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Background: Myb‐like, SWIRM, and MPN domains 1 (MYSM1) is a histone H2A deubiquitinase, has been discovered as one of the transcriptional regulators, and regulates the expression of specific transcription factors, which are essential for immunohematology development. Mutation in MYSM1 in humans leads to a rare autosomal recessive disease that has recently been known as inherited bone marrow failure syndrome 4 (BMFS4) associated with congenital bone marrow failure, immunodeficiency, and developmental aberrations. Allogeneic hematopoietic stem cell transplantation (HSCT) is the only curative option for immunohematology defects. Methods: In this paper, we report a pediatric patient with BMFS4 who suffered from pancytopenia and immunodeficiency affecting B cells and was successfully treated with HSCT from an HLA‐identical father at 6 years old of age. Fludarabine‐based reduced intensity conditioning was used and resulted in full donor chimerism. Results: Acute graft versus host disease (GVHD) grade II involving skin and gastrointestinal tract was observed, which was controlled with prednisolone. Conclusion: She achieved B‐cell recovery, and no blood or platelet transfusion was reported 1 year after HSCT. [ABSTRACT FROM AUTHOR]

β-thalassemia, a disease that results from defects in β-globin synthesis, leads to an imbalance of β- and α-globin chains and an excess of α chains. Defective erythroid maturation, ineffective erythropoiesis, and shortened red blood cell survival are commonly observed in most β-thalassemia patients. In severe cases, blood transfusion is considered as a mainstay therapy; however, regular blood transfusions result in chronic iron overload with life-threatening complications, e.g., endocrine dysfunction, cardiomyopathy, liver disease, and ultimately premature death. Therefore, transplantation of healthy hematopoietic stem cells (HSCs) is considered an alternative treatment. Patients with a compatible human leukocyte antigen (HLA) matched donor can be cured by allogeneic HSC transplantation. However, some recipients faced a high risk of morbidity/mortality due to graft versus host disease or graft failure, while a majority of patients do not have such HLA match-related donors. Currently, the infusion of autologous HSCs modified with a lentiviral vector expressing the β-globin gene into the erythroid progenitors of the patient is a promising approach to completely cure β-thalassemia. Here, we discuss a history of β-thalassemia treatments and limitations, in particular the development of β-globin lentiviral vectors, with emphasis on clinical applications and future perspectives in a new era of medicine. [ABSTRACT FROM AUTHOR]

Mutations in the gene encoding tRNA nucleotidyltransferase 1 (TRNT1) are associated with heterogeneous phenotypes and multisystem involvement of variable severity and progression. Immunodeficiency and inflammation are recurrent-associated features. The use of cytokine inhibitors in suppressing the inflammatory phenotype has been recently reported, with a 3-year follow-up for patients treated with Etanercept. We report on two unrelated patients sharing the same clinical condition, who had been referred to our Pediatric Rheumatology Unit because of recurrent fever associated with cutaneous lesions and increased levels of inflammatory markers since their first months of life. Whole exome sequencing allowed to identify compound heterozygosity for functionally relevant variants in TRNT1 as the only molecular event shared by the two patients. Both patients have been treated with Etanercept during 11 years, documenting normalization of inflammatory indexes and resolution of recurrent fever and associated symptoms. This is the longest follow-up assessment of Etanercept treatment in patients with TRNT1 mutations. Our findings confirm efficacy and safety of the treatment. Key Points • Mutations in TRNT1 have been associated with phenotypic heterogeneity. • We report on two patients with early-onset autoinflammatory syndrome. • Whole exome sequencing led to reveal compound heterozygosity for two variants in TRNT1 in both patients. • The patients were successfully treated with Etanercept for more than 10 years, the longest follow-up described in literature. [ABSTRACT FROM AUTHOR]

CD8+ T cells are major components of adaptive immunity and confer robust protective cellular immunity, which requires adequate T-cell numbers, targeted migration, and efficient T-cell proliferation. Altered CD8+ T-cell homeostasis and impaired proliferation result in dysfunctional immune response to infection or tumorigenesis. However, intrinsic factors controlling CD8+ T-cell homeostasis and immunity remain largely elusive. Here, we demonstrate the prominent role of Brd4 on CD8+ T cell homeostasis and immune response. By upregulating Myc and GLUT1 expression, Brd4 facilitates glucose uptake and energy production in mitochondria, subsequently supporting naïve CD8+ T-cell survival. Besides, Brd4 promotes the trafficking of naïve CD8+ T cells partially through maintaining the expression of homing receptors (CD62L and LFA-1). Furthermore, Brd4 is required for CD8+ T cell response to antigen stimulation, as Brd4 deficiency leads to a severe defect in clonal expansion and terminal differentiation by decreasing glycolysis. Importantly, as JQ1, a pan-BRD inhibitor, severely dampens CD8+ T-cell immune response, its usage as an anti-tumor agent or latency-reversing agent for human immunodeficiency virus type I (HIV-1) should be more cautious. Collectively, our study identifies a previously-unexpected role of Brd4 in the metabolic regulation of CD8+ T cell-mediated immune surveillance and also provides a potential immunomodulation target. [ABSTRACT FROM AUTHOR]

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Since the first description of the syndrome of sideroblastic anemia with immunodeficiency, fevers and development delay (SIFD), clinical pictures lacking both neurological and hematological manifestations have been reported. Moreover, prominent skin involvement, such as with relapsing erythema nodosum, is not a common finding. Up to this moment, no genotype and phenotype correlation could be done, but mild phenotypes seem to be located in the N or C part. B-cell deficiency is a hallmark of SIFD syndrome, and multiple others immunological defects have been reported, but not high levels of double negative T cells. Here we report a Brazilian patient with a novel phenotype of SFID syndrome, carrying multiple immune defects and harboring a novel mutation on TRNT1 gene. [ABSTRACT FROM AUTHOR]

