Two subsets of stem-like CD8+ memory T cell progenitors with distinct fate commitments in humans.

T cell memory relies on the generation of antigen-specific progenitors with stem-like properties. However, the identity of these progenitors has remained unclear, precluding a full understanding of the differentiation trajectories that underpin the heterogeneity of antigen-experienced T cells. We used a systematic approach guided by single-cell RNA-sequencing data to map the organizational structure of the human CD8+ memory T cell pool under physiological conditions. We identified two previously unrecognized subsets of clonally, epigenetically, functionally, phenotypically and transcriptionally distinct stem-like CD8+ memory T cells. Progenitors lacking the inhibitory receptors programmed death-1 (PD-1) and T cell immunoreceptor with Ig and ITIM domains (TIGIT) were committed to a functional lineage, whereas progenitors expressing PD-1 and TIGIT were committed to a dysfunctional, exhausted-like lineage. Collectively, these data reveal the existence of parallel differentiation programs in the human CD8+ memory T cell pool, with potentially broad implications for the development of immunotherapies and vaccines.

Antibody response to three-dose anti-SARS-CoV-2 mRNA-vaccination in treated solid cancer patients.

Solid cancer patients are at higher risk of SARS-CoV-2 infection and severe complications. Moreover, vaccine-induced antibody response is impaired in patients on anticancer treatment. In this retrospective, observational, hypothesis-generating, cohort study, we assessed the antibody response to the third dose of mRNA vaccine in a convenience sample of patients on anticancer treatment, comparing it to that of the primary two-dose cycle. Among 99 patients included, 62.6% were ≥60 years old, 32.3% males, 67.7% with advanced disease. Exactly 40.4% were receiving biological therapy, 16.2% chemotherapy only and 7.1% both treatments. After the third dose, seroconversion rate seems to increase significantly, especially in non-responders to two doses. Heterologous vaccine-type regimen (two-dose mRNA-1273 and subsequent tozinameran or vice versa) results in higher antibody levels. This explorative study suggests that repeated doses of mRNA-vaccines could be associated with a better antibody response in this population. Furthermore, heterologous vaccine-type three-dose vaccination seems more effective in this population. Since this is a hypothesis-generating study, adequately statistically powered studies should validate these results.

What do we mean by long COVID? A scoping review of the cognitive sequelae of SARS-CoV-2 infection.

BACKGROUND AND PURPOSE: Many COVID-19 patients report persistent symptoms, including cognitive disturbances. We performed a scoping review on this topic, focusing primarily on cognitive manifestations. METHODS: Abstracts and full texts of studies published on PubMed (until May 2023) addressing cognitive involvement persisting after SARS-CoV-2 infection were reviewed, focusing on terms used to name the cognitive syndrome, reported symptoms, their onset time and duration, and testing batteries employed. Reported psychiatric symptoms, their assessment tools, and more general manifestations were also extracted. RESULTS: Among the 947 records identified, 180 studies were included. Only one third of them used a label to define the syndrome. A minority of studies included patients according to stringent temporal criteria of syndrome onset (34%), whereas more studies reported a minimum required symptom duration (77%). The most frequently reported cognitive symptoms were memory and attentional-executive disturbances, and among psychiatric complaints, the most frequent were anxiety symptoms, depression, and sleep disturbances. Most studies reported fatigue among general symptoms. Thirty-six studies employed cognitive measures: screening tests alone (n = 19), full neuropsychological batteries (n = 25), or both (n = 29); 30 studies performed psychiatric testing. Cognitive deficits were demonstrated in 39% of subjects, the most frequently affected domains being attention/executive functions (90%) and memory (67%). CONCLUSIONS: Currently, no agreement exists on a label for post-COVID-19 cognitive syndrome. The time of symptom onset after acute infection and symptom duration are still discussed. Memory and attention-executive complaints and deficits, together with fatigue, anxiety, and depression symptoms, are consistently reported, but the objective evaluation of these symptoms is not standardized.

NK cells to cure cancer.

