Omicron BA.1 Mutations in SARS-CoV-2 Spike Lead to Reduced T-Cell Response in Vaccinated and Convalescent Individuals

Omicron BA.1 variant can readily infect people with vaccine-induced or naturally acquired SARS-CoV-2 immunity facilitated by escape from neutralizing antibodies. In contrast, T-cell reactivity against the Omicron BA.1 variant seems relatively well preserved. Here, we studied the preexisting T cells elicited by either vaccination with the mRNA-based BNT162b2 vaccine or by natural infection with ancestral SARS-CoV-2 for their cross-reactive potential to 20 selected CD4⁺ T-cell epitopes of spike-protein-harboring Omicron BA.1 mutations. Although the overall memory CD4⁺ T-cell responses primed by the ancestral spike protein was still preserved generally, we show here that there is also a clear loss of memory CD4⁺ T-cell cross-reactivity to immunodominant epitopes across the spike protein due to Omicron BA.1 mutations. Complete or partial loss of preexisting T-cell responsiveness was observed against 60% of 20 nonconserved CD4⁺ T-cell epitopes predicted to be presented by a broad set of common HLA class II alleles. Monitoring such mutations in circulating strains helps predict which virus variants may escape previously induced cellular immunity and could be of concern.     

Human T-Cell Leukemia Virus Type 1 Envelope Protein: Post-Entry Roles in Viral Pathogenesis

Human T-cell leukemia virus type 1 (HTLV-1) is an oncogenic retrovirus that is the causative infectious agent of adult T-cell leukemia/lymphoma (ATL), an aggressive and fatal CD4⁺ T-cell malignancy, and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a chronic neurological disease. Disease progression in infected individuals is the result of HTLV-1-driven clonal expansion of CD4⁺ T-cells and is generally associated with the activities of the viral oncoproteins Tax and Hbz. A closely related virus, HTLV-2, exhibits similar genomic features and the capacity to transform T-cells, but is non-pathogenic. In vitro, HTLV-1 primarily immortalizes or transforms CD4⁺ T-cells, while HTLV-2 displays a transformation tropism for CD8⁺ T-cells. This distinct tropism is recapitulated in infected people. Through comparative studies, the genetic determinant for this divergent tropism of HTLV-1/2 has been mapped to the viral envelope (Env). In this review, we explore the emerging roles for Env beyond initial viral entry and examine current perspectives on its contributions to HTLV-1-mediated disease development.     

High and Sustained Ex Vivo Frequency but Altered Phenotype of SARS-CoV-2-Specific CD4+ T-Cells in an Anti-CD20-Treated Patient with Prolonged COVID-19

Here, we longitudinally assessed the ex vivo frequency and phenotype of SARS-CoV-2 membrane protein (aa145–164) epitope-specific CD4⁺ T-cells of an anti-CD20-treated patient with prolonged viral positivity in direct comparison to an immunocompetent patient through an MHC class II DRB1*11:01 Tetramer analysis. We detected a high and stable SARS-CoV-2 membrane-specific CD4⁺ T-cell response in both patients, with higher frequencies of virus-specific CD4⁺ T-cells in the B-cell-depleted patient. However, we found an altered virus-specific CD4⁺ T-cell memory phenotype in the B-cell-depleted patient that was skewed towards late differentiated memory T-cells, as well as reduced frequencies of SARS-CoV-2-specific CD4⁺ T-cells with CD45RA⁻ CXCR5⁺ PD-1⁺ circulating T follicular helper cell (cT_FH) phenotype. Furthermore, we observed a delayed contraction of CD127⁻ virus-specific effector cells. The expression of the co-inhibitory receptors TIGIT and LAG-3 fluctuated on the virus-specific CD4⁺ T-cells of the patient, but were associated with the inflammation markers IL-6 and CRP. Our findings indicate that, despite B-cell depletion and a lack of B-cell—T-cell interaction, a robust virus-specific CD4⁺ T-cell response can be primed that helps to control the viral replication, but which is not sufficient to fully abrogate the infection.     

