Mesenchymal Stem/Stromal Cells Overexpressing CXCR4R334X Revealed Enhanced Migration: A Lesson Learned from the Pathogenesis of WHIM Syndrome.

C-X-C chemokine receptor type 4 (CXCR4), initially recognized as a co-receptor for HIV, contributes to several disorders, including the WHIM (Warts, Hypogammaglobulinemia, Infections, and Myelokathexis) syndrome. CXCR4 binds to its ligand SDF-1 to make an axis involved in the homing property of stem cells. This study aimed to employ WHIM syndrome pathogenesis as an inspirational approach to reinforce cell therapies. Wild type and WHIM-type variants of the CXCR4 gene were chemically synthesized and cloned in the pCDH-513B-1 lentiviral vector. Molecular cloning of the synthetic genes was confirmed by DNA sequencing, and expression of both types of CXCR4 at the protein level was confirmed by western blotting in HEK293T cells. Human adipose-derived mesenchymal stem cells (Ad-MSCs) were isolated, characterized, and subjected to lentiviral transduction with Wild type and WHIM-type variants of CXCR4. The presence of copGFP-positive MSCs confirmed the high efficiency of transduction. The migration ability of both groups of transduced cells was then assessed by transwell migration assay in the presence or absence of a CXCR4-blocking agent. Our qRT-PCR results showed overexpression of CXCR4 at mRNA level in both groups of transduced MSCs, and expression of WHIM-type CXCR4 was significantly higher than Wild type CXCR4 (P<0.05). Our results indicated that the migration of genetically modified MSCs expressing WHIM-type CXCR4 had significantly enhanced towards SDF1 in comparison with Wild type CXCR4 (P<0.05), while it was reduced after treatment with CXCR4 antagonist. These data suggest that overexpression of WHIM-type CXCR4 could lead to enhanced and sustained expression of CXCR4 on human MSCs, which would increase their homing capability; hence it might be an appropriate strategy to improve the efficiency of cell-based therapies.

DL4-µbeads induce T cell lineage differentiation from stem cells in a stromal cell-free system.

T cells are pivotal effectors of the immune system and can be harnessed as therapeutics for regenerative medicine and cancer immunotherapy. An unmet challenge in the field is the development of a clinically relevant system that is readily scalable to generate large numbers of T-lineage cells from hematopoietic stem/progenitor cells (HSPCs). Here, we report a stromal cell-free, microbead-based approach that supports the efficient in vitro development of both human progenitor T (proT) cells and T-lineage cells from CD34+cells sourced from cord blood, GCSF-mobilized peripheral blood, and pluripotent stem cells (PSCs). DL4-µbeads, along with lymphopoietic cytokines, induce an ordered sequence of differentiation from CD34+ cells to CD34+CD7+CD5+ proT cells to CD3+αß T cells. Single-cell RNA sequencing of human PSC-derived proT cells reveals a transcriptional profile similar to the earliest thymocytes found in the embryonic and fetal thymus. Furthermore, the adoptive transfer of CD34+CD7+ proT cells into immunodeficient mice demonstrates efficient thymic engraftment and functional maturation of peripheral T cells. DL4-µbeads provide a simple and robust platform to both study human T cell development and facilitate the development of engineered T cell therapies from renewable sources.

A patient with Silver-Russell syndrome with multilocus imprinting disturbance, and Schimke immuno-osseous dysplasia unmasked by uniparental isodisomy of chromosome 2.

Silver-Russell syndrome (SRS) is a congenital disorder characterized by prenatal and postnatal growth failure and craniofacial features. Hypomethylation of the H19/IGF2:IG-differential methylated region (H19LOM) is observed in 50% of SRS patients, and 15% of SRS patients with H19LOM had multilocus imprinting disturbance (MLID). Schimke immuno-osseous dysplasia (SIOD), characterized by spondyloepiphyseal dysplasia and nephropathy, is an autosomal recessive disorder caused by mutations in SMARCAL1 on chromosome 2. We report a patient with typical SRS-related features, spondyloepiphyseal dysplasia, and severe nephropathy. Molecular analyses showed H19LOM, paternal uniparental isodisomy of chromosome 2 (iUPD(2)pat), and a paternally inherited homozygous frameshift variant in SMARCAL1. Genome-wide methylation analysis showed MLID in this patient, although it showed no MLID in another patient with SIOD without SRS phenotype. These results suggest that iUPD(2)pat unmasked the recessive mutation in SMARCAL1 and that the SMARCAL1 gene mutation may have no direct effect on the patient's methylation defects.

