Sedgefield, United Kingdom
Sedgefield, United Kingdom

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Maltman D.J.,Durham University | Maltman D.J.,ReInnervate | Przyborski S.A.,Durham University | Przyborski S.A.,ReInnervate
Biochemical Society Transactions | Year: 2010

Drug discovery programmes require accurate in vitro systems for drug screening and testing. Traditional cell culture makes use of 2D (two-dimensional) surfaces for ex vivo cell growth. In such environments, cells are forced to adopt unnatural characteristics, including aberrant flattened morphologies. Therefore there is a strong demand for new cell culture platforms which allow cells to grow and respond to their environment in a more realistic manner. The development of 3D (three-dimensional) alternative substrates for in vitro cell growth has received much attention, and it is widely acknowledged that 3D cell growth is likely to more accurately reflect the in vivo tissue environments from which cultured cells are derived. 3D cell growth techniques promise numerous advantages over 2D culture, including enhanced proliferation and differentiation of stem cells. The present review focuses on the development of scaffold technologies for 3D cell culture. ©The Authors.


Caldwell S.,Durham University | Johnson D.W.,Durham University | Didsbury M.P.,Durham University | Murray B.A.,Durham University | And 4 more authors.
Soft Matter | Year: 2012

Emulsion templating has been used to prepare highly porous polyHIPE materials by thiol-ene photoinitiated network formation. Commercially available multifunctional thiols and acrylates were formulated into water-in-oil high internal phase emulsions (HIPEs) using an appropriate surfactant, and the HIPEs were photo-cured. The temperature of the HIPE aqueous phase was found to influence the morphology of the resulting materials. In agreement with previous work, a higher aqueous phase temperature (80 °C) gave rise to a larger mean void and interconnect diameter. The influence of temperature on morphology was found to be reduced at higher porosity, but still significant. The Young's modulus of the porous materials was shown to be related to the functionality of the acrylate comonomer used. A mixture of penta- and hexa-acrylate gave rise to a 100-fold increase in modulus, compared to an analogous tri-functional acrylate. The materials could be functionalised conveniently by addition of mono-acrylates or thiols to the organic phase of the precursor HIPE. Degradation was observed to occur at a rate depending on the degradation conditions. Under cell culture conditions at 37 °C, 19% mass loss occurred over 15 weeks. The scaffolds were found to be capable of supporting the growth of keratinocytic cells (HaCaTs) over 11 days in culture. Some penetrative in-growth of the cells into the scaffold was observed. © The Royal Society of Chemistry 2012.


Grant
Agency: GTR | Branch: BBSRC | Program: | Phase: Training Grant | Award Amount: 91.93K | Year: 2011

Provision of a barrier against the outside environment and prevention of inside-out evaporation is a major function of epidermal differentiation. The key cellular and molecular components of the epidermal barrier are cell-cell junctions between keratinocytes, cornified envelopes (remnants of flattened cells in stratum corneum), and the lipid envelope consisting of lipids and free fatty acids. The fundamental importance of barrier function in human health is illustrated by examples where a compromised barrier leads to disease. There are several important reasons to develop in vitro models of the epidermal barrier: 1) A key factor in transdermal drug delivery systems is to be able to assess permeability and skin deposition of the drugs and carrier agents; 2) Both the pharmaceutical and cosmetic industries require reliable means for testing for skin irritation and toxicity caused by novel compounds and formulations especially given recent changes in legislation that prohibit certain animal tests; 3) The contribution of barrier function to diseases, such as eczema and psoriasis, that cause considerable health economic burden has highlighted the need for in vitro disease models to investigate both normal and pathological barrier physiology. In recent years several epidermal models have been developed. However, most organotypic cultures suffer from several impracticalities: (i) variability in the dermal substrates that reduces reproducibility of the model; (ii) long set up time required by each experiment; (iii) restricted life-span that makes most models unsuitable for long term experiments. Such limitations are not practical for the routine use of such technology. One major drawback of organotypic co-cultures based on collagen gel is their limited lifespan due to dramatic shrinkage and reduced stability of the reconstructed tissues, indicating the need for further optimisation. In addition, as the major applied use of these culture models is in skin irritant and toxicity testing, only some existing models have been evaluated for their barrier properties. The industrial and academic partners for this proposal currently collaborate and have together generated a prototype 3D culture model for human and mouse epidermal keratinocytes. This proof of concept model utilises Alvetex technology, a novel polystyrene scaffold that offers genuine routine 3D cell culture (see www.reinnervate.com). The model can be used to generate organotypic skin cultures at air-liquid interface either 1) as a stratified epidermis within the scaffold with flattened stratum corneum forming on the top surface of the scaffolds or 2) as a composite skin model where the scaffold harbours collagen gel and fibroblasts with the epidermis subsequently seeded on top. Initial characterisation shows that even without a dermal component the epidermis grown in Alvetex completes terminal differentiation with mature, lipid-coated cornified envelopes and abundant desmosomes in the underlying cell layers. The aim of the proposed CASE studentship is to take this model forward and: 1) Further develop and optimise the skin construct that has an established cornified layer that mimics highly keratinized plantar skin as closely as possible in terms of its cellular and extracellular structure and conformation; 2) Validate the skin model in terms of its structure and protein composition in comparison to known published structure of existing collagen gel systems and native skin; 3) Investigate the extent of epidermal terminal differentiation and barrier assembly. Optimized epidermal cultures will be mounted within Franz cell system to investigate the function and permeability of the epidermal barrier to test compounds; 4) Generate disease a model for barrier deficient epidermis through the culture of primary keratinocytes with selective knockdown (RNAi) of key cornified envelop scaffold proteins (involucrin, envoplakin, periplakin) to replicate changes seen in atopic eczema.


