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PubMed2002

A tough biodegradable elastomer.

Biodegradable polymers have significant potential in biotechnology and bioengineering. However, for some applications, they are limited by their inferior mechanical properties and unsatisfactory compatibility with cells and tissues. A strong, biodegradable, and biocompatible elastomer could be useful for fields such as tissue engineering, drug delivery, and in vivo sensing. We designed, synthesized, and characterized a tough biodegradable elastomer from biocompatible monomers. This elastomer forms a covalently crosslinked, three-dimensional network of random coils with hydroxyl groups attached to its backbone. Both crosslinking and the hydrogen-bonding interactions between the hydroxyl groups likely contribute to the unique properties of the elastomer. In vitro and in vivo studies show that the polymer has good biocompatibility. Polymer implants under animal skin are absorbed completely within 60 days with restoration of the implantation sites to their normal architecture.

3T3 Cells↗
PubMed2002

Generation of histocompatible tissues using nuclear transplantation.

Nuclear transplantation (therapeutic cloning) could theoretically provide a limitless source of cells for regenerative therapy. Although the cloned cells would carry the nuclear genome of the patient, the presence of mitochondria inherited from the recipient oocyte raises questions about the histocompatibility of the resulting cells. In this study, we created bioengineered tissues from cardiac, skeletal muscle, and renal cells cloned from adult bovine fibroblasts. Long-term viability was demonstrated after transplantation of the grafts into the nuclear donor animals. Reverse transcription-PCR (RT-PCR) and western blot analysis confirmed that the cloned tissues expressed tissue-specific mRNA and proteins while expressing a different mitochondrial DNA (mtDNA) haplotype. In addition to creating skeletal muscle and cardiac "patches", nuclear transplantation was used to generate functioning renal units that produced urinelike fluid and demonstrated unidirectional secretion and concentration of urea nitrogen and creatinine. Examination of the explanted renal devices revealed formation of organized glomeruli- and tubule-like structures. Delayed-type hypersensitivity (DTH) testing in vivo and Elispot analysis in vitro suggested that there was no rejection response to the cloned renal cells. The ability to generate histocompatible cells using cloning techniques addresses one of the major challenges in transplantation medicine.

Animals↗
PubMed2003

Sialylated endogenous glycoconjugates in plant cells.

Bioengineered plants are emerging as promising systems for the production of therapeutically valuable proteins. It has been commonly accepted that plants do not perform mammalian-like post-translational modifications, particularly sialylation of glycoconjugates, and no evidence has previously been reported to suggest that they have such capabilities. Here we report the presence of sialylated glycoconjugates in suspension-cultured cells of Arabidopsis thaliana and suggest that a genetic and enzymatic basis for sialylation exists in plants.

Arabidopsis↗
PubMed2006

Sweet spots in functional glycomics.

Information contained in the mammalian glycome is decoded by glycan-binding proteins (GBPs) that mediate diverse functions including host-pathogen interactions, cell trafficking and transmembrane signaling. Although information on the biological roles of GBPs is rapidly expanding, challenges remain in identifying the glycan ligands and their impact on GBP function. Protein-glycan interactions are typically low affinity, requiring multivalent interactions to achieve a biological effect. Though many glycoproteins can carry the glycan structure recognized by the GBP, other factors, such as recognition of protein epitopes and microdomain localization, may restrict which glycoproteins are functional ligands in situ. Recent advances in development of glycan arrays, synthesis of multivalent glycan ligands, bioengineering of cell-surface glycans and glycomics databases are providing new tools to identify the ligands of GBPs and to elucidate the mechanisms by which they participate in GBP function.

Biomedical Engineering↗
PubMed2006

Biosynthesis of Dictyostelium discoideum differentiation-inducing factor by a hybrid type I fatty acid-type III polyketide synthase.

