Ccl21a RECRUITS CCR7+ DC PROGENITORS TO THE THYMUS

During αβ T cell development in the thymus, migration of newly selected CD4+ and CD8+ thymocytes into medullary areas enables tolerance mechanisms to purge the newly selected αβ TCR repertoire of autoreactive specificities. Thymic dendritic cells (DC) play key roles in this process and consist of three distinct subsets that differ in their developmental origins. Thus, plasmacytoid DC and Sirpα+ conventional DC type 2 are extrathymically derived and enter into the thymus via their respective expression of the chemokine receptors CCR9 and CCR2. In contrast, although Sirpα- conventional DC type 1 (cDC1) are known to arise intrathymically from immature progenitors, the precise nature of such thymus-colonizing progenitors and the mechanisms controlling their thymus entry are unclear. In this article, we report a selective reduction in thymic cDC1 in mice lacking the chemokine receptor CCR7. In addition, we show that the thymus contains a CD11c+MHC class II-Sirpα-Flt3+ cDC progenitor population that expresses CCR7, and that migration of these cells to the thymus is impaired in Ccr7-/- mice. Moreover, thymic cDC1 defects in Ccr7-/- mice are mirrored in plt/plt mice, with further analysis of mice individually lacking the CCR7 ligands CCL21Ser (Ccl21a-/-) or CCL19 (Ccl19-/-) demonstrating an essential role for CCR7-CCL21Ser during intrathymic cDC1 development. Collectively, our data support a mechanism in which CCR7-CCL21Ser interactions guide the migration of cDC progenitors to the thymus for correct formation of the intrathymic cDC1 pool

Chemical evolution of primary and formation of secondary biomass burning aerosols during daytime and nighttime

Organic matter (OM) can constitute more than half of fine particulate matter (PM) and affect climate and human health. Natural and man-made biomass burning is an important contributor to primary and secondary OM (POA and SOA) with an increasing trend. Aerosol mass spectrometry (AMS) and Fourier transform infrared spectroscopy (FTIR) are two complementary methods of identifying the complex chemical composition of OM in terms of mass fragments and functional groups, respectively. AMS offers a relatively higher temporal resolution compared to FTIR (performed on PTFE filters). However, the interpretation of AMS mass spectra remains complicated due to the extensive molecular fragmentation. In this study, we used collocated AMS and FTIR measurements to better understand the evolution of biomass burning POA and SOA due to different mechanisms of chemical aging (e.g., homogeneous gasphase oxidation and heterogeneous reactions). Primary emissions from wood and pellet stoves were injected into a 10 m3 environmental chamber located at the Center for Studies of Air Qualities and Climate Change (CSTACC) at ICE-HT/FORTH. Primary emissions were aged using hydroxyl and nitrate radicals with atmospherically relevant exposures. PM1 was analyzed by a highresolution time-of-flight (HR-ToF) and was also collected on PTFE filters over 20-minute periods before and after aging for off-line FTIR analysis. AMS and FTIR measurements agreed well with regards to the concentration of OM and some biomass burning tracers (levoglucosan and lignin; Yazdani A., 2020b) and the OM:OC ratio. Chamber wall loss rates were estimated using AMS OM concentration and were used to split the contribution of POA and SOA. The estimated FTIR and AMS spectra of SOA produced by reactions of biomass burning volatile organic compounds (VOCs) with OH were found to have prominent acid signatures. Organonitrates, on the other hand, appeared to be important for SOA produced by the nitrate radical. We found that with continued aging, SOA evolves and becomes similar to the oxygenated OA (OOA) in the atmosphere. We also found that POA composition also evolves with aging. Our estimates show that up to 10 % of POA mass undergoes aging. Biomass burning tracers such as lignin and levoglucosan in addition to hydrocarbons are among the POA compounds that are lost the most in biomass burning POA (up to 6 times more than OM decrease due to chamber wall losses; Fig. 1). This diminution is observed for both semi-volatile (levoglucosan and hydrocarbons) and non-volatile (lignin) POA species, implying the importance of gasparticle partitioning, heterogeneous reactions, and photolysis for POA evolution in the atmosphere. This result can be important since chemical transport models usually do not consider POA heterogeneous reactions. Figure 1. Trends of individual AMS mass fragments (with contribution to OM> 0.3 %) during aging with UV (starting from time zero). All mass fragments have been normalized by their concentration before the with start of aging and corrected for the chamber wall losses. Important mass fragments are shown in color. This work was supported by the project PyroTRACH (ERC- 2016-COG) funded from H2020-EU.1.1. - Excellent Science - European Research Council (ERC), project ID 726165 and funding from the Swiss National Science Foundation (200021_172923). References Yazdani, A., Dudani, N., Takahama, S., Bertrand, A., Prévôt, A. S. H., El Haddad, I., and Dillner, A. M.: Characterization of Primary and Aged Wood Burning and Coal Combustion Organic Aerosols in Environmental Chamber and Its Implications for Atmospheric Aerosols, Atmospheric Chemistry and Physics Discussions, pp. 1– 32

