Biology subjects

Daga, M.

Publications and source records attributed to Daga, M..

4 recordsLinked to original sources

bioRxiv2024

Spatial characterization of interface dermatitis in cutaneous lupus reveals novel chemokine ligand-receptor pairs that drive disease

BackgroundChemokines play critical roles in the recruitment and activation of immune cells in both homeostatic and pathologic conditions. Here, we examined chemokine ligand-receptor pairs to better understand the immunopathogenesis of cutaneous lupus erythematosus (CLE), a complex autoimmune connective tissue disorder. ObjectivesOur objectives were to (1) characterize the cellular and proteomic constitution of interface dermatitis in CLE using blister biopsies, (2) map chemokine:ligand receptor pairs that govern recruitment of immune cells to form interface dermatitis in CLE, and (3) perform unbiased analyses in tandem on different clinical subtypes to identify novel genes and proteins underlying discoid versus subacute CLE. MethodsWe used suction blister biopsies to measure cellular infiltrates with spectral flow cytometry in the interface dermatitis reaction, as well as 184 protein analytes in interstitial skin fluid using 96-plex immunoassay targeted proteomics. Flow and 96-plex immunoassay data concordantly demonstrated significant increases in T cells and antigen presenting cells (APCs). We also performed spatial transcriptomics and spatial proteomics of punch biopsies using digital spatial profiling (DSP) technology on CLE skin and healthy margin controls to examine discreet locations within the tissue. ResultsSpatial and 96-plex immunoassay data confirmed elevation of interferon (IFN) and IFN-inducible CXCR3 chemokine ligands. Comparing involved versus uninvolved keratinocytes in CLE samples revealed upregulation of essential inflammatory response genes in areas near interface dermatitis, including AIM2. 96-plex immunoassay data confirmed upregulation of Caspase 8, IL-18 which is the final product of AIM2 activation, and induced chemokines including CCL8 and CXCL6 in CLE lesional samples. Chemotaxis assays using PBMCs from healthy and CLE donors revealed that T cells are equally poised to respond to CXCR3 ligands, whereas CD14+CD16+ APC populations are more sensitive to CXCL6 via CXCR1 and CD14+ are more sensitive to CCL8 via CCR2. ConclusionsTaken together, our data map a pathway from keratinocyte injury to lymphocyte recruitment in CLE via AIM2-Casp8-IL-18-CXCL6/CXCR1 and CCL8/CCR2, and IFNG/IFNL1-CXCL9/CXCL11-CXCR3, and identify potential novel biomarkers of disease. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=168 SRC="FIGDIR/small/574422v2_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@1e86dbeorg.highwire.dtl.DTLVardef@7d0208org.highwire.dtl.DTLVardef@1108aa8org.highwire.dtl.DTLVardef@ff255f_HPS_FORMAT_FIGEXP M_FIG C_FIG Model of chemokine systems governing recruitment of immune cell subsets to form interface dermatitis in cutaneous lupus. A. Summary of fresh tissue blister biopsy studies demonstrating increases in HLADR+ cells in nonlesional lupus biopsies as assessed by flow cytometry, and increased CXCL6 and CCL8 as assessed by 96 plex immunoassay. Lesional lupus biopsies also demonstrated significant increases in T cells and CXCL9/10/11 production. B. Model of chemokine-directed formation of interface dermatitis in cutaneous lupus. 1. Whole transcriptome atlas (WTA) digital spatial profiling (DSP) revealed increased AIM2 in keratinocytes proximal to inflammation, which is reported to be induced by environmental triggers including UV light and toxins. We also noted increased Caspase 8 (Casp8) and IL18 at the protein level, which can be induced downstream of AIM2. Chemokines including CXCL6 and CCL8 can be induced downstream of IL18, explaining how CCL8 and CXCL6 might be induced. 2. Recruitment of myeloid cell first responders by CCL8 and CXCL6. CD14+CD16+ myeloid cells, which were recently described in nonlesional lupus skin, express CXCR1 and migrate towards CXCL6, whereas CD14+CD16-myeloid cells express more CCR2 and migrate towards CCL8. 3. The CXCR3 ligands CXCL9/10/11 are expressed by keratinocytes, but more strongly in CD45+ immune cell and T cell regions of interest (ROIs). 4. T cells express CXCR3 and migrate towards CXCL9 and to a greater extent CXCL11. 5. The recruited HLADR+ myeloid populations and T cells contribute to formation of interface dermatitis. Thus, we propose a model in which keratinocyte/myeloid crosstalk can reinforce chemokine systems to optimally recruit lymphocytes and other immune cells to form interface dermatitis. Created with Biorender.com. Plain language summaryLupus skin rashes arise during flares, after exposure to medications or sunlight, or in response to other triggers of inflammation. To understand how white blood cells enter the skin to cause these rashes, we used new technologies to look at proteins that attract them into the skin. We found proteins that are expressed by skin cells in lesions that can recruit specific types of white blood cells that are thought to be the initiators of skin rashes. Once in the skin, these and other white blood cells can make additional proteins that bring in more and more cells. We hope that our findings will be used to test new topical treatments for lupus and other autoimmune skin rashes.

