Identification of bone morphogenetic proteins 2 and 4 in commercial demineralized freeze-dried bone allograft preparations : pilot study

BACKGROUND: Demineralized freeze-dried bone allografts (DFDBAs) have been proposed as a useful adjunct in periodontal therapy to induce periodontal regeneration through the induction of new bone formation. The presence of bone morphogenetic proteins (BMPs) within the demineralized matrix has been proposed as a possible mechanism through which DFDBA may exert its biologic effect. However, in recent years, the predictability of results using DFDBA has been variable and has led to its use being questioned. One reason for the variability in tissue response may be attributed to differences in the processing of DFDBA, which may lead to loss of activity of any bioactive substances within the DFDBA matrix. Therefore, the purpose of this investigation was to determine whether there are detectable levels of bone morphogenetic proteins in commercial DFDBA preparations. METHODS: A single preparation of DFDBA was obtained from three commercial sources. Each preparation was studied in triplicate. Proteins within the DFDBA samples were first extracted with 4M guanidinium HCI for seven days at 40 degrees celsius and the residue was further extracted with 4M guanidinium HCL/EDTA for seven days at 40 degrees celsius. Two anti-human BMP-2 and -4 antibodies were used for the detection of the presence of BMP's in the extracts. RESULTS: Neither BMP-2 nor BMP-4 was detected in any of the extracts. When recombinant human BMP-2 and -4 were added throughout the extraction process of DFDBA extraction, not only were intact proteins detected but smaller molecular weight fragments were also noted in the extract. CONCLUSIONS: These results indicate that all of the DFDBA samples tested had no detectable amounts of BMP-2 and -4. In addition, an unknown substance present in the DFDBA may be responsible for degradation of whatever BMPs might be present.

The potential for production of freeze-dried oral vaccines using alginate hydrogel microspheres as protein carriers

Oral administration of dry vaccine formulations is acknowledged to offer major clinical and logistical benefits by eliminating the cold chain required for liquid preparations. A model antigen, bovine serum albumin (BSA) was encapsulated in alginate microspheres using aerosolisation. Hydrated microspheres 25 to 65 μm in size with protein loading of 3.3 % w/w were obtained. Environmental scanning electron microscopy indicated a stabilizing effect of encapsulated protein on alginate hydrogels revealed by an increase in dehydration resistance. Freeze drying of alginate microspheres without use of a cryoprotectant resulted in fragmentation and subsequent rapid loss of the majority of the protein load in simulated intestinal fluid in 2 h, whereas intact microspheres were observed following freeze-drying of BSA-loaded microspheres in the presence of maltodextrin. BSA release from freeze-dried preparations was limited to less than 7 % in simulated gastric fluid over 2 h, while 90 % of the protein load was gradually released in simulated intestinal fluid over 10 h. SDS-PAGE analysis indicated that released BSA largely preserved its molecular weight. These findings demonstrate the potential for manufacturing freeze-dried oral vaccines using alginate microspheres.

Interfacial microstructure in Y1Ba2Cu3O7-δ high Tc superconductors

The microstructures of YBa2Cu3O7-δ ceramics prepared from freeze dried powders and containing an excess of CuO have been studied by analytical electron microscopy. Special attention has been paid to the interfacial microstructure. It was found that a liquid phase formed during sintering between 890°C and 920°C and this promoted grain growth and densification. Both clean grain boundaries and boundaries containing an amorphous intergranular film, which was rich in Cu, have been observed. Both CuO and BaCuO2 were present as secondary phases.

