Effects of dissolved oxygen availability and culture biomass at induction upon the intracellular expression of monoamine oxidase by recombinant E. coli in fed batch bioprocesses

The effects of oxygen availability and induction culture biomass upon production of an industrially important monoamine oxidase (MAO) were investigated in fed-batch cultures of a recombinant E. coli. For each induction cell biomass 2 different oxygenation methods were used, aeration and oxygen enriched air. Induction at higher biomass levels increased the culture demand for oxygen, leading to fermentative metabolism and accumulation of high levels of acetate in the aerated cultures. Paradoxically, despite an almost eight fold increase in acetate accumulation to levels widely reported to be highly detrimental to protein production, when induction wet cell weight (WCW) rose from 100% to 137.5%, MAO specific activity in these aerated processes showed a 3 fold increase. By contrast, for oxygenated cultures induced at WCW's 100% and 137.5% specific activity levels were broadly similar, but fell rapidly after the maxima were reached. Induction at high biomass levels (WCW 175%) led to very low levels of specific MAO activity relative to induction at lower WCW's in both aerated and oxygenated cultures. Oxygen enrichment of these cultures was a useful strategy for boosting specific growth rates, but did not have positive effects upon specific enzyme activity. Based upon our findings, consideration of the amino acid composition of MAO and previous studies on related enzymes, we propose that this effect is due to oxidative damage to the MAO enzyme itself during these highly aerobic processes. Thus, the optimal process for MAO production is aerated, not oxygenated, and induced at moderate cell density, and clearly represents a compromise between oxygen supply effects on specific growth rate/induction cell density, acetate accumulation, and high specific MAO activity. This work shows that the negative effects of oxygen previously reported in free enzyme preparations, are not limited to these acellular environments but are also discernible in the sheltered environment of the cytosol of E. coli cells.

Effect of culture conditions and calcium phosphate coating on ectopic bone formation

This study investigated the effect of a calcium phosphate (CaP) coating onto a polycaprolactone melt electrospun scaffold and in vitro culture conditions on ectopic bone formation in a subcutaneous rat model. The CaP coating resulted in an increased alkaline phosphatase activity (ALP) in ovine osteoblasts regardless of the culture conditions and this was also translated into higher levels of mineralisation. A subcutaneous implantation was performed and increasing ectopic bone formation was observed over time for the CaPcoated samples previously cultured in osteogenic media whereas the corresponding non-coated samples displayed a lag phase before bone formation occurred from 4 to 8 weeks post-implantation. Histology and immunohistochemistry revealed bone fill through the scaffolds 8 weeks post-implantation for coated and non-coated specimens and that ALP, osteocalcin and collagen 1 were present at the ossification front and in the bone tissues. Vascularisation in the vicinity of the bone tissues was also observed indicating that the newly formed bone was not deprived of oxygen and nutrients.We found that in vitro osteogenic induction was essential for achieving bone formation and CaP coating accelerated the osteogenic process. We conclude that high cell density and preservation of the collagenous and mineralised extracellular matrix secreted in vitro are factors of importance for ectopic bone formation.

Enhancing learning in large undergraduate classes using custom made audio-visual cases

This is an exploratory study into the effective use of embedding custom made audiovisual case studies (AVCS) in enhancing the student’s learning experience. This paper describes a project that used AVCS for a large divergent cohort of undergraduate students, enrolled in an International Business course. The study makes a number of key contributions to advancing learning and teaching within the discipline. AVCS provide first hand reporting of the case material, where the students have the ability to improve their understanding from both verbal and nonverbal cues. The paper demonstrates how AVCS can be embedded in a student-centred teaching approach to capture the students’ interest and to enhance a deep approach to learning by providing real-world authentic experience.

Toward a sustainable culture of peer partnership

This project is a two phase design working in partnership with five universities to develop, implement and systematically embed a distributive leadership model that aims to embed peer partnership (review, development) within the culture of teaching and learning excellence. This presentation will posit a ‘prototype’ peer review leadership model based on ongoing research that brings together both the fundamentals of peer review with the broader importance of context and persons. It will be argued that essential to teaching development is a need to address not only the implementation of peer partnership programs but also strategies to influence and change both the contexts of teaching and the advantages for colleagues. Peer review as a strategy to develop excellence in teaching needs to be considered from a holistic perspective encompassing all elements of the teaching environment. The emphasis is on working to foster the type of conditions needed for leadership and change to begin and be sustained. The work has implications for policy, research, leadership development and student outcomes and has potential application world-wide. Phase 1 has collected focus interview and questionnaire data to inform the research and is being analysed using a thematic qualitative approach and statistical analysis Evidence is emerging currently as the project is ongoing