CXCR4 signaling controls dendritic cell location and activation at steady state and in inflammation.

Dendritic cells (DCs) encompass several cell subsets that collaborate to initiate and regulate immune responses. Proper DC localization determines their function and requires the tightly controlled action of chemokine receptors. All DC subsets express CXCR4, but the genuine contribution of this receptor to their biology has been overlooked. We addressed this question using natural CXCR4 mutants resistant to CXCL12-induced desensitization and harboring a gain of function that cause the warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome (WS), a rare immunodeficiency associated with high susceptibility to the pathogenesis of human papillomavirus (HPV). We report a reduction in the number of circulating plasmacytoid DCs (pDCs) in WHIM patients, whereas that of conventional DCs is preserved. This pattern was reproduced in an original mouse model of WS, enabling us to show that the circulating pDC defect can be corrected upon CXCR4 blockade and that pDC differentiation and function are preserved, despite CXCR4 dysfunction. We further identified proper CXCR4 signaling as a critical checkpoint for Langerhans cell and DC migration from the skin to lymph nodes, with corollary alterations of their activation state and tissue inflammation in a model of HPV-induced dysplasia. Beyond providing new hypotheses to explain the susceptibility of WHIM patients to HPV pathogenesis, this study shows that proper CXCR4 signaling establishes a migration threshold that controls DC egress from CXCL12-containing environments and highlights the critical and subset-specific contribution of CXCR4 signal termination to DC biology.

Immunodeficiency in a patient with microcephalic osteodysplastic primordial dwarfism type I as compared to Roifman syndrome.

BACKGROUND: Microcephalic osteodysplastic primordial dwarfism type I (MOPD I, also known as Taybi-Linder syndrome) is a rare genetic disorder associated with severe intrauterine growth retardation, short stature, microcephaly, brain anomalies, stunted limbs, and early mortality. RNU4ATAC, the gene responsible for this disorder, does not encode a protein but instead the U4atac small nuclear RNA (snRNA), a crucial component of the minor spliceosome. Roifman syndrome is an allelic disorder of MOPD I that is characterized by immunodeficiency complications. CASE REPORT: The patient described herein is an 18-year-old woman exhibiting congenital dwarfism and microcephaly with structural brain anomaly. She suffered human herpesvirus 6 (HHV-6)-associated acute necrotizing encephalopathy at the age of one, thereafter resulting in severe psychomotor disabilities. Genetic analysis using gene microarray and whole-exome sequencing could not identify the cause of her congenital anomalies. However, Sanger sequencing revealed a compound heterozygous mutation within RNU4ATAC (NR_023343.1:n.[50G > A];[55G > A]). Immunological findings showed decreases in total lymphocytes, CD4+ T cells, and T cell regenerative activity. Furthermore, antibodies against varicella-zoster, rubella, measles, mumps, and influenza were very low or negative despite having received vaccinations for these viruses. HHV-6 IgG antibodies were also undetected. DISCUSSION: The patient here exhibited a marked MOPD I phenotype complicated by various immunodeficiencies. Previous studies have not demonstrated immunodeficiency comorbidities within MOPD I subjects, but this report suggests an evident immunodeficiency in MOPD I. Patients with MOPD I should be treated with one of the immunodeficiency syndromes.

Alpha-1 Antitrypsin Deficiency and Pulmonary Morbidity in Patients with Primary Immunodeficiency Disease: A Single-Center Experience.

Background: Alpha-1 antitrypsin deficiency (AATD) is of importance in the pathogenesis of pulmonary emphysema, chronic obstructive pulmonary diseases (COPD), and bronchiectasis. Various pulmonary disorders are a typical feature of primary immunodeficiency disease (PID). This includes recurrent pulmonary infections, immunodysregulation, and autoinflammatory diseases. As a result, incidence of acute and chronic pulmonary diseases is higher. Interestingly, pulmonary morbidity in PID and AATD share similar features. To study the coexistence of AATD in patients suffering from PID, we performed the underlying investigation. Methods: We evaluated a study group of 149 patients (n = 149) with PID. In total, serum AAT concentrations were available for 110 patients (n = 110). For the identified patients, we analyzed both clinical associations and interactions. Results: Among the investigated patients, reduced serum AAT levels were detected in 7 patients. With regard to the genotype, PI∗ZZ was found in 2 patients, whereas PI∗MZ was observed in 5 patients. Independent of the underlying phenotype, obstructive lung diseases were found in 2 patients with PI∗ZZ and 2 patients with PI∗MZ. Conclusions: In Germany, the estimated percentage for PI∗ZZ and PI∗MZ is 0.01% and 1.9%, respectively. As demonstrated, the ratio in our study group was even higher. We identified seven patients with AATD. Since AATD contributes to pulmonary morbidity in PID patients, systematic underdiagnosis of the coexistence might yield a strong clinical impact. Hence, AAT analysis should be offered to all patients with confirmed PID diagnoses. To strengthen this finding, we suggest the investigation of larger databases.

