Osteogenic potential of human adipose derived stem cell co-culture with human osteoblast on titanium dioxide nanofibrous surface

The study aims to evaluate the osteogenic potential of human adipose derived stem cell (HADSC) co-culture with human osteoblast (HOB) using selected HADSC/HOB ratio of 2:1, 1:1 and 1:2, respectively. The HADSC/HOB was seeded on Titanium dioxide (TiO2) coated with or without nanofibre substrate. The non-coated TiO2 was used as control. The effects of TiO2 based scaffolds on cell adhesion were characterized by scanning electron microscopy (SEM). Cell viability, differentiation and mineralization were assessed by Alamar Blue, alkaline phosphatase (ALP) and Alizarin Red assays, respectively. The combination of HADSC/HOB, 2:1 ratio, seeded on nanofibrous-coated TiO2 showed better cell adhesion, viability, differentiation and mineralization than the other groups. This study offers opportunity to assess in vitro cellular development of HADSC through direct cell to cell contact with HOB. This study indicates that the co-cultured HADSC/HOB seeded on TiO2 based scaffolds may serve as a promising approach to facilitate osteogenic differentiation activity

Protocol for a mixed-methods investigation of quality improvement in early childhood education and care in Australia

International recognition of the early years as a crucial foundational period has led to the design and implementation of quality rating and improvement systems (QRIS) that define, communicate, and monitor the components of quality in early childhood education and care (ECEC). The aim of these policies is to achieve effective quality assurance and improvement through a system-oriented approach to assessment and evaluation. Informed by ecological systems theory, this paper outlines a three-phase, mixed-methods design for researching a national sample of child care centres that showed overall improvement on the Australian National Quality Standard (NQS) assessment and rating (A&R) criteria. The study samples are drawn from a national dataset of centre-based child care services with two or more A&R rounds and an initial rating of Working Towards NQS (N = 1,935). The results of this study will provide insights into the macro-, exo-, meso- and micro-systems level factors and strategies that support quality in ECEC services

Microglial MAC1 receptor and PI3K are essential in mediating β-amyloid peptide-induced microglial activation and subsequent neurotoxicity

<p>Abstract</p> <p>Background</p> <p>β-Amyloid peptide (Aβ) is a major protein in the brain associated with Alzheimer's and Parkinson's diseases. The purpose of this study was to investigate the role of macrophage antigen-1 (MAC1) receptor, an integrin scavenger receptor in microglia, and subsequent signaling events in mediating Aβ-induced neurotoxicity. We have previously reported that NADPH oxidase (PHOX) on microglia and superoxide produced by PHOX are critical for Aβ-induced loss of dopaminergic neurons. However, the upstream signaling pathway of superoxide production remains unclear.</p> <p>Methods</p> <p>For the <it>in vitro </it>study, mesencephalic neuron-glia cultures and microglia-enriched cultures from mice deficient in the MAC1 receptor (MAC1^-/-) and wild type controls were used to investigate the role of MAC1 receptor in Aβ-induced neurotoxicity and the role of phosphoinositide-3 kinase (PI3K) in the signal pathway between MAC1 receptor and PHOX. For the <it>in vivo </it>study, Aβ was injected into the substantia nigra of MAC1^-/-mice and wild type mice to confirm the role of MAC1 receptor.</p> <p>Results</p> <p>We found that Aβ-induced activation of microglia, activation of PHOX, generation of superoxide and other reactive oxygen species, and loss of dopaminergic neurons were decreased in MAC1^-/-cultures compared to MAC1^+/+cultures. In MAC1^-/-mice, dopaminergic neuron loss in response to Aβ injection into the substantia nigra was reduced relative to MAC1^+/+mice. Thus, MAC1 receptor-mediated PHOX activation and increased superoxide production are associated with Aβ-induced neurotoxicity. PI3K activation was one downstream step in MAC1 signaling to PHOX and played an important role in Aβ-induced neurotoxicity. In microglia-enriched cultures from MAC1^-/-mice, Aβ-induced activation of PI3K (phosphorylation of target proteins and PIP₃production) was reduced relative to MAC1^+/+cultures.</p> <p>Conclusions</p> <p>Taken together, our data demonstrate that Aβ activates MAC1 receptor to increase the activity of PI3K, which in turn phosphorylates p47^{<it>phox</it>}, triggers the translocation of cytosolic subunits of PHOX to microglia membrane, increases PHOX activation and the subsequent production of superoxide and causes neurotoxicity.</p

