Development of new ex-vivo models is essential as early as possible in research areas such as pharmacology, toxicology, immunology, virology and biochemistry. Implementation of procedures which would allow to define the state of the art and validation of these models, is urgent for enabling their incorporation as in-vitro tests into regulatory procedures for biological and clinical studies, and industrial applications.
Each organ and tissue is characterised by specific metabolic pathways which lead to qualitative and quantitative differences in sensitivity to chemicals or toxic agents and to different cell injury reactions. Moreover, although there is a high degree of similarities between different mammalian species regarding various functional parameters for a given tissue, extrapolation of studies between species is hampered by quantitative and sometimes qualitative differences. Therefore, development of cell systems from target organs and from different species including humans, is necessary to make possible the evaluation of the relevance of observations and to provide confidence in the results obtained with in-vitro tests.
At the present time a limited number of in-vitro cell systems is available. They can be grouped in two main categories: primary cultures of normal differentiated cells from various tissues which have a limited lifespan period, and till raise problems of variability and therefore of reliability, mainly when cells from human origin are concerned; and permanent cell lines which often exhibit only a fraction of specific functions. Furthermore, establishment of new immortalised cell lines by transfection of transforming agents such as SV40 sequences, was recently obtained but with only partial maintenance of the cell differentiated status and poor stability. Then, the difficulty resides in obtaining highly differentiated cells without inducing terminal proliferation and cell death, or highly proliferative cells without losing tissue-specific gene expression.
Recent progress in knowledge of cell biology and development of new powerful methodological strategies lead to the assumption that the behavior of the cells is dependent on the balance between proliferation, differentiation and cell death, and evidences have been provided that groups of different molecules (including oncogenes like c-myc, P53, bc12... or transcription factors like HLH- Id...) are involved in the control of this dynamic equilibrium by a process of reciprocal activation and inhibition accurately regulated by the time and the level at which they are expressed. On the other hand, new original methods have been developed which allow tissue targeting or ablation of one or two genes, induction or inhibition of these targeted genes in a time- and level dependent manner, in-vivo as well as in-vitro.
The objectives of this project are: 1) to evaluate in-vivo in transgenic animals, and in-vitro in various cell systems, the assumption that the balance between proliferation, differentiation and cell death can be stably modified by inducing changes in the expression of one (or more) of the regulating proteins or by modifying the transcriptional regulation of tissue-specific genes, without losing the capability of the cells to undergo one of these biological events; 2) to develop, on the basis of this assumption, new stable immortal differentiated cell lines from different species, mainly mouse, rat, rabbit and especially human, and different tissues mainly liver, kidney, intestine or lung which are main targets for toxic agents, and also cartilage because this tissue is frequently involved in inflammatory reactions and aging process. To reach this goal, the project proposes to ally researchers who will get benefit of the combination of their high diversity in experience and expertise: cell biologists, molecular biologists, pharmacologists and biochemists, specialists since many years of several different tissues or organs and experts in a wide range of powerful methodological approaches such as transgenesis, cell transfection or cell hybrids formation, primary culture and co-culture systems management...
Important benefits are expected in pharmaco- toxicology, virology but also by a better understanding of some immune reactions, apoptosis or tumour initiation and progression, in development of related clinical and industrial applications. The likely outcome is the settlement of new in-vitro tests based on the use of more accurate cell models selected on either their high expression of differentiated or targeted functions, or their proliferative and apoptotic properties. Of additional and considerable importance, the project will help to reduce the number of animals required currently in the development of these research tasks.
All these proposal objectives comply with the priorities of the programme Workplan Area 7, and fit well into the section 1-4: "cell cultures for the development of in- vitro tests".
Funding SchemeCSC - Cost-sharing contracts
WC2A 3PX London