Spontaneous large volume adipose tissue generation from a vascularized pedicled fat flap inside a chamber space

A novel method of spontaneous generation of new adipose tissue from an existing fat flap is described. A defined volume of fat flap based on the superficial inferior epigastric vascular pedicle in the rat was elevated and inset into a hollow plastic chamber implanted subcutaneously in the groin of the rat. The chamber walls were either perforated or solid and the chambers either contained poly(D,L-lactic-co-glycolic acid) (PLGA) sponge matrix or not. The contents were analyzed after being in situ for 6 weeks. The total volume of the flap tissue in all groups except the control groups, where the flap was not inserted into the chambers, increased significantly, especially in the perforated chambers (0.08 ± 0.007 mL baseline compared to 1.2 ± 0.08 mL in the intact ones). Volume analysis of individual component tissues within the flaps revealed that the adipocyte volume increased and was at a maximum in the chambers without PLGA, where it expanded from 0.04 ± 0.003 mL at insertion to 0.5 ± 0.08 mL (1250% increase) in the perforated chambers and to 0.16 ± 0.03 mL (400% increase) in the intact chambers. Addition of PLGA scaffolds resulted in less fat growth. Histomorphometric analysis rather than simple hypertrophy documented an increased number of adipocytes. The new tissue was highly vascularized and no fat necrosis or atypical changes were observed.

Contact with existing adipose tissue is inductive for adipogenesis in matrigel

The effect of adipose tissue on inductive adipogenesis within Matrigel (BD Biosciences) was assessed by using a murine chamber model containing a vascular pedicle. Three-chamber configurations that varied in the access to an adipose tissue source were used, including sealed- and open-chamber groups that had no access and limited access, respectively, to the surrounding adipose tissue, and a sealed-chamber group in which adipose tissue was placed as an autograft. All groups showed neovascularization, but varied in the amount of adipogenesis seen in direct relation to their access to preexisting adipose tissue: open chambers showed strong adipogenesis, whereas the sealed chambers had little or no adipose tissue; adipogenesis was restored in the autograft chamber group that contained 2- to 5-mg fat autografts. These showed significantly more adipogenesis than the sealed chambers with no autograft (p < 0.01). Autografts with 1 mg of fat were capable of producing adipogenesis but did so less consistently than the larger autografts. These findings have important implications for adipose tissue engineering strategies and for understanding de novo production of adipose tissue.

Host rather than graft origin of Matrigel-induced adipose tissue in the murine tissue-engineering chamber

We have recently shown that Matrigel-filled chambers containing fibroblast growth factor-2 (FGF2) and placed around an epigastric pedicle in the mouse were highly adipogenic. Contact of this construct with pre-existing tissue or a free adipose graft was required. To further investigate the mechanisms underpinning formation of new adipose tissue, we seeded these chambers with human adipose biopsies and human adipose-derived cell populations in severe combined immunodeficient mice and assessed the origin of the resultant adipose tissue after 6 weeks using species-specific probes. The tissues were negative for human-specific vimentin labeling, suggesting that the fat originates from the murine host rather than the human graft. This was supported by the strong presence of mouse-specific Cot-1 deoxyribonucleic acid labeling, and the absence of human Cot-1 labeling in the new fat. Even chambers seeded with FGF2/Matrigel containing cultured human stromal-vascular fraction (SVF) labeled strongly only for human vimentin in cells that did not have a mature adipocyte phenotype; the newly formed fat tissue was negative for human vimentin. These findings indicate that grafts placed in the chamber have an inductive function for neo-adipogenesis, rather than supplying adipocyte-precursor cells to generate the new fat tissue, and preliminary observations implicate the SVF in producing inductive factors. This surprising finding opens the door for refinement of current adipose tissue-engineering approaches.

