Genetic basis of non syndromic hypodontia: a DNA investigation performed on three couples of monozygotic twins about PAX9 mutation.

INTRODUCTION: Hypodontia, agenesis of one or of more teeth, is a common developmental dental anomaly. To date, over 200 candidate genes have been demonstrated to be active in tooth development. The genes Pax9 plays an important role in the initial stage of odontogenesis. Mutations of Pax9 are associated with autosomal dominant forms of oligodontia, the agenesis of more than six teeth and occasionally of premolars (MIM 604625) in humans. The aim of the present study was to screen the candidate gene causing the non syndromic hypodontia, with agenesis of upper third molars and upper lateral incisors, in three couples of twins. MATERIALS AND METHODS: Peripheral blood samples taken for routine laboratory investigations were used for genotyping. Total genomic DNA was extracted from the buffy coat of 1 ml of EDTA blood samples using phenol-chloroform and the salting out procedure. RESULTS: The insC mutation (nt793, exon4) was observed in the sequencing results by the use of the primers hPAX9ex4F and hPAX9ex4R. InsC raises a frameshift mutation that introduces a nonsense codon so the mRNA activity results impaired. CONCLUSION: In this work, it is described how the same mutation is responsible for a form of dental agenesis--less severe in the number of missing teeth--leading to hypodontia instead of oligodontia. Therefore, it is possible that mutations of the same gene cause different phenotypes; so we can presume that some modifier genes moderate the effect of the first mutation.

A gene for familial isolated chronic nail candidiasis maps to chromosome 11p12-q12.1.

Chronic mucocutaneous candidiases (CMC) are a group of rare disorders where an altered immune response against Candida leads to persistent and/or recurrent infections of the skin, nails, and mucous membranes. We analysed a five-generation Italian family with an isolated form of CMC, affecting nails only, in the presence of low serum concentration of intercellular adhesion molecule I (ICAM-1). We excluded linkage to candidate regions on chromosomes 2p (CMC with thyroid disease), 21q22.3 (APECED), and 19q13 (ICAM-1). We then carried out a genome-wide scan and assigned the CMC locus to a 19 cM pericentromeric region on chromosome 11.

Mitochondrial DNA mutation at np 3243 in a family with maternally inherited diabetes mellitus.

Mitochondrial DNA (mtDNA) gene defects may play a role in the development of maternally inherited diabetes mellitus and deafness (MIDD). A family from Southern Italy who showed maternal transmission of type 2 diabetes mellitus with three individuals affected is described. A 10.4 kb deletion and mutations at nucleotide positions (np) 3243, 7445 and 11778 in the mtDNA of six relatives were sought. The mitochondrial np 3243 mutation of the tRNA Leu (UUR) gene was identified in a boy affected by optic atrophy and mental retardation, as well as in his diabetic mother. No other mutations or deletions were found. Our study points out the variable phenotypic expression of the np 3243 mtDNA mutation. This may suggest the presence of other mitochondrial or nuclear mutations required to modulate the phenotype. A clinical and metabolic follow-up of all family members was necessary to understand the role of the np 3243 mutation, especially in one child affected by optic atrophy and mental retardation. Further studies will be aimed at investigating the prevalence of mutations and deletions of mtDNA in type 2 diabetes mellitus.

Epidemiology and treatment of otitis media with effusion in children in the first year of primary school.

In this multicentre study we evaluated the prevalence and risk factors of otitis media with effusion (OME) in Italian school-children and the effectiveness of medical treatment of chronic OME with a new cephalosporin, ceftibuten. During two winter periods, 3413 children, aged 5 to 7 years, were examined for the presence of OME by means of pneumotoscopy and a portable, hand-held tympanometer. The prevalence of asymptomatic OME was 14.2%, with no difference as regards sex, age, month of examination or geographic area. Younger children had significantly more bilateral than unilateral effusion. A recent episode of acute otitis media and previous tonsillectomy or adenoidectomy were associated with an increased risk of OME in multivariate logistic regression models. The presence of OME was unrelated to such factors as birthweight, prematurity, sibling or parental history of allergy, duration of daycare attendance, family history of ear infections. After 12 weeks, 26.6% of children with OME still had middle-ear fluid: 52 were randomized to ceftibuten (9 mg/kg q.d. for 14 days) and 59 to no treatment (nasal saline drops allowed). Children treated with ceftibuten had a significantly better resolution of middle-ear effusion after 4 and 8 weeks. As mass screening programmes for OME in the year of school entry are questioned, a focus only on children with known risk factors seems advisable. Ceftibuten can be useful in reducing the duration of middle-ear effusion.

Reduced plasma c-AMP levels in children with atopic dermatitis.

Plasma levels of cAMP and serum concentrations of IgE have been determined in children with acute atopic dermatitis (AD) and in a healthy control group, to illuminate the pathophysiological mechanisms that cause AD. There were significantly lower plasma levels of cAMP (P < 0.001) and significantly higher levels of serum IgE (P < 0.004) in children with AD, in comparison with a healthy control group. It is possible that defective control of c-AMP levels could contribute to the immunopathogenesis of AD and monitoring levels may be of value in the clinical evaluation of the disease.