Trademark
ReInnervate | Date: 2012-02-21

Biological preparations for use in the induction of cell differentiation in cell cultures and synthetic small molecules for use in the induction of in-vitro cell differentiation, other than medical; cell growth media for growing cells for use in scientific research; media for cell culture for use, namely, in non-medical research and biotechnological laboratories; chemical preparations for biotechnological purposes, namely, for the induction of cell differentiation, in particular stem cell differentiation; reagents for use in scientific apparatus for chemical or biological analysis; chemical preparations, agents and reagents for use in cellular research; biochemicals, namely, monoclonal antibodies other than for medical or veterinary purposes; biochemicals, namely, antibodies for diagnostic, laboratory, industrial or scientific use other than medical; biochemical products containing antibodies for use in scientific research. Pharmaceutical preparations for the treatment of skin and neuro-degenerative diseases; medical biological reagents for use in molecular biology; medical diagnostic reagents; medical diagnostic reagents for in vitro laboratory tests; chemical reagents for medical and veterinary purposes; chemical preparations, agents and reagents for medical and veterinary use, namely, in the differentiation of cell types; biological preparations for the treatment of skin and neuro-degenerative diseases; cell growth media for growing cells for medical or clinical use; diagnostic agents, preparations and substances for medical purposes, namely, for use in binding or detecting antibodies, and in the development of antibodies. Cell culture apparatus and instruments for laboratory or research medical use; laboratory apparatus, namely, cell culture vessels, multiwell plates, petri dishes, cell culture well inserts, cell factories, cell culture flasks and bioreactors for use in the separation of antibodies; reaction units, namely, cell based assays, routine cell cultures, cell passaging and perfusion cell culture systems containing chemical or biological reagents; cell culture scaffolds, namely, microspun fibres and microporous polystyrene substrates coated with extra-cellular matrices; deep well plates, namely, deep reservoir petri dishes with and without well insert cradles used in laboratory research. Batch biological and chemical treatment services of materials; batch biological and chemical material treatment involving cell culture processes and the differentiation of cell types; treatment of human, animal and plant cells, bacteria and antibody materials by culturing such cells in three dimensions; material treatment involving three dimensional cell culture processes and in-vitro stem cell differentiation. Education services, namely, provision of training, arranging and conducting of seminars and workshops all in the fields of biology, biotechnology, cell culture, cell biology, toxicity and safety testing, tissue and disease modelling and drug efficacy; on-line non-downloadable electronic publications, namely, bulletins and journal articles in the fields of biology, biotechnology, cell culture, cell biology, toxicity and safety testing, tissue and disease modelling and drug efficacy. Scientific research and design services in the field of three dimensional cell culture and analytical processes; industrial analysis and research services in the field of cell culture, cell biology, toxicity and safety testing, tissue and disease modelling and drug efficacy; laboratory services, namely, analysis and research in the field of cell and tissue cultures and their treatment; biotechnology research; laboratory services relating to the production of antibodies; growing cell cultures for research purposes; testing pharmaceutical, medical and biological agents for others; consultancy relating to biotechnology.


