Grafting of encapsulated BDNF-producing fibroblasts into the injured spinal cord without immune suppression in adult rats.

Grafting of genetically modified cells that express therapeutic products is a promising strategy in spinal cord repair. We have previously grafted BDNF-producing fibroblasts (FB/BDNF) into injured spinal cord of adult rats, but survival of these cells requires a strict protocol of immune suppression with cyclosporin A (CsA). To develop a transplantation strategy without the detrimental effects of CsA, we studied the properties of FB/BDNF that were encapsulated in alginate-poly-L-ornithine, which possesses a semipermeable membrane that allows production and diffusion of a therapeutic product while protecting the cells from the host immune system. Our results show that encapsulated FB/BDNF, placed in culture, can survive, secrete bioactive BDNF and continue to grow for at least one month. Furthermore, encapsulated cells that have been stored in liquid nitrogen retain the ability to grow and express the transgene. Encapsulated FB/BDNF survive for at least one month after grafting into an adult rat cervical spinal cord injury site in the absence of immune suppression. Transgene expression decreased within two weeks after grafting but resumed when the cells were harvested and re-cultured, suggesting that soluble factors originating from the host immune response may contribute to the downregulation. In the presence of capsules that contained FB/BDNF, but not cell-free control capsules, there were many axons and dendrites at the grafting site. We conclude that alginate encapsulation of genetically modified cells may be an effective strategy for delivery of therapeutic products to the injured spinal cord and may provide a permissive environment for host axon growth in the absence of immune suppression.

Contrast agents for diagnostic ultrasound: development and evaluation of polymer-coated microbubbles.

Although the concept of an ultrasound contrast agent dates from Gramiak's work in 1968 in which indocyanine green was injected into the ascending aorta and heart, no universally accepted contrast agent for ultrasound now exists. This is primarily due to problems with stability, size and/or toxicity of the agents which have been investigated. Development of an effective ultrasound contrast agent would be highly significant for the health care industry, since it would greatly expand the scope of ultrasound (a noninvasive and safe procedure) as a diagnostic technique. While encapsulated gas bubbles offer particular advantages in stability over hand-agitated systems, they frequently present problems with size. Capsules larger than 10 microns in diameter become entrapped in the capillary bed of the lung. This paper describes the use of ionotropic gelation of the naturally occurring polysaccharide, alginate, for microencapsulation of air. Two procedures have been investigated. A novel jet head has been developed which allows co-extrusion of a solution of sodium alginate and air to produce nascent microencapsulated air bubbles which fall into a hardening solution of calcium ions. A second method employs ultrasound to introduce cavitation-induced bubbles into the alginate before capsule formation by spraying. Power spectra of these preparations demonstrate echogenicity (that is strong scatter of the incident ultrasound wave back to the emitting transducer, which also acts as a receiver), with resonant peaks that are a function of capsule size and wall characteristics.

The importance of social and cultural effects of mercury on aboriginal peoples.

Environmental contaminants, including mercury, often by-products of industrialization land development projects, frequently have far-reaching consequences for Aboriginal people, who often receive little benefit from such projects. In trying to understand the full impact of environmental mercury on Aboriginal people, therefore, research endeavours must consider the social and cultural impacts, and not simply focus on the direct clinical effects resulting from exposure. This paper explores some of the key areas in developing such an understanding. Aboriginal peoples' understanding of mercury contamination is influenced by their holistic concepts of health and environment. Whether or not scientific assessment reveals direct clinical health effects from exposure to mercury, this holistic viewpoint may lead to an effect on both individuals and communities. Perceptions of changes and the disruption of the special relationship that Aboriginal people perceive themselves to have with the environment have a considerable impact on their social, cultural, spiritual and economic well-being. These qualitative impacts are not easily measured with standard social indicators. The involvement of communities in the identification and measurement of Aboriginal community indicators which reflect the reality of their situation is essential in order to establish a better understanding of the extent and importance of social and cultural effects of exposure to mercury. Toxicologists, policy makers and others concerned with setting risk levels which may lead to formal guidelines should be aware of the impacts that their actions and decisions may have on the lifestyle, and therefore on the health, of Aboriginal peoples.

Carcinoma of the laryngeal glottis therapy and end results.