Allelic association of gene markers on chromosome 11q in Italian families with atopy.

UNLABELLED: In our study, the genetic linkage of the Fcepsilon RIbeta gene with atopy in 77 affected sibling pairs recruited from an Italian panel of 201 subjects has been examined. Atopy was defined by the presence of a positive skin prick test to one or more common aeroallergens, a positive RAST test to one or more common aeroallergens and an elevated circulating total IgE. Genotype analysis was performed by PCR amplification of Fcepsilon RIbeta CA and CI11-319 CA microsatellites. All the family members were also tested for the Ilepsilon 181 mutation with the ARMS method and for Leu181/Leu183 polymorphism. Seventy-two point five percent (72.5%) of the affected sibling pairs shared their maternal allele and 27.5% did not. Therefore, an increased maternal allele sharing was observed: chi2 = 8.10, p < 0.01. Comparing paternal versus maternal allele sharing, a significant difference was observed for the C1II-319 CA marker (chi2 = 4.32, p < 0.05). Atopy phenotype with positive skin prick test, RASTs, and high total serum IgE also showed greater sharing of maternal than paternal alleles in affected sibling pairs. Of the 201 subjects studied, 17 (8.4%) were positive for Leu181. Ten of these were children and seven (70%) had inherited the variant maternally. The seven children had maternally inherited Leu181/Leu183 and were atopic. Within this sample the maternal inheritance of Fcepsilon RIbeta Leu181/Leu183 was associated with an increased risk of IgE responses to common allergens, raised eosinophil counts and increased skin prick test reactions. Therefore, the variant identified a genetic risk factor for atopy. CONCLUSION: The central role of Fcepsilon RIbeta in atopy and the linkage data presented here point to the possibility that genetic variation in Fcepsilon RIbeta or its controlling element may cause differences in the extent of IgE responses between atopic and non-atopic subjects. A search for such mutations or polymorphisms will need to take into account some carriers of atopy among the normal population and genetic heterogeneity among atopic individuals.

Tolerance and pharmacokinetics of ciprofloxacin in the chick. Preliminary experience in subjects of pediatric age with urinary tract infections (UTI).

We studied the tolerance and pharmacokinetic parameters of ciprofloxacin in (2 mg/kg, i.v.) in 45 chicks. Serum concentrations of ciprofloxacin at various times of drawing (15 min, 30 min, 1 hr, 3 hr, 6 hr, 8 hr and 12 hr) peaked in the first 15 min with a trough after 6 hr, and were completely absent after 8 hr. We found no neurotoxic effects or behavioral changes. Encouraged by a good tolerance for this quinolone, we conducted a preliminary study concerning the therapeutic efficacy of ciprofloxacin in 8 subjects of pediatric age (8-14 years) with urinary tract infections (UTI) that are resistant to classic antibiotics. The results and normality of hematologic parameters, checked before and after therapy, allow us to recommend the monitored use of ciprofloxacin in children with UTI resulting from bacteria resistant to other antibiotics.

Serum IgD levels in children with atopic asthma. A longitudinal study.

BACKGROUND: Serum IgD and IgE levels were measured in children with atopic asthma and in control Group in order to determine their relationship with clinical status. METHODS: Samples of venous blood (of 5 cc) were drawn from 25 asthmatic children (Group A) and 25 healthy children (Group B) at the moment of first diagnosis (T0), after 6 months (T180) and after 18 months (T540). To measure IgD, an ELISA assay based on the sandwich principle was used. RESULTS: At T0, IgD were significantly higher in Group A (182.7 5+/-88.18 IU/ml) in comparison with Group B (69.58+/-4.93 IU/ml, p<0.0001); IgD levels decreased in Group A at T540. CONCLUSIONS: In conclusion, a significant increase of IgD levels observed in children at first signs of asthma and the following normalization of these same levels after 18 months, may represent a non specific response or an attempt of the organism to block asthma, favouring therefore immunologic tolerance.

Preliminary data suggest that mutations in the CgRP pathway are not involved in human sporadic cryptorchidism.

In testicular descent to the scrotum, a multistep process, many anatomical and hormonal factors play a role. Cryptorchidism occurs in about 1-2% of males and may cause secondary degeneration of the testes. Animal models have shown that abnormalities, in the calcitonin gene-related peptide (CgRP) activity, could be relevant in the pathogenesis of cryptorchidism. We performed a mutation screening by PCR exon amplification, single-strand conformation polymorphism (SSCP) and sequencing in four candidate genes, CgRPs (alphaCgRP, betaCgRP), their receptor (CgRPR) and the receptor component protein (CgRP-RCP), in 90 selected cases of idiopathic unilateral or bilateral cryptorchidism. Mutation screening of the coding regions and intron-exon boundaries revealed some polymorphic variants but no pathogenic sequence changes. These preliminary data suggest that these genes are not major factors for cryptorchidism in humans.