Developing a culture of peer review of teaching through a distributive leadership approach

Teaching is a core function of higher education and must be effective if it is to provide students with learning experiences that are stimulating, challenging and rewarding Obtaining feedback on teaching is indispensable to enhancing the quality of learning design, facilitating personal and/or professional development and maximising student learning outcomes. Peer review of teaching has the potential to improve the quality of teaching at tertiary level, by encouraging critical reflection on teaching, innovation in teaching practice and scholarship of teaching at all academic levels. However, embedding peer review within the culture of teaching and learning is a significant challenge that requires sustained commitment from senior leadership as well as those in leadership roles within local contexts.

Culture makes you stronger : Aboriginal women’s voices from the South Coast of NSW

This paper will describe a community based research project examining the health and wellbeing of a sample of Aboriginal women in Australia, and present preliminary findings of a community needs analysis. The Shoalhaven Koori Women’s Study (SKWS) is being led by an Aboriginal woman based within Waminda, an Aboriginal women’s community controlled service located on the South Coast of NSW. The community needs analysis is the first stage of the SKWS, and aims to explore Aboriginal women’s perceptions and experiences of wellness and wellbeing, including issues related to their personal strengths, health and social priorities, support needs and that of their families. Thirty Aboriginal women were interviewed using a survey that included closed and open ended questions. Methods used to administer the survey included yarning and Dadirri (deep listening), two valid and culturally safe approaches for data collection with Aboriginal people. Adopting these approaches ensured Aboriginal protocols were maintained and upheld throughout the research process. This enabled scientific rigour while also ensuring activities were culturally safe. Key findings of the survey will be presented, and how Waminda is modifying service delivery to better respond to the health and social priorities of Aboriginal women in the Shoalhaven region will be discussed. Community feedback of survey results will occur to validate the analysis from the community perspective.

A visual profile of Queensland indigenous and non-indigenous school children, and the association between vision and reading

This research investigated the prevalence of vision disorders in Queensland Indigenous primary school children, creating the first comprehensive visual profile of Indigenous children. Findings showed reduced convergence ability and reduced visual information processing skills were more common in Indigenous compared to non-Indigenous children. Reduced visual information processing skills were also associated with reduced reading outcomes in both groups of children. As early detection of visual disorders is important, the research also reviewed the delivery of screening programs across Queensland and proposed a model for improved coordination and service delivery of vision screening to Queensland school children.

Paracrine interactions between LNCaP prostate cancer cells and bioengineered bone in 3D in vitro culture reflect molecular changes during bone metastasis

As microenvironmental factors such as three-dimensionality and cell–matrix interactions are increasingly being acknowledged by cancer biologists, more complex 3D in vitro models are being developed to study tumorigenesis and cancer progression. To better understand the pathophysiology of bone metastasis, we have established and validated a 3D indirect co-culture model to investigate the paracrine interactions between prostate cancer (PCa) cells and human osteoblasts. Co-culture of the human PCa, LNCaP cells embedded within polyethylene glycol hydrogels with human osteoblasts in the form of a tissue engineered bone construct (TEB), resulted in reduced proliferation of LNCaP cells. LNCaP cells in both monoculture and co-culture were responsive to the androgen analog, R1881, as indicated by an increase in the expression (mRNA and/or protein induction) of androgen-regulated genes including prostate specific antigen and fatty acid synthase. Microarray gene expression analysis further revealed an up-regulation of bone markers and other genes associated with skeletal and vasculature development and a significant activation of transforming growth factor β1 downstream genes in LNCaP cells after co-culture with TEB. LNCaP cells co-cultured with TEB also unexpectedly showed similar changes in classical androgen-responsive genes under androgen-deprived conditions not seen in LNCaP monocultures. The molecular changes of LNCaP cells after co-culturing with TEBs suggest that osteoblasts exert a paracrine effect that may promote osteomimicry and modulate the expression of androgen-responsive genes in LNCaP cells. Taken together, we have presented a novel 3D in vitro model that allows the study of cellular and molecular changes occurring in PCa cells and osteoblasts that are relevant to metastatic colonization of bone. This unique in vitro model could also facilitate cancer biologists to dissect specific biological hypotheses via extensive genomic or proteomic assessments to further our understanding of the PCa-bone crosstalk.