Adipose tissue engineering based on the controlled release of fibroblast growth factor-2 in a collagen matrix

Adipose tissue forms when basement membrane extract (Matrigel™) and fibroblast growth factor-2 (FGF-2) are added to our mouse tissue engineering chamber model. A mouse tumor extract, Matrigel is unsuitable for human clinical application, and finding an alternative to Matrigel is essential. In this study we generated adipose tissue in the chamber model without using Matrigel by controlled release of FGF-2 in a type I collagen matrix. FGF-2 was impregnated into biodegradable gelatin microspheres for its slow release. The chambers were filled with these microspheres suspended in 60 μL collagen gel. Injection of collagen containing free FGF-2 or collagen containing gelatin microspheres with buffer alone served as controls. When chambers were harvested 6 weeks after implantation, the volume and weight of the tissue obtained were higher in the group that received collagen and FGF-2 impregnated microspheres than in controls. Histologic analysis of tissue constructs showed the formation of de novo adipose tissue accompanied by angiogenesis. In contrast, control groups did not show extensive adipose tissue formation. In conclusion, this study has shown that de novo formation of adipose tissue can be achieved through controlled release of FGF-2 in collagen type I in the absence of Matrigel.

Endothelial precursor cells home to a vascularized tissue engineering chamber by application of the angiogenic chemokine CXCl12

Tissue engineering of vascularized constructs has great utility in reconstructive surgery. While we have been successful in generating vascularized granulation-like tissue and adipose tissue in an in vivo tissue engineering chamber, production of other differentiated tissues in a stable construct remains a challenge. One approach is to utilize potent differentiation factors, which can influence the base tissue. Endothelial precursor cells (EPCs) have the ability to both carry differentiation factors and home to developing vasculature. In this study, proof-of-principle experiments demonstrate that such cells can be recruited from the circulation into an in vivo tissue engineering chamber. CXC chemokine ligand 12 (CXCL12)/stromal cell-derived factor 1 was infused into the chamber through Alzet osmotic pumps and chamber cannulation between days 0 and 7, and facilitated recruitment of systemically inoculated exogenous human EPCs injected on day 6. CXCL12 infusion resulted in an eightfold increase in EPC recruitment, 2 (p = 0.03) and 7 days postinfusion (p = 0.008). Delivery of chemotactic/proliferation and/or differentiation factors and appropriately timed introduction of effective cells may allow us to better exploit the regenerative potential of the established chamber construct. © Copyright 2009, Mary Ann Liebert, Inc. 2009.

The inter-relationships between ovarian-independent growth, tumorigenicity, invasiveness and antioestrogen resistance in the malignant progression of human breast cancer

Among the processes contributing to the progressive acquisition of the highly malignant phenotype in breast cancer are ovarian-independent growth, antioestrogen resistance and increased metastatic potential. We have previously observed that increased invasiveness and development of ovarian-independent growth occur independently. In an attempt to define the inter-relationships between these processes further, we have compared the phenotypes of ovarian-independent, invasive and antioestrogen-resistant sublines of the ovarian-dependent human breast cancer cell line MCF-7. Cells acquiring ovarian-independent growth can retain sensitivity to anti-oestrogens. One clone of MCF-7 cells selected for stable antioestrogen resistance has become non-tumorigenic but its invasive potential remains unaltered. Thus, acquisitions of some characteristics of the progressed phenotype can occur independently. This phenomenon of independent parameters in phenotypic progression could partly explain the considerable intra- and intertumour heterogeneity characteristic of breast tumours.

Epithelial-to-mesenchymal transitions and circulating tumor cells

Epithelial-to-mesenchymal transition (EMT) phenomena endow epithelial cells with enhanced migratory and invasive potential, and as such, have been implicated in many physiological and pathological processes requiring cell migration/invasion. Although their involvement in the metastatic cascade is still a subject of debate, data are accumulating to demonstrate the existence of EMT phenotypes in primary human tumors, describe enhanced metastatic potential of EMT derivatives in animal models, and report EMT attributes in circulating tumor cells (CTCs). The relationships between EMT and CTCs remain largely unexplored, and we review here in vitro and in vivo data supporting a putative role of EMT processes in CTC generation and survival.

