Effect of bone morphogenetic protein-4 (BMP-4) on the expression of Sox2, Oct-4 and c-Myc in human periodontal ligament cells during long-term culture

Recent studies demonstrated endogenous expression level of Sox2, Oct-4 and c-Myc is correlated with the pluripotency and successful induction of induced pluripotent stem cells (iPSCs). Periondontal ligament cells (PDLCs)have multi-lineage diferentiation capability and ability to maintain undifferentiated stage, which makes PDLCs a suitable cell source for tissue repair and regeneration. To elucidate the effect of in vitro culture condition on the stemness potential of PDLCs, we explored the cell growth, proliferation, cell cycle, and the expression of Sox2, Oct-4 and c-Myc in PDLCs from passage 1 to 7 with or without the addition of recombinant human BMP4(rhBMP4). Our results revealed that BMP-4 promoted cell growth and proliferation, arrested PDLCs in S phase of cell cycle and upregulated PI value. It was revealed that without the addition of rhBMP4, the expression of Sox2, Oct-4 and c-Myc in PDLCs only maintained nucleus location until passage 3, then lost nucleus location subsequently. The mRNA expression in PDLCs further confirmed that the level of Sox2 and Oct-4 peaked at passage 3, then decreased afterwards, whereas c-Myc maintained consistently upregulation along passages. after the treatment with rhBMP4, the expression of Sox2, Oct-4 and c-Myc in PDLCs maintained nucleus location even at passage 7 and the mRNA expression of Sox2 and Oct-4 significantly upregulated at passage 5 and 7. These results demonstrated that addition of rhBMP-4 in the culture media could improve the current culture condition for PDLCs to maintain in an undifferentiated stage.

Organotypic culture of normal, dysplastic and squamous cell carcinoma-derived oral cell lines reveals loss of spatial regulation of CD44 and p75(NTR) in malignancy

Oral squamous cell carcinomas (OSCC) often arise from dysplastic lesions. The role of cancer stem cells in tumour initiation is widely accepted, yet the potential existence of pre-cancerous stem cells in dysplastic tissue has received little attention. Cell lines from oral diseases ranging in severity from dysplasia to malignancy provide opportunity to investigate the involvement of stem cells in malignant progression from dysplasia. Stem cells are functionally defined by their ability to generate hierarchical tissue structures in consortium with spatial regulation. Organotypic cultures readily display tissue hierarchy in vitro; hence, in this study, we compared hierarchical expression of stem cell-associated markers in dermis-based organotypic cultures of oral epithelial cells from normal tissue (OKF6-TERT2), mild dysplasia (DOK), severe dysplasia (POE-9n) and OSCC (PE/CA P J15). Expression of CD44, p75NTR, CD24 and ALDH was studied in monolayers by flow cytometry and in organotypic cultures by immunohistochemistry. Spatial regulation of CD44 and p75NTR was evident for organotypic cultures of normal (OKF6-TERT2) and dysplasia (DOK and POE-9n) but was lacking for OSCC (PE/CA PJ15)-derived cells. Spatial regulation of CD24 was not evident. All monolayer cultures exhibited CD44, p75NTR, CD24 antigens and ALDH activity (ALDEFLUOR® assay), with a trend towards loss of population heterogeneity that mirrored disease severity. In monolayer, increased FOXA1 and decreased FOXA2 expression correlated with disease severity, but OCT3/4, Sox2 and NANOG did not. We conclude that dermis-based organotypic cultures give opportunity to investigate the mechanisms that underlie loss of spatial regulation of stem cell markers seen with OSCC-derived cells.

An in-vitro 3-D cell culture model for studying pathomechanisms in AMD

Purpose To develop a novel 3-D cell culture model with the view to studying the pathomechanisms underlying the development of age-related macular degeneration (AMD). Our central hypothesis is that the silk structural protein fibroin used in conjunction with cultured human cells can be used to mimic the structural relationships between the RPE and choriocapillaris in health and disease. Methods Co-cultures of human RPE cells (ARPE-19 cells grown in Miller’s medium) and microvascular endothelial cells (HMEC-1 cells grown in endothelial culture medium) were established on opposing sides of a synthetic Bruch’s membrane (3 microns thick) constructed from B mori silk fibroin. Cell attachment was facilitated by pre-coating the fibroin membrane with vitronectin (for ARPE-19 cells) and gelatin (for HMEC-1 cells) respectively. The effects of tropoelastin on attachment of ARPE-19 cells was also examined. Barrier function was examined by measurement of trans-epithelial resistance (TER) using a voltohmmeter (EVOM-2). The phagocytic activity of the synthetic RPE was tested using vitronectin-coated microspheres (2 micron diameter FluoSpheres). In some cultures, membrane defects were created by puncturing within a 24 G needle. The architecture of the synthetic tissue before and after wounding was examined by confocal microscopy after staining for ZO-1 and F-actin. Results The RPE layer of the 3D model developed a cobblestoned morphology (validated by staining for ZO-1 and F-actin), displayed barrier function (validated by measurement of TER) and demonstrated cytoplasmic uptake of vitronectin-coated microspheres. Attachment of ARPE-19 cells to fibroin was unaffected by tropoelastin. Microvascular endothelial cells attached well to the gelatin-coated surface of the fibroin membrane and remained physically separated from the overlaying RPE layer. The fibroin membranes were amenable to puncturing without collapse thus providing the opportunity to study transmembrane migration of the endothelial cells. Conclusions Synthetic Bruch’s membranes constructed from silk fibroin, vitronectin and gelatin, support the co-cultivation of RPE cells and microvascular endothelial cells. The resulting RPE layer displays functions similar to that of native RPE and the entire tri-layered structure displays potential to be used as an in vitro model of choroidal neovascularization.