Natural Killer (NK) cells are innate lymphocytes able to mediate immune-surveillance and clearance of viral infected and tumor-transformed cells. Growing experimental and clinical evidence highlighted a dual role of NK cells either in the control of cancer development/progression or in promoting the onset of immune-suppressant tumor microenvironments. Indeed, several mechanisms of NK cell-mediated tumor escape have been described and these includes cancer-induced aberrant expression of activating and inhibitory receptors (i.e. NK cell immune checkpoints), impairments of NK cell migration to tumor sites and altered NK cell effector-functions. These phenomena highly contribute to tumor progression and metastasis formation. In this review, we discuss the latest insights on those NK cell receptors and related molecules that are currently being implemented in clinics either as possible prognostic factors or therapeutic targets to unleash NK cell anti-tumor effector-functions in vivo. Moreover, we address here the major recent advances in regard to the genetic modification and ex vivo expansion of anti-tumor specific NK cells used in innovative adoptive cellular transfer approaches.

Interaction of Bacteria, Immune Cells, and Surface Topography in Periprosthetic Joint Infections.

The incidence of periprosthetic joint infections (PJIs) is ~2% of total procedures and it is expected to rise due to an ageing population. Despite the large burden PJI has on both the individual and society, the immune response to the most commonly isolated pathogens, i.e., Staphylococcus aureus and Staphylococcus epidermidis, remains incompletely understood. In this work, we integrate the analysis of synovial fluids from patients undergoing hip and knee replacement surgery with in-vitro experimental data obtained using a newly developed platform, mimicking the environment of periprosthetic implants. We found that the presence of an implant, even in patients undergoing aseptic revisions, is sufficient to induce an immune response, which is significantly different between septic and aseptic revisions. This difference is confirmed by the presence of pro- and anti-inflammatory cytokines in synovial fluids. Moreover, we discovered that the immune response is also dependent on the type of bacteria and the topography of the implant surface. While S. epidermidis seems to be able to hide better from the attack of the immune system when cultured on rough surfaces (indicative of uncemented prostheses), S. aureus reacts differently depending on the contact surface it is exposed to. The experiments we performed in-vitro also showed a higher biofilm formation on rough surfaces compared to flat ones for both species, suggesting that the topography of the implant could influence both biofilm formation and the consequent immune response.

Comprehensive Phenotyping of Dendritic Cells in Cancer Patients by Flow Cytometry.

Dendritic cells (DCs) play a crucial role in the complex interplay between tumor cells and the immune system. During the elimination phase of cancer immunoediting, immunostimulatory DCs are critical for the control of tumor growth. During the escape phase, regulatory DCs sustain tumor tolerance and contribute to the development of the immunosuppressive tumor microenvironment that characterizes this phase. Moreover, increasing evidence indicates that DCs are also critical for the success of cancer immunotherapy. Hence, there is increasing need to fully characterize DC subsets and their activatory/inhibitory profile in cancer patients. In this review, we describe the role played by different DC subsets in the different phases of cancer immunoediting, the function exerted by different activatory and inhibitory molecules expressed on DC surface, and the cytokines produced by distinct DC subsets, in order to provide an overview on the DC features that may be useful to be assessed when dealing with the flow cytometric characterization of DCs in cancer patients. © 2020 International Society for Advancement of Cytometry.

CXCR3 Identifies Human Naive CD8+ T Cells with Enhanced Effector Differentiation Potential.

In mice, the ability of naive T (TN) cells to mount an effector response correlates with TCR sensitivity for self-derived Ags, which can be quantified indirectly by measuring surface expression levels of CD5. Equivalent findings have not been reported previously in humans. We identified two discrete subsets of human CD8+ TN cells, defined by the absence or presence of the chemokine receptor CXCR3. The more abundant CXCR3+ TN cell subset displayed an effector-like transcriptional profile and expressed TCRs with physicochemical characteristics indicative of enhanced interactions with peptide-HLA class I Ags. Moreover, CXCR3+ TN cells frequently produced IL-2 and TNF in response to nonspecific activation directly ex vivo and differentiated readily into Ag-specific effector cells in vitro. Comparative analyses further revealed that human CXCR3+ TN cells were transcriptionally equivalent to murine CXCR3+ TN cells, which expressed high levels of CD5. These findings provide support for the notion that effector differentiation is shaped by heterogeneity in the preimmune repertoire of human CD8+ T cells.

