Characteristics of patients with C1 esterase inhibitor deficiency: a single center study.

Summary: Hereditary angioedema (HAE) is a primary complement factor deficiency, characterized by recurrent submucosal/subcutaneous swelling episodes. SERPING1 gene defects encoding C1 esterase inhibitor (C1INH) are responsible from the disease. Fifteen patients with HAE are retrospectively evaluated in this study. All patients (n = 15) had HAE type I, 13 were from the same family, other two from two different families. Median age at onset of symptoms was 7 years (2-20); median age on diagnosis, 12 (0,5-41) and median delay in diagnosis, 3 years (0-33). Clinical symptoms were extremity edema(92,3%), facial edema(46%), abdominal pain (46%), genital edema (46%), and laryngeal edema (23%). Some patients suffered from recurrent abdominal pain, had been empirically given colchicine with familial mediterranean fever (FMF) when they admitted. One presented with bullous skin eruption, soon after developed extremity edema. Both resolved spontaneously after C1INH concentrate therapy. Two females suffered from recurrent genital edema after sexual activity. One patient experienced compartment syndrome-like swelling of extremity after playing football. One patient diagnosed with panic attack due to fear of death by asphyxiation, and was diagnosed with HAE disease. A nonsense mutation in exon 8, a missense mutation in exon 2 in SERPING1 gene was present in Family 1 and another patient (P14) from the other family, respectively. Sporadic/autosomal dominant inheritance ratio was 2/3 in the families present in our series. Patients with HAE presents with a large spectrum of symptoms. In mediterranean countries, patients with abdominal attacks may be misdiagnosed with FMF. Thus, health-care professionals should be alert, and put HAE in the first line of differential diagnoses when the disease symptoms are present. Consequently, morbidity/mortality will decrease with effective treatment options.

Primary Immunodeficiency Disorders in children with Non-Cystic Fibrosis Bronchiectasis.

Summary: Introduction. Primary immunodeficiency diseases (PID) are common in patients with non-cystic fibrosis bronchiectasis (NCFB). Our objective was to determine ratio/types of PID in NCFB. Methods. Seventy NCFB patients followed up in a two-year period were enrolled. Results. Median age was 14 years (min-max: 6-30). Male/female ratio was 39/31; parental consanguinity, 38.6%. Most patients with NCFB (84.28%) had their first pulmonary infection within the first year of their lives. Patients had their first pulmonary infection at a median age of 6 months (min-max: 0.5-84), were diagnosed with bronchiectasis at about 9 years (114 months, min-max: 2-276). PID, primary ciliary dyskinesia (PCD), bronchiolitis obliterans, rheumatic/autoimmune diseases, severe congenital heart disease and tuberculosis were evaluated as the most common causes of NCFB. About 40% of patients (n=16) had bronchial hyperreactivity (BH) and asthma. Twenty-nine patients (41.4%) had a PID, and nearly all (n=28) had primary antibody deficiency, including patients with combined T and B cell deficiency. PID and non-PID groups did not differ according to gender, parental consanguinity, age at first pneumonia, age of onset of chronic pulmonary symptoms, bronchiectasis, presence of gastroesophageal reflux disease (GERD), BH and asthma (p greater-than 0.05). Admission to immunology clinic was about 3 years later in PID compared with non-PID group (p less-than 0.001). Five patients got molecular diagnosis, X-linked agammaglobulinemia (n=2), LRBA deficiency (n=1), RASGRP1 deficiency (n=1), MHC Class II deficiency (n=1). They were given monthly IVIG and HSCT was performed for three patients. Conclusions. PID accounted for about 40% of NCFB. Early diagnosis/appropriate treatment have impact on clinical course of a PID patient. Thus, follow-up in also immunology clinics should be a routine for patients who experience pneumonia in the first year of their lives and those with NCFB.

Clinical and genetic features of the patients with X-Linked agammaglobulinemia from Turkey: Single-centre experience.

X-linked agammaglobulinemia is a primary immunodeficiency disorder resulting from BTK gene mutations. There are many studies in the literature suggesting contradictory ideas about phenotype-genotype correlation. The aim of this study was to identify the mutations and clinical findings of patients with XLA in Turkey, to determine long-term complications related to the disease and to analyse the phenotype-genotype correlation. Thirty-two patients with XLA diagnosed between 1985 and 2016 in Pediatric Immunology Department of Hacettepe University Ihsan Dogramaci Children's Hospital were investigated. A clinical survey including clinical features of the patients was completed, and thirty-two patients from 26 different families were included in the study. Getting early diagnosis and regular assessment with imaging techniques seem to be the most important issues for improving the health status of the patients with XLA. Early molecular analysis gives chance for definitive diagnosis and genetic counselling, but not for predicting the clinical severity and prognosis.

