CLDN1 Arg81His founder variant causes ichthyosis, leukocyte vacuoles, alopecia, and sclerosing cholangitis (ILVASC) syndrome in Moroccan Jews.

Neonatal ichthyosis and sclerosing cholangitis syndrome (NISCH), also known as ichthyosis, leukocyte vacuoles, alopecia, and sclerosing cholangitis (ILVASC), is an extremely rare disease of autosomal recessive inheritance, resulting from loss of function of the tight junction protein claudin-1. Its clinical presentation is highly variable, and is characterized by liver and ectodermal involvement. Although most ILVASC cases described to date were attributed to homozygous truncating variants in CLDN1, a single missense variant CLDN1 p.Arg81His, associated with isolated skin ichthyosis phenotype, has been recently reported in a family of Moroccan Jewish descent. We now describe seven patients with ILVASC, originating from four non consanguineous families of North African Jewish ancestry (including one previously reported family), harboring CLDN1 p.Arg81His variant, and broaden the phenotypic spectrum attributed to this variant to include teeth, hair, and liver/bile duct involvement, characteristic of ILVASC. Furthermore, we provide additional evidence for pathogenicity of the CLDN1 p.Arg81His variant by transmission electron microscopy of the affected skin, revealing distorted tight junction architecture, and show through haplotype analysis in the vicinity of the CLDN1 gene, that this variant represents a founder variant in Jews of Moroccan descent with an estimated carrier frequency of 1:220.

Early diagnosis of ichthyosis, leukocyte vacuoles, alopecia, and sclerosing cholangitis syndrome: A case report.

Congenital ichthyosis is a genodermatosis characterized by abnormal epidermal differentiation. The neonatal period is critical for patients with ichthyosis because of the risk for significant comorbidities and associated mortality, with most complications resulting from impaired barrier function. Early recognition can significantly alter the clinical course of this rare disease. Here we present a neonate with ichthyosis, leukocyte vacuoles, alopecia, and sclerosing cholangitis syndrome (ILVASC), a rare inherited disease, to highlight how an interdisciplinary approach led to prompt assessment, confirmation of a genetic diagnosis and management of potential complications.

A Novel CEBPE Variant Causes Severe Infections and Profound Neutropenia.

PURPOSE: Specific granule deficiency (SGD) is a rare inborn error of immunity resulting from loss-of-function variants in CEBPE gene (encoding for transcription factor C/EBPε). Although this genetic etiology has been known for over two decades, only a few patients with CEBPE variant-proven SGD (type I) have been reported. Herein, we describe two siblings with a novel homozygous CEBPE deletion who were noted to have profound neutropenia on initial evaluation. We aimed to evaluate the immunohematological consequences of this novel variant, including profound neutropenia. METHODS: Light scatter characteristics of granulocytes were examined on various automated hematology analyzers. Phagocyte immunophenotype, reactive oxygen species generation, and Toll-like receptor (TLR) signaling were assessed using flow cytometry. Relative expression of genes encoding various granule proteins was studied using RT-PCR. Western blot analysis and luciferase reporter assay were performed to explore variant C/EBPε expression and function. RESULTS: Severe infections occurred in both siblings. Analysis of granulocyte light scatter plots revealed automated hematology analyzers can provide anomalously low neutrophil counts due to abnormal neutrophil morphology. Neutrophils displayed absence/marked reduction of CD15/CD16 expression and overexpression (in a subset) of CD14/CD64. Three distinct populations of phagocytes with different oxidase activities were observed. Impaired shedding of CD62-ligand was noted on stimulation with TLR-4, TLR-2/6, and TLR-7/8 agonists. We demonstrated the variant C/EBPε to be functionally deficient. CONCLUSION: Homozygous c.655_665del variant in CEBPE causes SGD. Anomalous automated neutrophil counts may be reported in patients with SGD type I. Aberrant TLR signaling might be an additional pathogenetic mechanism underlying immunodeficiency in SGD type I.