The OSMR Gene Is Involved in Hirschsprung Associated Enterocolitis Susceptibility through an Altered Downstream Signaling.

Hirschsprung (HSCR) Associated Enterocolitis (HAEC) is a common life-threatening complication in HSCR. HAEC is suggested to be due to a loss of gut homeostasis caused by impairment of immune system, barrier defense, and microbiome, likely related to genetic causes. No gene has been claimed to contribute to HAEC occurrence, yet. Genetic investigation of HAEC by Whole-Exome Sequencing (WES) on 24 HSCR patients affected (HAEC) or not affected (HSCR-only) by enterocolitis and replication of results on a larger panel of patients allowed the identification of the HAEC susceptibility variant p.H187Q in the Oncostatin-M receptor (OSMR) gene (14.6% in HAEC and 5.1% in HSCR-only, p = 0.0024). Proteomic analysis on the lymphoblastoid cell lines from one HAEC patient homozygote for this variant and one HAEC patient not carrying the variant revealed two well distinct clusters of proteins significantly up or downregulated upon OSM stimulation. A marked enrichment in immune response pathways (q < 0.0001) was shown in the HAEC H187 cell line, while proteins upregulated in the HAEC Q187 lymphoblasts sustained pathways likely involved in pathogen infection and inflammation. In conclusion, OSMR p.H187Q is an HAEC susceptibility variant and perturbates the downstream signaling cascade necessary for the gut immune response and homeostasis maintenance.

APOL1 polymorphism modulates sphingolipid profile of human podocytes.

Apolipoprotein L1 (APOL1) wild type (G0) plays a role in the metabolism of sphingolipids, glycosphingolipids, sphingomyelin and ceramide, which constitute bioactive components of the lipid rafts (DRM). We asked whether APOL1 variants (APOL1-Vs) G1 and G2 carry the potential to alter the metabolism of sphingolipids in human podocytes. The sphingolipid pattern in HPs overexpressing either APOL1G0 or APOL1-Vs was analysed by using a thin mono- and bi-dimensional layer chromatography, mass-spectrometry and metabolic labelling with [1-3H]sphingosine. HP G0 and G1/G2-Vs exhibit a comparable decrease in lactosylceramide and an increase in the globotriaosylceramide content. An analysis of the main glycohydrolases activity involved in glycosphingolipid catabolism showed an overall decrease in the activeness of the tested enzymes, irrespective of the type of APOL1-Vs expression. Similarly, the high throughput cell live-based assay showed a comparable increased action of the plasma membrane glycosphingolipid-glycohydrolases in living cells independent of the genetic APOL1 expression profile. Importantly, the most significative modification of the sphingolipid pattern induced by APOL1-Vs occurred in DRM resulted with a drastic reduction of radioactivity associated with sphingolipids. G1/G2-Vs present a decrease amount of globotriaosylceramide and globopentaosylceramide compared to G0. Additionally, ceramide at the DRM site and lactosylceramide in general, showed a greatest fall in G1/G2 in comparison with G0. Additionally, the levels of glucosylceramide decreased only in the DRM of human podocytes overexpressing G1/G2-Vs. These findings suggest that altered sphingolipidsprofiles may contribute to the deranged functionality of the plasma membrane in APOL1 risk milieu.

The challenges of primary biliary cholangitis: What is new and what needs to be done.