Gene therapy is an innovative treatment for Primary Immune Deficiencies (PIDs) that uses autologous hematopoietic stem cell transplantation to deliver stem cells with added or edited versions of the missing or malfunctioning gene that causes the PID. Initial studies of gene therapy for PIDs in the 1990–2000's used integrating murine gamma-retroviral vectors. While these studies showed clinical efficacy in many cases, especially with the administration of marrow cytoreductive conditioning before cell re-infusion, these vectors caused genotoxicity and development of leukoproliferative disorders in several patients. More recent studies used lentiviral vectors in which the enhancer elements of the long terminal repeats self-inactivate during reverse transcription ("SIN" vectors). These SIN vectors have excellent safety profiles and have not been reported to cause any clinically significant genotoxicity. Gene therapy has successfully treated several PIDs including Adenosine Deaminase Severe Combined Immunodeficiency (SCID), X-linked SCID, Artemis SCID, Wiskott-Aldrich Syndrome, X-linked Chronic Granulomatous Disease and Leukocyte Adhesion Deficiency-I. In all, gene therapy for PIDs has progressed over the recent decades to be equal or better than allogeneic HSCT in terms of efficacy and safety. Further improvements in methods should lead to more consistent and reliable efficacy from gene therapy for a growing list of PIDs. [ABSTRACT FROM AUTHOR]

The thymus prevents autoimmune diseases through mechanisms that operate in the cortex and medulla, comprising positive and negative selection and the generation of regulatory T-cells (Tregs). Egress from the thymus through the perivascular space (PVS) to the blood is another possible checkpoint, as shown by some autoimmune/immunodeficiency syndromes. In polygenic autoimmune diseases, subtle thymic dysfunctions may compound genetic, hormonal and environmental cues. Here, we cover (a) tolerance-inducing cell types, whether thymic epithelial or tuft cells, or dendritic, B- or thymic myoid cells; (b) tolerance-inducing mechanisms and their failure in relation to thymic anatomic compartments, and with special emphasis on human monogenic and polygenic autoimmune diseases and the related thymic pathologies, if known; (c) polymorphisms and mutations of tolerance-related genes with an impact on positive selection (e.g. the gene encoding the thymoproteasome-specific subunit, PSMB11), promiscuous gene expression (e.g. AIRE, PRKDC, FEZF2, CHD4), Treg development (e.g. SATB1, FOXP3), T-cell migration (e.g. TAGAP) and egress from the thymus (e.g. MTS1, CORO1A); (d) myasthenia gravis as the prototypic outcome of an inflamed or disordered neoplastic 'sick thymus'. [ABSTRACT FROM AUTHOR]

Objective: STK4 (serine-threonine protein kinase 4) deficiency is categorized under combined immunodeficiencies, which are a subgroup of primary immunodeficiencies that profoundly affect T cells. Autosomal recessive STK4 deficiency is characterized by recurrent bacterial and viral infections, mucocutaneous candidiasis, and CD4 + T cell lymphopenia. Autoimmune comorbidities including hemolytic anemia and idiopathic thrombocytopenic purpura, and cardiac problems have been reported in STK4-deficient individuals. Current diagnostic tools for STK4 deficiency include next-generation sequencing (NGS) and Sanger sequencing that detect DNA sequence variations, and western blotting, which evaluates altered protein expression. However, all of these assays are timeconsuming; and in particular, NGS is a costly clinical use tool for a diagnostic laboratory. In comparison, flow cytometry is a rapid and reliable system commonly employed in the diagnosis of a wide range of primary immunodeficiency disorders. Materials and Methods: This study aimed to evaluate STK4 protein expression by flow cytometry among four patients with genetically confirmed STK4 gene mutations and seven healthy individuals. We calculated ΔMFI/cell values to investigate differences in protein expression among the subjects. Results: STK4 protein expression was reduced in the peripheral blood mononuclear cells obtained from all STK4-deficient patients compared to those from the healthy controls. Flow cytometry data was validated by western blotting. Conclusion: Flow cytometry is a rapid and reliable method to detect STK4 protein expression, qualified as a diagnostic tool to study STK4 deficiency. [ABSTRACT FROM AUTHOR]

X-linked severe immunodeficiency disease (SCID-X1) is an inherited, rare, and life-threating disease. The genetic origin is a defect in the interleukin 2 receptor γ chain (IL2RG) gene and patients are classically characterized by absence of T and NK cells, as well as presence of partially-functional B cells. Without any treatment the disease is usually lethal during the first year of life. The treatment of choice for these patients is hematopoietic stem cell transplantation, with an excellent survival rate (>90%) if an HLA-matched sibling donor is available. However, when alternative donors are used, the success and survival rates are often lower. Gene therapy has been developed as an alternative treatment initially using γ-retroviral vectors to correct the defective γ chain in the absence of pre-conditioning treatment. The results were highly promising in SCID-X1 infants, showing long-term T-cell recovery and clinical benefit, although NK and B cell recovery was less robust. However, some infants developed T-cell acute lymphoblastic leukemia after the gene therapy, due to vector-mediated insertional mutagenesis. Consequently, considerable efforts have been made to develop safer vectors. The most recent clinical trials using lentiviral vectors together with a low-dose pre-conditioning regimen have demonstrated excellent sustained T cell recovery, but also B and NK cells, in both children and adults. This review provides an overview about the different gene therapy approaches used over the last 20 years to treat SCID-X1 patients, particularly focusing on lymphoid immune reconstitution, as well as the developments that have improved the process and outcomes. [ABSTRACT FROM AUTHOR]