Differential escape of HCV from CD8+ T cell selection pressure between China and Germany depends on the presenting HLA class I molecule

Adaptation of hepatitis C virus (HCV) to CD8⁺ T cell selection pressure is well described; however, it is unclear if HCV differentially adapts in different populations. Here, we studied HLA class I‐associated viral sequence polymorphisms in HCV 1b isolates in a Chinese population and compared viral substitution patterns between Chinese and German populations. We identified three HLA class I‐restricted epitopes in HCV NS3 with statistical support for selection pressure and found evidence for differential escape pathways between isolates from China and Germany depending on the HLA class I molecule. The substitution patterns particularly differed in the epitope VTLTHPITK_1635‐1643, which was presented by HLA‐A*03 as well as HLA‐A*11, two alleles with highly different frequencies in the two populations. In Germany, a substitution in position seven of the epitope was the most frequent substitution in the presence of HLA‐A*03, functionally associated with immune escape and nearly absent in Chinese isolates. In contrast, the most frequent substitution in China was located at position two of the epitope and became the predominant consensus residue. Moreover, substitutions in position one of the epitope were significantly enriched in HLA‐A*11‐positive individuals in China and associated with different patterns of CD8⁺ T cell reactivity. Our study confirms the differential escape pathways selected by HCV that depended on different HLA class I alleles in Chinese and German populations, indicating that HCV differentially adapts to distinct HLA class I alleles in these populations. This result has important implications for vaccine design against highly variable and globally distributed pathogens, which may require matching antigen sequences to geographic regions for T cell‐based vaccine strategies.     

Identification of HLA-A2 restricted T-cell epitopes within the conserved region of the immunoglobulin G heavy-chain in patients with multiple myeloma

Objective: The aim of this study is the identification of HLA‐A2 restricted T‐cell epitopes in the conserved region of the immunoglobulin‐G‐heavy‐chain (IgG_H) that can be used for immunotherapy in multiple myeloma (MM) patients. Methods: After the IgG_H gene sequence was scanned for HLA‐A2 restricted T‐cell epitopes with a high binding affinity to the MHC‐I‐complex, promising nona‐peptides were synthesized. Peptide specific CD8 ⁺ T‐cells were generated from peripheral blood mononuclear cells (PBMC) of healthy donors (HD) and patients with MM using peptide pulsed dendritic cells (DC) in vitro. The activation and cytotoxicity of CD8 ⁺ T‐cells was analyzed by IFN‐α ELISpot‐assay and ⁵¹Chromium release‐assay. HLA‐A2 restriction was proven by blocking T‐cell activation with anti‐HLA‐A2 antibodies. Results: Two HLA‐A2 restricted T‐cell epitopes–TLVTVSSAS derived from the IgG_H‐framework‐region 4 (FR4) and LMISRTPEV from the constant region (CR)‐induced expansion of specific CD8 ⁺ T‐cells from PBMC in two of three (TLVTVSSAS) and one of three (LMISRTPEV) HD respectively. Specific T‐cells were induced from PBMC in two of six (TLVTVSSAS) and eight of 19 (LMISRTPEV) patients with MM. Specific CD8 ⁺ T‐cells also lysed peptide‐pulsed target cells in ⁵¹Chromium release‐assay. LMISRTPEV specific CD8 ⁺ T‐cells from MM patients lysed specifically the HLA‐A2 ⁺ IgG myeloma cell line XG‐6. Conclusion: We identified two HLA‐A2 restricted T‐cell epitopes–TLVTVSSAS and LMISRTPEV–which can yield an expansion of CD8 ⁺ T‐cells with the ability to kill peptide‐loaded target cells and HLA‐A2 ⁺ IgG⁺ myeloma cells. We conclude that TLVTVSSAS and LMISRTPEV could be T‐cell epitopes for immunotherapy in MM patients.     

