Interactions between mammalian cells in tissue culture

Simms, James William (1973) Interactions between mammalian cells in tissue culture. PhD thesis, University of Glasgow.

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Abstract

In any animal the individual cells must interact to co-ordinate and control their combined development, growth and functions. These interactions are both direct, between adjacent cells, and indirect by means of humoral factors. Humoral or hormonal responses have been extensively studied, but direct interactions are only poorly understood. One type of direct cell-cell interaction recently observed, both vivo and in tissue culture, in many animal cells from a wide variety of species has been termed metabolic co-operation. Variant cells, locking an enzyme function such as inosinic pyrophosphorylase (IPP) or thymidine kinase (TK) and unable to incorporate a nucleotide precursor, do incorporate the precursor, when cultured in contact with wild-type cells. However, the change of the variant phenotype is not permanent, and reversion is observed after separation of the two cell types. The possible explanations for this phenotypic modification of variant cells, growing in contact with wild-type cells, fall into two classes:- 1) the transfer of the enzyme itself from wild-type to variant cells or transfer of information (DKA, mRNA) to allow synthesis of the appropriate enzyme in the variant cells. 2) the transfer of nucleotides, beyond the enzyme block, or of nucleic acid from wild-type to variant cells. In the first class, the ability to incorporate the nucleotide precursor is transferred, in the second it is not. However, earlier work showed that, after separation of the variant cells from wild-type cells, no enzyme activity could be detected in variant cells. This eliminates explanation 1). It was therefore suggested, as DNA was not thought to be transferred between cells, that the most probable basis of metabolic co-operation was nucleotide transfer. However, a recent report suggested that RNA, but not nucleotides, is transferred between cells, so the clarification of the basis of metabolic co-operation (nucleotide transfer or nucleic acid transfer) required further investigation. In this present study the nature of the molecules which can be transferred from cell to cell in tissue culture has been examined in detail. Radioactive tracers have been used to prelabel specific classes of molecules in donor cells, and the transfer of these labelled molecules to unlabelled recipient cells during co-culture has been investigated by autoradiography. When [3H]uridine is used to label the uridine nucleotides and the RNA of donor cells, and such donor cells are co-cultured with unlabelled cells, label is transferred to the recipient cells. However, if the labelled uridine nucleotide pool is chased into RNA, prior to addition of recipient cells, the extent of label transferred to recipient cells is reduced by 97%. The label still transferred (3%) v/as shown to be due to transfer of residual labelled nucleotide pool in the donor cells, produced by RNA turnover. This strongly suggests that RNA is not transferred from cell to cell, but that RKA precursors are. These precursors must be nucleotides, and not nucleosides, to explain the observations on metabolic co-operation. A similar conclusion can be drawn from quantitative observations of the effect of actinomycin D on the transfer of label [3H]uridine labelled donor cells to unlabelled recipient cells. Actinomycin D prevents the appearance of labelled RNA in recipient cells by blocking the synthesis of RNA from the transferred nucleotide Similar observations on the transfer of label from [3H]thymidine labelled donor cells to unlabelled recipient cells, before and after a chase to reduce the label in the DKA precursor pools, showed that thymine nucleotides, but not DNA, are transferred between cells. When donor cells were prelabelled with glucose-2-h3, labelled acid soluble pools, but not labelled acid insoluble material, were transferred to recipient cells. This suggests that phosphorylated sugars are also transferred between cells. Using donor cells prelabelled with [3E] amnino acids, it was also shown that protein, like RNA, is not transferred in detectable amounts from donor to recipient cells. To summarise, nucleotides, but not RNA, DM or protein, appear therefore to be transferred between cells. (Abstract shortened by ProQuest.).

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: J D Pitts
Keywords: Cellular biology
Date of Award: 1973
Depositing User: Enlighten Team
Unique ID: glathesis:1973-73150
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 14 Jun 2019 08:56
Last Modified: 14 Jun 2019 08:56
URI: http://theses.gla.ac.uk/id/eprint/73150

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