Investigation of Genetic Defects in Severe Combined Immunodeficiency Patients from Turkey by Targeted Sequencing.

Primary immunodeficiencies (PIDs) represent a large group of disorders with an increased susceptibility to infections. Severe combined immunodeficiency (SCID) is the most severe form of primary immunodeficiencies (PIDs) with marked T-cell lymphopenia. Investigation of the genetic aetiology using classical Sanger sequencing is associated with considerable diagnostic delay. We here established a custom-designed, next-generation sequencing (NGS)-based panel to efficiently identify disease-causing genetic defects in PID patients and applied this method in SCID patients of Turkish origin with previously undefined genetic aetiology. We used HaloPlex enrichment technology, a targeted, NGS-based method which was designed to diagnose patients with SCID and other PIDs. Our HaloPlex panel included a total of 356 PID-related genes, and we searched disease-causing mutations in 19 Turkish SCID patients without a genetic diagnosis. The coverage of targeted regions ranged from 97.47% to 99.62% with an average of 98.31% for all patients. All known SCID genes were covered with a percentage of at least 97.3%. We made a genetic diagnosis in six of 19 (33%) patients, including four novel disease-causing mutations identified in RAG1, JAK3 and IL2RG, respectively. We showed that this NGS-based method can provide rapid genetic diagnosis for patients suffering from SCID, potentially facilitating clinical treatment decisions.

Association of interleukin-1 beta (+3954) gene polymorphism and gingival crevicular fluid levels in patients with aggressive and chronic periodontitis.

BACKGROUND AND OBJECTIVE: The immune mechanisms and genetic variations that regulate genetic expression, production and biological activity of IL-1beta, are thought to play an important role in the pathogenesis of periodontal disease. The aims of the present study were to analyse interleukin (IL)-1beta (+3954) genotype and allele frequency in both chronic and aggressive periodontitis patients, and also to investigate whether this polymorphism is associated with gingival crevicular fluid (GCF) IL-1beta levels, periodontal disease severity and clinical parameters in subjects of Turkish origin. METHODS: A total of 147 individuals were enrolled in the study including 56 aggressive periodontitis (AP), 44 chronic periodontitis (CP) patients and 47 healthy controls (C). Single nucleotide polymorphism at IL-1beta (+3954) is analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). GCF samples were analyzed for IL-1beta, using enzyme linked immunosorbent assay (ELISA). RESULTS: The distribution of genotypes and allele frequencies for IL-1beta (+3954) were similar among the groups, in spite of a trend toward a higher frequency of allele 2 in the patient groups. The genotype distribution and allele frequencies were also not different after stratification of subjects according to the clinical attachment level (CAL < 4 mm and CAL > 4mm). No differences were found between the GCF IL-1beta levels of the different genotypes. Allele 2 was associated with increased bleeding on probing (BOP) sites in chronic periodontitis patients. CONCLUSION: The results of this study do not support that genetic polymorphism in the IL-1beta (+3954) could be identified as a susceptibility or severity factor in aggressive periodontitis, in the present population. The association of allele 2 frequency and higher percentage of BOP sites in chronic periodontitis suggest that IL-1beta (+3954) potentially play a significant but not major role in the clinical outcome.

Naturally acquired hepatitis A antibodies after haematopoetic stem cell transplantation.

Haematopoietic stem cell transplant (HSCT) recipients lose immune memory of exposure to infectious agents and vaccines accumulated throughout their lifetime and therefore need to be revaccinated. We aimed to evaluate the influence of different factors on hepatitis A virus (HAV) immunity in both child and adult HSCT recipients living in an intermediate endemic region, Turkey. Eighty patients (age range 2·5-57 years) who had HAV serology prior to HSCT were evaluated. The prevalence of HAV seropositivity was 85% (n=68) before HSCT. There was no history of HAV vaccination before HSCT in children and HAV vaccine was not available in Turkey 10 years ago, so it was assumed that all seropositive patients reflected natural immunity. After the exclusion of six patients with autologous HSCT, the remaining 62 seropositive and allogeneic patients were included in this retrospective study. The duration of HAV seropositivity was estimated using the Kaplan-Meier method, log-rank analysis and Cox regression models. Estimated mean time to loss of HAV seropositivity was 48·6 months after transplantation. Patients who were older (⩾18 years) at transplantation and who had older (⩾18 years) donors became seronegative later (P<0·05). Cox backward-stepwise regression confirmed that older age of recipient at transplantation was the only significant parameter for HAV seropositivity (P<0·05). HAV-inactivated vaccine might be recommended later to older HSCT recipients in intermediate endemic regions.