MicroRNA-Mediated Restriction of HIV-1 in Resting CD4+ T Cells and Monocytes

In contrast to activated CD4⁺ T cells and differentiated macrophages, resting CD4⁺ T cells and monocytes are non-permissive for HIV-1 replication. The mediators which regulate the resting or quiescent phenotype are often actively involved in the restriction of viral replication and the establishment and maintenance of viral latency. Recently, certain microRNAs which are highly expressed in resting cells have been implicated in this capacity, inhibiting the expression of cellular proteins that are also viral co-factors; following activation these microRNAs exhibit decreased expression, while their targets are correspondingly up-regulated, contributing to a favorable milieu for virus replication. Other microRNAs exhibiting a similar expression pattern in resting and activated cells have been shown to directly target the HIV-1 genome. In this review we will discuss the resting state and the causes behind viral restriction in resting cells, with emphasis on the role of microRNAs.     

Heightened self-reactivity associated with selective survival,but not expansion,of na?ve virus-specific CD8+ T cells in aged mice

In advanced age, decreased CD8⁺ cytotoxic T-lymphocyte (CTL) responses to novel pathogens and cancer is paralleled by a decline in the number and function of naïve CTL precursors (CTLp). Although the age-related fall in CD8⁺ T-cell numbers is well established, neither the underlying mechanisms nor the extent of variation for different epitope specificities have been defined. Furthermore, naïve CD8⁺ T cells expressing high levels of CD44 accumulate with age, but it is unknown whether this accumulation reflects their preferential survival or an age-dependent driver of CD8⁺ T-cell proliferation. Here, we track the number and phenotype of four influenza A virus (IAV)-specific CTLp populations in naïve C57BL/6 (B6) mice during aging, and compare T-cell receptor (TCR) clonal diversity for the CD44hi and CD44lo subsets of one such population. We show differential onset of decline for several IAV-specific CD8⁺ T-cell populations with advanced age that parallel age-associated changes in the B6 immunodominance hierarchy, suggestive of distinct impacts of aging on different epitope-specific populations. Despite finding no evidence of clonal expansions in an aged, epitope-specific TCR repertoire, nonrandom alterations in TCR usage were observed, along with elevated CD5 and CD8 coreceptor expression. Collectively, these data demonstrate that naïve CD8⁺ T cells expressing markers of heightened self-recognition are selectively retained, but not clonally expanded, during aging.Given that CD8⁺ cytotoxic T-lymphocyte (CTL) immunity is critical for the control of viruses and tumors, specific defects are likely to contribute substantially to overall immune dysfunction. Normal aging is associated with increased health risk, as vaccine efficacy wanes and susceptibility to, and severity of, a variety of infections and malignancies is enhanced (1, 2). Whereas aging compromises a number of arms of the immune system (3), studies in mice and humans have demonstrated deficits that are intrinsic to naïve CTL populations (4–6). Thus, a complete understanding of both age-related CTL deficiencies and the underlying mechanisms is critical.The magnitude of the response, the diversity of T-cell receptor (TCR)-defined clonal recruitment, and the avidity of TCR binding to cognate peptides in complex with MHC class I glycoproteins (pMHCI) are all key determinants of CTL response efficacy (7). Each of these factors is substantially constrained by their respective characteristics in the naive CTL precursor (CTLp) pool (7–10). During aging, both the number and TCR diversity of naïve polyclonal and epitope-specific CD8⁺ T-cell sets decreases (11–13). In addition, a large proportion (∼60–80%) of the remaining naïve CD8⁺ T cells, termed virtual memory (T_VM) cells, express high levels of CD44, traditionally regarded as a marker of T-cell activation and suggestive of proliferation (14). It is unclear whether the accumulation of T_VM cells with age (15) represents preferential retention, de novo generation, or expansion of the T_VM subset. TCR repertoire analyses show emerging TCR bias with age (11, 13, 16), limiting the diversity of the TCR repertoire beyond that defined by reduced CTLp numbers. Although it is clear that TCR biases parallel age-related T-cell loss, the relative impact of selective clonal decay versus clonal expansion remains unresolved, as do the key determinants of naïve CTLp survival.The frequency and TCR usage of naïve and immune CD8⁺ T cells specific for a range of influenza A virus (IAV) epitopes is well characterized for B6 mice (17–20), providing a convenient experimental system for investigating the characteristics and drivers of age-related CD8⁺ T-cell decline. Primary responses to the D^bNP_366–374 and D^bPA_224–233 epitopes (derived from nucleoprotein and acid polymerase proteins, respectively) are immunodominant in young adult mice, whereas those to a polymerase B subunit 1 epitope (K^bPB1_703–711) and an epitope derived from a shifted reading frame of PB1 (D^bPB1-F2_62–70) are subdominant (21). This numerical immunodominance hierarchy shifts with aging, with a decrease in D^bNP₃₆₆-specific responses alongside an increase in K^bPB1₇₀₃- and D^bPB1-F2₆₂–specific responses (13, 22). It has been suggested that this reflects the stochastic decay of CTLp (13), but the rate of decline of individual epitope-specific populations has not been directly assessed during aging.Here, we directly track the numbers of immune and naive IAV epitope-specific CD8⁺ T-cell populations through the course of aging, together with the phenotypic and clonotypic characteristics of a naïve CTLp set. We show that the shift in immunodominance across D^bNP₃₆₆, D^bPA₂₂₄, K^bPB1₇₀₃, and D^bPB1-F2₆₂ for 12-mo-old versus 3-mo mice can be accounted for by differential onset of decline across naïve CTLp populations. Moreover, phenotypic changes in a naïve CTLp set indicates that age-related clonal persistence is associated with the capacity to recognize self-pMHCI, which may, in turn, alter the capacity of CTLps to responding to novel challenges.     

