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Osteomyelitis is a chronic inflammatory bone disease, which can present management difficulties owing to its complex pathogenic process and increasingly virulent micro-organisms. The incidence of the condition has decreased in recent years, primarily due to the availability of antimicrobial therapy; however, this alone does not always resolve symptoms. Once established osteomyelitis can be challenging to manage. This case demonstrates that in certain instances a more aggressive form of multidisciplinary treatment is required to improve the condition and alleviate the patient's symptoms, allowing for a better quality of life. CPD/Clinical Relevance: Knowledge of osteomyelitis is relevant to both general dental practitioners and specialists involved in the management of patients with complex medical needs.

Introduction: This paper reports on the outcomes from an analysis of 500 noncompliant regulations drawn from 1563 Early Years Services inspections that took place in Ireland between January and Dec...

Osteonecrosis of the jaws (ONJ) can be a serious complication of radiation and anti-resorptive therapies. At present, patients due to undergo radiotherapy are required to have a dental assessment to treat current or anticipated dental disease prior to commencing treatment. However no agreed guidelines are in place for those due to start anti-resorptive therapy for cancer. More patients are now being treated with various drug therapies and new combinations of drugs with unknown long-term effects, with many patients now surviving their disease for longer periods of time. It is important to increase awareness among patients and dental professionals of the possibility of them developing medication-related osteonecrosis of the jaw (MRONJ) and to put in place effective preventative measures to help to stop such complications developing. Case Report A 52-year-old patient with a complex medical history, including metastatic breast cancer, presented with generalised oral pain and tooth mobility (see Figure 1). The patient developed extensive stage III MRONJ following concurrent use of multiple anti-resorptive therapies for treatment of secondary bone disease. Dental treatment consisted of provision of antibiotics, hydrogen peroxide and chlorhexidine mouthwashes prior to the extraction of symptomatic teeth and the multiple bony sequestra. Conclusion Once established, stage III MRONJ can be challenging to manage for both the dental team and for the patient. This case demonstrates the importance of a comprehensive dental examination and need for preventative dental treatment prior to starting anti-resorptive therapy in order to prevent such instances.

Pseudoexfoliation syndrome (PXF) is the most common cause of secondary open angle glaucoma worldwide. Single nucleotide polymorphisms (SNPs) in the gene Lysyl oxidase like 1 (LOXL1) are strongly associated with the development of pseudoexfoliation glaucoma (PXFG). However, these SNPs are also present in 50-80% of the general population, suggestive of other factors being involved in the pathogenesis of PXFG. In this study, we aimed to investigate the influence of epigenetic regulation, specifically DNA methylation, on LOXL1 expression in PXFG using human tenons fibroblasts (HTFs), aqueous humour and serum samples from donors with and without PXFG. LOXL1 expression in HTFs was measured by qPCR and Western Blotting and LOXL1 concentration in aqueous humour was determined by ELISA. Global DNA methylation levels were quantified using an ELISA for 5-methylcytosine. MeDIP assays assessed the methylation status of the LOXL1 promoter region. Expression of methylation-associated enzymes (DNMT1, DNMT3a and MeCP2) were determined by qPCR and inhibited by 0.3 μM 5-azacytidine (5-aza). Results showed that LOXL1 expression was significantly decreased in PXFG HTFs compared with Control HTFs at gene (Fold change 0.37 ± 0.05, P 0.01) level and showed a decrease, when measured at the protein level (Fold change 0.65 ± 0.42, P = 0.22), however this was not found to be significant. LOXL1 concentration was increased in the aqueous of PXFG patients compared with Controls (2.76 ± 0.78 vs. 1.79 ± 0.33 ng/ml, P 0.01). Increased global methylation (56.07% ± 4.87% vs. 32.39% ± 4.29%, P 0.01) was observed in PXFG HTFs compared with Control HTFs, as was expression of methylation-associated enzymes (DNMT1 1.58 ± 0.30, P 0.05, DNMT3a 1.89 ± 0.24, P 0.05, MeCP2 1.63 ± 0.30, P 0.01). Methylation-associated enzymes were also increased when measured at protein level (DNMT1 5.70 ± 2.64, P = 0.04, DNMT3a 1.79 ± 1.55, P = 0.42, MeCP2 1.64 ± 1.33, P = 0.45). LOXL1 promoter methylation was increased in patients with PXFG compared to Control patients in both blood (3.98 ± 2.24, 2.10 ± 1.29, P 0.05) and HTF cells (37.31 ± 22.0, 8.66 ± 10.40, P 0.01). Treatment of PXFG HTFs with in 5-azacytidine increased LOXL1 expression when compared with untreated PXFG HTFs (Fold change 2.26 ± 0.67, P 0.05). These data demonstrate that LOXL1 expression is altered in PXFG via DNA methylation and that reversal of these epigenetic changes may represent future potential therapeutic targets in the management of PXFG.