Cernunnos deficiency: a case report.

B cell-negative severe combined immunodeficiency (SCID) is caused by molecules involved in the variable (diversity) joining (V[D]J) recombination process. Four genes involved in the nonhomologous end joining pathway--Artemis, DNA-PKcs, DNA ligase 4, and Cernunnos--are involved in B cell-negative radiosensitive SCID. Deficiencies in DNA ligase 4 and the recently described Cernunnos gene result in microcephaly, growth retardation, and typical bird-like facies. Lymphopenia and hypogammaglobulinemia with normal or elevated immunoglobulin (Ig) M levels indicate a defect in V(D)J recombination. We present a case with recurrent postnatal pulmonary infections leading to chronic lung disease, disseminated molluscum contagiosum, lymphopenia, low IgG, IgA and normal IgM levels. Our patient had phenotypic features such as microcephaly and severe growth retardation. Clinical presentation in patients with the B cell-negative subtype ranges from SCID to atypical combined immunodeficiency, occasionally associated with autoimmune manifestations and cytomegalovirus infection. Our patient survived beyond infancy with combined immunodeficiency and no autoimmune manifestations.

Six different CYBA mutations including three novel mutations in ten families from Turkey, resulting in autosomal recessive chronic granulomatous disease.

BACKGROUND: One of the rarest forms of autosomal recessive chronic granulomatous disease (AR-CGD) is attributable to mutations in the CYBA gene, which encodes the alpha polypeptide of cytochrome b(558), (also known as p22-phox), a key transmembrane protein in the phagocyte NADPH oxidase system. This gene is localized on chromosome 16q24, encompasses 8.5 kb and contains six exons. MATERIALS AND METHODS: We report here the clinical and molecular characterization of 12 AR-CGD patients from 10 consanguineous, unrelated Turkish families with clinical CGD and positive family history. The ages of the six male and six female patients were between 1and 18 years. Before mutation analysis, subgroup analysis of patients was made by flow cytometry with antibodies against NADPH-oxidase components and with the DHR assay (flow cytometric assay of NADPH oxidase activity in leucocytes). RESULTS: Mutation analysis of CYBA showed six different mutations: a frameshift insertion in exon 3 (C after C166); a missense mutation in exon 2 (p.Gly24Arg), a splice-site deletion in intron 1 (4-bp deletion +4_+7 AGTG), a novel nonsense mutation in exon 6 (p.Cys113X), a novel large deletion of exons 3-6 and a novel 1-bp deletion in exon 6 (c.408delC). All mutations were present in homozygous form and all parents investigated were found to be heterozygotes for these mutations. CONCLUSIONS: In our series of 40 CGD families, approximately 25% of the families have p22-phox defects, with six different mutations, including three novel mutations. The high rate of consanguineous marriages seems to be the underlying aetiology.

Four different NCF2 mutations in six families from Turkey and an overview of NCF2 gene mutations.

BACKGROUND: One of the rarest forms of autosomal recessive chronic granulomatous disease (AR-CGD) is attributable to mutations in the NCF2 gene, which encodes the polypeptide p67(phox), a key cytoplasmic protein in the phagocyte NADPH oxidase system. NCF2 is localized on chromosome 1q25, encompasses 40 kb and contains 16 exons. MATERIALS AND METHODS: We report here the clinical and molecular characterization of six patients with CGD from six consanguineous Turkish families. The ages of the five female patients were between 3 and 22 years and a male patient was 2 years old; all patients showed clear clinical symptoms of CGD. RESULTS: The mothers of the patients did not show a bimodal histogram pattern specific for X-CGD in the dihydrorhodamine-1,2,3 (DHR) assay. Moreover, p67(phox) protein expression was not detectable using flow cytometric analysis of the patients' neutrophils except in those from patient 6, which had a diminished expression. Mutation analysis of NCF2 revealed four different homozygous mutations: a novel nonsense mutation in exon 3 c.229C>T, p.Arg77X; a novel missense mutation in exon 4 c.279C>G, p.Asp93Glu; a nonsense mutation in exon 4 c.304C>T, p.Arg102X; and a novel missense mutation in exon 6 c.605C>T, p.Ala202Val. The parents were found to be heterozygotes for these mutations. CONCLUSIONS: The prevalence of NCF2 mutant families is approximately 15% in our series of 40 CGD families. This high incidence of A67 CGD in Turkey is undoubtedly caused by the high incidence of consanguineous marriages. We found three new mutations in NCF2 and one previously described. These are presented together with an overview of all NCF2 mutations now known.