High levels of CD34+CD38low/-CD123+ blasts are predictive of an adverse outcome in acute myeloid leukemia: a Groupe Ouest-Est des Leucemies Aigues et Maladies du Sang (GOELAMS) study

Background

Acute myeloid leukemias arise from a rare population of leukemic cells, known as leukemic stem cells, which initiate the disease and contribute to frequent relapses. Although the phenotype of these cells remains unclear in most patients, these cells are enriched within the CD34⁺CD38^low/− compartment expressing the interleukin-3 alpha chain receptor, CD123. The aim of this study was to determine the prognostic value of the percentage of blasts with the CD34⁺CD38^low/−CD123⁺ phenotype.

Design and Methods

The percentage of CD34⁺CD38^low/−CD123⁺ cells in the blast population was determined at diagnosis using flow cytometry. One hundred and eleven patients under 65 years of age with de novo acute myeloid leukemia and treated with intensive chemotherapy were retrospectively included in the study. Correlations with complete response, disease-free survival and overall survival were evaluated with univariate and multivariate analyses.

Results

A proportion of CD34⁺CD38^low/−CD123⁺ cells greater than 15% at diagnosis and an unfavorable karyotype were significantly correlated with a lack of complete response. By logistic regression analysis, a percentage of CD34⁺CD38^low/−CD123⁺ higher than 15% retained significance with an odds ratio of 0.33 (0.1–0.97; P=0.044). A greater than 1% population of CD34⁺CD38^low/−CD123⁺ cells negatively affected disease-free survival (0.9 versus 4.7 years; P<0.0001) and overall survival (1.25 years versus median not reached; P<0.0001). A greater than 1% population of CD34⁺CD38^low/−CD123⁺ cells retained prognostic significance for both parameters after multivariate analysis.

Conclusions

The percentage of CD34⁺CD38^low/−CD123⁺ leukemic cells at diagnosis was significantly correlated with response to treatment and survival. This prognostic marker might be easily adopted in clinical practice to rapidly identify patients at risk of treatment failure.     

Quantifying Susceptibility of CD4+ Stem Memory T-Cells to Infection by Laboratory Adapted and Clinical HIV-1 Strains