Background. Clinical trials are at the cornerstone of evidence-based stem cell therapies, but the quality assessment for designing and conduct these sometimes-complex studies are scarce of evidence. This study is aimed at developing a handy quality assessment tool for stem cell clinical trials, enhancing capacity of the self-regulate overall quality, and participating protection. Methods. The framework of quality assessment tool was based on the PQRS (progress-quality-regulation-scientific) quality assessment tool, and detailed quality indicators were developed by leader group discussion, expert consulting, and literature review. Stem cell clinical trials were retrieved from the International Clinical Trials Registry Platform, and corresponding quality indicators were assessed and extracted. The validity and feasibility of conceptual quality assessment tool were further evaluated by using structural equation modeling. Results. The quality assessment tool for stem cell clinical trials contains four critical quality attributes, including participant protection, scientific value, quality control, and stem cell products, and 9 observed quality indicators. From 11 primary clinical trial registries in the International Clinical Trials Registry Platform, 9410 stem cell trial registrations were identified, and 1036 studies were eligible for publications and protocols screening. After reviewed full text, 37 studies were included in the validity and feasibility evaluation: 32 studies were completed, and 3 studies terminated early. Most of the studies (83.79%) were in the early phase, and 63.16% of the studies were investigator-initiated trial. To further tested for validity, the critical quality attributes and quality indicators (QIs) between expertise further validated by the SEM method, which showed a good fit for the model (chi − square = 26.008 ; P = 0.353 ; TLI = 0.967 ; CFI = 0.978 ; RMSEA = 0.048). Compared with exploratory trials, evaluating using the quality assessment tool, confirmatory trials performed similarly in participant protection, scientific value, and quality control, but lower in stem cell products. Conclusions. The results of critical quality attributes and quality indicators between expertise and confirmatory validation analysis are basically consistent, indicating the feasibility and validity of applying this quality assessment tool for overall quality evaluation of stem cell trials. [ABSTRACT FROM AUTHOR]

Purpose of Review: The advent of enhanced genetic testing has allowed for the discovery of gene defects underlying two broad categories of antibody deficiency in children: agammaglobulinemia and common variable immunodeficiency (CVID). This review describes the underlying gene defects and the clinical manifestations. Recent Findings: Because novel monogenetic defects have been discovered in both categories, a strict dichotomous classification of B cell disorders as either X-linked agammaglobulinemia or common variable immunodeficiency is no longer appropriate. Summary: Advances in genetic testing technology and the decreasing cost of such testing permit more precise diagnosis of B cell disorders, more helpful information for genetic counselors, and a better understanding of the complex process of B cell development and function. More disorders await discovery. [ABSTRACT FROM AUTHOR]

Acute graft-vs.-host disease (aGvHD) is one of the most frequent causes of transplant-related mortality (TRM) after allogeneic hematopoietic cell transplantation (alloHCT). Its treatment is complex and costly. The aim of this study was to retrospectively analyze the impact of country-specific socioeconomic factors on outcome of patients who experience severe aGvHD. Adults with hematological malignancies receiving alloHCT from either HLA-matched siblings (n = 1,328) or unrelated donors (n = 2,824) developing grade 3 or 4 aGvHD were included. In univariate analysis, the probability of TRM at 2 years was increased for countries with lower current Health Care Expenditure (HCE, p = 0.04), lower HCE as % of Gross Domestic Product per capita (p = 0.003) and lower values of the Human Development Index (p = 0.02). In a multivariate model, the risk of TRM was most strongly predicted by current HCE (HR = 0.76, p = 0.006). HCE >median was also associated with reduced risk of the overall mortality (HR 0.73, p = 0.0006) and reduced risk of treatment failure (either relapse or TRM; HR 0.77, p = 0.004). We conclude that country-specific socioeconomic factors, in particular current HCE, are strongly associated with survival of patients who experience severe aGvHD. [ABSTRACT FROM AUTHOR]

Purpose: Owing to recent technological advancements, using next-generation sequencing (NGS) and the accumulation of clinical experiences worldwide, more than 420 genes associated with inborn errors of immunity (IEI) have been identified, which exhibit large genotypic and phenotypic variations. Consequently, NGS-based comprehensive genetic analysis, including whole-exome sequencing (WES), have become more valuable in the clinical setting and have contributed to earlier diagnosis, improved treatment, and prognosis. However, these approaches have the following disadvantages that need to be considered: a relatively low diagnostic rate, high cost, difficulties in the interpretation of each variant, and the risk of incidental findings. Thus, the objective of this study is to review our WES results of a large number of patients with IEI and to elucidate patient characteristics, which are related to the positive WES result. Methods: We performed WES for 136 IEI patients with negative conventional screening results for candidate genes and classified these variants depending on validity of their pathogenicity. Results: We identified disease-causing pathogenic mutations in 36 (26.5%) of the patients which were found in known IEI-causing genes. Although the overall diagnostic rate was not high and was not apparently correlated with the clinical subcategories and severity, we revealed that earlier onset with longer duration of diseases were associated with positive WES results, especially in pediatric cases. Conclusions: Most of the disease-causing germline mutations were located in the known IEI genes which could be predicted using patients' clinical characteristics. These results may be useful when considering appropriate genetic approaches in the clinical setting. [ABSTRACT FROM AUTHOR]

The STE20-like serine/threonine kinases MST1 and MST2 (MST1/2) are mammalian homologs of Hippo in flies. MST1/2 regulate organ size by suppressing the transcription factor YAP, which promotes proliferation. MST1 is predominantly expressed in immune cells, where it plays distinct roles. Here, we review the functions of MST1/2 in immune cells, uncovered by a series of recent studies, and discuss the connection between MST1/2 function and immune responses. MST1/2 regulate lymphocyte development, trafficking, survival, and antigen recognition by naive T cells. MST1/2 also regulate the function of regulatory T cells and effector T cell differentiation, thus acting to balance immune activation and tolerance. Interestingly, MST1/2 elicit these functions not by the "canonical" Hippo pathway, but by the non-canonical Hippo pathway or alternative pathways. In these pathways, MST1/2 regulates cellular processes relating to immune response, such as chemotaxis, cell adhesion, immunological synapse, gene transcriptions. Recent advances in our understanding of the molecular mechanisms of these processes have revealed important roles of MST1/2 in regulating cytoskeleton remodeling, integrin activation, and vesicular transport in lymphocytes. We discuss the significance of the MST1/2 signaling in lymphocytes in the regulation of organelle dynamics. [ABSTRACT FROM AUTHOR]