Life-threatening neurological complications after bone marrow transplantation in children.

Neurological complications may occur in BMT recipients (11-59%), frequently contributing to morbidity or mortality. They are the main causes of death in 10-15%. Life-threatening neurological complications were seen in 11 out of 113 (9.7%) children who underwent BMT from HLA-matched family (n=7) or mismatched donors (n=4) at our institution. Diagnoses of patients with neurological complications were acute myeloblastic leukemia (AML) (five), thalassemia major (two), Fanconi anemia (two), Omenn syndrome (one) and leukodystrophy (one), and the neurological events were seen between days +13 and +85 after transplantation. Minor symptoms including reversible, nonrepetitive seizures were excluded. Cyclosporine A toxicity was diagnosed in six children. The rest of the complications were brain abscess/meningoencephalitis (two), severe hypomagnesemia (one), busulfan toxicity (one), sustained hypertension (three), and intracranial hemorrhage (three). Six patients with neurological complications suffered from >grade II graft-versus-host disease (GvHD), and all were high risk for transplant-related complications. In this study, risk status of the underlying disease, mismatched transplantation, a diagnosis of AML (advanced stage), older age and >grade II GvHD were important adverse factors for the development of severe life-threatening neurological complications.

Bruton tyrosine kinase gene mutations in Turkish patients with presumed X-linked agammaglobulinemia.

X-linked agammglobulinemia (XLA) is a ptototypical humoral immunodeficiency caused by mutations in the gene coding for Bruton tyrosine kinase (BTK). The genetic defect in XLA impairs early B cell development resulting in marked reduction of mature B cells in the blood. Studies from different countries have demonstrated that approximately 90% of males with presumed XLA bear mutations in BTK. In this study, we report for the first time the occurrence of BTK mutations in Turkey. We performed mutational analysis of the BTK gene in 16 Turkish male patients from 13 separate families with presumed XLA based on abnormally low peripheral blood B-cell numbers (lt; 1%), hypogammaglobulinemia, and recurrent bacterial infections. We found that in nine of the 13 families (69%) a Btk mutation caused XLA. Two of the mutations were previously described, but seven novel mutations were identified: two missense (Y39C, G584R), one nonsense (Q343X), and 4 deletions (1800-1821del, 1843-1847del, 1288-1292del, 291del) resulting in frameshift and premature stop codon. By contrast, no mutations in the BTK gene were identified in the other 4 families. A consanguinity in three of these families raises the possibility that mutations in other autosomal genes which affect early B cell development may contribute to their phenotype resembling XLA.

High levels of delayed radiation-induced apoptosis observed in lymphoblastoid cell lines from ataxia-telangiectasia patients.

PURPOSE: Cells from ataxia-telangiectasia (A-T) patients are extremely sensitive to radiation but display decreased apoptosis, as measured during the first 3 days following radiation. To explain this apparent contradiction, we examined apoptosis in normal and A-T cells at late time points following radiation, under the assumption that radiation-induced apoptosis is delayed in the A-T cells. METHODS AND MATERIALS: Blood cells and lymphoblastoid cell lines from A-T patients, as well as healthy donors, were irradiated with X-rays. Apoptosis was measured at different time points (up to 7 and 30 days for lymphocytes and lymphoblastoid cells, respectively) using a flow cytometric method based on the reduction of intracellular DNA content (sub-G1 population). RESULTS: Compared to normal cells, CD4 and CD8 A-T lymphocytes displayed constantly reduced levels of radiation-induced apoptosis for up to 7 days after treatment. A-T lymphoblastoid cells, however, displayed a delayed and prolonged apoptosis. CONCLUSION: A-T lymphoblastoid cells show high levels of delayed radiation-induced apoptosis, which may contribute to the high cellular radiosensitivity displayed by the A-T phenotype. ATM (the gene mutated in A-T) plays different roles in the apoptotic response to ionizing radiation in quiescent lymphocytes and proliferative lymphoblastoid cells.

Successful bone marrow transplantation in a case of Griscelli disease which presented in accelerated phase with neurological involvement.

Griscelli disease (GD) is a rare disorder characterized by pigment dilution, immunodeficiency and occurrence of accelerated phase consisting of hemophagocytosis, pancytopenia and neurological manifestations. Allogeneic BMT in the early period is an important modality of treatment for GD. We carried out an alloBMT from an HLA-identical sibling donor on a 4-year-old girl who presented in accelerated phase with neurological manifestations including convulsions, strabismus, severe dysarthria, ataxia and clonus. She was treated with etoposide, methylprednisolone and intrathecal methotrexate for 8 weeks and underwent alloBMT after receiving a conditioning regimen including ATG (rabbit, 10 mg/kg x 5 days), Bu/Cy. 8 x 108/kg nucleated bone marrow cells were given. Engraftment occurred early and the post-BMT period was uneventful. Currently, she is at 18 months post BMT with sustained engraftment and with a normal neurological examination except for minimal clonus. Long-term follow-up will determine the prognosis regarding the neurological findings.