A hierarchical structural model of assessing innovation and project performance

Research in construction innovation highlights construction industry as having many barriers and resistance to innovations and suggests that it needs champions. A hierarchical structural model is presented, to assess the impact of the role of the project manager (PM) on the levels of innovation and project performance. The model adopts the structural equation modelling technique and uses the survey data collected from PMs and project team members working for general contractors in Singapore. The model fits well to the observed data, accounting for 24%, 37% and 49% of the variance in championing behaviour, the level of innovation and project performance, respectively. The results of this study show the importance of the championing role of PMs in construction innovation. However, in order to increase their effectiveness, such a role should be complemented by their competency and professionalism, tactical use of influence tactics, and decision authority. Moreover, senior management should provide adequate resources and a sustained support to innovation and create a conducive environment or organizational culture that nurtures and facilitates the PM’s role in the construction project as a champion of innovation.

The new normativity : the innovation imperative

At a quite fundamental level, the very way in which Public Service Broadcasting (PSB) may envisage its future usually captured in the semantic shift from PSB to Public Service Media (PSM) is at stake when considering the recent history of public value discourse and the public value test. The core Reithian PSB idea assumed that public value would be created through the application of core principles of universality of availability and appeal, provision for minorities, education of the public, distance from vested interests, quality programming standards, program maker independence, and fostering of national culture and the public sphere. On the other hand, the philosophical import of the public value test is that potentially any excursion into the provision of new media services needs to be justified ex ante. In this era of New Public Management, greater transparency and accountability, and the proposition that resources for public value deliverables be contestable and not sequestered in public sector institutions, what might be the new Archimedean point around which a contemporised normativity for PSM be built? This paper will argue for the innovation imperative as an organising principle for contemporary PSM. This may appear counterintuitive, as it is precisely PSB’s predilection for innovating in new media services (in online, mobile, and social media) that has produced the constraining apparatus of the ex ante/public value/Drei-Stufen-Test in Europe, based on principles of competitive neutrality and transparency in the application of public funds for defined and limited public benefit. However, I argue that a commitment to innovation can define as complementary to, rather than as competitive ‘crowding out’, the new products and services that PSM can, and should, be delivering into a post-scarcity, superabundant all-media marketplace. The evidence presented in this paper for this argument is derived mostly from analysis of PSM in the Australian media ecology. While no PSB outside Europe is subject to a formal public value test, the crowding out arguments are certainly run in Australia, particularly by powerful commercial interests for whom free news is a threat to monetising quality news journalism. Take right wing opinion leader, herself a former ABC Board member, Judith Sloan: ‘… the recent expansive nature of the ABC – all those television stations, radio stations and online offerings – is actually squeezing activity that would otherwise be undertaken by the private sector. From partly correcting market failure, the ABC is now causing it. We are now dealing with a case of unfair competition and wasted taxpayer funds’ (The Drum, 1 August http://www.abc.net.au/unleashed/2818220.html). But I argue that the crowding out argument is difficult to sustain in Australia because of the PSB’s non-dominant position and the fact that much of innovation generated by the two PSBs, the ABC and the SBS, has not been imitated by or competed for by the commercials. The paper will bring cases forward, such as SBS’ Go Back to Where you Came From (2011) as an example of product innovation, and a case study of process and organisational innovation which also has resulted in specific product and service innovation – the ABC’s Innovation Unit. In summary, at least some of the old Reithian dicta, along with spectrum scarcity and market failure arguments, have faded or are fading. Contemporary PSM need to justify their role in the system, and to society, in terms of innovation.