Capsule Summary We present an extended clinical phenotype of a patient with STK4 deficiency associated with mycobacterial infection. Seven patients have been treated with HSCT; four patients died of a combination of infectious, toxicity related and graft-versus-host disease (GVHD) complications, and the remaining three patients are alive and apparently cured of their immunodeficiency [[5]]. We report the first Egyptian case with STK4 mutation who developed mycobacterial infection, expanding our knowledge on the spectrum of infections that these patients are susceptible to. [Extracted from the article]

Repair of double strand DNA breaks (DSBs) can result in gene disruption or gene modification via homology directed repair (HDR) from donor DNA. Altering cellular responses to DSBs may rebalance editing outcomes towards HDR and away from other repair outcomes. Here, we utilize a pooled CRISPR screen to define host cell involvement in HDR between a Cas9 DSB and a plasmid double stranded donor DNA (dsDonor). We find that the Fanconi Anemia (FA) pathway is required for dsDonor HDR and that other genes act to repress HDR. Small molecule inhibition of one of these repressors, CDC7, by XL413 and other inhibitors increases the efficiency of HDR by up to 3.5 fold in many contexts, including primary T cells. XL413 stimulates HDR during a reversible slowing of S-phase that is unexplored for Cas9-induced HDR. We anticipate that XL413 and other such rationally developed inhibitors will be useful tools for gene modification. Altering cellular responses to double-strand breaks in DNA could rebalance CRISPRediting outcomes. Here, the authors use a pooled CRISPR screen to identify inhibition of CDC7 as a strategy to improve HDR outcomes. [ABSTRACT FROM AUTHOR]

The EuroFlow PID consortium developed a set of flow cytometry tests for evaluation of patients with suspicion of primary immunodeficiency (PID). In this technical report we evaluate the performance of the SCID-RTE tube that explores the presence of recent thymic emigrants (RTE) together with T-cell activation status and maturation stages and discuss its applicability in the context of the broader EuroFlow PID flow cytometry testing algorithm for diagnostic orientation of PID of the lymphoid system. We have analyzed peripheral blood cells of 26 patients diagnosed between birth and 2 years of age with a genetically defined primary immunodeficiency disorder: 15 severe combined immunodeficiency (SCID) patients had disease-causing mutations in RAG1 or RAG2 (n = 4, two of them presented with Omenn syndrome), IL2RG (n = 4, one of them with confirmed maternal engraftment), NHEJ1 (n = 1), CD3E (n = 1), ADA (n = 1), JAK3 (n = 3, two of them with maternal engraftment) and DCLRE1C (n = 1) and 11 other PID patients had diverse molecular defects [ ZAP70 (n = 1), WAS (n = 2), PNP (n = 1), FOXP3 (n = 1), del22q11.2 (DiGeorge n = 4), CDC42 (n = 1) and FAS (n = 1)]. In addition, 44 healthy controls in the same age group were analyzed using the SCID-RTE tube in four EuroFlow laboratories using a standardized 8-color approach. RTE were defined as CD62L+CD45RO-HLA-DR-CD31+ and the activation status was assessed by the expression of HLA-DR+. Naïve CD8+ T-lymphocytes and naïve CD4+ T-lymphocytes were defined as CD62L+CD45RO-HLA-DR-. With the SCID-RTE tube, we identified patients with PID by low levels or absence of RTE in comparison to controls as well as low levels of naïve CD4+ and naïve CD8+ lymphocytes. These parameters yielded 100% sensitivity for SCID. All SCID patients had absence of RTE, including the patients with confirmed maternal engraftment or oligoclonally expanded T-cells characteristic for Omenn syndrome. Another dominant finding was the increased numbers of activated CD4+HLA-DR+ and CD8+HLA-DR+ lymphocytes. Therefore, the EuroFlow SCID-RTE tube together with the previously published PIDOT tube form a sensitive and complete cytometric diagnostic test suitable for patients suspected of severe PID (SCID or CID) as well as for children identified via newborn screening programs for SCID with low or absent T-cell receptor excision circles (TRECs). [ABSTRACT FROM AUTHOR]

Graft-vs.-host disease (GvHD) is a major complication after allogeneic hematopoietic stem cell transplantation that causes mortality and severe morbidity. Genetic disparities in human leukocyte antigens between the recipient and donor are known contributors to the risk of the disease. However, the overall impact of genetic component is complex, and consistent findings across different populations and studies remain sparse. To gain a comprehensive understanding of the genes responsible for GvHD, we combined genome-wide association studies (GWAS) from two distinct populations with previously published gene expression studies on GvHD in a single gene-level meta-analysis. We hypothesized that genes driving GvHD should be associated in both data modalities and therefore could be detected more readily through their combined effects in the integrated analysis rather than in separate analyses. The meta-analysis yielded a total of 51 acute GvHD-associated genes (false detection rate [FDR] <0.1). In support of our hypothesis, this number was significantly higher than that in a permutation meta-analysis involving the whole data set, as well as in separate meta-analyses on the GWAS and gene expression data sets. The genes indicated by the meta-analysis were significantly enriched in 277 Gene Ontology terms (FDR < 0.05), such as T cell function and cytokine-mediated signaling pathways, and the results highlighted several established immune mediators, such as interleukins and JAK-STAT signaling, and presented TRAF6 and TERT as potential effector candidates. Altogether, the results support the chosen methodological approach, implicate a role of gene-level variation in donors' key immunological regulators predisposing patients to acute GVHD, and present potential targets for therapeutic intervention. [ABSTRACT FROM AUTHOR]

