Periodic Reporting for period 1 - INTERCONNECTION (Inter-tissue communication during nematode infections: role of dermal γδ17 T cells)
Okres sprawozdawczy: 2022-01-01 do 2023-12-31
The immune system is constantly exposed to invading microorganisms. In order to promote pathogen killing without causing excessive self-damage, immune cells have to carefully choose how to respond to a given threat and generate a balanced response. Nevertheless, how this decision is made is still not fully understood. Moreover, during infection pathogens often travel through different organs, requiring local immune cells to establish an inter-tissue communication network, ensuring a concerted action throughout the body. This process, however, remains largely unexplored. In general, this knowledge gap can be attributed to the lack of suitable experimental models to address this question. Experimental infection with the parasite Nippostrongylus brasiliensis provides a great tool to overcome this problem. Infective larvae actively penetrate the skin, and then migrate through the lungs before establishing themselves in the gut. Later on, parasite elimination depends on cross-regulation of distinct and seemingly exclusive arms of the immune system. Of note, to date, little is known about the impact of the early immune responses starting in the skin in the regulation of the anti-parasitic responses taking place at later stages in the lungs and gut. In fact, the current knowledge on the skin phase of N. brasiliensis infection is fragmented and superficial, making it difficult to assess the extension of these initial immune events to an organismal level. Therefore, in this project, we employed state-of-the-art technologies to examine at an unprecedented depth the early events taking place in the skin of N. brasiliensis-infected mice. Moreover, by using different routes of infection and genetically-modified mouse models we set to investigate the cellular mechanisms underlying skin to lung communication during parasitic infections. The data generated in this project added to the field of parasitology research by generating knowledge transposable to various microorganisms, including human-threatening pathogens. Furthermore, preliminary results generated in the present project support the idea that the skin - lung axis can be extrapolated to other inflammatory conditions starting in the skin, such as allergies, by inducing similar immune mechanisms as the ones elicited during parasite infections. Overall, completion of INTERCONNECTION greatly advanced our basic knowledge of the immune events happening early in the skin during parasite infections, and their consequence in distal organs; these results might also serve as the basis for understanding complex processes in human health as the development of allergies.
The larvae of N. brasiliensis parasites actively penetrate the skin and reach the lung after 18 to 32 hours, where they finish their development before migrating to the intestines, where they become sexually mature. Although the initial contact of the mammalian immune system with the invading nematode occurs in the skin, our current knowledge of these initial events is fragmented. Furthermore, as most studies hitherto focused on infection strategies using the subcutaneous route, less is known about the cutaneous immune responses following natural infection. The low efficiency and high variability of published “natural” N. brasiliensis infection models does not allow for a robust measure of late immune responses, and needle inoculation does not fully recapitulate skin immune responses to invading larvae. We, therefore, optimized an alternative model of “natural” infection using the percutaneous route, with worm migratory kinetics comparable to the more common subcutaneous inoculation. In accordance with previous findings, we observe an accumulation of immune cells early after larvae penetration in the skin. The use of high parameter flow cytometry allowed us to dissect with great detail parasite-induced changes in skin-resident and recruited immune cells, including less abundant populations. Moreover, we performed both a high-throughput analysis of inflammatory mediators within the tissue and analysed the kinetic changes in gene expression in the skin of infected mice. Thus, we generated the most complete roadmap to date describing the changes in skin immune system in response to nematode larvae. In order to test the contribution of skin-derived events in the regulation of distal immunity, we initially compared percutaneously-infected mice with intravenously-infected animals where the entirety of the skin immune response is bypassed. Interestingly, although intravenously-infected mice present similar numbers of larvae in the lungs as mice infected percutaneously, they display a distinct pulmonary immune response, both at the cellular and transcriptional level. Of note, these changes also translate in increased tissue damage in the lungs intravenously-infected mice, due to the effects of migrating larvae. By blocking lymphocyte migration and using a transgenic mouse models to track and manipulate lymphocytes from the skin, we were able to find that a subset of T lymphocytes migrates from the skin to the lungs, where they are responsible to kickstart the immune response observed in percutaneously infected mice. As such, when lymphocyte migration is blocked we observed the same immune hallmarks of intravenous infections, including increased tissue damage. Furthermore, using skin (but not pulmonary) allergic inflammation models we also observed the migration of lymphocytes from the skin to the lungs. Overall, our results confirm that we have a robust model to study the cutaneous (and distal) immune responses to N. brasiliensis, that strongly supports our initial hypothesis that early events in the skin regulate type-2 immunity at distal sites.
During the course of this project, these results have been presented in international conferences in Italy, Portugal and Greece. In addition, these findings were presented in local UK conferences. Finally, I was invited to speak about these results in institutional seminar series in Dundee, UK and Copenhagen, Denmark. Whereas the main body of work generated in the INTERCONNECTION project is still being prepared for publication in a peer-reviewed journal, it is important to note that results derived from the models and techniques developed for the present project figure in two peer-reviewed publications in the journals Cell Reports and Mucosal Immunology.
During the course of this project, these results have been presented in international conferences in Italy, Portugal and Greece. In addition, these findings were presented in local UK conferences. Finally, I was invited to speak about these results in institutional seminar series in Dundee, UK and Copenhagen, Denmark. Whereas the main body of work generated in the INTERCONNECTION project is still being prepared for publication in a peer-reviewed journal, it is important to note that results derived from the models and techniques developed for the present project figure in two peer-reviewed publications in the journals Cell Reports and Mucosal Immunology.
The results generated in the INTERCONNECTION project provided the most complete characterization of the immune responses to nematode larvae in the skin to date, which can help to pave the way for new strategies of parasite control in the future. Also, the percutaneous N. brasiliensis infection model optimized during the course of this project might become a useful tool to many researchers working in parasitology and skin immunology. Finally, this was the first time that the migration of lymphocytes from the skin to the lungs was described in the scientific literature. Furthermore, our results stress the importance of the, often overlooked, skin – lung axis in the generation of pulmonary immune responses to pathogens and the control of tissue damage. With further work, we believe our findings can be extrapolated to other immune-mediated conditions along the skin – lung axis, including the development of allergies, providing the basis for a deeper understanding of the mechanisms controlling these responses and novel immunoregulation strategies.