Gap junctions in lymphocyte ontogeny

Carolan, Edward John (1985) Gap junctions in lymphocyte ontogeny. PhD thesis, University of Glasgow.

Abstract

The primary aim of this research was to examine the ability of lymphoid cells to form permeable intercellular communicating junctions (gap junctions). Established techniques, based on the detection of intercellular exchange of radiolabelled nucleotides, were used to analyse the junction formation between the adherent lymphoid tumour cell line RAJI and other cell lines of known junction forming ability. Results show that there is no detectable junction formation between RAJI cells, or between RAJI cells and any of the other cell types elaborately tested. Peripheral blood lymphocytes (PBL) were examined, to resolve the conflict in the literature as to the ability of PBL to form communicating junctions. This required the development of modified procedures to analyse metabolite transfer from non-adherent cells. The results show no detectable junction formation between PBL and a range of junction-forming cell lines and the adherent lymphoid line RAJI. Phytohaemagglutinin (PHA) treated PBL also show no detectable junction formation between the same range of cells, which is in conflict with reported electrophysiological findings. Attempts were made to improve the sensitivity of the assay system used for junction formation, by fixation of the acid soluble component of the incorporated radiolabelled material. Retention of the label is increased, but the increase was not sufficient to significantly improve the assay. The analysis was extended to immature lymphocytes of the thymus, and population of thymocytes was identified which was able to form junctions with murine fibroblasts, Swiss 3T3 and a BALB/c 3T3 and the hypoxanthine guanine phosphoribosyl transferase mutant (HGPRT-) subline of BHK/C13 baby hampster kidney fibroblast line TG2. Further analysis demonstrates that the size of this population varies with the age of the mouse. Isopycnic centrifugation shows that this population is proportionately higher in low density fractions. Peanut agglutinin (PNA) fractionation shows that the thymocytes which can form junctions can also be agglutinated with PNA. Thymocytes were fractionated using a range of monoclonal antibodies specific for a range of lymphocyte surface antigens using complement mediated cytotoxicity (CMC) and fluorescence activated cell sorting (FACS). These experiments show that the thymocytes which can form junctions lack expression of Lyt-2 and la antigens, but express Lyt-1 molecules. However, there is variability in the expression of Thy-1 and H-2K antigens and the frequency of thymocytes that can form junctions is of the order of 5x10-4 and 10-5 cells. The implications of the immature surface antigen expression and the ability to form junctions are discussed and the position of thymocytes which can form junctions in intrathymic lymphocyte differentiation and development are explored.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Cellular biology, Biochemistry
• Date of Award: 1985
• Depositing User: Enlighten Team
• Unique ID: glathesis:1985-76620
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 19 Nov 2019 14:02
• Last Modified: 19 Nov 2019 14:02
• URI: https://theses.gla.ac.uk/id/eprint/76620