immunology↗

bioRxiv2022

Increased Potency and Breadth of SARS-CoV-2 Neutralizing Antibodies After a Third mRNA Vaccine Dose

The omicron variant of SARS-CoV-2 infected very large numbers of SARS-CoV-2 vaccinated and convalescent individuals1-3. The penetrance of this variant in the antigen experienced human population can be explained in part by the relatively low levels of plasma neutralizing activity against Omicron in people who were infected or vaccinated with the original Wuhan-Hu-1 strain4-7. The 3rd mRNA vaccine dose produces an initial increase in circulating anti-Omicron neutralizing antibodies, but titers remain 10-20-fold lower than against Wuhan-Hu-1 and are, in many cases, insufficient to prevent infection7. Despite the reduced protection from infection, individuals that received 3 doses of an mRNA vaccine were highly protected from the more serious consequences of infection8. Here we examine the memory B cell repertoire in a longitudinal cohort of individuals receiving 3 mRNA vaccine doses9,10. We find that the 3rd dose is accompanied by an increase in, and evolution of, anti-receptor binding domain specific memory B cells. The increase is due to expansion of memory B cell clones that were present after the 2nd vaccine dose as well as the emergence of new clones. The antibodies encoded by these cells showed significantly increased potency and breadth when compared to antibodies obtained after the 2nd vaccine dose. Notably, the increase in potency was especially evident among newly developing clones of memory cells that differed from the persisting clones in targeting more conserved regions of the RBD. Overall, more than 50% of the analyzed neutralizing antibodies in the memory compartment obtained from individuals receiving a 3rd mRNA vaccine dose neutralized Omicron. Thus, individuals receiving 3 doses of an mRNA vaccine encoding Wuhan-Hu-1, have a diverse memory B cell repertoire that can respond rapidly and produce antibodies capable of clearing even diversified variants such as Omicron. These data help explain why a 3rd dose of an mRNA vaccine that was not specifically designed to protect against variants is effective against variant-induced serious disease.

immunology↗

bioRxiv2021

Antibody Evolution after SARS-CoV-2 mRNA Vaccination

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection produces B-cell responses that continue to evolve for at least one year. During that time, memory B cells express increasingly broad and potent antibodies that are resistant to mutations found in variants of concern1. As a result, vaccination of coronavirus disease 2019 (COVID-19) convalescent individuals with currently available mRNA vaccines produces high levels of plasma neutralizing activity against all variants tested1, 2. Here, we examine memory B cell evolution 5 months after vaccination with either Moderna (mRNA-1273) or Pfizer- BioNTech (BNT162b2) mRNA vaccines in a cohort of SARS-CoV-2 naive individuals. Between prime and boost, memory B cells produce antibodies that evolve increased neutralizing activity, but there is no further increase in potency or breadth thereafter. Instead, memory B cells that emerge 5 months after vaccination of naive individuals express antibodies that are similar to those that dominate the initial response. While individual memory antibodies selected over time by natural infection have greater potency and breadth than antibodies elicited by vaccination, the overall neutralizing potency of plasma is greater following vaccination. These results suggest that boosting vaccinated individuals with currently available mRNA vaccines will increase plasma neutralizing activity but may not produce antibodies with breadth equivalent to those obtained by vaccinating convalescent individuals.

immunology↗

bioRxiv2021

Vaccination boosts naturally enhanced neutralizing breadth to SARS-CoV-2 one year after infection

Over one year after its inception, the coronavirus disease-2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) remains difficult to control despite the availability of several excellent vaccines. Progress in controlling the pandemic is slowed by the emergence of variants that appear to be more transmissible and more resistant to antibodies1,2. Here we report on a cohort of 63 COVID-19-convalescent individuals assessed at 1.3, 6.2 and 12 months after infection, 41% of whom also received mRNA vaccines3,4. In the absence of vaccination antibody reactivity to the receptor binding domain (RBD) of SARS-CoV-2, neutralizing activity and the number of RBD-specific memory B cells remain relatively stable from 6 to 12 months. Vaccination increases all components of the humoral response, and as expected, results in serum neutralizing activities against variants of concern that are comparable to or greater than neutralizing activity against the original Wuhan Hu-1 achieved by vaccination of naive individuals2,5-8. The mechanism underlying these broad-based responses involves ongoing antibody somatic mutation, memory B cell clonal turnover, and development of monoclonal antibodies that are exceptionally resistant to SARS-CoV-2 RBD mutations, including those found in variants of concern4,9. In addition, B cell clones expressing broad and potent antibodies are selectively retained in the repertoire over time and expand dramatically after vaccination. The data suggest that immunity in convalescent individuals will be very long lasting and that convalescent individuals who receive available mRNA vaccines will produce antibodies and memory B cells that should be protective against circulating SARS-CoV-2 variants.

immunology↗