Novel spondyloepimetaphyseal dysplasia due to UFSP2 gene mutation.

Beukes hip dysplasia is an autosomal dominant disease which has to date been described only in a large South African family of Dutch origin. The patients presented with progressive epiphyseal dysplasia limited to femoral capital epiphysis and their height was not significantly reduced. A unique variant of the ubiquitin-fold modifier 1 (Ufm1)-specific peptidase 2 (UFSP2) gene (c.868T>C) has been reported in all individuals from Beukes family with clinical and radiological diagnosis of Beukes hip dysplasia. Three individuals, propositus, mother, and grandmother, presented with short stature, joint pain, genu vara and a novel spondyloepimetaphyseal dysplasia involving epiphyses predominantly at hips, but also at knees, ankles, wrists and hands, associated with variable degrees of metaphysis and spine involvement. Exome sequencing allowed us to identify the heterozygous variant c.1277A>C of the UFSP2 gene, leading to the missense change p.D426A, in all 3 patients. This mutation is predicted as damaging and, similarly to the mutation originally described in the Beukes family (p. Y290H), directly affects one of the catalytic residues participating in the active site of the protein. This supports the novel notion that loss of catalytic UFSP2 activity, observed in association with different mutants and already experimentally proven in vitro, may have different clinical outcomes.

Clinical presentation and pathogenesis of cold-induced autoinflammatory disease in a family with recurrence of an NLRP12 mutation.

OBJECTIVE: NLRP12 mutations have been described in patients affected with peculiar autoinflammatory symptoms. This study was undertaken to characterize NLRP12 mutations in patients with autoinflammatory syndromes, particularly a novel missense mutation, p.D294E, affecting a protein sequence crucial for ATP binding, which was identified in a Caucasian family with familial cold-induced autoinflammatory syndrome in some family members. METHODS: Fifty patients were tested for NLRP12 mutations. A Caucasian family with the p.D294E missense mutation of NLRP12 in some family members was clinically characterized. In vitro analysis of the effects of the mutation on NF-κB activity was performed in HEK 293 cells after cotransfection of the cells with a luciferase NF-κB-responsive element and mutant or wild-type (WT) NLRP12 expression plasmids. NF-κB activity was also evaluated 24 hours after stimulation with tumor necrosis factor α in monocytes from individual family members carrying the mutation. Furthermore, secretion of interleukin-1ß (IL-1ß), production of reactive oxygen species (ROS), and activation of antioxidant systems in patient and healthy donor monocytes, under resting conditions and after stimulation with pathogen-associated molecular patterns (PAMPs), were also assessed. RESULTS: In the family assessed, the p.D294E mutation segregated in association with a particular sensitivity to cold exposure (especially arthralgias and myalgia), but not always with an inflammatory phenotype (e.g., urticarial rash or fever). In vitro, the mutant protein maintained the same inhibitory activity as that shown by WT NLRP12. Consistently, NLRP12-mutated monocytes showed neither increased levels of p65-induced NF-κB activity nor higher secretion of IL-1ß. However, the kinetics of PAMP-induced IL-1ß secretion were significantly accelerated, and high production of ROS and up-regulation of antioxidant systems were demonstrated. CONCLUSION: Even with a variable range of associated manifestations, the extreme sensitivity to cold represents the main clinical hallmark in an individual carrying the p.D294E mutation of the NLRP12 gene. Although regulation of NF-κB activity is not affected in patients, redox alterations and accelerated secretion of IL-1ß are associated with this mild autoinflammatory phenotype.

Long-term clinical profile of children with the low-penetrance R92Q mutation of the TNFRSF1A gene.