Differentiation-inducing factors (DIFs) are well known to modulate formation of distinct communal cell types from identical Dictyostelium discoideum amoebas, but DIF biosynthesis remains obscure. We report complimentary in vivo and in vitro experiments identifying one of two approximately 3,000-residue D. discoideum proteins, termed 'steely', as responsible for biosynthesis of the DIF acylphloroglucinol scaffold. Steely proteins possess six catalytic domains homologous to metazoan type I fatty acid synthases (FASs) but feature an iterative type III polyketide synthase (PKS) in place of the expected FAS C-terminal thioesterase used to off load fatty acid products. This new domain arrangement likely facilitates covalent transfer of steely N-terminal acyl products directly to the C-terminal type III PKS active sites, which catalyze both iterative polyketide extension and cyclization. The crystal structure of a steely C-terminal domain confirms conservation of the homodimeric type III PKS fold. These findings suggest new bioengineering strategies for expanding the scope of fatty acid and polyketide biosynthesis.

Acyltransferases↗
PubMed2005

Technology insight: Applications of tissue engineering and biological substitutes in urology.

Patients suffering from diseased or injured organs may be treated with transplanted organs. However, there is a severe shortage of donor organs, which is worsening yearly owing to the ageing population. Scientists in the field of regenerative medicine and tissue engineering apply the principles of cell transplantation, materials science, and bioengineering to construct biological substitutes that will restore and maintain normal function in diseased and injured tissues. This article reviews recent advances in regenerative medicine and describes applications of biological substitutes that may offer novel therapies for patients with end-stage organ failure.

Bioartificial Organs↗
PubMed2006

Redox-controlled molecular permeability of composite-wall microcapsules.

Many smart materials in bioengineering, nanotechnology and medicine allow the storage and release of encapsulated drugs on demand at a specific location by an external stimulus. Owing to their versatility in material selection, polyelectrolyte multilayers are very promising systems in the development of microencapsulation technologies with permeation control governed by variations in the environmental conditions. Here, organometallic polyelectrolyte multilayer capsules, composed of polyanions and polycations of poly(ferrocenylsilane) (PFS), are introduced. Their preparation involved layer-by-layer self-assembly onto colloidal templates followed by core removal. PFS polyelectrolytes feature redox-active ferrocene units in the main chain. Incorporation of PFS into the capsule walls allowed us to explore the effects of a new stimulus, that is, changing the redox state, on capsule wall permeability. The permeability of these capsules could be sensitively tuned via chemical oxidation, resulting in a fast capsule expansion accompanied by a drastic permeability increase in response to a very small trigger. The substantial swelling could be suppressed by the application of an additional coating bearing common redox-inert species of poly(styrene sulfonate) (PSS(-)) and poly(allylamine hydrochloride) (PAH(+)) on the outer wall of the capsules. Hence, we obtained a unique capsule system with redox-controlled permeability and swellability with a high application potential in materials as well as in bioscience.

Capsules↗
PubMed2002

The influence of genetics on contemporary art.

Contemporary visual artists are incorporating genetic concepts into their work, and this work has become prominently featured in numerous museum and gallery exhibitions. Such art uses visual images that represent the language of genomics, the values affected by genetic understanding of the body and the implications of bioengineering. Here, we present various examples of how artists depict aspects of genetics as cultural icons and symbols; in particular, their focus on DNA as information and on the commercialization of genetics research material.

Art↗
PubMed2006

The biosynthesis and regulation of bacterial prodiginines.

The red-pigmented prodiginines are bioactive secondary metabolites produced by both Gram-negative and Gram-positive bacteria. Recently, these tripyrrole molecules have received renewed attention owing to reported immunosuppressive and anticancer properties. The enzymes involved in the biosynthetic pathways for the production of two of these molecules, prodigiosin and undecylprodigiosin, are now known. However, the biochemistry of some of the reactions is still poorly understood. The physiology and regulation of prodiginine production in Serratia and Streptomyces are now well understood, although the biological role of these pigments in the producer organisms remains unclear. However, research into the biology of pigment production will stimulate interest in the bioengineering of strains to synthesize useful prodiginine derivatives.