CD4⁺ T cells are principal targets for human immunodeficiency virus type 1 (HIV-1) infection. CD4⁺ T cell subsets are heterogeneous cell populations, divided by functional and phenotypic differences into naïve and memory T cells. The memory CD4⁺ T cells are further segregated into central, effector and transitional memory cell subsets by functional, phenotypic and homeostatic characteristics. Defining the distribution of HIV-1 infection in different T cell subsets is important, as this can play a role in determining the size and composition of the viral reservoir. Both central memory and transitional memory CD4⁺ T cells have been described as long-lived viral reservoirs for HIV. Recently, the newly described stem memory T cell subset has also been implicated as a long-lived HIV reservoir. Using green fluorescent protein (GFP) reporter strains of HIV-1 and multi parameter flow cytometry, we developed an assay to simultaneously quantify the susceptibility of stem memory (TSCM), central memory, effector memory, transitional memory and naïve CD4⁺ T cell subsets, to HIV-1 infection in vitro. We show that TSCM are susceptible to infection with laboratory adapted and clinical HIV-1 strains. Our system facilitates the quantitation of HIV-1 infection in alternative T cell subsets by CCR5- and CXCR4-using viruses across different HIV-1 subtypes, and will be useful for studies of HIV-1 pathogenesis and viral reservoirs.     

Changes in peripheral CD19+Foxp3+ and CD19+TGFβ+ regulatory B cell populations in rheumatoid arthritis patients with interstitial lung disease

Objective

Although the role of regulatory B cells (Bregs) in rheumatic disease has gained increasing attention, two lesser-known Breg subsets that express either Foxp3 or transforming growth factor beta (TGFβ) are rarely examined in studies of rheumatic disease. This study investigates the association between the relative proportions of CD19⁺Foxp3⁺ and CD19⁺TGFβ⁺ Bregs, and clinical indicators of disease severity in rheumatoid arthritis (RA) patients with or without interstitial lung disease (ILD).

Methods

A total of 31 RA patients (14 with and 17 without ILD) and 26 healthy control subjects were included. All subjects did not have other autoimmune disease except RA, tumor, active infection, or a history of related drug administration. Peripheral blood mononuclear cells (PBMCs) were isolated and analyzed by flow cytometry (FCM). The relationship between the relative proportions of CD19⁺Foxp3⁺ and CD19⁺TGFβ⁺ Bregs and their associations to RA and ILD incidence, as well as disease severity assessed by common clinical indicators, were then examined.

Results

Our analyses revealed RA patients had significantly lower proportions of peripheral CD19⁺Foxp3⁺ and CD19⁺TGFβ⁺ Bregs as compared to healthy controls. While no association was observed between CD19⁺Foxp3⁺ Bregs and ILD incidence, patients with ILD had a substantially lower percentage of CD19⁺TGFβ⁺ Bregs compared to RA patients without ILD. In addition, CD19⁺Foxp3⁺ Bregs were negatively correlated with erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) levels in RA patients, whereas CD19⁺TGFβ⁺ Bregs were only correlated with CRP in RA patients with ILD. Furthermore, there was a negative association between CD19⁺Foxp3⁺ Bregs and disease severity scores, which was not found in analyses with CD19⁺TGFβ⁺ Bregs.

Conclusions

The proportions CD19⁺Foxp3⁺ and CD19⁺TGFβ⁺ Bregs were significantly decreased in RA patients, particularly in those with ILD complications, suggesting that Breg phenotypes may have different functions in the pathogenesis of RA and ILD.     

Monitoring of donor chimerism in sorted CD34+ peripheral blood cells allows the sensitive detection of imminent relapse after allogeneic stem cell transplantation

Analysis of donor chimerism is an important diagnostic tool to assess the risk of relapse after allogeneic stem cell transplantation, especially in patients lacking a specific marker suitable for monitoring of minimal residual disease. We prospectively investigated the predictive value of donor chimerism analyses in sorted CD34⁺ peripheral blood cells in 90 patients with acute leukemia and myelodysplastic syndrome. The cumulative incidence of relapse after four years was significantly increased in cases with decreasing or incomplete CD34⁺ donor chimerism (57% vs. 18%, p=0.0001). Multivariate analysis confirmed decreasing CD34⁺ donor chimerism as an independent predictor of relapse and inferior survival. The interval between a decrease of CD34⁺ chimerism of less than 80% and hematologic relapse was 61 days (range 0–567). Monitoring of CD34⁺ donor chimerism in the peripheral blood allows prediction of imminent relapse after allogeneic stem cell transplantation even when a disease-specific marker is lacking.     