Mapping the leap: Differences in quality improvement in relation to assessment rating outcomes

Introduction: Australia’s National Quality Standard (NQS) outlines the criteria to assess the quality of early childhood services. A four-point rating scale: (i) Exceeding NQS; (ii) Meeting NQS; (iii) Working Toward NQS; and (iv) Significant Improvement Required is applied to services following a regular assessment and rating process. Settings rated as Working Toward are reassessed within 12 months. Most settings achieved a one-step improvement in this Time 2 reassessment, moving to a Meeting rating but some settings made a two-step improvement, moving to an Exceeding rating. The QIP is a key document used by authorities to assess the quality of a service. Methods: A grounded theory, data driven approach was taken to deepen understanding of quality rating improvements in long day care services in Australia of quality rating improvements by early childhood education and care [ECEC] services in Australia. This study, part of the second phase of a three phase study involved a document analysis of the Time 2 Quality Improvement Plans (QIPs) of a representative sample of Long Day Care (LDC) services (n = 60) from all Australian states and territories to determine what factors may have contributed to these different levels of improvement, with a focus on Quality Area 1 (QA1) (Educational programs and practices) and Quality Area 7 (QA7) (Governance and leadership). The study utilized the semantic analysis tool Leximancer 4.5. Leximancer 4.5 statistically analyses the semantic relationships between concepts in documents by measuring word proximity and correlation. The software creates visual maps of concepts and their connections to each other in texts. Concepts located near one another on the map are more likely to be contextually related. This tool is particularly useful when there are multiple, complex documents to analyze, reducing the potential biases that can arise from documents that use language with which these researchers are very familiar with. Results: The analysis found clear differences between the Time 2 QIPs of services who had made a two-step rating improvement and those who made a one-step improvement. Two-step (Exceeding NQS) category improvers for QA1 placed attention in their QIPs on improvement to the program and overall practice, with an orientation to the role of the educational leader. Two-step (Exceeding NQS) category improvers for QA7 seemed to be more oriented to a systemic view of the processes encompassed by QA7; how the management of the service and information supports the work of educators, with stronger links made between leadership roles (the manager and nominated supervisor) and the work of educators. Discussion: The QIPs demonstrated how the intentional and systemic processes in these quality areas related to practice, management, and leadership

Polycaprolactone-based scaffolds facilitates osteogenic differentiation of human adipose-derived stem cells in a co-culture system

Background: Stem cells in combination with scaffolds and bioactive molecules have made significant contributions to the regeneration of damaged bone tissues. A co-culture system can be effective in enhancing the proliferation rate and osteogenic differentiation of the stem cells. Hence, the aim of this study was to investigate the osteogenic differentiation of human adipose derived stem cells when co-cultured with human osteoblasts and seeded on polycaprolactone (PCL):hydroxyapatite (HA) scaffold; (2) Methods: Human adipose-derived stem cells (ASC) and human osteoblasts (HOB) were seeded in three different ratios of 1:2, 1:2 and 2:1 in the PCL-HA scaffolds. The osteogenic differentiation ability was evaluated based on cell morphology, proliferation rate, alkaline phosphatase (ALP) activity, calcium deposition and osteogenic genes expression levels using quantitative RT-PCR; (3) Results: The co-cultured of ASC/HOB in ratio 2:1 seeded on the PCLHA scaffolds showed the most positive osteogenic differentiation as compared to other groups, which resulted in higher ALP activity, calcium deposition and osteogenic genes expression, particularly Runx, ALP and BSP. These genes indicate that the co-cultured ASC/HOB seeded on PCL-HA was at the early stage of osteogenic development; (4) Conclusions: The combination of co-culture system (ASC/HOB) and PCL-HA scaffolds promote osteogenic differentiation and early bone formation

