagen types. Identification of structural and functional collagen abnormalities in cells and tissues from patients and murine models of the autosomal recessive multisystem disorder Arthrogryposis, Renal dysfunction and Cholestasis syndrome caused by VIPAR and VPS33B deficiencies confirmed our findings. Thus, regulation of post-Golgi LH3 trafficking is essential for collagen homeostasis and for the development and function of multiple organs and tissues.

TiO2-coated CoCrMo: Improving the osteogenic differentiation and adhesion of mesenchymal stem cells in vitro

The current gold standard material for orthopedic applications is titanium (Ti), however, other materials such as cobalt-chromium-molybdenum (CoCrMo) are often preferred due to their wear resistance and mechanical strength. This study investigates if the bioactivity of CoCrMo can be enhanced by coating the surface with titanium oxide (TiO2 ) by atmospheric pressure chemical vapor deposition (CVD), thereby replicating the surface oxide layer found on Ti. CoCrMo, TiO2-coated CoCrMo (CCMT) and Ti substrates were used for this study. Cellular f-actin distribution was shown to be noticeably different between cells on CCMT and CoCrMo after 24 h in osteogenic culture, with cells on CCMT exhibiting greater spread with developed protrusions. Osteogenic differentiation was shown to be enhanced on CCMT compared to CoCrMo, with increased calcium ion content per cell (p < 0.05), greater hydroxyapatite nodule formation (p < 0.05) and reduced type I collagen deposition per cell (p < 0.05). The expression of the focal adhesion protein vinculin was shown to be marginally greater on CCMT compared to CoCrMo, whereas AFM results indicated that CCMT required more force to remove a single cell from the substrate surface compared to CoCrMo (p < 0.0001). These data suggest that CVD TiO2 coatings may have the potential to increase the biocompatibility of CoCrMo implantable devices.

Development of a surgically optimized graft insertion suture technique to accommodate a tissue-engineered tendon in vivo.

Abstract The traumatic rupture of tendons is a common clinical problem. Tendon repair is surgically challenging because the tendon often retracts, resulting in a gap between the torn end and its bony insertion. Tendon grafts are currently used to fill this deficit but are associated with potential complications relating to donor site morbidity and graft necrosis. We have developed a highly reproducible, rapid process technique to manufacture compressed cell-seeded type I collagen constructs to replace tendon grafts. However, the material properties of the engineered constructs are currently unsuitable to withstand complete load bearing in vivo. A modified suture technique has been developed to withstand physiological loading and off load the artificial construct while integration occurs. Lapine tendons were used ex vivo to test the strength of different suture techniques with different sizes of Prolene sutures and tissue-engineered collagen constructs in situ. The data were compared to standard modified Kessler suture using a standard tendon graft. Mechanical testing was carried out and a finite element analysis stress distribution model constructed using COMSOL 3.5 software. The break point for modified suture technique with a tissue-engineered scaffold was significantly higher (50.62 N) compared to a standard modified Kessler suture (12.49 N, p<0.05). Distributing suture tension further proximally and distally from the tendon ends increased the mechanical strength of the repairs. We now have ex vivo proof of concept that this suture technique is suitable for testing in vivo, and this will be the next stage of our research.

Spatial and temporal evaluation of cell attachment to printed polycaprolactone microfibres

Surface topography plays a crucial role in influencing cellular responses and has therefore been utilized in the development of numerous implantable devices. Whilst numerous studies have either investigated cell attachment or migration post-attachment, few have looked at the early-stages of this process temporally. The aim of this study was to evaluate the use of time-lapse microscopy to study the behaviour of fibroblasts cultured with polycaprolactone microfibres and to assess spatially and temporally the cell-structure interaction over a 24h period. Ordered polymeric structures were printed (predetermined) onto glass substrates using an electrohydrodynamic direct write process to produce fine (3-5 μm wide) structures. Fibroblast attachment and migration were characterized as a function of distance perpendicular from structures (∼17.3, 34.6 and 51.9 μm). The use of time-lapse microscopy revealed a gradual decrease in cell attachment as the distance from the microfibres was increased. The technique also revealed that some cells were attaching and detaching from the microfibre multiple times. Our findings demonstrate that time-lapse microscopy is a useful technique for evaluating early-stage cell-biomaterial interaction that is capable of recording important events that might otherwise be overlooked.