The culture of the digital : Pinterest and digital sociology

As the boundaries between public and private, human and technology, digital and social, mediated and natural, online and offline become increasingly blurred in modern techno-social hybrid societies, sociology as a discipline needs to adapt and adopt new ways of accounting for these digital cultures. In this paper I use the social networking site Pinterest to demonstrate how people today are shaped by, and in turn shape, the digital tools they are assembled with. Digital sociology is emerging as a sociological subdiscipline that engages with the convergence of the digital and the social. However, there seems to be a focus on developing new methods for studying digital social life, yet a neglect of concrete explorations of its culture. I argue for the need for critical socio-cultural ‘thick description’ to account for the interrelations between humans and technologies in modern digitally mediated cultures.

Soluble laminin and arginine-glycine-aspartic acid containing peptides differentially regulate type IV collagenase messenger RNA, activation, and localization in testicular cell culture

Rat testicular cells in culture produce several metalloproteinases including type IV collagenases (Sang et al. Biol Reprod 1990; 43:946-955, 956-964). We have now investigated the regulation of testicular cell type IV collagenase and other metalloprotemases in vitro. Soluble laminin stimulated Sertoli cell type IV collagenase mRNA levels. However, three peptides corresponding to different domains of the laminin molecule (CSRAKQAASIKVASADR, FALRGDNP, CLQDGDVRV) did not influence type IV collagenase mENA levels. Zyniographic analysis of medium collected from these cultures revealed that neither soluble laminin nor any of the peptides influenced 72-Wa type IV collagenase protein levels. However, peptide FALRGDNP resulted in both, a selective increase in two higher molecular-weight metalloprotemnases (83 kDa and 110 Wa and in an activation of the 72-Wa rat type IV collagenase. Interleukin-1, phorbol ester, testosterone, and FSH did not affect collagenase activation, lmmunocytochemical studies demonstrated that the addition of soluble laminin resulted in a redistribution of type IV collagenase from intracellular vesicles to the cell-substrate region beneath the cells. Peptide FALRGDNP induced a change from a vesicular to peripheral plasma membrane type of staining pattern. Zymography of plasma membrane preparations demonstrated triton-soluble gelatinases of 76 Wa, 83 Wa, and 110 Wa and a triton-insoluble gelatinase of 225 Wa, These results indicate that testicular cell type IV collagenase mRNA levels, enzyme activation, and distribution are influenced by laminin and RGD-containing peptides.

Characterization and novel activation of 72-kDa metalloproteinase in retinal interphotoreceptor matrix and Y-79 cell culture medium

Analysis of bovine interphotoreceptor matrix and conditioned medium from human Y-79 retinoblastoma cells by gelatin SDS-PAGE zymography reveals abundant activity of a 72-kDa M(r) gelatinase. The 72-kDa gelatinase from either source is inhibited by EDTA but not aprotinin or NEM, indicating that it is a metalloproteinase (MMP). The 72-kDa MMP is converted to a 62-kDa species with APMA treatment after gelatin sepharose affinity purification typical of previously described gelatinase MMP-2. The latent 72-kDa gelatinase from either bovine IPM or Y-79 media autoactivates without APMA in the presence of calcium and zinc after 72 hr at 37°C, producing a fully active mixture of proteinase species, 50 (48 in Y-79 medium), 38 and 35 kDa in size. The presence of inhibitory activity was examined in both whole bovine IPM and IPM fractions separated by SDS-PAGE. Whole IPM inhibited gelatinolytic activity of autoactivated Y-79-derived MMP in a dose-dependent manner. Inhibitory activities are observed in two protein fractions of 27-42 and 20-25 kDa. Western blots using antibodies to human tissue inhibitor of metalloproteinase 1 and 2 (TIMP-1 and -2) reveal the presence of two TIMP-1-like proteins at 21 and 29 kDa in inhibitory fractions of the bovine IPM. TIMP-2 was not detected in the inhibitory IPM fractions, consistent with the observed autoactivation of bovine IPM 72-kDa gelatinase. Potential roles for this IPM MMP-TIMP system include physiologic remodelling of the neural retina-RPE cell interface and digestion of shed rod outer segment as well as pathological processes such as retinal detachment, PE cell migration, neovascularization and tumor progression. Cultured Y-79 cells appear to be a good model for studying the production and regulation of this proteinase system.