Targeting CBLB as a potential therapeutic approach for disseminated candidiasis

We thank J.M. Penninger (University of Toronto) for providing Cblb−/− mice, Y. Iwakura (Tokyo University of Science) for providing Clec4n−/− mice, S. Lipkowitz (National Cancer Institute, US National Institutes of Health) for providing Cblb constructs, X. Lin (MD Anderson Cancer Center) for providing the antibody to mouse dectin-3 and Card9−/− bone marrow cells, P.R. Sundstrom (Dartmouth University) for providing the C. albicans cap1 mutant, and L.D. Chaves (University at Buffalo) for flow cytometric analysis of myeloid cells in the kidneys. We also thank A. Lovett-Racke (Ohio State University) for her advice on in vivo Cblb-knockdown experiments. This work was supported by the US National Institutes of Health (grants R01 AI090901, R01 AI123253, and R21 AI117547; all to J.Z.), the American Heart Association (AHA Great Rivers Associate Grant-in-Aid grant 16GRNT26990004; J.Z.), a start-up fund from the Ohio State University College of Medicine (J.Z.), and the Wellcome Trust (G.D.B.).Peer reviewedPostprin

Identification of phenotypically and functionally heterogeneous mouse mucosal-associated invariant T cells using MR1 tetramers

Studies on the biology of mucosal-associated invariant T cells (MAIT cells) in mice have been hampered by a lack of specific reagents. Using MR1-antigen (Ag) tetramers that specifically bind to the MR1-restricted MAIT T cell receptors (TCRs), we demonstrate that MAIT cells are detectable in a broad range of tissues in C57BL/6 and BALB/c mice. These cells include CD4(-)CD8(-), CD4(-)CD8(+), and CD4(+)CD8(-) subsets, and their frequency varies in a tissue- and strain-specific manner. Mouse MAIT cells have a CD44(hi)CD62L(lo) memory phenotype and produce high levels of IL-17A, whereas other cytokines, including IFN-gamma, IL-4, IL-10, IL-13, and GM-CSF, are produced at low to moderate levels. Consistent with high IL-17A production, most MAIT cells express high levels of retinoic acid-related orphan receptor gamma t (ROR gamma t), whereas ROR gamma t(lo) MAIT cells predominantly express T-bet and produce IFN-gamma. Most MAIT cells express the promyelocytic leukemia zinc finger (PLZF) transcription factor, and their development is largely PLZF dependent. These observations contrast with previous reports that MAIT cells from V alpha 19 TCR transgenic mice are PLZF(-) and express a naive CD44(lo) phenotype. Accordingly, MAIT cells from normal mice more closely resemble human MAIT cells than previously appreciated, and this provides the foundation for further investigations of these cells in health and disease

α-Synuclein, a chemoattractant, directs microglial migration via H 2 O 2 -dependent Lyn phosphorylation

α-Synuclein (α-syn) aggregates released from neurons activate microglia, leading to chronic neuroinflammation that causes damage to neurons in brains with synucleinopathies, such as Parkinson’s disease (PD). However, little is known about the mechanism by which α-syn affects microglial activity, especially motility, and why microglia migrate toward the injured neurons and preferentially accumulate along with α-syn aggregates in the affected areas, e.g., in the substantia nigra of PD brains. Here we show that neuron-derived α-syn aggregates are chemoattractants that direct microglial migration by acting on NADPH oxidase and several specific downstream proteins. Blocking the targets involved in α-syn–mediated microglial directional migration may represent a therapeutic strategy to protect against progressive neuronal loss in PD and related synucleinopathies

The quest for continuous quality improvement in Australian long day care services: getting the most out of the Assessment and Rating process