Primary Biliary Cholangitis (PBC) is an uncommon, chronic, cholangiopathy of autoimmune origin and unknown etiology characterized by positive anti-mitochondrial autoantibodies (AMA), female preponderance and progression to cirrhosis if left untreated. The diagnosis is based on AMA- or PBC-specific anti-nuclear antibody (ANA)-positivity in the presence of a cholestatic biochemical profile, histologic confirmation being mandatory only in seronegative cases. First-line treatment is ursodeoxycholic acid (UDCA), which is effective in preventing disease progression in about two thirds of the patients. The only approved second-line treatment is obeticholic acid. This article summarizes the most relevant conclusions of a meeting held in Lugano, Switzerland, from September 23rd-25th 2018, gathering basic and clinical scientists with various background from around the world to discuss the latest advances in PBC research. The meeting was dedicated to Ian Mackay, pioneer in the field of autoimmune liver diseases. The role of liver histology needs to be reconsidered: liver pathology consistent with PBC in AMA-positive individuals without biochemical cholestasis is increasingly reported, raising the question as to whether biochemical cholestasis is a reliable disease marker for both clinical practice and trials. The urgent need for new biomarkers, including more accurate markers of cholestasis, was also widely discussed during the meeting. Moreover, new insights in interactions of bile acids with biliary epithelia in PBC provide solid evidence of a role for impaired epithelial protection against potentially toxic hydrophobic bile acids, raising the fundamental question as to whether this bile acid-induced epithelial damage is the cause or the consequence of the autoimmune attack to the biliary epithelium. Strategies are needed to identify difficult-to-treat patients at an early disease stage, when new therapeutic approaches targeting immunologic pathways, in addition to bile acid-based therapies, may be effective. In conclusion, using interdisciplinary approaches, groundbreaking advances can be expected before long in respect to our understanding of the etiopathogenesis of PBC, with the ultimate aim of improving its treatment.

Proceedings From the Fourth Haploidentical Stem Cell Transplantation Symposium (HAPLO2016), San Diego, California, December 1, 2016.

The resurgence of haploidentical stem cell transplantation (HaploSCT) over the last decade is one of the most important advances in the field of hematopoietic stem cell transplantation (HSCT). The modified platforms of T cell depletion either ex vivo (CD34+ cell selection, "megadoses" of purified CD34+ cells, or selective depletion of T cells) or newer platforms of in vivo depletion of T cells, with either post-transplantation high-dose cyclophosphamide or intensified immune suppression, have contributed to better outcomes, with survival similar to that in HLA-matched donor transplantation. Further efforts are underway to control viral reactivation using modified T cells, improve immunologic reconstitution, and decrease the relapse rate post-transplantation using donor-derived cellular therapy products, such as genetically modified donor lymphocytes and natural killer cells. Improvements in treatment-related mortality have allowed the extension of haploidentical donor transplants to patients with hemoglobinopathies, such as thalassemia and sickle cell disease, and the possible development of platforms for immunotherapy in solid tumors. Moreover, combining HSCT from a related donor with solid organ transplantation could allow early tapering of immunosuppression in recipients of solid organ transplants and hopefully prevent organ rejection in this setting. This symposium summarizes some of the most important recent advances in HaploSCT and provides a glimpse in the future of fast growing field.

MFSD2A Promotes Endothelial Generation of Inflammation-Resolving Lipid Mediators and Reduces Colitis in Mice.

BACKGROUND & AIMS: Alterations in signaling pathways that regulate resolution of inflammation (resolving pathways) contribute to pathogenesis of ulcerative colitis (UC). The resolution process is regulated by lipid mediators, such as those derived from the ω-3 docosahexaenoic acid (DHA), whose esterified form is transported by the major facilitator superfamily domain containing 2A (MFSD2A) through the endothelium of brain, retina, and placenta. We investigated if and how MFSD2A regulates lipid metabolism of gut endothelial cells to promote resolution of intestinal inflammation. METHODS: We performed lipidomic and functional analyses of MFSD2A in mucosal biopsies and primary human intestinal microvascular endothelial cells (HIMECs) isolated from surgical specimens from patients with active, resolving UC and healthy individuals without UC (controls). MFSD2A was knocked down in HIMECs with small hairpin RNAs or overexpressed from a lentiviral vector. Human circulating endothelial progenitor cells that overexpress MFSD2A were transferred to CD1 nude mice with dextran sodium sulfate-induced colitis, with or without oral administration of DHA. RESULTS: Colonic biopsies from patients with UC had reduced levels of inflammation-resolving DHA-derived epoxy metabolites compared to healthy colon tissues or tissues with resolution of inflammation. Production of these metabolites by HIMECs required MFSD2A, which is required for DHA retention and metabolism in the gut vasculature. In mice with colitis, transplanted endothelial progenitor cells that overexpressed MFSD2A not only localized to the inflamed mucosa but also restored the ability of the endothelium to resolve intestinal inflammation, compared with mice with colitis that did not receive MFSD2A-overexpressing endothelial progenitors. CONCLUSIONS: Levels of DHA-derived epoxides are lower in colon tissues from patients with UC than healthy and resolving mucosa. Production of these metabolites by gut endothelium requires MFSD2A; endothelial progenitor cells that overexpress MFSD2A reduce colitis in mice. This pathway might be induced to resolve intestinal inflammation in patients with colitis.

