New insights on type 2 immunity key drivers : mast cells and group 2 innate lymphoid cells

Type 2 immune responses are characterized by the production of cytokines IL-4, IL-5 and IL-13. They are known to have crucial physiological and pathophysiological functions in the body. Data from recent studies indicate their importance in the containment and clearance of helminth infections. They are induced to generate rapid response against helminth infections even before the elicitation of specific adaptive responses. On the other hand, Th2 responses are also involved in the promotion of inflammatory conditions such as allergy and asthma. Studies from the past have probed the mounting of type 2 immune responses and Th2 cytokine production by innate immune cells. In this thesis, we set out to study two innate cells, which are considered to be one of the key drivers of allergic conditions – mast cells and group 2 innate lymphoid cells.

In paper I, an optimized protocol to efficiently isolate mast cells from human lung tissue with high yield and cell viability is described. Mast cells are heterogeneous, tissue-resident inflammatory cells characterized by the expression of high-affinity IgE receptor, FcεRI, and CD117/KIT, the receptor for stem cell factor (SCF). The isolation of mast cells was performed by a sequential combination of washing, enzymatic digestion, mechanical disruption and Percol centrifugation (WEMP). The yield from WEMP protocol was significantly high when compared to the conventional enzyme-based method. The isolated cells were further used for flow cytometry-based characterization studies and single cell RNA sequencing. Paper II is a study of human lung mast cells at the single cell level using RNA sequencing. The classic marker expression by mast cells revealed the integrity of the sort and the sequencing reactions. Following this, analyses of sequencing data was performed to depict highly variable genes, highly abundant genes and the presence of subpopulation. Preliminary data displayed homogeneity within the sorted population of the isolated lung mast cells with no clear subpopulations. In paper III, the fundamental aim was to determine the effects of prostaglandin D2 on ILC2. ILC2 play critical roles in the initiation and promotion of type 2 immune responses. The results indicated the constitutive expression of prostaglandin D2 synthase by ILC2 and the endogenous PGD2 production was involved in the activation of ILC2 through CRTH2 receptors in a para/autocrine fashion. In paper IV, the study was designed to elucidate the effect of PGE2 on ILC2. ILC2 were isolated, cultured and expanded under different treatment conditions to study the effect. PGE2 was found to have suppressive effect on GATA3 and IL-2 receptor (CD25) expression and cytokine production by ILC2. The suppressive effects were mediated by EP2 and EP4 receptors. The use of EP2 and EP4 agonists can therefore serve as potential therapeutic target in controlling ILC2 mediated inflammation.

In conclusion, the results from our studies add to the already known information about initiation, maintenance and regulation of type 2 immune responses. They serve as basis for future mast cell-ILC2 interaction studies that could provide potential insights into their contribution in allergic and other TH2-driven conditions.

List of scientific papers

I. Ravindran A, Rönnberg E, Dahlin JS, Mazzurana L, Säfholm J, Orre AC, Al-Ameri M, Peachell P, Adner M, Dahlén SE, Mjösberg J and Nilsson G. An Optimized Protocol for the Isolation and Functional Analysis of Human Lung Mast Cells. Front Immunol. 2018 Oct 5;9:2193.
https://doi.org/10.3389/fimmu.2018.02193

II. Ravindran A, Mazzurana L, Rönnberg E, Dahlin J, Säfholm J, Ann-Charlotte Orre4, Al-Ameri M, Adner M, Dahlén SE, Mjösberg J and Nilsson G. Deciphering the heterogeneity of human lung mast cells by single cell RNA sequencing. [Manuscript]

III. Maric J, Ravindran A, Mazzurana L, Van Acker A, Rao A, Ekoff M, Thomas D, Fauland A, Nilsson G, Wheelock C, Dahlén SE, Ferreiros Bouzas N, Geisslinger G, Friberg D, Heinemann A, Konya V and Mjösberg J. Cytokine-induced endogenous production of PGD2 is essential for ILC2 activation. J Allergy Clin Immunol. 2018 Dec 19.
https://doi.org/10.1016/j.jaci.2018.10.069

IV. Maric J, Ravindran A, Mazzurana L, Björklund ÅK, Van Acker A, Rao A, Friberg D, Dahlén SE, Heinemann A, Konya V and Mjösberg J. Prostaglandin E2 suppresses human group 2 innate lymphoid cell function. J Allergy Clin Immunol. 2018 May;141(5): 1761-1773.
https://doi.org/10.1016/j.jaci.2017.09.050