Space within the surface : the patterned surfaces of Florence Broadhurst and Jim Thompson

The Woods Bagot 2007 refurbishment of the Qantas and British Airways Bangkok Business lounge in the Survarnabhumi Airport features wall finishes designed by wallpaper designer, Florence Broadhurst (1899-1977) and Thai Silk trader, Jim Thompson (1906-1967). This distinctive selection, which is proclaimed on the airport’s website, of patterned wall surfaces side by side draws attention to their striking similarities and their defining differences . Thompson and Broadhurst would appear to be worlds apart, but here in the airport their work brings them together. Thompson, the son of a wealthy cotton family in America, worked as an architect before joining the army. He moved to Bangkok to start The Thai Silk Company in 1948. Broadhurst was born on a farm in Mt. Perry, Queensland. She began her career as a performance artist, as part of an Australian troupe in Shanghai, moving onto pursue a career in fashion design, catering to the middle and upper classes in London. Upon her return to Australia, Broadhurst started a print design company in 1959. Both Broadhurst and Thompson pursued multiple careers, lived many lives, and died under mysterious circumstances. Broadhurst was murdered in 1977 at her Sydney print warehouse, which remains an unsolved crime. Thompson disappeared in Malaysia in 1967 and his body has never been found. This chapter investigates the parallels between Thompson and Broadhurst and what lead them to design such popular patterns for wall surfaces towards the end of their careers. While neither designer was a household name, their work is familiar to most, seen in the costume and set design of films, on the walls of restaurants and cafes and even in family homes. The reason for the popularity of their patterns has not previously been analysed. However, this chapter suggests that the patterns are intriguing because they contain something of their designers’ identities. It suggests that the coloured surface provides a way of camouflaging and hiding its subjects’ histories, such that Broadhurst and Thompson, consciously or unconsciously, used the patterned surface as a plane in which their past lives could be buried. The revealing nature of the stark white wall, compared with the forgiveness provided by the pattern in which to hide, is elaborated by painter and advocate for polychromatic architecture, Fernand Léger in his essay, “The Wall, The Architect, The Painter (1965).” Léger writes that, “the modern architect has gone too far in his magnificent attempts to cleanse through emptiness,” and that the resultant white walls of modernity create ‘an impalpability of air, of slick, brilliant new surfaces where nothing can be hidden any longer …even shadows don’t dare to enter’. To counter the exposure produced by the white wall, Thompson and Broadhurst designed patterned surfaces that could harbour their personal histories. Broadhurst and Thompson’s works share a number of commonalities in their design production, even though their work in print design commenced a decade apart. Both designers opted to work more with traditional methods of pattern making. Broadhurst used hand-operated screens, and Thompson outsourced work to local weavers and refrained from operating out of a factory. Despite humble beginnings, Broadhurst and Thompson enjoyed international success with their wall patterns being featured in a number of renowned international hotels in Bahrain, Singapore, Sydney, and London in the 1970s and 1980s. Their patterns were also transferred to fabric for soft furnishings and clothing. Thompson’s patterns were used for costumes in films including the King and I and Ben Hur. Broadhurst’s patterns were also widely used by fashion designers and artists, such as Akira Isogowa‘s costume design for Salome, a 1998 production by the Sydney Dance Company. Most recently her print designs have been used by skin illustrator Emma Hack, in a series of works painting female bodies into Broadhurst’s patterns. Hack’s works camouflage the models’ bodies into the patterned surface, assimilating subject and surface, hinting at there being something living within the patterned wall. More than four decades after Broadhurst’s murder and five decades since Thompson’s disappearance, their print designs persist as more than just a legacy. They are applied as surface finishes with the same fervour as when the designs were first released. This chapter argues that the reason for the ongoing celebration of their work is that there is the impalpable presence of the creator in the patterns. It suggests that the patterns blur the boundary between subject and surface.