The National Quality Framework (NQF) was intended to drive continuous improvement in education and care services in Australia. Ten years into implementation, the effectiveness of the NQF is demonstrated by steady improvements in quality as measured against the National Quality Standard (NQS). The process of assessing and rating services is a key element in the NQF, drawing together regulatory compliance and quality assurance. This paper draws on findings from a national Quality Improvement Research Project investigating the characteristics, processes, challenges and enablers of quality improvement in long day care services, concentrating on Quality Area 1 Educational program and practice and Quality Area 7 Governance and leadership. This was a mixed-method study focusing on long day care services that had improved their rating from Working toward NQS to Meeting NQS or to Exceeding NQS. The study comprised three phases, and in this paper, we draw on Phase 3 to understand the contribution of the NQS Assessment and Rating (A&amp;R) process to continuous quality improvement from the standpoint of providers and professionals delivering these services. Phase 3 involved qualitative case studies of 15 long day care services to investigate factors that enabled and challenged quality improvement. Data was collected during two-day site visits, using professional conversations and field notes to elicit the views and experiences of service providers, leaders and educators. In this paper, we look at how the A&amp;R process is experienced by those involved in service provision, with a focus on the factors that enabled and challenged quality improvement. Recognizing the interchangeability of enablers and challenges, three broad themes emerged: (i) curriculum knowledge, pedagogical skills and agency; (ii) collaborative leadership and teamwork; and (iii) meaningful engagement in the A&amp;R process. The study found that meaningful engagement in the A&amp;R process informed priorities for ongoing learning and acted as a catalyst for continuous quality improvement. Apprised by stakeholder views and experiences of A&amp;R, we offer a model to foster stakeholder participation in quality assurance matters through affordances of meaningful engagement

Dysfunction of the noradrenergic system drives inflammation, α-synucleinopathy, and neuronal loss in mouse colon

Clinical and pathological evidence revealed that α-synuclein (α-syn) pathology seen in PD patients starts in the gut and spreads via anatomically connected structures from the gut to the brain. Our previous study demonstrated that depletion of central norepinephrine (NE) disrupted brain immune homeostasis, producing a spatiotemporal order of neurodegeneration in the mouse brain. The purpose of this study was 1) to determine the role of peripheral noradrenergic system in the maintenance of gut immune homeostasis and in the pathogenesis of PD and 2) to investigate whether NE-depletion induced PD-like α-syn pathological changes starts from the gut. For these purposes, we investigated time-dependent changes of α-synucleinopathy and neuronal loss in the gut following a single injection of DSP-4 (a selective noradrenergic neurotoxin) to A53T-SNCA (human mutant α-syn) over-expression mice. We found DPS-4 significantly reduced the tissue level of NE and increased immune activities in gut, characterized by increased number of phagocytes and proinflammatory gene expression. Furthermore, a rapid-onset of α-syn pathology was observed in enteric neurons after 2 weeks and delayed dopaminergic neurodegeneration in the substantia nigra was detected after 3-5 months, associated with the appearance of constipation and impaired motor function, respectively. The increased α-syn pathology was only observed in large, but not in the small, intestine, which is similar to what was observed in PD patients. Mechanistic studies reveal that DSP-4-elicited upregulation of NADPH oxidase (NOX2) initially occurred only in immune cells during the acute intestinal inflammation stage, and then spread to enteric neurons and mucosal epithelial cells during the chronic inflammation stage. The upregulation of neuronal NOX2 correlated well with the extent of α-syn aggregation and subsequent enteric neuronal loss, suggesting that NOX2-generated reactive oxygen species play a key role in α-synucleinopathy. Moreover, inhibiting NOX2 by diphenyleneiodonium or restoring NE function by salmeterol (a β2-receptor agonist) significantly attenuated colon inflammation, α-syn aggregation/propagation, and enteric neurodegeneration in the colon and ameliorated subsequent behavioral deficits. Taken together, our model of PD shows a progressive pattern of pathological changes from the gut to the brain and suggests a potential role of the noradrenergic dysfunction in the pathogenesis of PD