Rishma Shah

Northcroft Memorial Lecture 2005: Muscling in on malocclusions: Current concepts on the role of muscles in the aetiology and treatment of malocclusion

The British Orthodontic Society invites outstanding contributors from the field of Orthodontics to give the guest lecture in memory of George Northcroft. In 2005 the guest lecturer was Professor Nigel Hunt. The article that follows was presented as the Northcroft Memorial Lecture 2005 at the World Orthodontic Congress, Paris.

Development of a novel smart scaffold for human skeletal muscle regeneration.

Skeletal muscle defects are notoriously difficult to manage and the current methods used are associated with many limitations. Engineered skeletal muscle tissue has the potential to provide a solution that circumvents these disadvantages. Our previous work has identified a novel three‐dimensionally aligned degradable phosphate glass fibre scaffold that can support myoblast differentiation and maturation. This current study has further developed the scaffold by encasing the fibres within a collagen gel to produce a smart composite scaffold that provides key biomimetic cues and supports the formation of a tissue that may be implanted in vivo. The constructs formed were approximately 30 mm long and microscopic examination confirmed favourable unidirectional cell alignment. There was good cell survival, and gene expression studies demonstrated upregulation of the myogenic regulatory factors and developmental and adult myosin heavy chain isoforms indicating myofibre formation and maturation respectively. Compared with the three‐dimensional glass fibre scaffolds, the composite scaffolds had later gene upregulation, however, the use of collagen gels reinforced with degradable aligned glass fibres offers the opportunity to create a tissue analogue that can be easily manipulated. Furthermore, the glass fibre ends could support tendon/bone formation, and the channels formed as the fibres degrade could allow for vascular ingrowth. Copyright

Implementation of the virtual learning environment into a UK orthodontic training programme: the postgraduate and lecturer perspective

The virtual learning environment (VLE) was formally introduced into the orthodontic postgraduate teaching programme at the UCL Eastman Dental Institute in October 2006 with the main role to support the existing didactic teaching. The aim of this study was to establish the programme co-ordinator, lecturer and postgraduate perspective on the introduction of the VLE as an adjunct to other teaching methods. All participants were interviewed utilising open-ended questions to ascertain their response. The interviews were semi-structured and were continued until no new information was elicited. The interviews suggested that the programme co-ordinator, lecturers and the majority of the postgraduates were confident that the VLE could produce a better learning experience. The VLE has a number of advantages and disadvantages, however, there is great potential for the VLE to encourage a constructivist approach to teaching and learning. Moves have been made to align aspects, such as assessment, with the VLE and the rest of the curriculum.

Strontium- and calcium-containing, titanium-stabilised phosphate-based glasses with prolonged degradation for orthopaedic tissue engineering

Strontium- and calcium-releasing, titanium-stabilised phosphate-based glasses with a controlled degradation rate are currently under development for orthopaedic tissue engineering applications. Ca and/or Sr were incorporated at varying concentrations in quaternary phosphate-based glasses, in order to promote osteoinduction. Ti was incorporated at a fixed concentration in order to prolong degradation. Glasses of the general formula (P2O5)–(Na2O)–(TiO2)–(CaO)–(SrO) were prepared via the melt-quench technique. The materials were characterised by energy-dispersive X-ray spectroscopy, X-ray diffraction, 31P magic angle spinning nuclear magnetic resonance, Fourier transform infrared spectroscopy, differential thermal analysis and density determination. The dissolution rate in distilled water was determined by measuring mass loss, ion release and pH change over a two-week period. In addition, the cytocompatibility and alkaline phosphatase activity of an osteoblast-like cell line cultured on the surface of glass discs was assessed. The glasses were shown to be amorphous and contained Q1, Q2 and Q3 species. Fourier transform infrared spectroscopy revealed small changes in the glass structure as Ca was substituted with Sr and differential thermal analysis confirmed a decrease in crystallisation temperature with increasing Sr content. Degradation and ion release studies also showed that mass loss was positively correlated with Sr content. These results were attributed to the lower electronegativity of Sr in comparison to Ca favouring the formation of phosphate-based mineral phases. All compositions supported cell proliferation and survival and induced at least 2.3-fold alkaline phosphatase activity relative to the control. Glass containing 17.5 mol% Sr had 3.6-fold greater alkaline phosphatase activity than the control. The gradual release of Ca and Sr supported osteoinduction, indicating their potential suitability in orthopaedic tissue engineering applications.

Small molecule stimulation enhances bone regeneration but not titanium implant osseointegration.

The osteogenic and osseointegrative potential of a small molecule was examined to assess its usefulness in regenerative procedures. Purmorphamine was used to stimulate bone growth and repair in an in vitro cell-based assay and an in vivo chick embryo CAM-assay with and without the presence of an implant. Purmorphamine adhered to precipitated hydroxyapatite coating, could activate the sonic hedgehog pathway and thereby stimulated osteodifferentiation. Porous calcium phosphate beads were used to deliver this small molecule in vivo and showed that purmorphamine increased the trabecular bone to bone area significantly. The assay showed purmorphamine failed to induce any significant difference in osseointegration on titanium coated PTFE implants. This suggests that, while a small molecule can enhance osteogenesis and might be useful in regenerative procedures, it failed to enhance the osseointegration of a Ti coated implant, suggesting that this sort of stimulation might be useful for enhancing bone regeneration where bone loss due to disease exists, but not for enhancing early stability of an implant.