Curtailed T-cell activation curbs effector differentiation and generates CD8+ T cells with a naturally-occurring memory stem cell phenotype.

Human T memory stem (TSCM ) cells with superior persistence capacity and effector functions are emerging as important players in the maintenance of long-lived T-cell memory and are thus considered an attractive population to be used in adoptive transfer-based immunotherapy of cancer. However, the molecular signals regulating their generation remain poorly defined. Here we show that curtailed T-cell receptor stimulation curbs human effector CD8+ T-cell differentiation and allows the generation of CD45RO- CD45RA+ CCR7+ CD27+ CD95+ -phenotype cells from highly purified naïve T-cell precursors, resembling naturally-occurring human TSCM . These cells proliferate extensively in vitro and in vivo, express low amounts of effector-associated genes and transcription factors and undergo considerable self-renewal in response to IL-15 while retaining effector differentiation potential. Such a phenotype is associated with a lower number of mitochondria compared to highly-activated effector T cells committed to terminal differentiation. These results shed light on the molecular signals that are required to generate long-lived memory T cells with potential application in adoptive cell transfer immunotherapy.

The yin-yang of the interaction between myelomonocytic cells and NK cells.

NK cells are innate lymphoid cells, which play a key role in the immune response to cancer and pathogens and participate in the shaping of adaptive immunity. NK cells engage in a complex bidirectional interaction with myelomonocytic cells. In particular, macrophages, dendritic cells and neutrophils promote differentiation and effector function of NK cells and, on the other hand, myelomonocytic cells express triggers of checkpoint blockade (eg PD-L1) and other immunosuppressive molecules, which negatively regulate NK cell function. In addition, NK cells express high levels of IL-1R8, which acts as a checkpoint for IL-18 driven differentiation and activation of NK cells. Evidence suggests that targeting the myeloid cell-NK cell crosstalk unleashes effective anti-tumour and anti-viral resistance.

The early expansion of anergic NKG2Apos/CD56dim/CD16neg natural killer represents a therapeutic target in haploidentical hematopoietic stem cell transplantation.

Natural killer cells are the first lymphocyte population to reconstitute early after non-myeloablative and T cell-replete haploidentical hematopoietic stem cell transplantation with post-transplant infusion of cyclophosphamide. The study herein characterizes the transient and predominant expansion starting from the second week following haploidentical hematopoietic stem cell transplantation of a donor-derived unconventional subset of NKp46neg-low/CD56dim/CD16neg natural killer cells expressing remarkably high levels of CD94/NKG2A. Both transcription and phenotypic profiles indicated that unconventional NKp46neg-low/CD56dim/CD16neg cells are a distinct natural killer cell subpopulation with features of late stage differentiation, yet retaining proliferative capability and functional plasticity to generate conventional NKp46pos/CD56bright/CD16neg-low cells in response to interleukin-15 plus interleukin-18. While present at low frequency in healthy donors, unconventional NKp46neg-low/CD56dim/CD16neg cells are greatly expanded in the seven weeks following haploidentical hematopoietic stem cell transplantation, and express high levels of the activating receptors NKG2D and NKp30 as well as of the lytic granules Granzyme-B and Perforin. Nonetheless, NKp46neg-low/CD56dim/CD16neg cells displayed a markedly defective cytotoxicity that could be reversed by blocking the inhibitory receptor CD94/NKG2A. These data open new and important perspectives to better understand the ontogenesis/homeostasis of human natural killer cells and to develop a novel immune-therapeutic approach that targets the inhibitory NKG2A check-point, thus unleashing natural killer cell alloreactivity early after haploidentical hematopoietic stem cell transplantation.