Antineoplastic Agents↗
PubMed2005

Effect of Herceptin on the development and progression of skeletal metastases in a xenograft model of human breast cancer.

We examined the effects of Herceptin, a bioengineered monoclonal antibody directed against Her-2/neu oncogene on skeletal metastasis using a xenograft model of breast cancer. Treatment of Her-2 overexpressing human breast cancer cells BT-474 with Herceptin caused a dose-dependent decrease in cell proliferation. In in vivo studies, BT-474 cells (1 x 10(5)) were injected into the left ventricle of female BALB/c nu/nu mice. Intraperitoneal (i.p.) infusion of Herceptin (1 mg/kg twice a week for 5 weeks) from the day of tumor cell inoculation or at the time of radiologically detectable skeletal metastasis either slowed the development or prevented the progression of skeletal metastasis as compared to control groups of animals receiving nonspecific IgG. Bone histological analysis of long bones showed the ability of Herceptin to reduce the ratio of tumor volume to bone volume as well as mitotic index, effects that were more pronounced when Herceptin treatment was initiated from the day of tumor cell inoculation. While immunohistochemical analysis of long bones showed no difference in the production of Her-2, phosphorylated (P) Her-2 and MAPK, a significantly lower level of P-MAPK was seen in bones of Herceptin treated animals. These studies demonstrate the ability of Herceptin to inhibit the development and abrogate the progression of skeletal metastases associated with breast cancer by blocking the HER-2-mediated signaling pathways.

Animals↗
PubMed2000

Paraplegic adaptation to assisted-walking: energy expenditure during wheelchair versus orthosis use.

STUDY DESIGN: To study the energy cost of locomotion during ambulation with different orthoses (HIP Guidance Orthosis Orlau Parawalker (PW), n=4; Reciprocating Gait Orthosis (RGO), n=6; RGO + FNS, n=4). OBJECTIVES: Since high energy costs of locomotion have been proposed as a major reason for early rejection of orthotic use, our aims were (a) to evaluate the impact of functional neuromuscular stimulation (FNS) on energy expenditure during orthosis-assisted ambulation; (b) to study whether energy expenditure data can predict the poor long-term patients' compliance and (c) to assess selection criteria for the assignment of the different types of orthosis. SETTING: The study was completed at the Center of Sports Medicine and Center of Bioengineering, don C. Gnocchi Foundation, IRCCS, Milan, Italy. METHODS: The HR/VO2 relationship and the energy cost of locomotion (C) were studied in 14 patients (lesion level C7-T11) during wheelchair (WHCH) use at various speeds and during locomotion with different orthotic devices. Patients' short- and long-term compliance were assessed by questionnaires evaluating duration of and problems related to orthotic use. RESULTS: (a) In patients using RGO + FNS the slope difference of HR/VO2 curves (deltaslHR/VO2) between WHCH and orthosis was significantly lower than in other groups (-3 beats l(-1) in RGO + FNS vs 43 and 52 beats l(-1) in RGO and PW, respectively); (b) neither C, nor VO2 peak, or deltaslHR/VO2 correlated with orthosis duration of use; (c) in the RGO + FNS group, C was lower at maximal walking speed, which linearly correlated with maximal WHCH speed. CONCLUSION: (a) Electrical stimulation seems to improve locomotion, as a consequence of hemodynamic effects, but does not decrease energy expenditure, which remains high; (b) the poor long-term compliance to orthosis use cannot be predicted by the energy expenditure parameters; (c) the subjects who can reach high speeds by WHCH seem to be the most appropriate for RGO + FNS locomotion.

Adaptation, Physiological↗
PubMed2003

Energy consumption of locomotion with orthosis versus Parastep-assisted gait: a single case study.

