Immunoregulatory functions of neutrophil serine proteases
Examensarbete för masterexamen
Biotechnology (MPBIO), MSc
Neutrophils are the most abundant white blood cells in humans and play an impor-tant role in the innate immune system as pathogen-eliminators. In recent years, it has been shown that neutrophils have other functions in the body as immunoregula-tors in the immune system. These immunoregulative functions are in part conveyed by the release of cytokines and chemokines as well as neutrophil derived proteases. This thesis is a part of a larger project, and is based on previous results regarding the act of neutrophil serine proteases. It has been suggested that these proteases are re-sponsible for the cleavage of di˙erent proteins, that act as regulators of the immune system. When cleaved, such proteins can become more or less potent and stimulate the activation of responder cells, hence enhancing/dampening the immune response. The aim of this thesis was to look further into the processing of such a protein by in-vestigating the activity of neutrophil serine proteases and to evaluate the processing of a protein X operated by the serine protease, Proteinase 3. Neutrophil cytoplasts, which are neutrophils devoid of intracellular granular compartments, were prepared and used in the experiments as models for neutrophils with limited serine protease activity. Furthermore, MS analysis was performed on the cleavage products and one fragment was selected for production in E. coli. Serine protease activity was eval-uated with the help of activity assays. ELISAs were used to analyse supernatants from responder cells and thus determine the potency of protein X after processing. MS analysis was performed in collaboration with Proteomics core facility and the expression of protein X fragments in E.coli was performed in collaboration with the Protein Production unit at SciLifelab in Stockholm. Results indicated that 1 ng/ml of Proteinase 3 can process protein X into a more potent protein that increases the activity of responder cells. Further results suggested that neutrophil cytoplasts also have the capacity to process protein X to become more potent.
Immunologi inom det medicinska området , Läkemedelskemi , Medicinsk bioteknologi (med inriktning mot cellbiologi) , Livsvetenskaper , Immunology in the medical area , Medicinal Chemistry , Medical Biotechnology (with a focus on Cell Biology) , Life Science