Trademark
ReInnervate | Date: 2011-10-04

Cell culture apparatus and instruments for laboratory or research medical use; laboratory apparatus, namely, cell culture laboratory vessels, plates having multi-well arrays, petri dishes, cell culture well inserts, laboratory glassware, cell factories, cell culture flasks and bioreactors for use in the separation of antibodies; reaction units for cell based assay research, routine cell cultures, cell passaging research and perfusion cell culture systems research containing chemical or biological reagents; cell culture scaffolds, namely, microspun fibres and microporous polystyrene substrates coated with extra-cellular matrices for use in three-dimensional cell culturing; deep well plates, namely, deep reservoir petri dishes with and without well insert cradles used in laboratory research. Treatment of human, animal and plant cells, bacteria and antibody materials by culturing such cells in three dimensions; material treatment involving three dimensional cell culture processes and in-vitro stem cell differentiation. Scientific research and design services in the field of three dimensional cell culture and analytical processes; industrial analysis and research services in the field of cell culture, cell biology, toxicity and safety testing, tissue and disease modelling and drug efficacy; laboratory services, namely, analysis and research in the field of cell and tissue cultures and their treatment; biotechnology research; laboratory services relating to the production of antibodies; growing cell cultures for research purposes; testing pharmaceutical, medical and biological agents for others; consultancy relating to biotechnology.


Trademark
ReInnervate | Date: 2011-11-15

Biological preparations for use in the induction of cell differentiation in cell cultures and synthetic small molecules for use in the induction of in-vitro cell differentiation, other than medical; cell growth media for growing cells for use in scientific research; media for cell culture for use, namely, in non-medical research and biotechnological laboratories; chemical preparations for biotechnological purposes, namely, for the induction of cell differentiation, in particular stem cell differentiation; reagents for use in scientific apparatus for chemical or biological analysis; chemical preparations, agents and reagents for use in cellular research; biochemicals, namely, monoclonal antibodies other than for medical or veterinary purposes; biochemicals, namely, antibodies for diagnostic, laboratory, industrial or scientific use other than medical; biochemical products containing antibodies for use in scientific research. Pharmaceutical preparations for the treatment of skin and neuro-degenerative diseases; medical biological reagents for use in molecular biology; medical diagnostic reagents; medical diagnostic reagents for in vitro laboratory tests; chemical reagents for medical and veterinary purposes; chemical preparations, agents and reagents for medical and veterinary use, namely, in the differentiation of cell types; biological preparations for the treatment of skin and neuro-degenerative diseases; cell growth media for growing cells for medical or clinical use; diagnostic agents, preparations and substances for medical purposes, namely, for use in binding or detecting antibodies, and in the development of antibodies. Batch biological and chemical treatment services of materials; batch biological and chemical material treatment involving cell culture processes and the differentiation of cell types.


Trademark
ReInnervate | Date: 2012-06-06

Cell culture apparatus and instruments for laboratory or medical research use; laboratory apparatus, namely, cell culture laboratory vessels, plates having multi-well arrays, petri dishes, cell culture well inserts, laboratory glassware, cell factories, cell culture flasks and bioreactors for use in the separation of antibodies; reaction units for cell based assay research, routine cell cultures, cell passaging research and perfusion cell culture systems research containing chemical or biological reagents; cell culture scaffolds, namely, microspun fibres and microporous polystyrene substrates coated with extra-cellular matrices for use in three-dimensional cell culturing; deep well plates, namely, deep reservoir petri dishes with and without well insert cradles used in laboratory research.


Trademark
ReInnervate | Date: 2012-06-06

Cell culture apparatus and instruments for laboratory or research medical use; laboratory apparatus, namely, cell culture laboratory vessels, plates having multi-well arrays, petri dishes, cell culture well inserts, laboratory glassware, cell factories, cell culture flasks and bioreactors for use in the separation of antibodies; reaction units for cell based array research, routine cell cultures, cell passaging research and perfusion cell culture systems research containing chemical or biological reagents; cell culture scaffolds, namely, microspun fibres and microporous polystyrene substrates coated with extra-cellular matrices for use in three-dimensional cell culturing; deep well plates, namely, deep reservoir petri dishes with and without well insert cradles used in laboratory research.


Grant
Agency: GTR | Branch: Innovate UK | Program: | Phase: Feasibility Study | Award Amount: 24.78K | Year: 2012

Currently in the laboratory cells are routinely cultured on two dimensional (2D) substrates which limits their functionality. Reinnervate has developed alvetex which is the market leading scaffold-based technology for three dimensional (3D) cell culture that significantly enhances cell growth and function. Cells in these static culture systems are fed by passive diffusion exchange with the culture medium. In this project, we will design and build a unique perfusion plate that combines the advantages of 3D cell culture with a dynamic medium perfusion system to further enhance the performance of cultured mammalian cells. The perfusion plate will be innovatively designed and will incorporate well-to-well perfusion. Importantly, the design will be compatible with Reinnervates existing 3D products and will be developed for simple routine use thus opening new opportunities in the conventional cell culture laboratory.


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