STUDY DESIGN: Single case study. OBJECTIVES: To evaluate the energy expenditure during ambulation with the Advanced Reciprocating Gait Orthosis (ARGO), with and without functional electrical stimulation (FES), and with the Parastep system in a single subject, in order to avoid the effect of inter-subject variability. SETTING: The Centre of Sport Medicine and Bioengineering Centre 'Don C Gnocchi' Foundation ONLUS IRCCS, Milano, Italy. METHODS: A single patient (lesion level T5-T6) was trained specifically for each walking system and was evaluated after each training period. The effects of FES on muscle conditioning, spasticity and bone density were also evaluated. The HR/VO2 relationship and the energy cost of locomotion were measured during wheelchair (WHCH) use, during locomotion with ARGO (with and without FES) and Parastep system at different speeds. RESULTS: The following was observed at the end of the whole training: (a) circumferences of both lower limbs and quadriceps forces were increased, whereas fatigue index was slightly decreased, spasticity and bone density were unchanged; (b) compared to WHCH locomotion, the slope of HR/VO2 curves with ARGO was higher (slope difference=51.1 b 1O2(-1)), with ARGO+FES was similar (slope difference=-5.3 b 1O2(-1)) and with Parastep was smaller (slope difference=-55.6 b 1O2(-1)); (c) HR increased linearly with all locomotion systems, but did not rise above 125 bpm with Parastep; (d) the cost of locomotion was higher with Parastep than with ARGO (with and without FES), tested at each velocity; (e) Parastep appears to be easier to use for the subject. CONCLUSIONS: (a) FES can improve ambulation with orthosis, but the cost of locomotion remains very high; (b) the Parastep assisted gait elicits a higher energy expenditure than other orthoses, probably due to the lower speed of locomotion and to the high isometric effort of the stimulated muscles. SPONSORSHIP: This work has been partially supported by the Italian Minister of Public Health (Ricerca Finalizzata IRCCS no ICS030.7/RF97.25).

Adaptation, Physiological↗
PubMed2005

Effects of electrically induced muscle contraction on flexion reflex in human spinal cord injury.

STUDY DESIGN: Flexion reflex study in motor complete human spinal cord injury (SCI). OBJECTIVES: To examine changes in the magnitude of the flexion reflex following functional electrical stimulation (FES) of the rectus femoris (RF) muscle. SETTING: Bioengineering Unit, University of Strathclyde, Glasgow, Scotland, UK. METHODS: The flexion reflex was evoked by electrical stimulation of the sural nerve, and was recorded in the tibialis anterior (TA) muscle. RF muscle conditioning stimulation was performed at 0.7, 1, and 2 times motor threshold ( x MT) over a range of conditioning test intervals. RESULTS: The incidence of the early component of the flexion reflex (<100 ms) was low, suggesting that this reflex component might be suppressed in SCI. The long latency flexion reflex component (>120 ms) was observed in all subjects during control conditions and following sensorimotor conditioning. FES applied to the RF muscle (above and below MT) in the main induced a significant early and long lasting depression of the long latency flexion reflex. CONCLUSION: The depression of the flexion reflex was a result of multisensory actions on flexion reflex pathways resulting from the direct and indirect (mechanical) consequences of electrically induced muscle contraction on cutaneous and muscle afferents. Our findings emphasize the importance of sensory feedback mechanisms in modulating flexion reflex excitability, and highlight the need for rehabilitation professionals to consider the central actions of FES-induced afferent feedback when incorporating FES into a rehabilitation program. SPONSORSHIP: State Scholarships Foundation (IKY) of Hellas.

Adult↗
PubMed2003

Polyhydroxyalkanoate (PHA) hemeostasis: the role of PHA synthase.

Polyhydroxyalkanoates (PHAs) are biodegradable polymers with properties of thermoplastics Production of these polymers in an economically competitive fashion via bioengineering requires an understanding of the biosynthetic pathway and its regulation. This review summarizes our current knowledge of the mechanism of the class I and III PHA synthases: the initiation, elongation and termination processes. It also summarizes our current understanding of the phase transition from soluble substrates (coenzyme A esters of beta-hydroxyalkanoates) to insoluble granules and our understanding of the requirement for a transcription factor, phasin proteins, and depolymerases in PHA homeostasis.