Differentiating between primary and secondary organic aerosols of biomass burning in an environmental chamber with FTIR and AMS

Fine particulate matter (PM) affects visibility, climate and public health. Organic matter (OM), which is hard to characterize due to its complex chemical composition, can constitute more than half of the PM. Biomass burning from residential wood burning, wildfires, and prescribed burning is a major source of OM with an ever-increasing importance. Aerosol mass spectrometry (AMS) and Fourier transform infrared spectroscopy (FTIR) are two complementary methods of identifying the chemical composition of OM. AMS measures the bulk composition of OM with relatively high temporal resolution but provides limited parent compound information. FTIR, carried out on samples collected on Teflon filters, provides detailed functional groupinformation at the expense of relatively low temporal resolution. In this study, we used these two methods to better understand the evolution of biomass burning OM in the atmosphere with aging. For this purpose, primary emissions from wood and pellet stoves were injected into the Center for Studies of Air Qualities and Climate Change (C-STACC) environmental chamber at ICE-HT/FORTH. Primary emissions were aged using hydroxyl and nitrate radicals (with atmospherically relevant exposures) simulating atmospheric day-time and night-time oxidation. A time-of-flight (ToF) AMS reported the composition of non-refractory PM1 every three minutes and PM1 was collected on PTFE filters over 20-minute periods before and after aging for off-line FTIR analysis. We found that AMS and FTIR measurements agreed well in terms of measured OM mass concentration, the OM:OC ratio, and concentration of biomass burning tracers – lignin and levoglucosan. AMS OM concentration was used to estimate chamber wall loss rates which were then used separate the contribution of primary and secondary organic aerosols (POA and SOA) to the aged OM. AMS mass spectra and FTIR spectra of biomass burning SOA and estimates of bulk composition were obtained by this procedure. FTIR and AMS spectra of SOA produced by OH oxidation of biomass burning volatile organic compounds (VOCs) were dominated by acid signatures. Organonitrates, on the other hand, appeared to be important in the SOA aged by the nitrate radical. The spectra from the two instruments also indicated that the signatures of certain compounds such as levoglucosan, lignin and hydrocarbons, which are abundant in biomass burning POA, diminish with aging significantly more than what can be attributed to chamber wall losses. The latter suggests biomass burning POA chemical composition might change noticeably due to heterogeneous reactions or partitioning in the atmosphere. Therefore, the common assumption of stable POA composition is only partially true. In addition, more stable biomass burning tracers should be used to be able to identify highly aged biomass burning aerosols in the atmosphere

Diversity in medullary thymic epithelial cells controls the activity and availability of iNKT cells

The thymus supports multiple αβ T cell lineages that are functionally distinct, but mechanisms that control this multifaceted development are poorly understood. Here we examine medullary thymic epithelial cell (mTEC) heterogeneity and its influence on CD1d-restricted iNKT cells. We find three distinct mTEClow subsets distinguished by surface, intracellular and secreted molecules, and identify LTβR as a cell-autonomous controller of their development. Importantly, this mTEC heterogeneity enables the thymus to differentially control iNKT sublineages possessing distinct effector properties. mTEC expression of LTβR is essential for the development thymic tuft cells which regulate NKT2 via IL-25, while LTβR controls CD104+ CCL21+ mTEClow that are capable of IL-15-transpresentation for regulating NKT1 and NKT17. Finally, mTECs regulate both iNKT-mediated activation of thymic dendritic cells, and iNKT availability in extrathymic sites. In conclusion, mTEC specialization controls intrathymic iNKT cell development and function, and determines iNKT pool size in peripheral tissues

Diversity in medullary thymic epithelial cells controls the activity and availability of iNKT cells

The thymus supports multiple αβ T cell lineages that are functionally distinct, but mechanisms that control this multifaceted development are poorly understood. Here we examine medullary thymic epithelial cell (mTEC) heterogeneity and its influence on CD1d-restricted iNKT cells. We find three distinct mTEClow subsets distinguished by surface, intracellular and secreted molecules, and identify LTβR as a cell-autonomous controller of their development. Importantly, this mTEC heterogeneity enables the thymus to differentially control iNKT sublineages possessing distinct effector properties. mTEC expression of LTβR is essential for the development thymic tuft cells which regulate NKT2 via IL-25, while LTβR controls CD104+ CCL21+ mTEClow that are capable of IL-15-transpresentation for regulating NKT1 and NKT17. Finally, mTECs regulate both iNKT-mediated activation of thymic dendritic cells, and iNKT availability in extrathymic sites. In conclusion, mTEC specialization controls intrathymic iNKT cell development and function, and determines iNKT pool size in peripheral tissues

