Understanding heterogeneity of human bone marrow plasma cell maturation and survival pathways by single-cell analyses.

Human bone marrow (BM) plasma cells are heterogeneous, ranging from newly arrived antibody-secreting cells (ASCs) to long-lived plasma cells (LLPCs). We provide single-cell transcriptional resolution of 17,347 BM ASCs from five healthy adults. Fifteen clusters are identified ranging from newly minted ASCs (cluster 1) expressing MKI67 and high major histocompatibility complex (MHC) class II that progress to late clusters 5-8 through intermediate clusters 2-4. Additional ASC clusters include the following: immunoglobulin (Ig) M predominant (likely of extra-follicular origin), interferon responsive, and high mitochondrial activity. Late ASCs are distinguished by G2M checkpoints, mammalian target of rapamycin (mTOR) signaling, distinct metabolic pathways, CD38 expression, utilization of tumor necrosis factor (TNF)-receptor superfamily members, and two distinct maturation pathways involving TNF signaling through nuclear factor κB (NF-κB). This study provides a single-cell atlas and molecular roadmap of LLPC maturation trajectories essential in the BM microniche. Altogether, understanding BM ASC heterogeneity in health and disease enables development of new strategies to enhance protective ASCs and to deplete pathogenic ones.

The microbiome and rheumatic heart disease: current knowledge and future perspectives.

Rheumatic heart disease (RHD) is a cardiovascular disease caused by an autoimmune response to group A Streptococcus (GAS) infection resulting in the damage of heart valves. RHD is the most commonly acquired heart disease among children and young adults with a global burden of over 40 million cases accounting for 306,000 deaths annually. Inflammation in the heart valves caused due to molecular mimicry between the GAS antigens and host cardiac proteins is facilitated by cytokines, cross-reactive antibodies and CD4+ T cells. The complex interaction between genetic and environmental factors linked with erratic events leads to the loss of immunological tolerance and autoimmunity in RHD. Despite extensive research on the etiopathogenesis of RHD, the precise mechanism underpinning the initiation of acute rheumatic fever (ARF) to the progression of RHD still remains elusive. Mounting evidences support the contribution of the human microbiome in the development of several immune-mediated diseases including rheumatoid arthritis, juvenile idiopathic arthritis, Kawasaki disease, inflammatory bowel disease and type 1 diabetes. The microbiome and their metabolites could play a crucial role in the integrity of the epithelial barrier, development of the immune system, inflammation and differentiation of T cell subsets. Consequently, microbiome dysbiosis might result in autoimmunity by molecular mimicry, epitope spreading and bystander activation. This review discusses various aspects of the interaction between the microbiome and the immune system in order to reveal causative links relating dysbiosis and autoimmune diseases with special emphasis on RHD.

Human Bone Marrow Plasma Cell Atlas: Maturation and Survival Pathways Unraveled by Single Cell Analyses.

Human bone marrow (BM) plasma cells are heterogeneous, ranging from newly arrived antibody-secreting cells (ASC) to long-lived plasma cells (LLPC). We provide single cell transcriptional resolution of 17,347 BM ASC from 5 healthy adults. Fifteen clusters were identified ranging from newly minted ASC (cluster 1) expressing MKI67 and high MHC Class II that progressed to late clusters 5-8 through intermediate clusters 2-4. Additional clusters included early and late IgM-predominant ASC of likely extra-follicular origin; IFN-responsive; and high mitochondrial activity ASC. Late ASCs were distinguished by differences in G2M checkpoints, MTOR signaling, distinct metabolic pathways, CD38 expression, and utilization of TNF-receptor superfamily members. They mature through two distinct paths differentiated by the degree of TNF signaling through NFKB. This study provides the first single cell resolution atlas and molecular roadmap of LLPC maturation, thereby providing insight into differentiation trajectories and molecular regulation of these essential processes in the human BM microniche. This information enables investigation of the origin of protective and pathogenic antibodies in multiple diseases and development of new strategies targeted to the enhancement or depletion of the corresponding ASC. One Sentence Summary: The single cell transcriptomic atlas of human bone marrow plasma cell heterogeneity shows maturation of class-switched early and late subsets, specific IgM and Interferon-driven clusters, and unique heterogeneity of the late subsets which encompass the long-lived plasma cells.