In the last two decades, fetal amniotic fluid stem cells progressively attracted attention in the context of both basic research and the development of innovative therapeutic concepts. They exhibit broadly multipotent plasticity with the ability to differentiate into cells of all three embryonic germ layers and low immunogenicity. They are convenient to maintain, highly proliferative, genomically stable, non-tumorigenic, perfectly amenable to genetic modifications, and do not raise ethical concerns. However, it is important to note that among the various fetal amniotic fluid cells, only c-Kit+ amniotic fluid stem cells represent a distinct entity showing the full spectrum of these features. Since amniotic fluid additionally contains numerous terminally differentiated cells and progenitor cells with more limited differentiation potentials, it is of highest relevance to always precisely describe the isolation procedure and characteristics of the used amniotic fluid-derived cell type. It is of obvious interest for scientists, clinicians, and patients alike to be able to rely on up-todate and concisely separated pictures of the utilities as well as the limitations of terminally differentiated amniotic fluid cells, amniotic fluid-derived progenitor cells, and c-Kit+ amniotic fluid stem cells, to drive these distinct cellular models towards as many individual clinical applications as possible., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

We report the case of a young woman who developed, 3 years after stopping Rituximab (RTX) prescribed for immune thrombocytopenia (ITP), a severe immunodeficiency leading to fatal pulmonary Epstein–Barr virus-positive diffuse large B-cell lymphoma. Genetic analysis led us to identify four missense mutations known to affect immune-deficiency–associated genes (FAS-ligand (FASL) gene (p.G167R); perforin-1 (PRF1 (p.R55C) gene; the Bloom syndrome RecQ-Like helicase (BLM) gene and the Moesin (MSN) (p.A122T) gene). The heterozygous mutation in the FASL gene, not present in the Genome Aggregation Database or ClinVar database, could suggest atypical Autoimmune LymphoProliferative Syndrome and its role in this patient's immunodepression is discussed. This observation strengthens the role of FASL gene mutation in severe clinical phenotypes of primary immune deficiency and raises new questions about the genetic background of ITP occurring in young people in a context of immunodeficiency. [ABSTRACT FROM AUTHOR]

Summary: Stress erythropoiesis and chronic inflammation in subjects with sickle cell disease (SCD) may have an impact on the bone marrow (BM) haematopoietic stem and progenitor cell (HSPC) quality and yield necessary for effective autologous, ex vivo HSPC gene therapy. BM from 19 subjects with SCD and five volunteers without SCD (non‐SCD) was collected in different anticoagulants and processed immediately (day 0) or the following day (day 1). Inflammatory, contamination and aggregation markers within the mononuclear layer, and CD34, CD45 and Glycophorin‐A (GPA) expression on HSPCs after CD34+ selection were analysed by conventional and imaging flow cytometry. Compared to non‐SCD BM, multiple markers of inflammation, contamination (red cells, P < 0·01; platelets, P < 0·01) and aggregates (platelet/granulocytes, P < 0·01; mononuclear/red cells, P < 0·01) were higher in SCD BM. Total CD34+ cell count was lower in SCD BM (P < 0·05), however CD34+ count was higher in SCD BM when collected in acid citrate dextrose‐A (ACDA) versus heparin (P < 0·05). Greater than 50% of CD34+ HSPCs from SCD BM are CD34dim due to higher erythroid lineage expression (P < 0·01) as single cell CD34+CD45+GPA+ (P < 0·01) and CD34+CD45−GPA+ (P < 0·01) HSPCs. SCD BM is characterized by increased inflammation, aggregation and contamination contributing to significant differences in HSPC quality and yield compared to non‐SCD BM. [ABSTRACT FROM AUTHOR]

All nucleoside analogues for treating HIV infection, due to their capacity to integrate into and alter human DNA, are experimentally genotoxic to some extent. The long‐term oncogenic risk after in utero exposure remains to be determined. Cancer incidence in uninfected children exposed to nucleos(t)ide reverse transcriptase inhibitors (NRTIs) was evaluated, by cross‐checking against the National Cancer Registry, in the French perinatal study of children born to HIV+ mothers. Twenty‐one cancers were identified in 15,163 children (median age: 9.9 years [interquartile range (IQR): 5.8–14.2]) exposed to at least one NRTI in utero between 1990 and 2014. Five of these children were exposed to zidovudine monotherapy, and 15 to various combinations, seven of which included didanosine. Overall, the total number of cases was not significantly different from that expected for the general population (SIR = 0.8[0.47–1.24]), but the number of cases after didanosine exposure was twice that expected (SIR = 2.5 [1.01–5.19]). Didanosine accounted for only 10% of prescriptions but was associated with one‐third of cancers. In multivariate analysis, didanosine exposure was significantly associated with higher risk (HR = 3.0 [0.9–9.8]). This risk was specifically linked to first‐trimester exposure (HR = 5.5 [2.1–14.4]). Three cases of pineoblastoma, a very rare cancer, were observed, whereas 0.03 were expected. Two were associated with didanosine exposure. Despite reassuring data overall, there is strong evidence to suggest that didanosine displays transplacental oncogenicity. These findings cannot be extrapolated to other NRTIs, but they highlight the need for comprehensive evaluations of the transplacental genotoxicity of this antiretroviral class. Environ. Mol. Mutagen., 60:404–409, 2019. © 2017 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Adenylate kinase is a small, usually monomeric, enzyme found in every living thing due to its crucial role in energetic metabolism. This paper outlines the most relevant data about adenylate kinases isoforms, and the connection between dysregulation or mutation of human adenylate kinase and medical conditions. The following datadases were consulted: National Centre for Biotechnology Information, Protein Data Bank, and Mouse Genomic Informatics. The SmartBLAST tool, EMBOSS Needle Program, and Clustal Omega Program were used to analyze the best protein match, and to perform pairwise sequence alignment and multiple sequence alignment. Human adenylate kinase genes are located on different chromosomes, six of them being on the chromosomes 1 and 9. The adenylate kinases' intracellular localization and organ distribution explain their dysregulation in many diseases. The cytosolic isoenzyme 1 and the mitochondrial isoenzyme 2 are the main adenylate kinases that are integrated in the vast network of inflammatory modulators. The cytosolic isoenzyme 5 is correlated with limbic encephalitis and Leu673Pro mutation of the isoenzyme 7 leads to primary male infertility due to impairment of the ciliary function. The impairment of the mitochondrial isoenzymes 2 and 4 is demonstrated in neuroblastoma or glioma. The adenylate kinases are disease modifier that can assess the risk of diseases where oxidative stress plays a crucial role in pathogenesis like metabolic syndrome or neurodegenerative diseases. Because adenylate kinases has ATP as substrate, they are integrated in the global network of energetic process of any organism therefore are valid target for new pharmaceutical compounds. [ABSTRACT FROM AUTHOR]