Evaluation of neurodevelopmental symptoms in 10 cases of neonatal ichthyosis and sclerosing cholangitis syndrome.

Neonatal ichthyosis and sclerosing cholangitis (NISCH) syndrome is an extremely rare entity with only 19 patients described in the literature. We report an extended family with the disorder and investigate the association of neurodevelopmental symptoms. Patients with CLDN1 mutations, and specifically « the Moroccan¼ c.200_201delTT deletion, may be an increased risk for neurodevelopmental symptoms such as learning disabilities, mental retardation, and language delay.

Identification of a novel IRF8 homozygous mutation causing neutrophilia, monocytopenia and fatal infection in a female neonate.

Inborn errors of immunity (IEIs) result from mutations in genes involved in host immune defense and immune regulation. Herein, we report the identification of a novel IRF8 mutation in a neonate with an IEI. DNA samples from both the neonate and her parents were subjected to DNA sequencing, and the immune status of the patient was assessed. We identified a mutation (c.331C > T, p. Arg111*) in the interferon regulatory factor 8 (IRF8) gene that manifested as sever dysfunctional neutrophilia (96.53 × 109/l) and monocytopenia (0.02 × 109/l). The patient's CD3+ T cell and CD8+ T cell counts were decreased. Her levels of IFN-γ were low even during severe infection. The mRNA expression levels of IRF8 were lower than normal. Her clinical manifestations included a recurrent and progressively fatal infection. Since IRF8 plays a key role in the differentiation and development of immune cells, we suspected that the novel mutation (c.331C > T, p. Arg111*) may be consistent with a severe loss of IRF8 function and result in a failure of immune cells to differentiate and maturation, and lead to a severe infection with early onset.

Autosomal dominant familial periodic fever patient with a missense variant c.215G>A (p.Cys72Tyr) in TNFRSF1A gene presenting as neutrophilia.

Familial periodic fever (FPF) is an uncommonly diagnosed autosomal dominant disorder caused by a genetic alteration in the TNFRSF1A gene. These patients usually present with fever which is usually under-investigated and under-diagnosed. In untreated cases, amyloidosis is a frequent complication. We present a 24 years male who had a history of fever from childhood, however, remained undiagnosed short of genetic testing. He has recurrent episodes of fever. During the episodes of fever, he was found to have leukocytosis (total leukocyte count- 25.7 x10^9/L) and neutrophilia (absolute neutrophil count- 22.7 x10^9/L) both of which came back to normal limits as the fever subsided. On further evaluation for neutrophilia, the exclusion of common causes of neutrophilia was done. Next-generation sequencing detected a missense variant in TNFRSF1A: c.215G > A (p.Cys72Tyr) which was confirmed by Sanger sequencing. This variant has been described in the literature in anecdotal cases of FPF. This is a first case report from the Indian subcontinent reporting TNFRSF1A: c.215G > A (p.Cys72Tyr) variant in a patient of FPF. Short of genetic testing, the fever would remain a diagnostic dilemma in this patient. This report highlights the importance of targeted resequencing in clinching diagnosis in such patients.

Molecular Genetics of Keratinization Disorders - What's New About Ichthyosis.

The heritable forms of keratinization disorders, including various forms of ichthyosis and keratodermas, comprise a phenotypically heterogeneous group of diseases which can be divided into syndromic and non-syndromic forms. In the non-syndromic forms, the clinical manifestations are limited to the cutaneous structures while the syndromic ones are associated with a spectrum of extracutaneous manifestations. The inheritance in different families can be autosomal dominant, autosomal recessive or either X-linked dominant or recessive. Currently at least 67 distinct genes have been associated with different forms of ichthyosis. These genes can be grouped on the basis of their physiological involvement, including genes encoding structural components of epidermis, those involved in epidermal lipid metabolism, or those critical for cell-cell adhesion, and keratinocyte differentiation. This overview highlights some of the recent progress made in understanding the molecular genetics of keratinization disorders, and presents selected, recently characterized cases as representative of different forms of heritable ichthyosis.