OBJECTIVE: To analyze the long-term impact of the R92Q mutation of TNFRSF1A in children with periodic fever, in comparison with children with tumor necrosis factor receptor-associated periodic syndrome (TRAPS) with TNFRSF1A structural mutations and children with periodic fever of unknown origin fulfilling the criteria for periodic fever, aphthosis, pharyngitis, and adenitis syndrome (PFAPA). METHODS: The extracellular region of TNFRSF1A was analyzed in 720 consecutive children with periodic fever, using denaturing high-performance liquid chromatography and DNA sequencing. Followup data on 11 pediatric patients with TNFRSF1A structural mutations (cysteine or T50M), 23 pediatric patients with an R92Q substitution, and 64 pediatric patients with PFAPA were collected during routine clinic visits. The 50-item Child Health Questionnaire was used to assess health-related quality of life (HRQOL). RESULTS: The frequency of typical TRAPS-related clinical manifestations was significantly lower and the impact of the disease on HRQOL was significantly reduced in patients with the R92Q mutation compared with TRAPS patients carrying structural mutations of TNFRSF1A. Followup data on 11 TRAPS patients with TNFRSF1A structural mutations (mean followup 7.9 years), 16 patients with theR92Q substitution (mean followup 7.3 years), and 64 patients with PFAPA (mean followup 5.2 years) were available. Patients with R92Q mutations and patients with PFAPA displayed a higher rate of self-resolution or amelioration of the fever episodes than did TRAPS patients with structural mutations. CONCLUSION: Although some cases may progress to a more chronic disease course, the majority of children with an R92Q mutation of the TNFRSFA1 gene show a milder disease course than that in children with TNFRSFA1 structural mutations and have a high rate of spontaneous resolution and amelioration of the recurrent fever episodes.

Major surface protein complex of Treponema denticola induces the production of tumor necrosis factor alpha, interleukin-1 beta, interleukin-6 and matrix metalloproteinase 9 by primary human peripheral blood monocytes.

BACKGROUND AND OBJECTIVE: Treponema denticola is a micro-organism that is involved in the pathogenesis of periodontitis. Major surface protein complex (MSPc), which is expressed on the envelope of this treponeme, plays a key role in the interaction between T. denticola and gingival cells. The peptidoglycan extracted from T. denticola induces the production of a large variety of inflammatory mediators by macrophage-like cells, suggesting that individual components of T. denticola cells induce the inflammatory response during periodontal disease. This study was designed to demonstrate that MSPc of T. denticola stimulates release of proinflammatory mediators in primary human monocytes. MATERIAL AND METHODS: Primary human monocytes were separated from the blood of healthy donors and incubated for up to 24 h with varying concentrations of MSPc. The production of tumor necrosis factor alpha (TNF-alpha), interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6) and matrix metalloproteinase 9 (MMP-9) was measured at different time points with commercially available enzyme-linked immunosorbent assays. RESULTS: T. denticola MSPc induced the synthesis of TNF-alpha, IL-1 beta, IL-6 and MMP-9 in a dose- and time-dependent manner. Similar patterns of TNF-alpha, IL-1 beta and IL-6 release were observed when cells were stimulated with 100 and 1000 ng/mL of MSPc. The production of MMP-9 was significant only when cells were treated with 1000 ng/mL of MSPc. CONCLUSION: These results indicate that T. denticola MSPc, at concentrations ranging from 100 ng/mL to 1.0 microg/mL, activates a proinflammatory response in primary human monocytes.

[Persons suffering from schizophrenia and relapses]. / Les personnes atteintes de schizophrénie et la rechute.