Individualized Constellation of Killer Cell Immunoglobulin-Like Receptors and Cognate HLA Class I Ligands that Controls Natural Killer Cell Antiviral Immunity Predisposes COVID-19.

Background: The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection causes coronavirus disease-2019 (COVID-19) in some individuals, while the majority remain asymptomatic. Natural killer (NK) cells play an essential role in antiviral defense. NK cell maturation and function are regulated mainly by highly polymorphic killer cell immunoglobulin-like receptors (KIR) and cognate HLA class I ligands. Herein, we tested our hypothesis that the individualized KIR and HLA class I ligand combinations that control NK cell function determine the outcome of SARS-CoV-2 infection. Methods: We characterized KIR and HLA genes in 200 patients hospitalized for COVID-19 and 195 healthy general population controls. Results: The KIR3DL1+HLA-Bw4+ [Odds ratio (OR) = 0.65, p = 0.03] and KIR3DL2+HLA-A3/11+ (OR = 0.6, p = 0.02) combinations were encountered at significantly lower frequency in COVID-19 patients than in the controls. Notably, 40% of the patients lacked both of these KIR+HLA+ combinations compared to 24.6% of the controls (OR = 2.04, p = 0.001). Additionally, activating receptors KIR2DS1+KIR2DS5+ are more frequent in patients with severe COVID-19 than patients with mild disease (OR = 1.8, p = 0.05). Individuals carrying KIR2DS1+KIR2DS5+ genes but missing either KIR3DL1+HLA-Bw4+ combination (OR = 1.73, p = 0.04) or KIR3DL2+HLA-A3/11+ combination (OR = 1.75, p = 0.02) or both KIR3DL1+HLA-Bw4+ and KIR2DL2+HLA-A3/11+ combinations (OR = 1.63, p = 0.03) were more frequent in the COVID-19 cohort compared to controls. Conclusions: The absence of KIR3DL1+HLA-Bw4+ and KIR3DL2+HLA-A3/11+ combinations presumably yields inadequate NK cell maturation and reduces anti-SARS-CoV-2 defense, causing COVID-19. An increased frequency of KIR2DS1+KIR2DS5+ in severe COVID-19 patients suggests vigorous NK cell response triggered via these activating receptors and subsequent production of exuberant inflammatory cytokines responsible for severe COVID-19. Our results demonstrate that specific KIR-HLA combinations that control NK cell maturation and function are underlying immunogenetic variables that determine the dual role of NK cells in mediating beneficial antiviral and detrimental pathologic action. These findings offer a framework for developing potential host genetic biomarkers to distinguish individuals prone to COVID-19.

Concurrent typing of over 4000 samples by long-range PCR amplicon-based NGS and rSSO revealed the need to verify NGS typing for HLA allelic dropouts.