Immunodeficiency in cartilage-hair hypoplasia: Pathogenesis, clinical course and management.

Cartilage-hair hypoplasia (CHH) is an autosomal recessive syndromic immunodeficiency with skeletal dysplasia, short stature, hypotrichosis, variable degree of immune dysfunction and increased incidence of anaemia, Hirschsprung disease and malignancy. CHH is caused by variants in the RMRP gene, encoding the untranslated RNA molecule of the mitochondrial RNA-processing endoribonuclease, which participates in for example cell cycle regulation and telomere maintenance. Recent studies have expanded our understanding of the complex pathogenesis of CHH. Immune dysfunction has a major impact on clinical course and prognosis. Clinical features of immune dysfunction are highly variable, progressive and include infections, lung disease, immune dysregulation and malignancy. Mortality is increased compared with the general population, due to infections, malignancy and pulmonary disease. Several risk factors for early mortality have been reported in the Finnish CHH cohort and can be used to guide management. Newborn screening for severe combined immunodeficiency can possibly be of prognostic value in CHH. Regular follow-up by a multidisciplinary team should be implemented to address immune dysfunction in all patients with CHH, also in asymptomatic cases. Haematopoietic stem cell transplantation can cure immune dysfunction, but its benefits in mildly symptomatic patients with CHH remain debatable. Further research is needed to understand the mechanisms behind the variability of clinical features, to search for potential molecular treatment targets, to examine and validate risk factors for early mortality outside the Finnish CHH cohort and to develop management guidelines. This review focuses on the pathogenesis, clinical course and management of CHH.

Schimke immuno-osseous dysplasia, two new cases with peculiar EEG pattern.

INTRODUCTION: Schimke Immuno-Osseous Dysplasia (SIOD) is an autosomal recessive multisystem disorder caused by pathogenic variants in the gene SMARCAL1. The clinical picture is characterized by spondyloepiphyseal dysplasia resulting in growth failure, nephropathy and T-cell deficiency. Neurologic manifestations include microcephaly, cognitive impairment, migraine-like headaches and cerebrovascular manifestations such as cerebral atherosclerotic vascular disease and reversible cerebral vasoconstriction. The role of SMARCAL1 deficiency in non-vascular neurological complications is still under debate. Epilepsy has been reported in a few patients, even in the absence of brain abnormalities. Data regarding electroencephalographic (EEG) patterns in SIOD are scarce METHODS: We describe the clinical, neuroradiological and EEG findings in two unrelated patients with SIOD showing a peculiar pseudo-periodic EEG pattern apparently not related to the cerebrovascular complications, since it was recognized both before and after cerebrovascular events CONCLUSION: Our observations support the hypothesis that SMARCAL1plays an important role in neurodevelopment and brain function and expand the spectrum of neurological abnormalities related to SIOD.

Pulmonary function evaluation in pediatric patients with primary immunodeficiency complicated by bronchiectasis.

BACKGROUND: Primary immunodeficiency (PID) accompanying with recurrent respiratory infections is thought to have a devastating effect on lung function. However, the associations between the airway structural abnormalities on chest computed tomography (CT), severity of dyspnea, and deterioration of pulmonary function test (PFT) have not been fully addressed. METHODS: Children diagnosed with PID in a tertiary referred center in northern Taiwan were enrolled. Demographic and clinical data including age, sex, age at diagnosis of PID, and follow-up period were collected. Chest CT images (modified Reiff scores), parameters of PFT, and life quality questionnaires (mMRC dyspnea scale) were analyzed and correlated using Spearman's rank correlation test. RESULTS: A total of nineteen children with PID were enrolled and thirteen patients were diagnosed as having bronchiectasis based on chest CT scans. Modified Reiff scores of chest CT scan were negatively correlated with FEV1 (% predicted) and FEV1/FVC ratio (P < 0.05). A strongly negative correlation was found between the mMRC dyspnea scale and FEV1 (% predicted) and FVC (% predicted), but positively correlated with RV (% predicted) and RV/TLC ratio (P < 0.05). Furthermore, there was a negative correlation between FVC (% predicted) with increasing follow-up period (P < 0.05). CONCLUSIONS: In pediatric patients with PID, chest CT scan appears to be a good tool for not only the diagnosis of bronchiectasis, but also the degree of pulmonary function impairment. Further quality of life impairments could be particularly due to the airflow obstruction and air trapping related to bronchiectasis.