A critical analysis of sperm donation practices : the personal and social effects of disrupting the unity of biological and social relatedness for the offspring

This thesis critically analyses sperm donation practices from a child-centred perspective. It examines the effects, both personal and social, of disrupting the unity of biological and social relatedness in families affected by donor conception. It examines how disruption is facilitated by a process of mediation which is detailed using a model provided by Sunderland (2002). This model identifies mediating movements - alienation, translation, re-contextualisation and absorption - which help to explain the powerful and dominating material, and social and political processes which occur in biotechnology, or in reproductive technology in this case. The understanding of such movements and mediation of meanings is inspired by the complementary work of Silverstone (1999) and Sunderland. This model allows for a more critical appreciation of the movement of meaning from previously inalienable aspects of life to alienable products through biotechnology (Sunderland, 2002). Once this mediation in donor conception is subjected to critical examination here, it is then approached from different angles of investigation. The thesis posits that two conflicting notions of the self are being applied to fertility-frustrated adults and the offspring of reproductive interventions. Adults using reproductive interventions receive support to maximise their genetic continuity, but in so doing they create and dismiss the corresponding genetic discontinuity produced for the offspring. The offspring’s kinship and identity are then framed through an experimental postmodernist notion, presenting them as social rather than innate constructs. The adults using the reproductive intervention, on the other hand, have their identity and kinship continuity framed and supported as normative, innate, and based on genetic connection. This use of shifting frameworks is presented as unjust and harmful, creating double standards and a corrosion of kinship values, connection and intelligibility between generations; indeed, it is put forward as adult-centric. The analysis of other forms of human kinship dislocation provided by this thesis explores an under-utilised resource which is used to counter the commonly held opinion that any disruption of social and genetic relatedness for donor offspring is insignificant. The experiences of adoption and the stolen generations are used to inform understanding of the personal and social effects of such kinship disruption and potential reunion for donor offspring. These examples, along with laws governing international human rights, further strengthen the appeal here for normative principles and protections based on collective knowledge and standards to be applied to children of reproductive technology. The thesis presents the argument that the framing and regulation of reproductive technology is excessively influenced by industry providers and users. The interests of these parties collide with and corrode any accurate assessments and protections afforded to the children of reproductive technology. The thesis seeks to counter such encroachments and concludes by presenting these protections, frameworks, and human experiences as resources which can help to address the problems created for the offspring of such reproductive interventions, thereby illustrating why these reproductive interventions should be discontinued.

The effect of rhBMP-2 on canine osteoblasts seeded onto 3D bioactive polycaprolactone scaffolds

Our strategy entails investigating the influence of varied concentrations (0, 10, 100 and 1000 ng/ml) of human recombinant bone morphogenetic protein-2 (rhBMP-2) on the osteogenic expression of canine osteoblasts, seeded onto poly-caprolactone 20% tricalcium phosphate (PCL-TCP) scaffolds in vitro. Biochemical assay revealed that groups with rhBMP-2 displayed an initial burst in cell growth that was not dose-dependent. However, after 13 days, cell growth declined to a value similar to control. Significantly less cell growth was observed for construct with 1000 ng/ml of rhBMP-2 from 20 days onwards. Confocal microscopy confirmed viability of osteoblasts and at day 20, groups seeded with rhBMP-2 displayed heightened cell death as compared to control. Phase contrast and scanning electron microscopy revealed that osteoblasts heavily colonized surfaces, rods and pores of the PCL-TCP scaffolds. This was consistent for all groups. Finally, Von Kossa and osteocalcin assays demonstrated that cells from all groups maintained their osteogenic phenotype throughout the experiment. Calcification was observed as early as four days after stimulation for groups seeded with rhBMP-2. In conclusion, rhBMP-2 seems to enhance the differentiated function of canine osteoblasts in a non-dose dependent manner. This resulted in accelerated mineralization, followed by death of osteoblasts as they underwent terminal differentiation. Notably, PCL-TCP scaffolds seeded only with canine osteoblasts could sustain excellent osteogenic expression in vitro. Hence, the synergy of PCL with bioactive TCP and rhBMP-2 in a novel composite scaffold, could offer an exciting approach for bone regeneration.