Reproducible selection of high avidity CD8+ T-cell clones following secondary acute virus infection

The recall of memory CD8⁺ cytotoxic T lymphocytes (CTLs), elicited by prior virus infection or vaccination, is critical for immune protection. The extent to which this arises as a consequence of stochastic clonal expansion vs. active selection of particular clones remains unclear. Using a parallel adoptive transfer protocol in combination with single cell analysis to define the complementarity determining region (CDR) 3α and CDR3β regions of individual T-cell receptor (TCR) heterodimers, we characterized the antigen-driven recall of the same memory CTL population in three individual recipients. This high-resolution analysis showed reproducible enrichment (or diminution) of particular TCR clonotypes across all challenged animals. These changes in clonal composition were TCRα− and β chain–dependent and were directly related to the avidity of the TCR for the virus-derived peptide (p) + major histocompatibility complex class I molecule. Despite this shift in clonotype representation indicative of differential selection, there was no evidence of overall repertoire narrowing, suggesting a strategy to optimize CTL responses while safeguarding TCR diversity.Virus-specific CD8⁺ cytotoxic T lymphocytes (CTLs) are key for effective pathogen clearance. To exert their antiviral effects, naïve CD8⁺ CTL precursors (CTLps) must first be activated through the specific recognition of virus-derived peptides (p) in the context of major histocompatibility complex class I molecules (MHCI) expressed on the surface of dendritic cells. Ligation of these pMHCI epitopes is mediated via specific T-cell receptor (TCR) αβ heterodimers, leading to the recruitment, proliferation, and activation of antigen-specific CTLs. Subsequent to virus clearance, CTL populations contract to form a stable pool of resting memory cells, typically at around 5–10% of their acute phase numbers (1, 2). On secondary virus encounter, this memory pool of CD8⁺ T cells is able to expand rapidly, providing potent immune protection (2).An understanding of CD8⁺ T-cell recruitment/expansion into the recall response has significant implications for effective vaccine strategies. If recruitment and expansion occur stochastically, a memory population in which highly effective clones predominate is desirable. Alternatively, if selection is deterministic, the basic requirement would be the presence of high-quality clones in the memory population for selective expansion by antigen-driven mechanisms. Moreover, different mechanisms of memory recruitment have different long-term implications for the clonal diversity of epitope-specific populations, especially with repeated virus exposure. Repeated selection of particular clones inherent in deterministic clonal selection has the potential consequence of narrowing the CTL repertoire, whereas stochastic recruitment/expansion seems more likely to maintain CTL diversity, shown to be beneficial for virus control (3–7). Despite the implications, there remains conjecture about how epitope-specific CD8⁺ T cells are recruited from the memory pool. Studies have indicated a focusing of the recall response relative to the memory pool, arising from only a subpopulation of cells being recruited or expanded (8), whereas others have indicated that primary and secondary responses are either highly similar or randomly different, prompting the conclusion that selection from the memory pool likely occurs as a result of stochastic T-cell selection (9–13).Using secondary virus challenge of influenza A virus–primed B6 mice as a model of acute localized infection, we investigated the recruitment/expansion of memory CD8⁺ T-cell clones specific for the immunodominant D^bNP₃₆₆ epitope. We used a parallel adoptive transfer method, in which a population of memory CD8⁺ T cells from one individual is rechallenged with virus in three independent recipients, to definitively ascertain whether changes in clonal prevalence between memory and recall populations occur in a stochastic or deterministic fashion. Using high-resolution single cell TCRβ or αβ analysis, our data show evidence of active selection for particular clones following secondary infection, without a reduction in overall TCR diversity. Further, the selection for particular T-cell clones appears to be based on the avidity of the TCR–pMHCI interaction. Thus, it seems that the recall response may be optimized without diminishing the breadth of TCR use that may be critical for effective virus control.     