Inmunodeficiencias primarias y citopenias / Primary immunodeficiencies and cytopenias

Introducción: La visión actual de las enfermedades por inmunodeficiencia primaria (IDP) incluye un número creciente de síndromes que están asociados con la desregulación inmune y la autoinmunidad como características predominantes. Las citopenias autoinmunes pueden ser el primer signo de desregulación que precede a la presentación clásica de inmunodeficiencia primaria, con infecciones recurrentes u oportunistas. El conocimiento de un espectro de enfermedades potencialmente involucradas (hematológicas, reumatológicas e inmunológicas) es crucial para la identificación de una cierta proporción de genotipos y fenotipos de otros diagnósticos descritos. También permitirá excluir desórdenes como lupus eritematoso sistémico, inmunodeficiencia variable común, síndrome linfoproliferativo autoinmune; así como realizar diagnósticos diferenciales noveles como la deficiencia de GATA2, deficiencia de CD27, deficiencia de sensibilidad a lipopolisacáridos, síndrome fosfoinositol-3-quinasa delta activada, inmunodeficiencia ligada a X con déficit de magnesio y otros. Objetivo: Proporcionar una sinopsis conceptual de la aparición de citopenias en las IDP con el propósito de actualizar el conocimiento actual sobre dicho tema y de aumentar la percepción, tanto de hematólogos como inmunólogos, en relación a la presentación de citopenias como manifestación de estas enfermedades. Métodos: Se revisaron artículos originales y de corte experimental publicados en la década 2009 - 2019, en algunas bases de datos de la Biblioteca Virtual de Salud (BVS) de Cuba. Conclusiones: Al igual que las formas benignas autolimitadas de citopenia autoimmune post o parainfecciosas, o la neutropenia autoimmune adquirida de la infancia, que generalmente ocurren independientemente de una IDP subyacente reconocida, muchas de las citopenias que acompañan a esta enfermedad (pero no todas) están mediadas por autoanticuerpos. Es esencial entonces, que los médicos valoren, ante la evidencia clara de citopenia, que esta puede ser autoinmune(AU)

Introduction: The current view of primary immunodeficiency diseases (IDP) includes an increasing number of syndromes that are associated with immune dysregulation and autoimmunity as predominant characteristics. Autoimmune cytopenias may be the first sign of dysregulation that precedes the classic presentation of primary immunodeficiency, with recurrent or opportunistic infections. The knowledge of a spectrum of potentially involved diseases (hematological, rheumatological and immunological) is crucial for the identification of a certain proportion of genotypes and phenotypes of other diagnoses described. It will also allow excluding disorders such as systemic lupus erythematosus, common variable immunodeficiency, autoimmune lymphoproliferative syndrome; as well as making novel differential diagnoses such as GATA2 deficiency, CD27 deficiency, lipopolysaccharide sensitivity deficiency, activated delta phosphoinositol-3-kinase syndrome, X-linked immunodeficiency with magnesium deficiency and others. Objective: This review provides a conceptual synopsis of the appearance of cytopenias in the IDPs with the purpose of updating current knowledge on this topic and increasing the perception, of both hematologists and immunologists, in relation to the presentation of cytopenias as manifestation of these diseases. Methodos: Original and experimental articles published in the 2009-2019 decade were reviewed in some databases of the Virtual Health Library (VHL) of Cuba. Conclusions: As the self-limited benign forms of post or parainfectious autoimmune cytopenia, or childhood acquired autoimmune neutropenia, which generally occur independently of a recognized underlying IDP, many of the cytopenias that accompany this disease (but not all) mediated by autoantibodies. It is essential, then, that doctors assess, given the clear evidence of cytopenia, that it may be autoimmune(AU)

Primary Immunodeficiency and Cancer in Children; A Review of the Literature.