Immunohistochemical detection of survivin in canine lymphoma

Survivin is a member of the family of proteins known as 'inhibitors of apoptosis proteins'. Survivin has a role in cellular decisions concerning division and survival and is frequently expressed in neoplastic cells. The aim of the present study was to investigate immunohistochemically the expression of survivin in normal canine tissues and in canine lymphoma. A representative range of fetal and adult normal tissues as well as biopsy samples from dogs with lymphoma were assembled in tissue arrays. The lymphomas were classified according to the revised Kiel and to the Revised European American Lymphoma - World Health Organization (REAL-WHO) schemes. Polyclonal and monoclonal antisera cross-reactive with canine survivin identified cytoplasmic expression of the molecule in a broad range of normal canine cells. The same reagents demonstrated cytoplasmic labelling of more than 5% of cells in all 83 lymphoma samples tested with polyclonal antiserum and in 67 of 82 (82%) of samples tested with monoclonal antiserum. Survivin was expressed by a wide range of canine lymphoma subtypes, but the expression of this molecule in normal canine tissues must be considered if novel therapies targeting survivin are applied to the management of canine lymphoma. © 2010 Elsevier Ltd.

A protocol for the rapid isolation of full geminivirus genomes from dried plant tissue

A high-throughput method of isolating and cloning geminivirus genomes from dried plant material, by combining an Extract-n-Amp™-based DNA isolation technique with rolling circle amplification (RCA) of viral DNA, is presented. Using this method an attempt was made to isolate and clone full geminivirus genomes/genome components from 102 plant samples, including dried leaves stored at room temperature for between 6 months and 10 years, with an average hands-on-time to RCA-ready DNA of 15 min per 20 samples. While storage of dried leaves for up to 6 months did not appreciably decrease cloning success rates relative to those achieved with fresh samples, efficiency of the method decreased with increasing storage time. However, it was still possible to clone virus genomes from 47% of 10-year-old samples. To illustrate the utility of this simple method for high-throughput geminivirus diversity studies, six Maize streak virus genomes, an Abutilon mosaic virus DNA-B component and the DNA-A component of a previously unidentified New Word begomovirus species were fully sequenced. Genomic clones of the 69 other viruses were verified as such by end sequencing. This method should be extremely useful for the study of any circular DNA plant viruses with genome component lengths smaller than the maximum size amplifiable by RCA. © 2008 Elsevier B.V. All rights reserved.

Uniform Civil Procedure Rules r150 : orders relating to "property" – whether sperm extracted and stored may be characterised as “property”

In Bazley v Wesley Monash IVF Pty Ltd [2010] QSC 118 an order was made under r 250 of the Uniform Civil Procedure Rules 1999 (Qld) (“UCPR”) requiring the respondent to continue to hold and maintain straws of semen belonging to the applicant’s deceased husband. The decision includes a useful analysis of the development of the common law regarding property rights in human bodies and body parts.

Effect of moisture and temperature distribution on dried food microstructure and porosity

Dehydration of food materials requires water removal from it. This removal of moisture prevents the growth and reproduction of microorganisms that cause decay and minimizes many of the moisture-driven deterioration reactions (Brennan, 1994). However, during food drying, many other changes occur simultaneously resulting in a modified overall quality (Kompany et al., 1993). Among the physical attributes of dried food material porosity and microstructure are the important ones that can dominant other quality of dried foods (Aguilera et al., 2000). In addition, this two concerned quality attributes affected by process conditions, material components and raw structure of food stuff. In this work, temperature moisture distribution within food materials during microwave drying will be taken into consideration to observe its participation on the microstructure and porosity of the finished product. Apple is the selective materials for this work. Generally, most of the food materials are found in non-uniformed moisture contained condition. To develop non uniform temperature distribution, food materials have been dried in a microwave oven with different power levels (Chua et al., 2000). First of all, temperature and moisture model is simulated by COMSOL Multiphysics. Later on, digital imaging camera and Image Pro Premier software have been deployed to observation moisture distribution and thermal imaging camera for temperature distribution. Finally, Microstructure and porosity of the food materials are obtained from scanning electron microscope and porosity measuring devices respectively . Moisture distribution and temperature during drying influence the microstructure and porosity significantly. Specially, High temperature and moisture contained regions show less porosity and more rupture. These findings support other literatures of Halder et al. (2011) and Rahman et al (1990). On the other hand, low temperature and moisture regions depict uniform microstructure and high porosity. This work therefore assists in better understanding of the role of moisture and temperature distribution to a prediction of micro structure and porosity of dried food materials.