INTRODUCTION: In schizophrenia, relapse is a common event that affects more than half the patients within 2 years after a first episode. It is a real setback for them and their relatives. Surprisingly, we do not have much information on how patients and their relatives experience the relapse. METHOD: A national survey was conducted among 316 schizophrenic outpatients treated with antipsychotics, and 82 of their relatives. The survey assessed the following four aspects: disease history, last relapse history, hospitalization experiences, and relapse prevention. RESULTS: Regarding the disease history, the average psychiatric follow-up was 13 years and patients had been hospitalized five times on average. Relatives reported approximately the same history. Regarding the last relapse, 9/10 of relatives reported that this relapse led to hospitalization and 69% of patients understood that their hospitalizations were due to relapse. 4% of patients and 7% of relatives identified the end of the treatment as a precursor to relapse. While a lack of compliance was found in about four relapses out of 10. It has also been shown that patients confided primarily in the medical team and the relatives thought to be the first confidant of patients. Regarding the experience of hospitalization, 87% of patients and 86% of relatives judged the hospitalization useful. For both, hospitalization represented a solving step to manifestations of relapse. Regarding the relapse prevention, almost three patients out of four thought they knew what to do in order to avoid a new relapse, while only 52% of the relatives thought patients knew what to do for this matter. For more than one third of the patients, the last relapse (3 years ago) was still a painful event. Avoiding a new relapse was considered very important or important by 91% of patients and 100% of relatives. Relatives felt that regular appointments with the medical team helped avoid relapses. Fifty-nine per cent of relatives have said it was difficult to verify whether or not the treatment was taken by a schizophrenic patient. Relatives' opinion on the injectable treatment was favorable and approximately 50% of the patients declared knowing of injectable treatments. Among these 72% felt that such treatment was reassuring, 69% said it was simpler than oral therapy, and 67% thought it was the most suitable to check the compliance. Only 31% considered it restricting for the patient, against 54% who were considering it not restricting. Finally 57% of patients were willing to take an injectable treatment in order to prevent further hospitalization. CONCLUSION: This study brings us a better understanding of patients' and relatives' experience of relapse. These results demonstrate the potential impact of relapse on the patients and their relatives and highlight their motivation to avoid further relapses. Also revealed, the lack of importance given to the link between compliance and relapse by patients and relatives. These results underscore the complexity of this disease management in which each player has a key role.

A diagnostic score for molecular analysis of hereditary autoinflammatory syndromes with periodic fever in children.

OBJECTIVE: To identify a set of clinical parameters that can predict the probability of carrying mutations in one of the genes associated with hereditary autoinflammatory syndromes. METHODS: A total of 228 consecutive patients with a clinical history of periodic fever were screened for mutations in the MVK, TNFRSF1A, and MEFV genes, and detailed clinical information was collected. A diagnostic score was formulated based on univariate and multivariate analyses in genetically positive and negative patients (training set). The diagnostic score was validated in an independent set of 77 patients (validation set). RESULTS: Young age at onset (odds ratio [OR] 0.94, P = 0.003), positive family history of periodic fever (OR 4.1, P = 0.039), thoracic pain (OR 4.6, P = 0.05), abdominal pain (OR 33.1, P < 0.001), diarrhea (OR 3.3, P = 0.028), and oral aphthosis (OR 0.2, P = 0.007) were found to be independently correlated with a positive genetic test result. These variables were combined in a linear score whose ability to predict a positive result on genetic testing was validated in an independent data set. In this latter set, the diagnostic score revealed high sensitivity (82%) and specificity (72%) for discriminating patients who were genetically positive from those who were negative. In patients with a high probability of having a positive result on genetic testing, a regression tree analysis provided the most reasonable order in which the genes should be screened. CONCLUSION: The proposed approach in patients with periodic fever will increase the probability of obtaining positive results on genetic testing, with good specificity and sensitivity. Our results further help to optimize the molecular analysis by suggesting the order in which the genes should be screened.

Uncommon association of germline mutations of RET proto-oncogene and CDKN2A gene.