Regulatory mechanisms of B cell responses and the implication in B cell-related diseases.

Terminally differentiated B cell, the plasma cell, is the sole cell type capable of producing antibodies in our body. Over the past 30 years, the identification of many key molecules controlling B cell activation and differentiation has elucidated the molecular pathways for generating antibody-producing plasma cells. Several types of regulation modulating the functions of the important key molecules in B cell activation and differentiation add other layers of complexity in shaping B cell responses following antigen exposure in the absence or presence of T cell help. Further understanding of the mechanisms contributing to the proper activation and differentiation of B cells into antibody-secreting plasma cells may enable us to develop new strategies for managing antibody humoral responses during health and disease. Herein, we reviewed the effect of different types of regulation, including transcriptional regulation, post-transcriptional regulation and epigenetic regulation, on B cell activation, and on mounting memory B cell and antibody responses. We also discussed the link between the dysregulation of the abovementioned regulatory mechanisms and B cell-related disorders.

C/EBPε ΔRS derived from a neutrophil-specific granule deficiency patient interacts with HDAC1 and its dysfunction is restored by trichostatin A.

CCAAT/enhancer binding protein epsilon (C/EBPε), a myeloid-specific transcription factor, plays an important role in granulopoiesis. A loss-of-function mutation in this protein can result in an abnormal development of neutrophils and eosinophils, known as neutrophil-specific granule deficiency (SGD). The transcriptional activity of C/EBPε is regulated by interactions with other transcription factors and/or post-translational modification, including acetylation. Previously, we reported a novel SGD patient who had a homozygous mutation for two amino acids, arginine (R247) and serine (S248), which were deleted in the basic leucine zipper domain of C/EBPε (ΔRS) and exhibited loss of transcriptional activity with aberrant protein-protein interactions. In the present study, we found that a single amino acid deletion of either R247 (ΔR) or S248 (ΔS) was sufficient for the loss of C/EBPε transcriptional activity, while an amino acid substitution at S248 to alanine in C/EBPε (SA) had comparable transcriptional activity with the wild-type C/EBPε (WT). Although acetylation at lysine residues (K121 and K198) is indispensable for C/EBPε transcriptional activity, an acetylation mimic form of ΔRS (ΔRS-K121/198Q) did not exhibit the transcriptional activity. Interestingly, we discovered that ΔRS, ΔR, ΔS, and ΔRS-K121/198Q interacted with histone deacetylase 1 (HDAC1), whereas WT and SA did not. Furthermore, the proteoglycan 2/eosinophil major basic protein induction activity of ΔRS, ΔR, and ΔS could be restored by the HDAC inhibitor, trichostatin A (TSA), and protein-protein interactions between ΔRS and Gata1 could also be recovered by TSA treatment. Taken together, our results show that TSA has the potential to restore the transcriptional activity of ΔRS, indicating that the inhibition of HDAC1 could be a molecularly targeted treatment for SGD with ΔRS.