Safety culture in supply chains : 'Customer is king'

Despite the presence of many regulations governing the operation of heavy vehicles and supply chains in Australia, the truck driving sector continues to have the highest incidence of fatal injuries compared to all other industries. The working environment has been the focus of attention by safety researchers during the past few decades, with particular consideration been given to the concept ‘safety culture’ and how to maintain, modify and advance responses to occupational risk. One important aspect of the heavy industry which sets it apart is the existence of cultural or sub-cultural influences at an industry wide and occupation-specific level rather than organisational level. This paper reports on the findings of stakeholder’s perceptions of the influences of power and control, and culture on industry safety. In-depth structured interviews were conducted during 2011 with Australian industry stakeholders (n=31). The questioning surrounded decision-making processes with regards to identifying risks, self-monitoring and reducing risky activities; as well as how power-affected relationships may influence the operational performance of supply chains and impacts on driver safety. One of the most significant findings from these interviews relates to the notion of power. The perception that the ‘Customer is King’ was widely viewed, with the majority of stakeholders believing that there exists a ‘master slave mentality’ in the industry. There appears to be great frustration in the industry as to the apparent immunity of customers (particularly retail supply chains) to their responsibilities. There was also a strong perception that the customer holds the balance of power by covertly employing remuneration-related incentives and pressures. Smaller trucking companies are perceived as being more vulnerable to the pressure of customer expectations.

Returning to the roots of culture : a review and re-conceptualisation of safety culture

Despite significant research, there is still little agreement over how to define safety culture or of what it is comprised. Due to this lack of agreement, much of the safety culture research has little more than safety management strategies in common. There is, however, a degree of acceptance of the close relationship between safety culture and organisational culture. Organisational culture can be described using traditional views of culture drawn from the anthropology and cultural psychology literature. However, the safety culture literature rarely ventures beyond organisational culture into discussions of these more traditional concepts of culture. There is a need to discuss how these concepts of culture can be applied to safety culture to provide greater understanding of safety culture and additional means by which to approach safety in the workplace. This review explores how three traditional conceptualisations of culture; the normative, anthropological and pragmatist conceptualisations, can and have been be applied to safety culture. Finally the review proposes a synthesised conceptualisation of safety culture which can be used to provide greater depth and practical applicability of safety culture, by increasing our understanding of the interactions between cultural and contextual variables in a given workplace and the effect they have on safety.

Culture as a driver of entrepreneurship : contrasting independent entrepreneurship versus employee entrepreneurship

Principal Topic Counties in Northern Europe, such as Sweden, Finland and Denmark, have comparatively low per capita rates of entrepreneurship as measured by independent new venture start-up rates – as for example measured by the Global Entrepreneurship Monitor (GEM) Total entrepreneurial activity (TEA) rate. However, the latest 2011 GEM data reveals that these same countries have comparatively very high Employee Entrepreneurship Activity (EEA) rates – that is a high rate per capita of employees involved in new product development or new enterprise activities. This observation has prompted us to investigate the role of national culture in driving independent versus employee entrepreneurial activities. Prior research has established that national (and regional) culture plays an important role in forming an “entrepreneurial culture” that encourages (or discourages) independent business start-ups and TEA (e.g. Davidsson, 1995; Beugelsdijk, 2007). However, the relationship of culture and EEA has not received research attention. Moreover, empirical relationships between elements of national culture and independent entrepreneurship have revealed some surprising results. For example, Wildeman et al. (1999) report an unexpected higher share of individual business ownership in countries that have higher uncertainty avoidance, higher power distance and lower individualism according to Hofstede’s dimensions of culture. They speculate that dissatisfaction can be a source of entrepreneurship: in countries with a high power distance, a high uncertainty avoidance and low individualism, there may be relatively more business owners since enterprising individuals cannot satisfy their needs within existing organizations. Yet it remains a rather open question whether entrepreneurial behaviour in existing organisations provides a satisfactory explanation for these empirical findings. Methods We will conduct a cross sectional study of the influence of national culture according to the five / six dimensions of Hofstede (1980; 2001) on both TEA and EEA for the 54 countries that participated in GEM 2011. Since it is well established that the opportunities for entrepreneurship vary substantially with a country’s level of economic development, we intend to conduct separate analyses for the three categories of development – innovation driven economies, efficient driven economies and factor driven economies. We also intend to restrict our assessment of TEA to opportunity driven entrepreneurship, as necessity driven entrepreneurship has a different relationship to the “entrepreneurial culture” that is the focus of our study. We will control for a range of factors such as GDP growth, ease of doing business index and unemployment. Results and Implications Descriptive analyses of the GEM TEA and EEA data reveal clusters of countries that appear to be have similar national culture. We are yet to conduct regression analyses.