Amniotic fluid (AF) contains a heterogeneous population of cells that have been identified to possess pluripotent and progenitor-like characteristics. These cells have been applied in various regenerative medicine applications ranging from in vitro cell differentiation to tissue engineering to cellular therapies for different organs including the heart, the liver, the lung, and the kidneys. In this review, we examine the different methodologies used for the derivation of amniotic fluid stem cells and renal progenitors, and their application in renal repair and regeneration. Moreover, we discuss the recent achievements and newly emerging challenges in our understanding of their biology, their immunoregulatory characteristics, and their paracrine-mediated therapeutic potential for the treatment of acute and chronic kidney diseases. [ABSTRACT FROM AUTHOR]

The development of mature blood cells from haematopoietic stem cells has long served as a model for stem-cell research, with the haematopoietic differentiation tree being widely used as a model for the maintenance of hierarchically organized tissues. Recent results and new technologies have challenged the demarcations between stem and progenitor cell populations, the timing of cell-fate choices and the contribution of stem and multipotent progenitor cells to the maintenance of steady-state blood production. These evolving views of haematopoiesis have broad implications for our understanding of the functions of adult stem cells, as well as the development of new therapies for malignant and non-malignant haematopoietic diseases. [ABSTRACT FROM AUTHOR]

The methylation status of the receptor for advanced glycation end products (RAGE) gene promoter in peripheral blood mononuclear cells (PBMCs) of type 2 diabetic retinopathy (DR) patients was evaluated to investigate the correlation between RAGE gene promoter methylation and diabetic retinal inflammation. Eighty patients admitted and diagnosed as type 2 DR in Qilu Hospital, Shandong University during the period from October, 2013 to October, 2015 were enrolled in this study. They were the observation group and 40 healthy subjects were enrolled in the control group. PBMCs were collected from patients using density gradient centrifugation, and the methylation status of RAGE gene promoters was detected using methylation-specific PCP (MSP). Interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) levels of in the serum were measured using enzyme-linked immunosorbent assay (ELISA). PBMCs in patients with positive RAGE gene promoter methylation were isolated and cultured and RAGE gene promoter methylation was inhibited using the demethylating agent, 5'-aza-2'-deoxycytidine (5-aza-dC). The methylation status of RAGE gene promoters in PBMCs was detected via MSP. IL-1β, IL-6 and TNF-α levels in the supernatant of PBMC culture solution were evaluated using ELISA. MSP results showed that there were 26 cases (32.50%) of RAGE gene promoter methylation in PBMCs in DR patients. RAGE gene promoters were methylated in all normal healthy subjects. IL-1β, IL-6 and TNF-α levels in serum for positive RAGE gene promoter methylation group were significantly lower than those in negative RAGE gene promoter methylation group (p<0.01). 5-aza-dC inhibited the RAGE gene promoter methylation of PBMCs in patients with positive RAGE gene promoter methylation. The inhibition of methylation in RAGE gene promoter increased the levels of IL-1β, IL-6 and TNF-α in supernatant of culture solution. In conclusion, RAGE gene promoter hypomethylation was detected in DR patients, indicating that RAGE gene promoter methylation could inhibit the diabetic retinal inflammation. [ABSTRACT FROM AUTHOR]

Poor graft function ( PGF) is a severe complication of haematopoietic stem cell transplantation ( HSCT) and administration of donor stem cell boosts ( SCBs) represents a therapeutic option. We report 50 paediatric patients with PGF who received 61 boosts with CD34+ selected peripheral blood stem cells ( PBSC) after transplantation from matched unrelated ( n = 25) or mismatched related ( n = 25) donors. Within 8 weeks, a significant increase in median neutrophil counts (0·6 vs. 1·516 × 109/l, P < 0·05) and a decrease in red blood cell and platelet transfusion requirement (median frequencies 1 and 7 vs. 0, P < 0·0001 and <0·001), were observed, and 78·8% of patients resolved one or two of their cytopenias. 36·5% had a complete haematological response. Median lymphocyte counts for CD3+, CD3+ CD4+, CD19+ and CD56+ increased 8·3-, 14·2-, 22.- and 1·6-fold. The rate of de novo acute graft-versus-host disease (Gv HD) grade I-III was only 6% and resolved completely. No Gv HD grade IV or chronic Gv HD occurred. Patients who responded to SCB displayed a trend toward better overall survival ( OS) ( P = 0·07). Thus, administration of CD34+ selected SCBs from alternative donors is safe and effective. Further studies are warranted to clarify the impact on immune reconstitution and survival. [ABSTRACT FROM AUTHOR]