ALS blood expression profiling identifies new biomarkers, patient subgroups, and evidence for neutrophilia and hypoxia.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a debilitating disease with few treatment options. Progress towards new therapies requires validated disease biomarkers, but there is no consensus on which fluid-based measures are most informative. METHODS: This study analyzed microarray data derived from blood samples of patients with ALS (n = 396), ALS mimic diseases (n = 75), and healthy controls (n = 645). Goals were to provide in-depth analysis of differentially expressed genes (DEGs), characterize patient-to-patient heterogeneity, and identify candidate biomarkers. RESULTS: We identified 752 ALS-increased and 764 ALS-decreased DEGs (FDR < 0.10 with > 10% expression change). Gene expression shifts in ALS blood broadly resembled acute high altitude stress responses. ALS-increased DEGs had high exosome expression, were neutrophil-specific, associated with translation, and overlapped significantly with genes near ALS susceptibility loci (e.g., IFRD1, TBK1, CREB5). ALS-decreased DEGs, in contrast, had low exosome expression, were erythroid lineage-specific, and associated with anemia and blood disorders. Genes encoding neurofilament proteins (NEFH, NEFL) had poor diagnostic accuracy (50-53%). However, support vector machines distinguished ALS patients from ALS mimics and controls with 87% accuracy (sensitivity: 86%, specificity: 87%). Expression profiles were heterogeneous among patients and we identified two subgroups: (i) patients with higher expression of IL6R and myeloid lineage-specific genes and (ii) patients with higher expression of IL23A and lymphoid-specific genes. The gene encoding copper chaperone for superoxide dismutase (CCS) was most strongly associated with survival (HR = 0.77; P = 1.84e-05) and other survival-associated genes were linked to mitochondrial respiration. We identify a 61 gene signature that significantly improves survival prediction when added to Cox proportional hazard models with baseline clinical data (i.e., age at onset, site of onset and sex). Predicted median survival differed 2-fold between patients with favorable and risk-associated gene expression signatures. CONCLUSIONS: Peripheral blood analysis informs our understanding of ALS disease mechanisms and genetic association signals. Our findings are consistent with low-grade neutrophilia and hypoxia as ALS phenotypes, with heterogeneity among patients partly driven by differences in myeloid and lymphoid cell abundance. Biomarkers identified in this study require further validation but may provide new tools for research and clinical practice.

Effect of Qinghuang Powder () Combined with Bupi Yishen Decoction () in Treating Patients with Refractory Cytopenia with Multilineage Dysplasia through Regulating DNA Methylation.

OBJECTIVE: To explore the effect of Qinghuang Powder (QHP,()combined with Bupi Yishen Decoction (BPYS, ) on myelodysplastic syndromes (MDS) patients with refractory cytopenia with multilineage dysplasia (RCMD) and determine the change of DNA methylation in MDS-RCMD patients after the treatment of Chinese medicine formula. METHODS: All 308 MDS-RCMD patients were treated with QHP combined with BPYS for 2 months at least, absolute neutrophil count (ANC), hemoglobin (Hb), platelets (PLT), primitive bone marrow cells and chromosome karyotype were chosen as the main evaluation indexes to analyze the treatment effect according to criteria from the MDS International Working Group. Then 43 bone marrow samples from 15 MDS-RCMD patients and 28 healthy donors were obtained for the examination of DNA methylation. Gene Ontology (GO) and Pathway analysis were applied to analyze the methylation data. RESULTS: The overall MDS response rate to QHP was 61.68% (190/360) including hematologic improvement-neutrophil (HI-N) or hematologic improvement-erythroid (HI-E) or hematologic improvement-platelet (HI-P). Patients with anemia had a better response rate than patients with neutropenia or thrombocypenia (55.88% vs 31.54% or 55.88% vs. 36.9%). The DNA methylation microarray analysis disclosed that 4,257 hypermethylated genes were demethylated upon the treatment with QHP and BPYS. GO analysis and Pathway analysis showed that these demethylated genes were involved in a lot of tumor-related pathways and functions. CONCLUSIONS: QHP combined with BPYS could effectively treat MDS-RCMD patients through hematologic improvement (HI-N, HI-P or HI-E) and PLT and RBC transfusion independence due to the demethylation, thereby providing another choice for the treatment of patients with MDS-RCMD.

Bacterial and Pneumocystis Infections in the Lungs of Gene-Knockout Rabbits with Severe Combined Immunodeficiency.