Elevated intraocular pressure (IOP) narrows Schlemm’s canal (SC), theoretically increasing luminal shear stress. Using engineered adenoviruses containing a functional fragment of the shear-responsive endothelial nitric oxide synthase (eNOS) promoter, we tested effects of shear stress and elevated flow rate on reporter expression in vitro and ex vivo. Cultured human umbilical vein endothelial cells (HUVECs) and SC cells were transduced with adenovirus containing eNOS promoter driving secreted alkaline phosphatase (SEAP) or green fluorescent protein (GFP) and subjected to shear stress. In parallel, human anterior segments were perfused under controlled flow. After delivering adenoviruses to the SC lumen by retroperfusion, the flow rate in one anterior segment of pair was increased to double pressure. In response to high shear stress, HUVECs and SC cells expressed more SEAP and GFP than control. Similarly, human anterior segments perfused at higher flow rates released significantly more nitrites and SEAP into perfusion effluent, and SC cells expressed increased GFP near collector channel ostia compared to control. These data establish that engineered adenoviruses have the capacity to quantify and localize shear stress experienced by endothelial cells. This is the first in situ demonstration of shear-mediated SC mechanobiology as a key IOP-sensing mechanism necessary for IOP homeostasis.

Due to their similarities in anatomy, physiology, and pharmacology to humans, mice are a valuable model system to study the generation and mechanisms modulating conventional outflow resistance and thus intraocular pressure. In addition, mouse models are critical for understanding the complex nature of conventional outflow homeostasis and dysfunction that results in ocular hypertension. In this review, we describe a set of minimum acceptable standards for developing, characterizing, and utilizing mouse models of open-angle ocular hypertension. We expect that this set of standard practices will increase scientific rigor when using mouse models and will better enable researchers to replicate and build upon previous findings.

The trabecular meshwork (TM) and Schlemm’s canal generate the majority of outflow resistance; however, the distal regions of the conventional outflow pathway account for 25–50% of total resistance. Sections of distal vessels are surrounded by α-smooth muscle actin-containing cells, indicating that they may be vasoregulated. This study examined the effect of a potent vasodilator, nitric oxide (NO), and its physiological antagonist, endothelin-1 (ET-1), on the regulation of outflow resistance in the distal regions of the conventional outflow pathway. Using a physiological model of the conventional outflow pathway, human and porcine anterior segments were perfused in organ culture under constant flow conditions, while intrachamber pressure was continually monitored. For porcine anterior segments, a stable baseline outflow facility with TM intact was first achieved before anterior segments were removed and a trabeculotomy was performed. For human anterior segments, a trabeculotomy was immediately performed. In human anterior segments, 100 nM ET-1 significantly decreased distal outflow facility from 0.49 ± 0.26 to 0.31 ± 0.18 (mean ± SD) µl·min−1·mmHg, P < 0.01. Perfusion with 100 µM diethylenetriamine-NO in the presence of 1 nM ET-1 immediately reversed ET-1 effects, significantly increasing distal outflow facility to 0.54 ± 0.35 µl·min−1·mmHg, P = 0.01. Similar results were obtained in porcine anterior segment experiments. Therefore, data show a dynamic range of resistance generation by distal vessels in both the human and the porcine conventional outflow pathways. Interestingly, maximal contraction of vessels in the distal outflow tract of trabeculotomized eyes generated resistance very near physiological levels for both species having an intact TM.

