Epigenetics in neutrophil biology: from transcriptome to granulocyte transfusions. (LSBR 1121)
Prof. Dr. Taco W. Kuijpers, Dept. of Pediatrics, Academic Medical Center, Amsterdam
Postdoc investigator: Farzin Pourfarzad, PhD (Sept. 2012 – Mar. 2014)
Technician: John van Hamme (Nov. 2011 – No. 2014)
Our study provides patterns of specific “programs” (epigenetic profiles) to explain the unique steps of differentiation from an early white blood cell type (i.e. promyelocyte) onto the last step of maturation into a cell with a segmented multilobular nucleus that is remarkably deformable (i.e. the neutrophilic granulocyte or neutrophil). Neutrophils are generated in the bone marrow. Normal “programming” of the progenitor cell type through different cell stages into the non-dividing neutrophil stage takes 10-14 days. When mature, neutrophils leave the bone marrow and enter the bloodstream to circulate for 1-2 days. Thereafter, they extravasate into the tissues, phagocytize infectious microbes and die.
The neutrophil is a highly toxic killer cell that offers host defense against bacterial and fungal pathogens. Without these cells or in case of a reduced production capacity of the bone marrow, the host would fall ill from often fatal infections. The development of the toxic machinery of a neutrophil needs to be strictly regulated during its development from the early stem cell onward, because a premature activation and toxic reaction within the bone marrow would result in serious damage to the bone marrow as such. We studied the programming of the circulating blood neutrophils and those obtained from donors that have received a stimulus to increase the number of cells in the blood, i.e. G-CSF and dexamethasone (GTX). These cells clearly differ in their “programming” from each other as well as from the immature bone marrow cells, as for the first time clearly indicated by our study. GTX donor neutrophils are being used for infusion into infected and often very sick patients due to prolonged shortage or severe dysfunction of neutrophils, e.g. due to cancer treatment or inherited immune defects. These GTX pre-activated donor neutrophils differ from normal blood neutrophils (i.e. by enhanced cell survival and less efficient yeast killing). Our data may help to invent a strategy on how to reprogram neutrophils to improve granulocyte transfusion products for use in severely infected patients with a shortage of neutrophils.
The information of our epigenetic study is unique. The study has resulted in the discovery of the expression of some highly specific proteins per stage of neutrophil development, which –sometimes- could be directly related to a specific cellular behavior or function. One such stage-specific RNA transcript has been directly linked to the capacity to form the typical segmented nucleus of the mature neutrophil. This nuclear membrane protein has now been shown to have a short isoform. This small isoform is only expressed at the final stage of neutrophil development. Our findings on this short nuclear membrane isoform have unraveled the mechanism of segmentation of the nucleus: 1) to obtain a hyperlobulated nucleus with considerable silencing of many of the neutrophil-expressed genes, and 2) to generate a loose nuclear chromatin, that can be released upon strong neutrophil activation as weblike strings of DNA, i.e. Neutrophil Extracellular Traps (NETs). With this ‘kamikaze’ behavior of nuclear DNA release the neutrophil can not only trap and kill invading pathogens but will also itself undergo cell death (NETosis). These NETs catch invading pathogens and kill by direct toxicity (DNA-histone proteins, and binding of granular proteins, proteases, peroxidase).
In collaboration with dedicated bioinformatics teams of the BLUEPRINT consortium, a EU-funded project on the integration of all information on programming of gene transcription (epigenome), on the repertoire of inactive ‘silenced’ genes (methylome), and on actively transcribed RNA transcript profiles (transcriptome) was performed. This has yielded highly variable information that is currently being analyzed to unveil pertinent biological questions in neutrophil biology in relation to health and inflammatory disease, and its potential use to “reprogram” neutrophils for more effective transfusion purposes.