The end results of 264 patients with vocal cord carcinoma treated by either surgery or radiation therapy at the University of Illinois from 1955 to 1972 are analyzed. Stage I glottic carcinoma can be cured by either surgery or radiation alone (86 percent and 80 percent). Stage II glottic cancer treated by surgery has achieved survival rates of 70 percent. The five-year end results of Stage III glottic cancer are 49 percent for surgery and 30 percent for radiation. The management of glottic carcinoma, according to anatomic site and indications for various modalities are discussed.

Influence of environmental conditions on a new surfactant-based contrast agent: ST68.

Environmental influences on the new surfactant-stabilized bubbles, ST68, were investigated. We have developed a new surfactant-based contrast agent ST68, which is prepared by insonating buffered mixtures of Span 60 and Tween 80 in the presence of either air, PFC, or SF(6) gas. The effect of dilution, shear, and sonication on size distribution of ST68 showed that PFC-containing bubbles (ST68-PFC) were most stable. ST68-PFC bubbles lasted more than 15 min with approximately 30 dB backscatter enhancement in degassed phosphate-buffered saline, (pH 7.4), and air bubbles lasted approximately 3 s, suggesting the effects of diffusion. Additionally, it was found that the ionic strength of the suspending medium (for example, PBS), did not have any effect on ST68 bubbles containing SF(6) or PFC, but had a dramatic impact on bubbles containing air.

Quantitative acoustic characterization of a new surfactant-based ultrasound contrast agent.

The acoustic properties of a new ultrasound contrast agent, ST68, have been investigated. ST68 is a sonicated mixture of nonionic surfactants (Span-type and Tween-type) consisting of stabilized microbubbles with a mean diameter of 3.8 microns and a concentrations of 7.1 x 10(8) bubbles/mL. A pulsatile flow system was used to acquire data in vitro. The acoustic properties of ST68, as a function of time, frequency and dose, were calculated. Enhancement changed nonlinearly with contrast agent dose; maximum was 13.1 dB +/- 1.0 dB for a dose of 0.30 microL/mL of suspending medium. Attenuation reached approximately 11 dB/cm for dosages above 0.27 microL/mL and for frequencies between 2.5 and 6.0 MHz. In vivo, i.v. injections of ST68 were given to 4 rabbits (doses from 0.01 to 0.23 mL/kg). A clear increase in flow signal intensity was observed for 1 to 2 min. An in vivo dose-response curve was calculated from audio Doppler signals obtained with a 10-MHz cuff transducer placed around the distal aorta. Maximum enhancement was 18.3 dB +/- 3.13 dB for a 0.13 mL/kg dose. Moreover, ST68 appears to follow a simple relationship between in vivo enhancement and dose. In conclusion, ST68 is capable of producing marked vascular enhancement. Its acoustic properties have been characterized in vitro and in vivo.

Effect of filling gases on the backscatter from contrast microbubbles: theory and in vivo measurements.

Two surfactant-based contrast agents, ST44 and ST68, were produced according to US Patent # 5,352,436 and filled with either air, C4F10 (perfluorobutane) or SF6 (sulfur hexaflouride). Ten rabbits received i.v. injections of each agent/gas combination with 5 repetitions of each dose (range: 0.005-0.13 mL/kg). A custom-made 10-MHz cuff transducer was placed around the surgically exposed distal aorta and audio Doppler signals were acquired in vivo. Quantitative in vivo dose responses were calculated off-line using spectral power analysis and compared to a theoretical model of microbubble dissolution and enhancement. For qualitative comparisons, 10 rabbits were imaged pre- and postcontrast administration (dose: 0.1 mL/kg) in gray-scale and colour. All agent/gas combinations produced marked Doppler enhancement with air bubbles enhancing least of all (p < 0.0001) and ST68-SF6 best of all (maximum: 27.6 +/- 2.04 dB; p < 0.012). There were no significant differences between other agent/gas combinations (0.30 < p < 0.70). Theoretical enhancement was within 1 order of magnitude of the experimental observations (i.e., deviations of up to 10 dB). The duration of contrast enhancement was 1-2 min for air-filled bubbles, 3-5 min for SF6-filled bubbles and more than 7 min for C4F10-filled bubbles. In conclusion, ST68-SF6 microbubbles produced most in vivo enhancement of the agent/gas combinations studied. Theory matched the measurements within an order of magnitude.