Glaucoma is an optic neuropathy that affects 60 million people worldwide. There is an underlying fibrosis associated with the lamina cribrosa (LC) in glaucoma. DNA methylation is well established in regulating fibrosis and may be a therapeutic target for glaucoma. The purpose of this study was to compare global DNA methylation levels in primary human normal (NLC) and glaucomatous (GLC) cells, and to investigate DNA methylation in driving fibrosis through regulation of transforming growth factor β1 (TGFβ1).LC cells were cultured from normal and glaucomatous human donors. Global methylation was assessed by ELISA. qPCR was conducted for DNA methyltransferases (DNMTs), methyl-CpG-binding protein 2 (MeCP2), TGFβ 1 and 2, collagen 1α1 (COL1A1), and α-smooth muscle actin (αSMA). TGFβ1 and DNMT1 were examined by immunofluorescence. Methylation of the TGFβ1 promoter was determined by methylation-specific PCR (MSP).Global DNA methylation demonstrated an increase in GLC compared with NLC cells (P0.05). The previously mentioned methylation and matrix genes were increased in GLC compared with NLC cells (P0.05). Immunofluorescence showed increased TGFβ1 and DNMT1 in GLC compared with NLC cells. MSP showed increased unmethylated DNA in the TGFβ1 promoter of GLC compared with NLC cells.We found increased expression of fibrotic genes in GLC cells and demonstrated an increase in global DNA methylation and in associated enzymes in GLC cells. Furthermore, we showed decreased promoter methylation of TGFβ1 in GLC cells. Determining a role for methylation in glaucoma and in regulating TGFβ1 may provide a novel therapeutic approach.

Cultured trabecular meshwork (TM) cells are a valuable model system to study the cellular mechanisms involved in the regulation of conventional outflow resistance and thus intraocular pressure; and their dysfunction resulting in ocular hypertension. In this review, we describe the standard procedures used for the isolation of TM cells from several animal species including humans, and the methods used to validate their identity. Having a set of standard practices for TM cells will increase the scientific rigor when used as a model, and enable other researchers to replicate and build upon previous findings.

Purpose Fibrosis and a hypoxic environment are associated with the trabecular meshwork (TM) region in the blinding disease glaucoma. Hypoxia has been shown to alter DNA methylation, an epigenetic mechanism involved in regulating gene expression such as the pro-fibrotic transforming growth factor (TGF) β1 and the anti-fibrotic Ras protein activator like 1 (RASAL1). The purpose of this study was to compare DNA methylation levels, and the expression of TGFβ1 and RASAL1 in primary human normal (NTM) with glaucomatous (GTM) cells and in NTM cells under hypoxic conditions. Methods Global DNA methylation was assessed by ELISA in cultured age-matched NTM and GTM cells. qPCR was conducted for TGFβ1, collagen 1α1 (COL1A1), and RASAL1 expression. Western immunoblotting was used to determine protein expression. For hypoxia experiments, NTM cells were cultured in a 1%O2, 5%CO2 and 37°C environment. NTM and GTM cells were treated with TGFβ1 (10ng/ml) and the methylation inhibitor 5-azacytidine (5-aza) (0.5μM) respectively to determine their effects on DNA Methyltransferase 1 (DNMT1) and RASAL1 expression. Results We found increased DNA methylation, increased TGFβ1 expression and decreased RASAL1 expression in GTM cells compared to NTM cells. Similar results were obtained in NTM cells under hypoxic conditions. TGFβ1 treatment increased DNMT1 and COL1A1, and decreased RASAL1 expression in NTM cells. 5-aza treatment decreased DNMT1, TGFβ1 and COL1A1 expression, and increased RASAL1 expression in GTM cells. Conclusions TGFβ1 and RASAL1 expression, global DNA methylation, and expression of associated methylation enzymes were altered between NTM and GTM cells. We found that hypoxia in NTM cells induced similar results to the GTM cells. Furthermore, DNA methylation, TGFβ1 and RASAL1 appear to have an interacting relationship that may play a role in driving pro-fibrotic disease progression in the glaucomatous TM.