Ex vivo genetic manipulation of autologous hematopoietic stem and progenitor cells (HSPCs) is a viable strategy for the treatment of hematologic and primary immune disorders. Targeted genome editing of HSPCs using the CRISPR-Cas9 system provides an effective platform to edit the desired genomic locus for therapeutic purposes with minimal off-target effects. In this chapter, we describe the detailed methodology for the CRISPR-Cas9 mediated gene knockout, deletion, addition, and correction in human HSPCs by viral and nonviral approaches. We also present a comprehensive protocol for the analysis of genome modified HSPCs toward the erythroid and megakaryocyte lineage in vitro and the long-term multilineage reconstitution capacity in the recently developed NBSGW mouse model that supports human erythropoiesis., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)

Patients with Wiskott-Aldrich syndrome (WAS) lacking a human leukocyte antigen-matched donor may benefit from gene therapy through the provision of gene-corrected, autologous hematopoietic stem/progenitor cells. Here, we present comprehensive, long-term follow-up results (median follow-up, 7.6 years) (phase I/II trial no. NCT02333760 ) for eight patients with WAS having undergone phase I/II lentiviral vector-based gene therapy trials (nos. NCT01347346 and NCT01347242 ), with a focus on thrombocytopenia and autoimmunity. Primary outcomes of the long-term study were to establish clinical and biological safety, efficacy and tolerability by evaluating the incidence and type of serious adverse events and clinical status and biological parameters including lentiviral genomic integration sites in different cell subpopulations from 3 years to 15 years after gene therapy. Secondary outcomes included monitoring the need for additional treatment and T cell repertoire diversity. An interim analysis shows that the study meets the primary outcome criteria tested given that the gene-corrected cells engrafted stably, and no serious treatment-associated adverse events occurred. Overall, severe infections and eczema resolved. Autoimmune disorders and bleeding episodes were significantly less frequent, despite only partial correction of the platelet compartment. The results suggest that lentiviral gene therapy provides sustained clinical benefits for patients with WAS., (© 2022. The Author(s).)

Increased morbidity and fetal growth restriction are reported in uninfected children born to human immunodeficiency virus type 1 (HIV-1)-infected women treated with antiretroviral (ARV) therapy. Viruses and/or pharmacological interventions such as ARVs can induce metabolic stress, skewing the cell's immune response and restricting (cell) growth. Novel metabolomic techniques provided the opportunity to investigate the impact of fetal HIV-1 and combination ARV therapy (cART) exposure on the infants' immune metabolome. Peroxidized lipids, generated by reactive oxygen species, were increased in cART/HIV-1-exposed infants, indicating altered mitochondrial functioning. The lipid metabolism was further dysregulated with increased triglyceride species and a subsequent decrease in phospholipids in cART/HIV-1-exposed infants compared to control infants. Proinflammatory immune mediators, lysophospholipids as well as cytokines such as CXCL10 and CCL3, were increased whereas anti-inflammatory metabolites from the cytochrome P450 pathway were reduced in cART/HIV-1-exposed infants. Taken together, these data demonstrate that the fetal metabolism is impacted by maternal factors (cART and HIV-1) and skews physiological immune responses toward inflammation in the newborn infant. [ABSTRACT FROM AUTHOR]

Many of the alterations that affect innate and adaptive immune cell compartments in HIV-infected patients are reminiscent of the process of immune aging, characteristic of old age. These alterations define the immunological age of individuals and are likely to participate to the decline of immune competence with HIV disease progression. It is therefore important to characterize these changes, which point toward the accumulation of highly differentiated immunocompetent cells, associated with overall telomere length shortening, as well as understanding their etiology, especially related to the impact of chronic immune activation. Particular attention should be given to the exhaustion of primary immune resources, including haematopoietic progenitors and naïve cells, which holds the key for effective hematopoiesis and immune response induction, respectively. The alteration of these compartments during HIV infection certainly represents the foundation of the immune parallel with aging. [ABSTRACT FROM AUTHOR]

Objective: Zidovudine (ZDV) has been extensively used in pregnant women to prevent vertical transmission of HIV but few studies have evaluated potential mutagenic effects of ZDV during fetal development., Design: Our study investigated clonal hematopoiesis in HIV-exposed uninfected (HEU) newborns, 94 of whom were ZDV-exposed and 91 antiretroviral therapy (ART)-unexposed and matched for potential confounding factors., Methods: Utilizing high depth sequencing and genotyping arrays, we comprehensively examined blood samples collected during the first week after birth for potential clonal hematopoiesis associated with fetal ZDV exposure, including clonal single nucleotide variants (SNVs), small insertions and deletions (indels), and large structural copy number or copy neutral alterations., Results: We observed no statistically significant difference in the number of SNVs and indels per person in ZDV-exposed children (adjusted ratio [95% confidence interval, CI] for expected number of mutations = 0.79 [0.50--1.22], P = 0.3), and no difference in the number of large structural alterations. Mutations in common clonal hematopoiesis driver genes were not found in the study population. Mutational signature analyses on SNVs detected no novel signatures unique to the ZDV-exposed children and the mutational profiles were similar between the two groups., Conclusion: Our results suggest that clonal hematopoiesis at levels detectable in our study is not strongly influenced by in-utero ZDV exposure; however, additional follow-up studies are needed to further evaluate the safety and potential long-term impacts of in-utero ZDV exposure in HEU children as well as better investigate genomic aberrations occurring late in pregnancy., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Pulmonary malignancies are a major source of morbidity and mortality in HIV-infected persons. Non-AIDS-defining lung cancers (mostly non-small cell lung cancers) are now a leading cause of cancer death among HIV-infected persons. HIV-associated factors appear to affect the risk of lung cancer and may adversely impact cancer treatment and outcomes. HIV infection also may modify the potential harms and benefits of lung cancer screening with computed tomography. AIDS-defining lung malignancies include pulmonary Kaposi sarcoma and pulmonary lymphoma, both of which are less prevalent with widespread adoption of antiretroviral therapy. [ABSTRACT FROM AUTHOR]

