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Characterization of the murine n-formyl peptide chemotactic receptor

Mitrophanous, Kyriacos Andreou; (1996) Characterization of the murine n-formyl peptide chemotactic receptor. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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The N-formyl peptide receptor (FPR) present on neutrophils is of importance in providing the host with a detection mechanism of broad specificity for invading microorganisms and damaged tissue. The aim of this project was to develop an accurate physiological model for the study of FPR. Since neutrophils isolated from blood leukocytes are heterogeneous, short lived and terminally differentiated they do not make good models for the study of FPR. Hence, the need for in vitro model systems. The current model used, the human leukaemic cell line (HL-60), does not produce fully mature neutrophils. In contrast, the murine pluripotent stem cell line (FDCP), can be fully differentiated to mature neutrophils. This cell line was therefore chosen for the characterization and development of a model system for the FPR. A detailed study of cytokine-mediated differentiation was undertaken. Differentiated FDCP cells, expressed FPR and showed cell adhesion and degranulation in response to N-formyl peptides. The kinetics of the expressed murine FPR and the efficacy of a number of synthetic N-formyl peptides was established. The peptide formyl-Norleu-Leu-Phe-Norleu-Tyr-Lys bound with high and low affinity dissociation constants of 3.7 and 22.6 nM, respectively. The number of receptors was estimated to be 79 000 per cell with 25[percent] being of high affinity. The differentiated FDCP cells, neutrophils had low affinity binding for the peptide fMet-Leu-Phe as compared to human and rabbit neutrophils. Attempts were first made to clone the human and then the murine FPR gene. However, the putative genes were cloned by another group before completion of this work. The muFPR gene, which was transcribed and expressed in murine FDCP cells differentiated to neutrophils, was identified from six putative genes by reverse transcriptase PCR. The time course of transcription was consistent with the appearance of functional FPR during differentiation.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Characterization of the murine n-formyl peptide chemotactic receptor
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Thesis digitised by ProQuest
URI: https://discovery.ucl.ac.uk/id/eprint/10099340
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