Purpose Alteration in the extracellular matrix (ECM) of the optic nerve head (ONH) causes lamina cribrosa (LC) fibrosis and affects the mechanical integrity of the ONH. Increased ECM tissue stiffness drives myofibroblast activation leading to tissue fibrosis throughout the body. Here using primary human LC cells, we investigate the effect of substrate stiffness on profibrotic changes, which might be a key molecular mechanism driving ECM remodeling of the LC in primary open-angle glaucoma (POAG) glaucoma. Methods Primary human LC cells from normal and age-matched POAG glaucoma donors were cultured on substrates with defined mechanical properties of 5 and 100 kPa to replicate the range of mechanical microenvironments that cells may experience in vivo. Cell morphology, spread area, actin stress fibers, vinculin-focal adhesion formation, and α-smooth muscle actin (α-SMA) signal were examined using immunofluorescence staining. The elastic modulus of cells was measured using atomic force microscopy (AFM). Results Significantly greater cell spread area along with increased actin filament development, and vinculin-focal adhesion formation (number and size) were found in both normal and glaucoma LC cells cultured on stiff substrates. These changes were positively associated with elevated cell stiffness measured by AFM. Changes in spreading and cytoskeleton organization of glaucoma LC cells were significantly more pronounced than those in normal cells. The transformation to a myofibroblast-like cell phenotype was identified in both LC cells exposed to stiffer substrates, as indicated by an increased α-SMA signal and its colocalization with the actin stress fibers. Conclusions These findings demonstrated that a stiffer cell microenvironment activates a myofibroblastic transformation in human LC cells, and therefore contributes to LC remodelling and fibrosis in glaucoma.

The authors of the three papers in this issue discuss and analyze the practice underlying “critical exploration,” a research pedagogy applied in common within their separate art, science, and teacher education classrooms. Eleanor Duckworth developed critical exploration as a method of teaching by involving students so actively and reflectively with a subject that they have “wonderful ideas” that arise from their own questioning. Teachers who encourage critical exploration support their students in encountering complex materials, experiencing confusion, considering multiple possibilities, and constructing new understandings. Teachers refrain from providing answers, or even implying that there is an acceptable answer or technique, and instead facilitate the personal process of development that Jean Piaget, Barbel Inhelder, and others documented and analyzed. Applying Piaget's findings requires teachers to sustain what David Hawkins described as “triangular relationships” of trust and respect among teacher, ...

Glaucoma is an optic neuropathy that affects 60 million people worldwide. The main risk factor for glaucoma is increased intraocular pressure (IOP), this is currently the only target for treatment of glaucoma. However, some patients show disease progression despite well-controlled IOP. Another possible therapeutic target is the extracellular matrix (ECM) changes in glaucoma. There is an accumulation of ECM in the lamina cribrosa (LC) and trabecular meshwork (TM) and upregulation of profibrotic factors such as transforming growth factorβ(TGFβ), collagen1α1 (COL1A1), andα-smooth muscle actin (αSMA). One method of regulating fibrosis is through epigenetics; the study of heritable changes in gene function caused by mechanisms other than changes in the underlying DNA sequence. Epigenetic mechanisms have been shown to drive renal and pulmonary fibrosis by upregulating profibrotic factors. Hypoxia alters epigenetic mechanisms through regulating the cell’s response and there is a hypoxic environment in the LC and TM in glaucoma. This review looks at the role that hypoxia plays in inducing aberrant epigenetic mechanisms and the role these mechanisms play in inducing fibrosis. Evidence suggests that a hypoxic environment in glaucoma may induce aberrant epigenetic mechanisms that contribute to disease fibrosis. These may prove to be relevant therapeutic targets in glaucoma.

This paper describes the development of a national assessment guide to assist the Irish Pre-School Inspectorate in evaluating support for child development in early childhood care and education settings. The development arose as a direct result of changes to the regulations governing pre-school childcare provision in Ireland. Revised regulations, published in 2006, built on previous regulations and made explicit the whole-child perspective within them. This places an important and overt focus on support for children's development within the inspection process, and is based on a socio-ecological understanding of children's lives. Using a collaborative approach with the Pre-School Inspectorate, an assessment guide was developed to enhance practitioners' understanding of the whole-child perspective element of the revised regulations, and to ensure consistency in the inspection process.