Myosin proteins identified from masseter muscle using quantitative reverse transcriptase–polymerase chain reaction—a pilot study of the relevance to orthodontics

There is a clearly established relationship between masticatory muscle structure and facial form. Human studies in this area, however, have been limited, especially in consideration of the myosin heavy chain (MyHC) family of contractile proteins. The aim of this pilot study was to assess if differences were detectable between genotype with respect to MyHC isoforms and the vertical facial phenotype in a sample of nine Caucasian female patients, age range 18-49 years, using a novel rapid technique. Masseter muscle biopsies were taken from patients with a range of vertical facial form. The levels of expression of the MyHC isoform genes MYH 1, 2, 3, 6, 7, and 8 were compared with the expression in a female calibrator patient aged 23 years with normal vertical facial form, using quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Statistical analysis was undertaken using Pearson correlation coefficient. The results showed that there were distinct differences in gene expression between patients with a wide range of variation although changes in MYH1 were consistent with one cephalometric variable, the maxillo-mandibular angle. The full procedure, from start to finish, can be completed within half a day. Rapid genotyping of patients in this way could reveal important information of relevance to treatment. This technology has potential as a diagnostic and prognostic aid when considering correction of certain malocclusions.

Sequential identification of a degradable phosphate glass scaffold for skeletal muscle regeneration

Tissue engineering has the potential to overcome limitations associated with current management of skeletal muscle defects. This study aimed to sequentially identify a degradable phosphate glass scaffold for the restoration of muscle defects. A series of glass compositions were investigated for the potential to promote bacterial growth. Thereafter, the response of human craniofacial muscle‐derived cells was determined. Glass compositions containing Fe4‐ and 5 mol% did not promote greater Staphylococcus aureus and Staphylococcus epidermidis growth compared to the control (p > 0.05). Following confirmation of myogenicity, further studies assessed the biocompatibility of glasses containing Fe5 mol%. Cells seeded on collagen‐coated disks demonstrated comparable cellular metabolic activity to control. Upregulation of genes encoding for myogenic regulatory factors (MRFs) confirmed myofibre formation and there was expression of developmental MYH genes. The use of 3‐D aligned fibre scaffolds supported unidirectional cell alignment and upregulation of MRF and developmental MYH genes. Compared to the 2‐D disks, there was also expression of MYH2 and MYH7 genes, indicating further myofibre maturation on the 3‐D scaffolds and confirming the importance of key biophysical cues. Copyright

Degradation of zinc containing phosphate-based glass as a material for orthopedic tissue engineering

Phosphate-based glasses have been examined in many studies as a potential biomaterial for bone repair because of its degradation properties, which can be controlled and allow the release of various elements to promote osteogenic tissue growth. However most of these experiments studied either tertiary or quaternary glass systems. This study investigated a qinternary system that included titanium dioxide for degradation rate control and zinc that is considered to have a role in bone formation. Zinc and titanium phosphate glass discs of different compositions were melt synthesized and samples of each composition was tested for different physical, chemical and biological characteristics via density measurement, X-ray diffraction, differential thermal analysis, mass loss, ion release, scanning electron microscopy, biocompatibility studies via live/dead assays at three time points (day 1, 4, and 7). The results showed that the glass was amorphous and that the all thermal variables decreased as zinc oxide amount raised, mass loss as well as ion release increased as zinc oxide increased, and the maximum rise was with ZnO15. The cellular studies showed that all the formulation showed similar cytocompatibility properties with MG63 except ZnO15, which displayed cytotoxic properties and this was confirmed also by the scanning electron microscope images. In conclusion, replacing calcium oxide with zinc oxide in proportion less than 10 % can have a positive effect on bone forming cells.

Muscle tissue engineering

Skeletal muscle is a classical example of “structure determining function.” Successful strategies for clinical applications of tissue-engineered skeletal muscle must recapitulate the processes that muscle undergoes during either embryonic development or adult regeneration. As discussed in this chapter, current approaches to tissue engineering of skeletal muscle broadly utilize different aspects of these processes, e.g. the generation of multinucleate, elongated myotubes (nascent myofibres) surrounded by a connective tissue sheath. The former are generated by encouraging muscle precursor cell fusion, in which alignment is vital, so that surface topography, fibre scaffolds and mechanical loading are key processes. Connective tissue sheaths can be generated by encasing constructs in materials such as collagen and fibrin. Finally, functional connections need to be made to the skeletal system via tendons, to the neural system via neuromuscular junctions and to the vascular system. The chapter that follows reviews the current situation as regards these developments.

Masticatory Muscle Structure and Function

The muscle group referred to as the muscles of mastication includes the temporalis, the medial and lateral pterygoid, and the masseter muscles on both sides of the face and jaws. These voluntary skeletal muscles are derived from the paraxial mesoderm of the first branchial arch whilst their connective tissue components are derived from mesenchymal cells of neural crest origin. They are innervated by the mandibular division of the fifth (trigeminal) cranial nerve (CNV). Precision in the control of jaw position and movement of the mandible is provided by the human mandibular locomotor system. It is important to remember that this control of both jaw position and function varies considerably throughout the life of an individual, principally to support the activities of providing nutrition, speaking, and swallowing. For example, in the newborn infant very fine movements of the jaw are associated with the important tongue activity necessary for breast or bottle-feeding. Subsequently, as the deciduous teeth erupt and later as the permanent teeth erupt, these changes in dentition are associated with periods of rapid growth of the individual in general, and therefore the need to create increased biting and chewing forces is developed. There may be altered demands in relation to the ravages of such conditions as dental caries and periodontal disease with the possibility that teeth may need to be extracted or lost leading to prosthetic replacement. The different embryological origin and innervation of these muscles compared to somatic skeletal muscle coupled with their changing and unique functions have led to the possibility of structural specialisation within the muscles of mastication. Of the muscles in this group, the masseter muscle has undergone most investigation particularly in relation to its ease of access compared to the other muscles of the group.