Acyltransferases↗
PubMed2006

First blueprint, now bricks: DNA as construction material on the nanoscale.

For the most part DNA was considered Nature's instruction manual for life leading to the popular description 'blueprint of life'. However, DNA is now taking on a new aspect where it is finding use as a construction element for architecture on the nanoscale. This tutorial review addresses the importance of building ordered structures with DNA on the nanoscale, the underlying principles and approaches to build such scaffolds, the current limitations and the anticipated trajectory of the area. This is would be of interest to the chemical biology, supramolecular and bioengineering communities in particular.

DNA↗
PubMed2006

A novel approach to the micropatterning of proteins using dewetting of polymer bilayers.

The ability to control protein and cell positioning on a microscopic scale is crucial in many biomedical and bioengineering applications, such as tissue engineering and the development of biosensors. We propose here a novel, simple, and versatile method for the micropatterning of proteins. Micropatterned substrates are produced by the dewetting of a metastable polymer film on top of another polymer film. Selective adsorption, or micropatterning, of proteins can be achieved on such substrates by choosing pairs of polymers which differ in protein affinity. In this study, patterns were produced in bilayers of poly(methylmethacrylate) (PMMA) and polystyrene (PS), and of PMMA and octadecyltrichlorosilane (OTS). Fluorescence microscopy and atomic force microscopy (AFM) provide evidence that model proteins adsorb preferentially on isolated bio-adhesive (PS and OTS) micropatches in a protein-resistant (PMMA) matrix. "Inverse" protein patterns, containing non-adhesive (PMMA) islands in a protein-adhesive (PS) matrix can also be produced. Such micropatterned substrates could potentially be used in the development of biosensors and bioassays, and in the study of cell growth and motility.

Animals↗
PubMed2004

Generation of a tissue-engineered tracheal equivalent.

In this Minireview we summarize efforts to generate an autologous tissue-engineered trachea (TET) shaped as a cylinder containing a helix of cartilage to form the structural component of a functional tracheal replacement. Our first step was to demonstrate that a composite engineered tracheal equivalent composed of cartilaginous cylinder could be lined with nasal epithelial cells in a nude-mouse model. We then demonstrated the feasibility of creating the cartilage and fibrous portion of the trachea using autologous tissue harvested from a single procedure in both nude-rat and sheep models. Gross morphology and tissue morphology of these TET series were similar to that of native trachea. Histological data indicated the presence of mature cartilage formation of a pseudostratified columnar epithelium and the presence of mature cartilage surrounded by connective tissue, as would be expected of native trachea. Regarding the biomechanical properties, the cartilage was excellent in nude model; however, sheep autologous TET provided less support and therefore collapsed easily. In addition to utilizing the angiogenic qualities of cytokines, we hope to exploit the growth potentials of stromal cells derived from bone marrow for the production of a clinically practical, bioengineered trachea.

Animals↗
PubMed2004

Stem-cell plasticity and therapy for injuries of the peripheral nervous system.

Numerous publications have investigated stem-cell biology and the possible therapeutic use of stem cells in a wide range of injuries and diseases. This interest has been fueled by recent reports suggesting that mesenchymal stem cells can show unorthodox plasticity, their being able to transdifferentiate into cells of different lineages, such as neuronal phenotypes. This capability has obvious implications for their potential application in tissue engineering and tissue regeneration. The peripheral nervous system has an inherent capacity for regeneration, but this is limited and not matched by the level of reinnervation of target organs, with a resulting loss of functional recovery. Several approaches have been attempted in order to overcome this deficiency, and transplant of cultured Schwann cells into bioengineered conduits has been shown to improve regeneration. An alternative may be the use of stem-cell technology, whereby cultured and differentiated stem cells can be transplanted to the site of injury in order to promote enhanced regeneration. The present review discusses the use of stem cells applied to the repair of peripheral nerve injury and their role in the regeneration process.

Animals↗