Methylmercury and the health of indigenous peoples: a risk management challenge for physical and social sciences and for public health policy.

Methylmercury in aquatic ecosystems and bio-accumulated in aquatic biota, especially fish, is a major public health concern internationally. Precautionary efforts are currently underway internationally to reduce the anthropogenic release of mercury, which in turn, over time, will reduce human exposure. However, at the present time, it is important to address the issue of management of the risks of exposure as they exist now. Of particular concern are the impacts of methylmercury on indigenous populations which depend on fish as a subsistence food source, both in remote areas of developed countries, such as Canada, and in developing countries such as Brazil. Research into these impacts over the past two or three decades has shown that, other than in very severe pollution situations such as occurred in Minamata, Japan, the direct impacts on human health are difficult to prove. On the other hand, the indirect negative effects of methylmercury on health, mediated through the disruption of lifestyle and eating patterns and the associated socio-cultural and socio-economic consequences among the affected native populations, have, in many cases, been significant. These social factors have raised serious challenges in determining practical public health policies on the issue. Policy development relating to environmental contaminants has been presented, with the problem of assessing the role of the various factors which contribute to the impact on health as a result of socio-cultural disruption. These factors include changes in diet and lifestyle due to methylmercury in the environment and its real or perceived risk. The standard physical sciences risk assessment process, based on the lowest observed adverse effects level (LOAEL) or no observed adverse effects level (NOAEL) used in defining health policies may be seen as over-simplistic theoretical extrapolations when viewed in the context of the concerns of the social sciences. Both approaches, however, have relevance to health policies that address the risks posed by environmental methylmercury. Therefore, the standard physical sciences approach of the past three decades now needs to be linked with the social sciences approach, with its focus on the indirect impacts of exposure to methylmercury, to provide a comprehensive approach to public health policy development. With this objective in mind, this paper reviews methylmercury-related data from both physical and social sciences. It attempts to draw on the findings in both disciplines to provide suggestions for an integrated approach in policy development relating to human health and human exposure to methylmercury, especially among indigenous peoples in remote areas and in developing countries. An integrated approach such as this may help to limit adverse health effects in the indigenous communities affected.

Radiation failures in cancer of the larynx.

Overwhelming statistics have been published regarding the success of radiation therapy in the management of laryngeal carcinoma. The fate of those patients whose cancers are not controlled by radiotherapy is often left to speculation. An analysis is made of 61 patients with early and advanced laryngeal cancer treated initially with radiation therapy and subsequently operated upon because of recurrent or persistent tumor. Frequently, diagnosis of recurrence is difficult or delayed. Preservation of voice can only be accomplished when recurrence of T1 and T2 lesions is detected early and strict criteria are followed. Conservation surgery is not feasible for early lesions which progress after radiation failure. Advanced T2 and T4 lesions which become radiation failures also require laryngectomy for salvage. In cancer of the larynx, there is a definite, identifiable group of patients in whom surgery would be advisable as the initial therapy.

Generation of ultraharmonics in surfactant based ultrasound contrast agents: use and advantages.

A unique distinction between surfactant stabilized ultrasound contrast agent ST68 and water (or tissue), is the enhanced ability of the agent to generate non-linear frequencies such as sub-harmonics (f0/2), higher harmonics (2fo, 3fo, 4fo,...), and ultraharmonics (3f0/2, Sf0/2, 7f0/2,...), when insonated with fundamental frequency f0. Currently, second harmonics (2f0) have been predominantly researched, to exploit the diagnostic benefits of the contrast-specific non-linear imaging. However, we found that at normal imaging pressures (100 kPa-1 MPa), ST68 agent-generated second harmonic enhancements dropped to approximately 8 dB at 100 kPa and approximately 2 dB at 1 MPa. Moreover, at these pressures water (or tissue) produced strong second harmonics due to non-linear propagation. Ultraharmonics and sub-harmonics on the other hand, were generated only by the agent, and were not produced due to the non-linear propagation of ultrasound in either water or tissue. Additionally, ultraharmonic (3f0/2) enhancements of approximately 23 dB at 100 kPa, approximately 35 dB at 0.5 MPa and approximately 41dB at 1.1 MPa for ST68-PFC, offer much greater signal to noise ratio than higher harmonics.