Identification of Novel Candidate CD8+ T Cell Epitopes of the SARS-CoV2 with Homology to Other Seasonal Coronaviruses

Cross-reactive T cell immunity to seasonal coronaviruses (HCoVs) may lead to immunopathology or protection during SARS-CoV2 infection. To understand the influence of cross-reactive T cell responses, we used IEDB (Immune epitope database) and NetMHCpan (ver. 4.1) to identify candidate CD8⁺ T cell epitopes, restricted through HLA-A and B alleles. Conservation analysis was carried out for these epitopes with HCoVs, OC43, HKU1, and NL63. 12/18 the candidate CD8⁺ T cell epitopes (binding score of ≥0.90), which had a high degree of homology (>75%) with the other three HCoVs were within the NSP12 and NSP13 proteins. They were predicted to be restricted through HLA-A*2402, HLA-A*201, HLA-A*206, and HLA-B alleles B*3501. Thirty-one candidate CD8⁺ T cell epitopes that were specific to SARS-CoV2 virus (<25% homology with other HCoVs) were predominantly identified within the structural proteins (spike, envelop, membrane, and nucleocapsid) and the NSP1, NSP2, and NSP3. They were predominantly restricted through HLA-B*3501 (6/31), HLA-B*4001 (6/31), HLA-B*4403 (7/31), and HLA-A*2402 (8/31). It would be crucial to understand T cell responses that associate with protection, and the differences in the functionality and phenotype of epitope specific T cell responses, presented through different HLA alleles common in different geographical groups, to understand disease pathogenesis.     

Rudimentary signs of immunosenescence in Cytomegalovirus-seropositive healthy young adults

Ageing is associated with a decline in immune competence termed immunosenescence. In the elderly, this process results in an accumulation of differentiated ‘effector’ phenotype memory T cells, predominantly driven by Cytomegalovirus (CMV) infection. Here, we asked whether CMV also drives immunity towards a senescent profile in healthy young adults. One hundred and fifty-eight individuals (mean ± SD; age 21 ± 3 years, body mass index 22.7 ± 2.7 kg m²) were assessed for CMV serostatus, the numbers/proportions of CD4⁺ and CD8⁺ late differentiated/effector memory cells (i.e. CD27⁻CD28⁻/CD45RA⁺), plasma interleukin-6 (IL-6) and antibody responses to an in vivo antigen challenge (half-dose influenza vaccine). Thirty percent (48/158) of participants were CMV⁺. A higher lymphocyte and CD8⁺ count (both p < 0.01) and a lower CD4/CD8 ratio (p < 0.03) were observed in CMV⁺ people. Eight percent (4/58) of CMV⁺ individuals exhibited a CD4/CD8 ratio <1.0, whereas no CMV⁻ donor showed an inverted ratio (p < 0.001). The numbers of CD4⁺ and CD8⁺CD27⁻CD28⁻/CD45RA⁺ cells were ~ fourfold higher in CMV⁺ people (p < 0.001). Plasma IL-6 was higher in CMV⁺ donors (p < 0.05) and showed a positive association with the numbers of CD8⁺CD28⁻ cells (p < 0.03). Finally, there was a significant negative correlation between vaccine-induced antibody responses to the A/Brisbane influenza strain and CMV-specific immunoglobulin G titres (p < 0.05). This reduced vaccination response was associated with greater numbers of total CD8⁺ and CD4⁺ and CD8⁺CD27⁻CD28⁻/CD45RA⁺ cells (p < 0.05). This study observed marked changes in the immune profile of young adults infected with CMV, suggesting that this virus may underlie rudimentary aspects of immunosenescence even in a chronologically young population.     

GDP-mannose pyrophosphorylase is a genetic determinant of ammonium sensitivity in Arabidopsis thaliana

Higher plant species differ widely in their growth responses to ammonium (NH₄⁺). However, the molecular genetic mechanisms underlying NH₄⁺ sensitivity in plants remain unknown. Here, we report that mutations in the Arabidopsis gene encoding GDP-mannose pyrophosphorylase (GMPase) essential for synthesizing GDP-mannose confer hypersensitivity to NH₄⁺. The in planta activities of WT and mutant GMPases all were inhibited by NH₄⁺, but the magnitude of the inhibition was significantly larger in the mutant. Despite the involvement of GDP-mannose in both l-ascorbic acid (AsA) and N-glycoprotein biosynthesis, defective protein glycosylation in the roots, rather than decreased AsA content, was linked to the hypersensitivity of GMPase mutants to NH₄⁺. We conclude that NH₄⁺ inhibits GMPase activity and that the level of GMPase activity regulates Arabidopsis sensitivity to NH₄⁺. Further analysis showed that defective N-glycosylation of proteins, unfolded protein response, and cell death in the roots are likely important downstream molecular events involved in the growth inhibition of Arabidopsis by NH₄⁺.     