Chemical evolution of primary and secondary biomass burning aerosols during daytime and nighttime

Primary emissions from wood and pellet stoves were aged in an atmospheric simulation chamber under daytime and nighttime conditions. The aerosol was analyzed with the online Aerosol Mass Spectrometer (AMS) and offline Fourier transform infrared spectroscopy (FTIR). Measurements using the two techniques agreed reasonably well in terms of the organic aerosol (OA) mass concentration, OA:OC trends, and concentrations of biomass burning markers – lignin-like compounds and anhydrosugars. Based on the AMS, around 15 % of the primary organic aerosol (POA) mass underwent some form of transformation during daytime oxidation conditions after 6–10 hours of atmospheric exposure. A lesser extent of transformation was observed during the nighttime oxidation. The decay of certain semi-volatile (e.g., levoglucosan) and less volatile (e.g., lignin-like) POA components was substantial during aging, highlighting the role of heterogeneous reactions and gas-particle partitioning. Lignin-like compounds were observed to degrade under both daytime and nighttime conditions, whereas anhydrosugars degraded only under daytime conditions. Among the marker mass fragments of primary biomass burning OA (bbPOA), heavy ones (higher m/z) were relatively more stable during aging. The biomass burning secondary OA (bbSOA) became more oxidized with continued aging and resembled those of aged atmospheric organic aerosols. The bbSOA formed during daytime oxidation was dominated by acids. Organonitrates were an important product of nighttime reactions in both humid and dry conditions. Our results underline the importance of changes to both the primary and secondary biomass burning aerosols during their atmospheric aging. Heavier AMS fragments seldomly used in atmospheric chemistry can be used as more stable tracers of bbPOA and in combination with the established levoglucosan marker, can provide an indication of the extent of bbPOA aging

Developmental conversion of thymocyte-attracting cells into self-antigen-displaying cells in embryonic thymus medulla epithelium

Thymus medulla epithelium establishes immune self-tolerance and comprises diverse cellular subsets. Functionally relevant medullary thymic epithelial cells (mTECs) include a self-antigen-displaying subset that exhibits genome-wide promiscuous gene expression promoted by the nuclear protein Aire and that resembles a mosaic of extrathymic cells including mucosal tuft cells. An additional mTEC subset produces the chemokine CCL21, thereby attracting positively selected thymocytes from the cortex to the medulla. Both self-antigen-displaying and thymocyte-attracting mTEC subsets are essential for self-tolerance. Here, we identify a developmental pathway by which mTECs gain their diversity in functionally distinct subsets. We show that CCL21-expressing mTECs arise early during thymus ontogeny in mice. Fate-mapping analysis reveals that self-antigen-displaying mTECs, including Aire-expressing mTECs and thymic tuft cells, are derived from CCL21-expressing cells. The differentiation capability of CCL21-expressing embryonic mTECs is verified in reaggregate thymus experiments. These results indicate that CCL21-expressing embryonic mTECs carry a developmental potential to give rise to self-antigen-displaying mTECs, revealing that the sequential conversion of thymocyte-attracting subset into self-antigen-displaying subset serves to assemble functional diversity in the thymus medulla epithelium

Leukocyte and lymphoid organ ontogeny

Autoimmune regulator+ (Aire) medullary thymic epithelial cells (mTECs) play a critical role in tolerance induction. Several studies demonstrated that Aire+mTECs differentiate further into Post-Aire cells. Yet, the identification of terminal stages of mTEC maturation depends on unique fate-mapping mouse models. Herein, we resolve this limitation by segmenting the mTEChi(MHCIIhiCD80hi) compartment into mTECA/hi (CD24−Sca1−), mTECB/hi (CD24+Sca1−), and mTECC/hi (CD24+Sca1+). While mTECA/hi included mostly Aire-expressing cells, mTECB/hi contained Aire+ and Aire− cells and mTECC/hi were mainly composed of cells lacking Aire. The differential expression pattern of Aire led us to investigate the precursor-product relationship between these subsets. Strikingly, transcriptomic analysis of mTECA/hi, mTECB/hi, and mTECC/hi sequentially mirrored the specific genetic program of Early-, Late- and Post-Aire mTECs. Corroborating their Post-Aire nature, mTECC/hi downregulated the expression of tissue-restricted antigens, acquired traits of differentiated keratinocytes, and were absent in Aire-deficient mice. Collectively, our findings reveal a new and simple blueprint to survey late stages of mTEC differentiation