Tissue-resident and memory properties of human T-cell and NK-cell subsets.

Efficient immune responses to invading pathogens are the result of the complex but coordinated synergy between a variety of cell types from both the innate and adaptive arms of the immune system. While adaptive and innate immune responses are highly complementary, some cells types within these two systems perform similar functions, underscoring the need for redundancy and increased flexibility. In this review, we will discuss the striking shared features of immunological memory and tissue residency recently discovered between T cells, a component of the adaptive immune system, and natural killer (NK) cells, members generally assigned to the innate compartment. Specifically, we will focus on the T-cell and NK-cell diversity at the single-cell level, on the discrete function of specific subsets, and on their anatomical location. Finally, we will discuss the implication of such diversity in the generation of long-term memory.

Role of naive-derived T memory stem cells in T-cell reconstitution following allogeneic transplantation.

Early T-cell reconstitution following allogeneic transplantation depends on the persistence and function of T cells that are adoptively transferred with the graft. Posttransplant cyclophosphamide (pt-Cy) effectively prevents alloreactive responses from unmanipulated grafts, but its effect on subsequent immune reconstitution remains undetermined. Here, we show that T memory stem cells (TSCM), which demonstrated superior reconstitution capacity in preclinical models, are the most abundant circulating T-cell population in the early days following haploidentical transplantation combined with pt-Cy and precede the expansion of effector cells. Transferred naive, but not TSCM or conventional memory cells preferentially survive cyclophosphamide, thus suggesting that posttransplant TSCM originate from naive precursors. Moreover, donor naive T cells specific for exogenous and self/tumor antigens persist in the host and contribute to peripheral reconstitution by differentiating into effectors. Similarly, pathogen-specific memory T cells generate detectable recall responses, but only in the presence of the cognate antigen. We thus define the cellular basis of T-cell reconstitution following pt-Cy at the antigen-specific level and propose to explore naive-derived TSCM in the clinical setting to overcome immunodeficiency. These trials were registered at www.clinicaltrials.gov as #NCT02049424 and #NCT02049580.

Priming of Human Resting NK Cells by Autologous M1 Macrophages via the Engagement of IL-1ß, IFN-ß, and IL-15 Pathways.

The cross talk between NK cells and macrophages is emerging as a major line of defense against microbial infections and tumors. This study reveals a complex network of soluble mediators and cell-to-cell interactions allowing human classically activated (M1) macrophages, but not resting (M0) or alternatively activated (M2) macrophages, to prime resting autologous NK cells. In this article, we show that M1 increase NK cell cytotoxicity by IL-23 and IFN-ß-dependent upregulation of NKG2D, IL-1ß-dependent upregulation of NKp44, and trans-presentation of IL-15. Moreover, both IFN-ß-dependent cis-presentation of IL-15 on NK cells and engagement of the 2B4-CD48 pathway are used by M1 to trigger NK cell production of IFN-γ. The disclosure of these synergic cellular mechanisms regulating the M1-NK cell cross talk provides novel insights to better understand the role of innate immune responses in the physiopathology of tumor biology and microbial infections.

Proceedings from the Second Haploidentical Stem Cell Transplantation Symposium-Haplo2014, San Francisco, California, December 4, 2014.

Significant progress has been made over the past decade in haploidentical transplantation, with the development of novel methods to control intense alloreactive reactions generated in the major HLA-mismatched setting. Application of post-transplantation cyclophosphamide has gained worldwide acceptance as an effective and low-cost way to perform this type of transplantation, with outcomes now similar to those from HLA-matched unrelated donors. These advances have resulted in improved treatment-related mortality, whereas disease relapse has emerged as the most common cause of treatment failure. In addition, improvements in immunologic reconstitution after transplantation are much needed, not only in haploidentical transplantation but in all forms of stem cell transplantation. This symposium has focused on some of the most promising methods to control alloreactivity in this form of transplantation and application of cellular therapy to prevent disease relapse after transplantation, as well as understanding immunologic reconstitution and foreseeable approaches to improve immune recovery after transplantation.