In vitro and in vivo MMP gene expression localisation by In Situ-RT-PCR in cell culture and paraffin embedded human breast cancer cell line xenografts

Background Members of the matrix metalloproteinase (MMP) family of proteases are required for the degradation of the basement membrane and extracellular matrix in both normal and pathological conditions. In vitro, MT1-MMP (MMP-14, membrane type-1-MMP) expression is higher in more invasive human breast cancer (HBC) cell lines, whilst in vivo its expression has been associated with the stroma surrounding breast tumours. MMP-1 (interstitial collagenase) has been associated with MDA-MB-231 invasion in vitro, while MMP-3 (stromelysin-1) has been localised around invasive cells of breast tumours in vivo. As MMPs are not stored intracellularly, the ability to localise their expression to their cells of origin is difficult. Methods We utilised the unique in situ-reverse transcription-polymerase chain reaction (IS-RT-PCR) methodology to localise the in vitro and in vivo gene expression of MT1-MMP, MMP-1 and MMP-3 in human breast cancer. In vitro, MMP induction was examined in the MDA-MB-231 and MCF-7 HBC cell lines following exposure to Concanavalin A (Con A). In vivo, we examined their expression in archival paraffin embedded xenografts derived from a range of HBC cell lines of varied invasive and metastatic potential. Mouse xenografts are heterogenous, containing neoplastic human parenchyma with mouse stroma and vasculature and provide a reproducible in vivo model system correlated to the human disease state. Results In vitro, exposure to Con A increased MT1-MMP gene expression in MDA-MB-231 cells and decreased MT1-MMP gene expression in MCF-7 cells. MMP-1 and MMP-3 gene expression remained unchanged in both cell lines. In vivo, stromal cells recruited into each xenograft demonstrated differences in localised levels of MMP gene expression. Specifically, MDA-MB-231, MDA-MB-435 and Hs578T HBC cell lines are able to influence MMP gene expression in the surrounding stroma. Conclusion We have demonstrated the applicability and sensitivity of IS-RT-PCR for the examination of MMP gene expression both in vitro and in vivo. Induction of MMP gene expression in both the epithelial tumour cells and surrounding stromal cells is associated with increased metastatic potential. Our data demonstrate the contribution of the stroma to epithelial MMP gene expression, and highlight the complexity of the role of MMPs in the stromal-epithelial interactions within breast carcinoma.

Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

Background Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. Methods To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. Results Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. Conclusion We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms.

Traffic safety culture : what can we learn from Australia?

-- The role of traffic safety culture in Australia -- A comparison of drink driving (a success story) and speeding (a work in progress) ―Countermeasure approaches ―Community attitudes, perceptions and behaviors -- Lessons from Australia for the further development of the traffic safety culture concept

The challenges of adopting design-led innovative strategies in not for profits : the role of consumers, culture and employees

Major changes to regulations, funding and consumer demand in the Australian aged care industry are driving not for profits in this sector to reshape and rethink the services they offer and the ways in which they deliver their services to consumers. Many not for profit organisations facing these new challenges are also facing organisational cultural barriers in the development and implementation of innovative strategies. This paper presents a case study where one organisation, using design led innovation, explored consumer insights and employee values to find new ways to facilitate change.

Research capacity and culture in podiatry : early observations within Queensland Health

Background Research is a major driver of health care improvement and evidence-based practice is becoming the foundation of health care delivery. For health professions to develop within emerging models of health care delivery, it would seem imperative to develop and monitor the research capacity and evidence-based literacy of the health care workforce. This observational paper aims to report the research capacity levels of statewide populations of public-sector podiatrists at two different time points twelve-months apart. Methods The Research Capacity & Culture (RCC) survey was electronically distributed to all Queensland Health (Australia) employed podiatrists in January 2011 (n = 58) and January 2012 (n = 60). The RCC is a validated tool designed to measure indicators of research skill in health professionals. Participants rate skill levels against each individual, team and organisation statement on a 10-point scale (one = lowest, ten = highest). Chi-squared and Mann Whitney U tests were used to determine any differences between the results of the two survey samples. A minimum significance of p < 0.05 was used throughout. Results Thirty-seven (64%) podiatrists responded to the 2011 survey and 33 (55%) the 2012 survey. The 2011 survey respondents reported low skill levels (Median < 4) on most aspects of individual research aspects, except for their ability to locate and critically review research literature (Median > 6). Whereas, most reported their organisation’s skills to perform and support research at much higher levels (Median > 6). The 2012 survey respondents reported significantly higher skill ratings compared to the 2011 survey in individuals’ ability to secure research funding, submit ethics applications, and provide research advice, plus, in their organisation’s skills to support, fund, monitor, mentor and engage universities to partner their research (p < 0.05). Conclusions This study appears to report the research capacity levels of the largest populations of podiatrists published. The 2011 survey findings indicate podiatrists have similarly low research capacity skill levels to those reported in the allied health literature. The 2012 survey, compared to the 2011 survey, suggests podiatrists perceived higher skills and support to initiate research in 2012. This improvement coincided with the implementation of research capacity building strategies.