Demonstration of Low-Regulatory CD25High+CD4+ and High-Pro-inflammatory CD28−CD4+ T-Cell Subsets in Patients with Ulcerative Colitis: Modified by Selective Granulocyte and Monocyte Adsorption Apheresis

Low-CD25^High+CD4⁺, a subset of regulatory CD25⁺CD4⁺ T cells and high-inflammatory CD28⁻CD4⁺ T cells can exacerbate ulcerative colitis (UC). This study sought to investigate the frequency of CD25^High+CD4⁺ and CD28⁻CD4⁺ T cells in patients with UC and the changes in these cells during Adacolumn granulocyte and monocyte adsorption apheresis (GMA). Subjects were 12 patients with active UC, 11 with quiescent UC, and 14 healthy volunteers (HVs). The mean clinical activity index was 15.7 ± 2.2 in active UC and 4.5 ± 1.1 in quiescent UC. Peripheral blood samples were stained with CD4, CD25, and CD28 antibodies for flow cytometry. Patients with active UC received GMA and blood samples were examined before and after the first GMA session. Patients with active UC (P < 0.04) or quiescent UC (P < 0.02) had a higher percentage of CD28⁻D4⁺T cells compared with HVs, while the percentage of CD28⁺CD4⁺ T cells was lower in both UC groups compared with HVs (P = 0.03 and P < 0.02). Patients with active UC had a lower percentage of CD25^High+CD4⁺T cells compared with quiescent UC patients (P < 0.001). A significant increase in CD25^High+CD4⁺ T cells was associated with GMA (P < 0.03). Low CD25^High+CD4⁺ and high CD28⁻CD4⁺ are prominent features in UC. The increase in CD25^High+CD4⁺ T cells induced by GMA should contribute to improved immune function. Additional studies are warranted, since a low frequency of CD25^High+CD4⁺ ₋ and a high frequency of CD28⁻CD4⁺ ₋ expressing T cells might be a predictor of clinical response to GMA.     

Ex vivo expansion CD34+/AC133+ - selected autologous peripheral blood progenitor cells (PBPC) in high-risk breast cancer patients receiving intensive chemotherapy

AC133 antibody provides an alternative to CD34 for the selection and characterization of cells necessary for engraftment in transplant situations. We studied the effect of stem cell factor (SCF), interleukin 3 (IL-3) and interleukin 11 (IL-11) on the ex vivo expansion of human CD34⁺/AC133⁺ progenitors isolated from leukapheresis products from chemotherapy plus granulocyte-colony-stimulating factor (G-CSF) -mobilized adult donors. MiniMACS AC133⁺ isolated cells contained a mean of 85% (80-90) AC133⁺ cells. Enriched AC133⁺ cells co-expressed CD34⁺ 80%, CD71^low 36.6 % and CD33⁺ 6.6 %. After a seven-day ex vivo expansion in the presence of SCF + IL-3 + IL-11, the number of cells increased 19 times. These cells expressed a mean 12% CD34⁺ and 74% CD71⁺ (23% CD 71^high) and 59% CD33⁺. This means that the absolute number of CD34⁺ cells increased slightly, the number of CD33⁺ increased 100 times and cells with high density CD71^high (23%) appeared. These cells represent the cells committed to erythroid differentiation. The increase in the number of CFU-GM with various combinations of cytokines SCF + Il-3 + IL-11 will be discussed. The number of CFU-GM in culture after a seven-day ex vivo expansion in the presence of SCF + IL-3 + IL-11 increased 45 times.