Molecular analysis of beta-thalassaemia patients in a high incidence area of southern Italy.

The prevalence of eight mutations in 84 patients with beta-thalassaemia major and in 16 subjects with thalassaemia intermedia was investigated. All of the patients were Italian, originating from Eastern Sicily (Messina area) and some Calabrian regions. Genomic DNA was amplified by polymerase chain reaction (PCR). DNA molecular investigations were performed by allele-specific oligonucleotide (ASO) hybridization, to identify the following beta-thalassaemia mutations: CD39 (C-T), IVS1-110 (G-A), IVS1-6 (T-C), IVS1-1 (G-A), IVS2-745 (C-G), IVS2-1 (G-A), -87 (C-G), CD6 A (-A). Our data underline that in thalassemia intermedia two mutations were statistically prevalent: IVS1-6 T-->C (P < 0.001) and CD 6-A (P < 0.05). CD 39 was statistically prevalent in beta-thalassaemia major patients (P < 0.01). The difference between the two groups was not statistically significant for all the other mutations. Five different genotypes were recorded among thalassaemia intermedia and 15 among beta-thalassaemia major patients. Twenty-five percent of the intermedia patients and 4.5% of the major patients had homozygosity for mild mutations (group I); 62.5% of the intermedia patients and 26.2% of the major patients had combinations of mild/severe mutations (group II). In addition, homozygosity or double heterozygosity for severe mutations (group III) was found in 12.5% of the intermedia patients and 69% of the major patients. Some genotypes were restricted to thalassaemia intermedia, including heterozygosity -87/IVS1-6 and IVS1-6/CD 6-A. It is essential to understand the distribution and frequency of the relevant mutations in each population where beta-thalassaemias exist. This is of particular importance for genotype-phenotype correlation and for carrier detection, genetic counselling and prenatal diagnosis.

Monocyte and neutrophil activity after minor surgical stress.

BACKGROUND/PURPOSE: Surgical stress produces changes in the immune status of patients. In adults, major surgery causes immunosuppression, whereas minor operations stimulate immune responses. In children, the immunologic response to surgery has not been elucidated completely. The authors investigated the effects of minor surgery on immune response by analyzing neutrophil and monocyte phagocytosis and oxidative burst activity. METHODS: Sixteen children undergoing elective minor surgery were enrolled. Blood samples were collected before the operation (at time of induction of anesthesia), at the end of operation, and 72 hours after surgery. Neutrophil and monocyte phagocytosis and oxidative burst activity were studied using a flow cytometric method. RESULTS: Phagocytosis and oxidative burst increased significantly at the end of the operation, both in neutrophils (7.4% and 14.3%, respectively) and monocytes (11.6% and 27%, respectively). The increase was only significant for monocytes (17.5%) 72 hours after surgery. White cell count did not show any significant changes. There was no significant correlation between phagocytosis, oxidative burst activity, and white cell count or neutrophil and monocyte count. CONCLUSIONS: This study shows that minor surgery in children induces immune activation by increasing neutrophil and monocyte phagocytosis and oxidative burst activity. Further studies are required to understand the molecular basis of these findings.

Serum levels of malondialdehyde and 4-hydroxy-2,3-nonenal in patients affected by familial chronic nail candidiasis.

Familial chronic nail candidiasis (FCNC.MIM 607644) is a rare disorder characterized by early onset infections caused by different species of Candida and restricted to the nails; this disorder is genetically associated with low serum concentration of intercellular adhesion molecule 1 (ICAM-1). Herein we report the evidence of high circulating levels of malondialdehyde (MDA) and 4-hydroxy-2,3-nonenal (HNE) in seven patients of a five-generation Italian family affected by FCNC.MIM 607644. The present data evidence, in these patients, an increase in circulating MDA and HNE levels. Only some merely speculative hypotheses may be suggested to explain the mechanisms subserving the oxidative stress condition observed in these genetically ICAM-1 deficient patients; however, one has to point out that a chronic oxidative stress condition could contribute to the development of concurrent pathological alterations in which an overproduction of free radicals may play a central role.

Protein carbonyl group content in patients affected by familiar chronic nail candidiasis.

Familiar chronic nail candidiasis (FCNC) is a rare disorder characterized by early-onset infections caused by different species of Candida, restricted to the nail of the hands and feet, and associated with a low serum concentration of intercellular adhesion molecule 1. Host defense mechanisms against candidiasis require the cooperation of many immune cells through several candidacidal mechanisms, including oxygen-dependent killing mechanisms, mediated by a superoxide anion radical myeloperoxidase--H2O2--halide system, and reactive nitrogen intermediates. We analyzed protein carbonyl groups (considered a useful marker of oxidative stress) in the serum of patients belonging to a five-generation Italian family with an isolated form of FCNC. Serum protein carbonyl groups in FCNC patients were significantly lower than those measured in healthy donors. Also, if this hypothesis is merely speculative, we could suggest that the decreased circulating level of protein carbonyl groups in these patients is not a marker of a lower oxidative stress condition, but might be linked to a lower protease activity.