Social and cultural impacts of environmental change on aboriginal Peoples in Canada.

Environmental change, often the result of Western industrial development, has had a major impact on Canadian Aboriginal people. Even when there are no provable direct health effects, Aboriginal peoples' holistic concepts of health, balance, and harmony, and the interrelatedness of health and environment lead them to regard social and cultural effects as health effects. This paper explores the links between environmental change and the social and cultural and, hence, health effects these changes produce. It is argued that factors such as Aboriginal holistic concepts of environment and health, perceptions of risk, and difficulties in communication contribute to these social and cultural effects and their subsequent health effects-effects which frequently present a greater problem in Canadian Aboriginal communities than do the direct health effects of environmental change.

Effects of pediatric head trauma for children, parents, and families.

Severe pediatric head injury has negative consequences for children of all ages. Even mild and moderate head injury results in residual impairment for school-age children and adolescents. Data are needed on the effects of these less severe insults, especially for preschoolers. Although research on the impact of the child's head injury on the parent-child relationship and family functioning is limited, the experience is likely to be very stressful for the parent and the family. Indeed, family integrity may be at risk. Research is needed that examines the effects of a child's head injury for the parent and the family over time and identifies factors related to these outcomes.

Delivery of encapsulated Doxorubicin by ultrasound-mediated size reduction of drug-loaded polymer contrast agents.

Low delivery efficiency combined with systemic toxicity of traditional chemotherapy provides a need for improved chemotherapeutic delivery. Within our laboratory, we have developed polymer ultrasound contrast agents (1.2-1.8 mum in diameter) containing doxorubicin (Dox) within the shell (100-150 nm). In vivo this platform is expected to circulate through the vasculature until activated at the tumor site with external focused ultrasound (US). In vitro, the agent is responsive to US and when insonated at peak positive pressure amplitudes of 0.69 MPa and above, shows dramatic size reduction, eventually reaching a mean particle size of 350 nm, presumably due to fragmentation of, or gas release from the agent. The resulting Dox-polymer particles retain the drug and are small enough to pass through the leaky pores (350-400 nm) within the tumor vasculature, providing a sustained intratumoral release of chemotherapeutic as the polymer degrades. In vivo studies using a VX2 liver tumor model have shown that the combination of the agent and US results in nearly 50% less drug delivered to the nontargeted, healthy liver ( p = 0.009) and a 110% increase ( p = 0.004) in Dox delivery to the viable peripheral tissue of the tumor, relative to the uninsonated controls. This study shows how US-mediated destruction of drug-loaded polymer contrast agent can be used to deliver encapsulated drug for potential sustained release. Penetration mechanisms of these resulting particles and their ability to provide a sustained release from the tumor interstia will be explored in the future.

Development and optimization of a doxorubicin loaded poly(lactic acid) contrast agent for ultrasound directed drug delivery.

An echogenic, intravenous drug delivery platform is proposed in which an encapsulated chemotherapeutic can travel to a desired location and drug delivery can be triggered using external, focused ultrasound at the area of interest. Three methods of loading poly(lactic acid) (PLA) shelled ultrasound contrast agents (UCA) with doxorubicin are presented. Effects on encapsulation efficiency, in vitro enhancement, stability, particle size, morphology and release during UCA rupture are compared by loading method and drug concentration. An agent containing doxorubicin within the shell was selected as an ideal candidate for future hepatocellular carcinoma studies. The agent achieved a maximal drug load of 6.2 mg Dox/g PLA with an encapsulation efficiency of 20.5%, showed a smooth surface morphology and tight size distribution (poly dispersity index=0.309) with a peak size of 1865 nm. Acoustically, the agent provided 19 dB of enhancement in vitro at a dosage of 10 microg/ml, with a half life of over 15 min. In vivo, the agent provided ultrasound enhancement of 13.4+/-1.6 dB within the ascending aorta of New Zealand rabbits at a dose of 0.15 ml/kg. While the drug-incorporated agent is thought to be well suited for future drug delivery experiments, this study has shown that agent properties can be tailored for specific applications based on choice of drug loading method.