Using the CRISPR/Cas9 gene-editing technology, we recently produced a number of rabbits with mutations in immune function genes, including FOXN1, PRKDC, RAG1, RAG2, and IL2RG. Seven founder knockout rabbits (F0) and three male IL2RG null (-/y) F1 animals demonstrated severe combined immunodeficiency (SCID), characterized by absence or pronounced hypoplasia of the thymus and splenic white pulp, and absence of immature and mature T and B-lymphocytes in peripheral blood. Complete blood count analysis showed severe leukopenia and lymphocytopenia accompanied by severe neutrophilia. Without prophylactic antibiotics, the SCID rabbits universally succumbed to lung infections following weaning. Pathology examination revealed severe heterophilic bronchopneumonia caused by Bordetella bronchiseptica in several animals, but a consistent feature of lung lesions in all animals was a severe interstitial pneumonia caused by Pneumocystis oryctolagi, as confirmed by histological examination and PCR analysis of Pneumocystis genes. The results of this study suggest that these SCID rabbits could serve as a useful model for human SCID to investigate the disease pathogenesis and the development of gene and drug therapies.

Decreased soluble RAGE in neutrophilic asthma is correlated with disease severity and RAGE G82S variants.

The advanced glycosylation end product-specific receptor (RAGE) has been demonstrated to be an important mediator of asthma pathogenesis. The soluble isoform of RAGE (sRAGE) acts as a 'decoy' to sequester RAGE ligands, and thus prevents their binding to the receptor. A number of reports have linked deficiency of sRAGE to the severity and outcomes of various human diseases, and association with RAGE G82S variants. However, whether sRAGE levels are increased or decreased in asthmatic patients is unclear. The aim of the present study was to determine plasma sRAGE levels in different asthma phenotypes and associations of plasma sRAGE levels with RAGE G82S variants. A total of 85 neutrophilic and 109 non­neutrophilic newly diagnosed asthmatic patients, and 118 healthy controls, were recruited. Plasma sRAGE levels were measured by ELISA analysis. RAGE G82S genotypes were detected using the Sanger sequencing method. Plasma sRAGE levels were decreased in neutrophilic asthmatics (443.67±208.9 pg/ml) and increased in non­neutrophilic asthmatics (677.63±300.75 pg/ml) compared with healthy controls (550.02±300.83 pg/ml) (P<0.001). Plasma sRAGE levels were positively correlated with FEV1% predicted (FEV1% Pre) (rp=0.258; P=0.023) in neutrophilic asthmatics. The frequency of G82S genotypes was significantly different between neutrophilic and non­neutrophilic asthmatics (P=0.009). Neutrophilic asthmatics with genotypes A/G or A/A (389.83±150.37 and 264.59±161.74 pg/ml, respectively) had significantly decreased sRAGE levels compared with the G/G genotype (498.64±235.37 pg/ml) (P=0.022). Those with the A/G and A/A genotype (60.14±22.36%) displayed a trend toward lower FEV1% Pre compared with those with the G/G genotype (64.51±27.37%). No significant difference in sRAGE levels or an association with FEV1% Pre was observed between the different genotypes in non­neutrophilic asthmatics. In conclusion, the results of the present study indicated that plasma sRAGE levels are altered in different asthma inflammatory phenotypes. Plasma sRAGE may be a biomarker of asthma severity and may be associated with G82S gene variants in neutrophilic asthmatics.

[NISCH syndrome, a rare cause of neonatal cholestasis: A case report]. / Le syndrome NISCH, une cause rare de cholestase néonatale : à propos d'un cas.

NISCH syndrome is a rare autosomal recessive disease. It is characterized by scalp hypotrichosis, scarring alopecia, ichthyosis, and neonatal sclerosing cholangitis. It is caused by mutations in the CLDN1 gene encoding the claudin-1 protein, which is located at tight junctions. Fifteen cases have been reported to date and three different mutations have been identified. We report on the case of a 2-year-old boy from a consanguineous Moroccan family, presenting with NISCH syndrome and carrying the so-called Moroccan homozygous mutation (c.200-201delTT). The patient presented with the characteristic symptoms of the syndrome and a favorable progression with normalization of hepatic analyses under symptomatic treatment (vitamin supplementation and ursodeoxycholic acid). The currently limited availability of clinical and therapeutic data does not allow accurate prediction of the course of the disease and short- and long-term prognosis. Moreover, substantial interindividual variability has been reported. Description of new cases will provide new insights into the understanding and the overall management of this syndrome, the course of which remains elusive.