Experimental colonization of the canine urinary tract with the asymptomatic bacteriuria Escherichia coli strain 83972

Establishment of asymptomatic bacteriuria (ABU) with Escherichia coli 83972 is a viable prophylactic alternative to antibiotic therapy for the prevention of recurrent bacterial urinary tract infection in humans. Approximately 2 x 108 viable E. coli 83972 cells were introduced into the bladder of six healthy female dogs via a sterile urinary catheter. The presence of pyuria, depression, stranguria, pollakiuria and haematuria was documented for 6 weeks and urinalysis and aerobic bacterial cultures were performed every 24–72 h. Pyuria was present in all dogs on day 1 post-inoculation and 4/6 dogs (67%) had a positive urine culture on this day. Duration of colonization ranged from 0 to 10 days (median 4 days). Four dogs were re-inoculated on day 20. Duration of colonization following the second inoculation ranged from 1 to 3 days. No dog suffered pyrexia or appeared systemically unwell but all dogs initially exhibited mild pollakiuria and a small number displayed gross haematuria and/or stranguria. By day 3 of each trial all clinical signs had resolved. Persistent bacteriuria was not achieved in any dog but two dogs were colonized for 10 days following a single inoculation. Further research is required to determine whether establishment of ABU in dogs with recurrent urinary tract infection is a viable alternative to repeated doses of antimicrobial agents.

Oncology service initiatives and research in regional Australia

Objective: This paper reflects on the recent growth of cancer research being conducted through some of Australia’s rural centres. It encompasses work being done across the fields of clinical, translational and health services research. Design: This is a collaborative piece with contributions from rural health researchers, clinical and cancer services staff from several different regions. Conclusion: The past decade has seen an expansion in cancer research in rural and regional Australia driven in part by the recognition that cancer patients in remote areas experience poorer outcomes than their metropolitan counterparts. This work has led to the development of more effective cancer networks and new models of care designed to meet the particular needs of the rural cancer patient. It is hoped that the growth of cancer research in regional centres will, in time, reduce the disparity between rural and urban communities and improve outcomes for cancer patients across both populations.

Reducing pressure injuries in critically ill patients by using a patient skin integrity care bundle (InSPiRE)

Purpose To test an interventional patient skin integrity bundle, InSPiRE protocol, on the impact of pressure injuries (PrIs) in critically ill patients in an Australian adult intensive care unit (ICU). Methods Before and after design was used where the group of patients receiving the intervention (InSPiRE protocol) was compared with a similar control group who received standard care. Data collected included demographic and clinical variables, skin assessment, PrI presence and stage, and a Sequential Organ Failure Assessment (SOFA) score. Results Overall, 207 patients were enrolled, 105 in the intervention group and 102 in the control group. Most patients were men, mean age 55. The groups were similar on major demographic variables (age, SOFA scores, ICU length of stay). Pressure injury cumulative incidence was significantly lower in the intervention group (18%) compared to the control group for skin injuries(30.4%) (χ2=4.271, df=1, p=0.039) and mucous injuries (t test =3.27, p=<0.001) . Significantly fewer PrIs developing over time in the intervention group (Logrank= 11.842, df=1, p=<0.001) and patients developed fewer skin injuries (>3 PrIs/patient = 1/105) compared with the control group (>3 injuries/patient = 10/102) (p=0.018). Conclusion The intervention group, recieving the InSPiRE protocol, had lower PrI cumulative incidence, and reduced number and severity of PrIs that developed over time. Systematic and ongoing assessment of the patient's skin and PrI risk as well as implementation of tailored prevention measures are central to preventing PrIs.