Ultrasound triggered cell death in vitro with doxorubicin loaded poly lactic-acid contrast agents.

Traditional chemotherapy generally results in systemic toxicity, which also limits drug levels at the area of need. Two ultrasound contrast agents (UCA), with diameters between 1-2 microm in diameter and shell thicknesses of 100-200 nm, composed of poly lactic-acid (PLA), one loaded by surface adsorption and the other loaded by drug incorporation in the shell, were compared in vitro for potential use in cancer therapy. These poly lactic-acid (PLA) UCA platforms contain a gas core that in an ultrasound (US) field can cause the UCA to oscillate or rupture. Following a systemic injection of drug loaded UCA with external application of US focused at the area of interest, this platform could potentially increase drug toxicity at the area of need, while protecting healthy tissue through microencapsulation of the drug. In vitro toxicity in MDA-MB-231 breast cancer cells of the surface-adsorbed and shell-incorporated doxorubicin (Dox) loaded UCA were examined at 5 MHz insonation using a pulse repetition frequency of 100 Hz at varying pressure amplitudes. Both platforms resulted in equivalent cell death compared to free Dox and US when insonated at peak positive pressure amplitudes of 1.26 MPa and above. While no significant changes in cell death were seen for surface adsorbed Dox-UCA with or without insonation, cell death using the platform with Dox incorporated within the shell increased from 16.12% to 25.78% (p=0.0272), approaching double the potency of the platform when insonated at peak positive pressure amplitudes of 1.26 MPa and above. This mechanism is believed to be the result of UCA rupture at higher insonation pressure amplitudes, resulting in more exposed drug and shell surface area as well as increased cellular uptake of Dox containing polymer shell fragments. This study has shown that a polymer UCA with drug housed within the shell may be used for US-triggered cell death. US activation can be used to make a carrier significantly more potent once in the area of need.

Developing an integrated traditional/clinical health system in the Yukon.

The introductory steps have been taken in Yukon. Elders have met and voiced their concerns and initial contacts have been made with government officials, medical and legal consultants. A proposal is now being developed to obtain funding to design a suitable model for an integrated Yukon Indian/clinical health care system. One of the next steps, following on the advice of the Elders, should be for communities to establish their own projects to record the plants and practices used, with the assistance of their Elders. The communities should also identify people who use traditional practices who are willing to come forward. They could then come together with the Elders to discuss their concerns. Beyond that, representatives from the traditional system will need to meet with representatives of the mainstream system, to discuss areas of co-operation. Once the "content" has been identified, the model for integrating the two health systems can be addressed. This will necessitate further meetings of Yukon Territorial Government officials, legal advisors, medical advisors, and Yukon First Nations representatives. The proposal currently being developed will build on the initial steps which have been taken. The Yukon Territorial Government has indicated a willingness to look at ways of including traditional health care practices for patients who wish to use them. The receptivity of government and Yukon medical profession and the expressed concerns of the Elders indicate that now is the time to proceed.

Degradation of polysaccharides by endo- and exoenzymes: dextran-dextranase model systems.

Experiments were carried out on dextran-dextranase systems to test the prediction of a mechanistic model recently proposed by us, for the synergistic effect of combined exo/endo enzymic action in the degradation of polymeric substrates. Soluble forms of the substrate were used. Preliminary experiments with an insoluble form of the substrate were also carried out to demonstrate the applicability of the analytical techniques to these cases. Molecular weight distributions of the degradation products were determined (by gel-permeation chromatography) and the rates of production of glucose and of other reducing sugars were also measured. It was found that the exodextranase alone had very little effect on the molecular weight distributions compared to a significant shift towards lower molecular weights obtained with the endodextranase which was synergistically enhanced by the action of the combined enzymes. Glucose was produced more rapidly by the exoenzyme compared to the endoenzyme, but combinations of the two enzymes gave a rate enhancement greater than the linear sum of the effects of the two individual enzymes. In comparing the degradation indices and polydispersities of the various degradation products, similar synergistic effects of the combined enzymes in accordance with the theoretical predictions, were observed. The practical implications of these findings to the design of fermentation processes which depend on the action of endo- and exoenzyme mixtures are noted.