Ubiquitinases are a select group of enzymes that modify target proteins through ubiquitination, which plays a crucial role in the regulation of protein degradation, location, and function. B lymphocytes that originated from bone marrow hematopoietic stem cells (HSC), exert humoral immune functions by differentiating into plasma cells and producing antibodies. Previous studies have shown that ubiquitination is involved in the regulation of the cell cycle and signal transduction important for B lymphocyte development and function. In this review, how ubiquitinases regulate B cell development, activation, apoptosis, and proliferation is discussed, which could help in understanding the physiological processes and diseases related to B cells and also provides potential new targets for further studies., (©2020 Society for Leukocyte Biology.)

Naturally occurring stem cells isolated from humans have been used therapeutically for decades. This has primarily involved the transplantation of primary cells such as haematopoietic and mesenchymal stem cells and, more recently, derivatives of pluripotent stem cells. However, the advent of cell-engineering approaches is ushering in a new generation of stem cell-based therapies, greatly expanding their therapeutic utility. These next-generation stem cells are being used as 'Trojan horses' to improve the delivery of drugs and oncolytic viruses to intractable tumours and are also being engineered with angiogenic, neurotrophic and anti-inflammatory molecules to accelerate the repair of injured or diseased tissues. Moreover, gene therapy and gene editing technologies are being used to create stem cell derivatives with improved functionality, specificity and responsiveness compared with their natural counterparts. Here, we review these engineering approaches and areas in which they will help broaden the utility and clinical applicability of stem cells.

Epstein-Barr virus (EBV) was discovered 50 years ago from an African Burkitt lymphoma cell line. EBV-associated lymphoproliferative disorders (LPDs) are lifethreatening diseases, especially in children. In this article, we review EBV-associated LPDs, especially in the area of primary immunodeficiency disease (PID). We searched PubMed for publications with key words including EBV infection, lymphoma, LPDs and PID, and selected the manuscripts written in English that we judged to be relevant to the topic of this review. On the basis of the data in the literature, we grouped the EBV-associated LPDs into four categories: nonmalignant disease, malignant disease, acquired immunodeficiency disease and PID. Each category has its own risk factor for LPD development. EBV-associated LPD is a complex disease, creating new challenges for diagnosis and treatment. [ABSTRACT FROM AUTHOR]

The risk of serious bacterial infection in human immunodeficiency virus–exposed but uninfected infants is inverse to the mother's CD4 count. This association remains significant during the first year of life.Background. Morbidity and mortality are higher among human immunodeficiency virus (HIV) exposed but uninfected (HEU) infants than unexposed infants, particularly if the mother had a low CD4 count. We investigated the possible association between maternal immune depression during pregnancy and the risk of infection in HEU infants in the national French Perinatal Cohort (EPF).Methods. All neonates, born alive, to HIV-1–infected women enrolled in the EPF between 2002 and 2010 were included. The primary outcome was the first serious (hospitalization or death) infection during the first year of life. The main exposure variable was maternal CD4 cell count near delivery. The Kaplan–Meier method and multivariate Cox models were applied, with the different types of infections managed as competing events.Results. Among 7638 HEU neonates, 699 had at least 1 serious infection (of which 159 were bacterial) with a Kaplan–Meier probability of 9.3% (95% confidence interval, 8.7–10.0) at 1 year. The risk of serious bacterial infection during the first year of life significantly increased with lower maternal CD4 cell count, before and after adjustment for maternal CD4 cell count <350 and 350–499 CD4/mm3 (adjusted hazard ratio = 1.7 [1.2–2.6] and 1.2 [0.8–1.9], respectively; P = .03). This association mainly concerned infections involving encapsulated bacteria (P = .03). The risk of serious viral infection was, by contrast, independent of the mother's CD4 cell count.Conclusions. Maternal CD4 count is significantly and specifically associated with the risk of serious infections with encapsulated bacteria in HEU infants. [ABSTRACT FROM AUTHOR]

Background. Biomarkers of T-cell receptor excision circles (TRECs) and immunoglobulin κ-deleting recombination excision circles (KRECs) reflect naïve T and B cell emigrants. This study assessed the biomarkers in patients with primary immunodeficiency diseases (PIDs) to determine the lymphocyte output disturbance and the correlation to lymphocytes. Methods. A standard plasmid was constructed to calculate TRECs and KRECs in 250 ng genomic DNA from whole blood of PIDs patients. These were correlated to naïve and memory lymphocytes for further classification and adequate treatment. Results. In 69 studied patients, the low TRECs mainly included those with severe combined T and B immunodeficiency (SCID, 7/8), combined immunodeficiency (CID, 4/4), and common variable immunodeficiency (CVID, 6/7). The diminished KRECs was in SCID (4/8), CID (4/4), CVID (7/7), Bruton's tyrosine kinase mutation (Btk, 3/4), anti-B cell deletion (by anti-CD20 antibody in 1), and Behçet syndrome under steroid treatment (1). The TRECs and KRECs positively correlated to absolute naïve T (CD4 + CD45RA+) and naïve B (CD19 + CD27−), and to memory B (CD19 + CD27+) numbers, respectively. Conclusion. This study validates that low TRECs and KRECs values reflect low naïve T and B lymphocytes in 'combined immunodeficiencies' and in some CVID patients with the potential to develop the CID phenotype. [ABSTRACT FROM AUTHOR]