Performance pay for Australian teachers: A critical policy historiography

In May 2011, the Australian Federal Education Minister announced there would be a unique, innovative and new policy of performance pay for teachers, Rewards for Great Teachers (Garrett, 2011a). In response, this paper uses critical policy historiography to argue that the unintended consequences of performance pay for teachers makes it unlikely it will deliver improved quality or efficiency in Australian schools. What is new, in the Australian context, is that performance pay is one of a raft of education policies being driven by the federal government within a system that constitutionally and historically has placed the responsibility for schooling with the states and territories. Since 2008, a key platform of the Australian federal Labor government has been a commitment to an Education Revolution that would promote quality, equity and accountability in Australian schools. This commitment has resulted in new national initiatives impacting on Australian schools including a high-stakes testing regime 14 National Assessment Program 13 Literacy and Numeracy (NAPLAN) 14a mandated national curriculum (the Australian Curriculum), professional standards for teachers and teacher accreditation 14Australian Institute for Teaching and School Leadership (AITSL) 14and the idea of rewarding excellent teachers through performance pay (Garrett, 2011b). These reforms demonstrate the increased influence of the federal government in education policy processes and the growth of a 1Ccoercive federalism 1D that pits the state and federal governments against each other (Harris-Hart, 2010). Central to these initiatives is the measuring, or auditing, of educational practices and relationships. While this shift in education policy hegemony from state to federal governments has been occurring in Australia at least since the 1970s, it has escalated and been transformed in more recent times with a greater emphasis on national human capital agendas which link education and training to Australia 19s international economic competitiveness (Lingard & Sellar, in press). This paper uses historically informed critical analysis to critique claims about the effects of such policies. We argue that performance pay has a detailed and complex historical trajectory both internationally and within Australian states. Using Gale 19s (2001) critical policy historiography, we illuminate some of the effects that performance pay policies have had on education internationally and in particular within Australia. This critical historical lens also provides opportunities to highlight how teachers have, in the past, tactically engaged with such policies.

Soluble mediators in platelet concentrates modulate dendritic cell inflammatory responses in an experimental model of transfusion

The transfusion of platelet concentrates (PCs) is widely used to treat thrombocytopenia and severe trauma. Ex vivo storage of PCs is associated with a storage lesion characterized by partial platelet activation and the release of soluble mediators, such as soluble CD40 ligand (sCD40L), RANTES, and interleukin (IL)-8. An in vitro whole blood culture transfusion model was employed to assess whether mediators present in PC supernatants (PC-SNs) modulated dendritic cell (DC)-specific inflammatory responses (intracellular staining) and the overall inflammatory response (cytometric bead array). Lipopolysaccharide (LPS) was included in parallel cultures to model the impact of PC-SNs on cell responses following toll-like receptor-mediated pathogen recognition. The impact of both the PC dose (10%, 25%) and ex vivo storage period was investigated [day 2 (D2), day 5 (D5), day 7 (D7)]. PC-SNs alone had minimal impact on DC-specific inflammatory responses and the overall inflammatory response. However, in the presence of LPS, exposure to PC-SNs resulted in a significant dose associated suppression of the production of DC IL-12, IL-6, IL-1a, tumor necrosis factor-a (TNF-a), and macrophage inflammatory protein (MIP)-1b and storage-associated suppression of the production of DC IL-10, TNF-a, and IL-8. For the overall inflammatory response, IL-6, TNF-a, MIP-1a, MIP-1b, and inflammatory protein (IP)-10 were significantly suppressed and IL-8, IL-10, and IL-1b significantly increased following exposure to PC-SNs in the presence of LPS. These data suggest that soluble mediators present in PCs significantly suppress DC function and modulate the overall inflammatory response, particularly in the presence of an infectious stimulus. Given the central role of DCs in the initiation and regulation of the immune response, these results suggest that modulation of the DC inflammatory profile is a probable mechanism contributing to transfusion-related complications.

A polymerase chain reaction-based method for isolating clones from a complimentary DNA library in sheep

The sheep (Ovis aries) is favored by many musculoskeletal tissue engineering groups as a large animal model because of its docile temperament and ease of husbandry. The size and weight of sheep are comparable to humans, which allows for the use of implants and fixation devices used in human clinical practice. The construction of a complimentary DNA (cDNA) library can capture the expression of genes in both a tissue- and time-specific manner. cDNA libraries have been a consistent source of gene discovery ever since the technology became commonplace more than three decades ago. Here, we describe the construction of a cDNA library using cells derived from sheep bones based on the pBluescript cDNA kit. Thirty clones were picked at random and sequenced. This led to the identification of a novel gene, C12orf29, which our initial experiments indicate is involved in skeletal biology. We also describe a polymerase chain reaction-based cDNA clone isolation method that allows the isolation of genes of interest from a cDNA library pool. The techniques outlined here can be applied in-house by smaller tissue engineering groups to generate tools for biomolecular research for large preclinical animal studies and highlights the power of standard cDNA library protocols to uncover novel genes.