Effects of intravenous immunoglobulin on clinical and immunological findings of patients with humoral immunodeficiency diseases.

We evaluated nine patients with humoral immunodeficiency (6 immunodeficiency with hyper-IgM, 2 X-linked agammaglobulinemia, 1 common variable immunodeficiency) who were being treated with intravenous immunoglobulins (IVIG). After the use of the IVIG regimen in a dose of 250-300 mg/kg/4 weeks for one year, the severity and frequency of infections, even in patients with chronic lung disease, decreased significantly. An improvement in pulmonary function tests was observed in four patients who had airway obstruction prior to IVIG therapy. Side effects such as chills and fever were observed in 21 of 91 infusions, particularly in the early months of therapy. Preinfusion administration of aspirin and diphenhydramine prevented these side effects. The inversion of the CD4+/CD8+ ratio was detected in most patients during both intramuscular gammaglobulins (IMIG) and IVIG therapy.

IgG subclasses in symptomatic IgA deficiency.

We evaluated IgG subclass levels in 11 symptomatic patients (ages between 3 and 22; mean 7.5 years) with IgA deficiency, seven with selective IgA deficiency, and four with low IgA levels. All patients had experienced three or more episodes of sinopulmonary infections a year. Combined IgG2-IgG4 deficiency was detected in two patients, IgG2 deficiency in one patient and IgG4 deficiency in two patients. Elevated IgG1 and IgG3 levels were detected in most of the IgG subclass deficient and sufficient patients. It is known that gammaglobulin replacement therapy reduces the frequency of infections significantly in IgG subclass deficiency. Although immunization against IgA is a risk in IgA deficiency, these patients can be treated with gammaglobulins containing low IgA.

Clinical and immunological aspects of hyper-IgM syndrome.

Eight patients with Hyper-IgM syndrome were subjected to clinical and immunological evaluation. There were seven males and one female. All the patients had recurrent pyogenic infections; one had lymphoid hyperplasia with centrally necrotic granulomas, and one had gingivitis with neutropenia. Isohemagglutinin titers were either high or normal in all the patients and five had group 0 blood. The percentage of IgM-bearing cells were normal in five patients. The percentage of T cells were normal in all the patients, helper T cells were decreased in two patients, and suppressor T cells were increased in four patients. These results suggest that at least in some patients, the imbalances of T cell subsets may play a role in the pathogenesis of the disease rather than it being attributed to an intrinsic B cell defect.

Serum IgD concentrations in immunodeficiency diseases.

Serum IgD levels were determined in 66 patients with well-defined primary immunodeficiency diseases. The two major groups of patients consisted of those with ataxia-telangiectasia (38 patients) and those with selective IgA deficiency (11 patients). The ataxia-telangiectasia patients tended to have higher serum IgD levels while no significant difference was found in the serum IgD levels of the selective IgA deficiency patients when compared with the controls. No correlation was found between the IgD levels and the presence of frequent infections in both patient groups or the associated disorders present in the selective IgA deficiency patients. The percentage of low serum IgD phenotype in the normal subjects was similar to that described in the literature.

Allogeneic bone marrow transplantation for children with myelodysplastic syndrome.

Six children with myelodysplastic syndrome underwent allogeneic bone marrow transplantation (BMT) from their HLA-identical siblings. Ages ranged from six to 16 years. French-American British (FAB) diagnosis was refractory anemia with excess blasts (RAEB) in three, RAEB in transformation (RAEB-t) in one and chronic myelomonocytic leukemia (CMML) in two cases. Two patients had progressed to leukemia before BMT. All patients received busulfan and cyclophosphamide as a conditioning regimen. Antithymocyte globulin (ATG) was administered to two of them due to the multiple transfusion history. Graft versus host disease (GvHD) prophylaxis consisted of cyclosporine-methotrexate. Engraftment was documented in all patients except one who underwent a second infusion of bone marrow cells. She died in the early post-transplant period with pancytopenia and veno-occlusive disease of the liver. Two patients died from disease recurrence. Three patients are alive > 12 months post-transplant, two are in remission and one just relapsed at +16 months and is now being prepared for a second bone marrow transplant. The only significant factor for favorable outcome was short duration between diagnosis to transplant in the two patients in remission.