The life span of patients with primary and secondary immunodeficiencies has increased due to recent advances in diagnostic and therapeutic strategies. Primary immune deficiencies (PIDs) are genetic disorders that predispose patients to frequent infections, autoimmunity and malignancies. Genomic instability due to defective DNA repair processes and other unknown mechanisms in patients with PID leads to an enhanced risk of cancer. PIDs were originally described as rare diseases occurring only in infants and young children, which are associated with severe clinical symptoms. However, advances in gene sequencing technologies, have revealed that they are much more common than originally appreciated and are present in older children, adolescents, and adults. After infection, malignancy is the most prevalent cause of death in both children and adults with PIDs. The overall risk of developing cancer in patients with PID is estimated to range from 4.7 to 5.7 percent. A 1.4 to 1.6-fold excess relative risk of cancer has been reported for PIDs. Increasing awareness among physicians regarding PID and cancer may lead to earlier diagnosis which may decrease morbidity and mortality. In this paper, we review the various categories of PIDs in children and highlight their association with various malignancies. MEDLINE was searched to identify articles for inclusion. Three authors have independently screened literature search results from MEDLINE and abstracted data from studies dealing with cancers of children among primary immune deficiencies.

Long-Term Outcome of WHIM Syndrome in 18 Patients: High Risk of Lung Disease and HPV-Related Malignancies.

BACKGROUND: In the warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome, variable phenotypic expression may delay diagnosis. Panleukopenia, malignancy, and chronic lung disease all affect morbidity and mortality risks. Routinely used treatments include immunoglobulins, granulocyte-colony stimulating factor (G-CSF), and antibiotics; recent trials with a target C-X-C chemokine receptor type 4 (CXCR4) antagonist show promising results. OBJECTIVE: We sought to characterize the largest cohort of patients with WHIM and evaluate their diagnostic and therapeutic management. METHODS: Data were collected from an international cohort of 18 patients with CXCR4 mutations. RESULTS: The clinical features manifested at 2.2 ± 2.6 years of age, whereas the disease diagnosis was delayed until 12.5 ± 10.4 years of age. Patients with WHIM commonly presented with a severe bacterial infection (78%). Pneumonia recurrence was observed in 61% of patients and was complicated with bronchiectasis in 27%. Skin warts were observed in 61% of patients at a mean age of 11 years, whereas human papilloma virus (HPV)-related malignancies manifested in 16% of patients. All the patients had severe neutropenia (195 ± 102 cells/mm3 at onset), whereas lymphopenia and hypogammaglobulinemia were detected in 88% and 58% of patients, respectively. Approximately 50% of patients received antibiotic prophylaxis, whereas G-CSF and immunoglobulin treatments were used in 72% and 55% of patients, respectively. CONCLUSIONS: The WHIM syndrome onsets early in life and should be suspected in patients with chronic neutropenia. Patients with WHIM need careful monitoring and timely intervention for complications, mainly lung disease and HPV-related malignancies. We suggest that immunoglobulin therapy should be promptly considered to control the frequency of bacterial infections and prevent chronic lung damage.

Gynecologic health in cartilage-hair hypoplasia: A survey of 26 adult females.

Cartilage-hair hypoplasia (CHH) is a rare metaphyseal chondrodysplasia significantly affecting adult height and quality of life. Immunodeficiency and increased risk for malignancies contribute to significant morbidity. Little is known about gynecologic health in CHH. We performed a questionnaire study of 26 women (mean age 42.3 years) with genetically confirmed CHH, inquiring about pubertal development, menstrual cycle, use of contraception, pregnancies, gynecologic infections, and gynecologic cancers. Mean age at menarche and menopause was 12.7 and 46.1 years, respectively. Mean length of menstrual cycle was 27 days. Contraception was used by 76%, most commonly condom (60%), and combined contraception (60%). Despite significant short stature (mean height 121 cm) and potentially small pelvic diameters, 10 CHH women (38%) had been pregnant. Six of these women reported miscarriages and three had, induced pregnancy terminations. Eight women had in total, 19 deliveries. Abnormal Pap smear was reported in five patients and cervical cancer once. Our findings of normal timing of puberty and menopause suggest a fairly normal length of the fertility period in women with CHH. However, many patients expressed concerns regarding the safety of pregnancy and lack of prepregnancy counseling. Immunodeficiency may predispose CHH women to prolonged HPV infections. This study highlights the importance of careful gynecologic follow-up for these patients.