Hematopoietic stem cell transplantation (HSCT) from HLA-matched donors significantly decreases the risks of graft-rejection and graft-versus-host disease. Long-range PCR- amplicon-based next-generation sequencing (NGS) is increasingly used as a standalone method in clinical laboratories to determine HLA compatibility for HSCT and solid-organ transplantation. We hypothesized that an allelic dropout is a frequent event in the long-range PCR amplicon-based NGS HLA typing method. To test the hypothesis, we typed 4,006 samples concurrently using a commercially available long-range PCR amplicon-based NGS-typing and short exon-specific amplicon-based reverse sequence-specific oligonucleotide (rSSO) methods. The concordance between the NGS and rSSO typing results was 100% at HLA-A, -B, -C, -DRB1, -DRB3, -DRB5, -DQA1, DPA1 loci. However, 4.5% of the samples (179/4006) showed allelic-dropouts at one of the other three loci: HLA-DRB4 (3.9%), HLA-DPB1 (0.4%), and HLA-DQB1*(0.15%). The allelic-dropouts are not associated with specific haplotypes, and some dropouts can be reagent lot-specific. Although DRB1-DRB3/4/5-DQB1 linkages help to diagnose these allelic-dropouts in some cases, the rSSO typing was crucial to identify the dropouts in DQB1 and DPB1 loci. These results uncover the critical limitations of using long-range PCR amplicon-based NGS as a standalone method in clinical histocompatibility laboratories and advocate the need for strategies to diagnose and resolve allelic-dropouts.

Inhibitory natural killer cell receptor KIR3DL1 with its ligand Bw4 constraints HIV-1 disease among South Indians.

OBJECTIVE: To investigate the role of genotypic and phenotypic characteristics of killer cell immunoglobulin-like receptors (KIRs) and their human leukocyte antigen (HLA) class-1 ligands in HIV-1 disease progression. STUDY DESIGN AND METHODS: This is a nested case-control study including 347 HIV seropositive (HIV-1+) individuals from South India constituting 45 long-term nonprogressors (LTNPs) and 302 disease progressors. KIR genotyping was performed by multiplex sequence-specific primer-directed PCR (SSP-PCR). Phenotypic expressions of KIR3DL1/S1 was studied using multiparametric flow cytometry assay. HLA-Bw4 and Bw6 epitopes were determined by ARMS-PCR. HLA-Bw4I80, HLA-Bw4T80, HLA-C1, HLA-C2, and HLA-Aw4 were genotyped using SSP-PCR. Serum levels of IFN-γ was quantified using ELISA method. RESULTS: Overall, 37 different KIR genotypes were observed and the distribution of genotypes with AB-AB (OR = 2.2, P = 0.033) constellations showed significant increase among LTNPs. The frequencies of 3DL1-2DL3-2DL5 (OR = 2.2, Pc = 0.031), 3DL1-Bw4/Aw4 (OR = 2.49, Pc = 0.019), homozygous Bw4 (OR = 2.422, Pc = 0.011) were observed higher in LTNPs and 2DS1-2DS2-2DS3 (OR =  0.475, Pc = 0.03), homozygous Bw6 (OR = 0.413, Pc = 0.011) were higher in the disease progressors. Flow cytometry assay showed the increased expression and maintenance of 3DL1/S1+NK cells in LTNPs (P = 0.0001). Further the expansion of 3DS1+NK cells was higher than 3DL1+NK cells in the heterozygous 3DL1/S1 LTNPs (P = 0.001). CONCLUSION: The inhibitory receptor 3DL1 with Bw4 and its A-haplotype defining KIR genes (2DL3/L5) confers protection against HIV-1 disease progression. An increased expression and maintenance of 3DL1/S1+ natural killer cells may contribute to the efficient activation of the natural killer cells and subsequent long-term nonprogression (LTNPn) to the disease.

Predominance of B haplotype associated KIR genes in Tamil Speaking Dravidians.

Killer cell immunoglobulin like receptors (KIRs) are a group of activating (aKIRs) and inhibitory receptors (iKIRs) expressed on subsets of lymphoid cells. Their interaction with HLA class I molecules modulate the innate and adaptive immune response against infections and malignancies. KIR haplotypes varies in gene content and also at allelic level, thereby, distinguishing individuals and populations. Hence, the present study is aimed to determine the KIR gene diversity in Piramalai Kallar (PK) population of South India. The PK population shows diverged KIR gene frequencies and novel haplotypes than other South Indian populations. 52 different KIR gene profiles were identified and 18 of them were new in this population. In phylogenetic analysis the study population is positioned between African and Iranian population in the clade, which supports the South African ancestry of Indian population.