INTRODUCTION: Calcitonin measurement is advised in the diagnosis of thyroid nodules, as it is an accurate marker of medullary thyroid carcinoma (MTC). C-cell hyperplasia (CCH)-induced hypercalcitoninemia cannot be distinguished from that induced by MTC, unless surgery is performed. CASE: We report the clinical and biological features of a patient with a family history of cancer, including melanoma and pancreatic cancer, who had previously undergone surgery for melanoma. He presented the unusual association of papillary thyroid carcinoma (PTC), normocalcemic hyperparathyroidism, and hypercalcitoninemia with a pathological response to pentagastrin, which was histologically deemed secondary to CCH. Multiple endocrine neoplasia (MEN) 2A was diagnosed. RET gene analysis showed a p.V804M missense mutation in exon 14, a low- but variably penetrant defect found in both sporadic and MEN2A-associated MTC/CCH, and a p.G691S polymorphism in exon 11. Furthermore, the germline P48T mutation was found in the CDKN2A gene exon 1, which is known to be associated with melanoma and pancreatic cancer. The patient showed the uncommon coexistence of a germline mutation in two suppressor genes, RET and CDKN2A; this finding, deemed to be a mere coincidence, did not modify the phenotype expected by each single mutation. CCH associated with V804M RET mutation is a precancerous condition and surgery is recommended. In order to exclude MTC, surgery is advised in patients with a pathological calcitonin response to pentagastrin, in the absence of thyroid autoimmunity. CCH-induced hypercalcitoninemia can be associated with thyroid cancers other than MTC (e.g., PTC). Family history is important in scheduling specific genetic screening in high-risk patients and their relatives.

Refractory thyrotoxicosis induced by iodinated contrast agents treated with therapeutic plasma exchange. A case report.

Excess free iodide in the blood (ingested or injected) may cause thyrotoxicosis in patients at risk. Iodinated contrast solutions contain small amounts of free iodide and may be of significance for patients affected by Graves' disease, multinodular goiter or living in areas of iodine deficiency. Herein, we report a 57 elderly woman with a clinical history of multinodular goiter presented with a thyrotoxicosis induced by an iodinate contrast agent used during computed tomography scan. Because of the patient's resistance to conventional antithyroid drugs, she was treated with therapeutic plasma exchange (TPE). TPE is used in the treatment of several immunologic and nonimmunologic disorders. Temporary improvement after TPE in cases with thyrotoxicosis has been reported. In our patient's case, we observed an improvement in the thyroid hormone laboratory values as well as clinical findings. TPE can be an addition treatment when standard therapies for thyrotoxicosis fail providing the clinician with an adjuvant tool for rapid preparation of such a patient for thyroidectomy surgery.

GFAP mutations and polymorphisms in 13 unrelated Italian patients affected by Alexander disease.

Alexander disease (AD), a rare neurodegenerative disorder of the central nervous system, is characterized by the accumulation of cytoplasmic protein aggregates (Rosenthal fibers) composed of glial fibrillary acidic protein (GFAP) and small heat-shock proteins within astrocytes. To date, more than 40 different GFAP mutations have been reported in AD. The present study is aimed at the molecular diagnosis of Italian patients suspected to be affected by AD. By analyzing the GFAP gene of 13 unrelated patients (eight with infantile form, two with juvenile form and three with adult form), we found 11 different alleles, including four new ones. Among the novel mutations, three (p.R70Q, p.R73K, and p.R79P) were identified in exon 1 and p.L359P in exon 6. The sequence analysis also detected six different single nucleotide polymorphic variants, including two previously unreported ones, spread throughout non-coding regions (introns 2, 3, 5, 6, and 3'UTR) of the gene. All patients were heterozygous for the mutations, thus confirming their dominant effect.

Clinical and genetic characterization of Italian patients affected by CINCA syndrome.

OBJECTIVE: We report the experience of the Italian Registry of patients affected by chronic infantile neurological, cutaneous, articular (CINCA) syndrome. The clinical and genetic features of 12 unrelated Italian patients with CINCA syndrome are described, focusing on the possible influence of the presence of CIAS1/cryopyrin mutations on the phenotype of the disease and on its prognosis. METHODS: The clinical features of 12 Italian CINCA patients were evaluated. Genomic DNA of the patients was sequenced using specific primers for CIAS1 and ASC genes. RESULTS: Our patients shared typical CINCA characteristics and, sometimes, remarkable perinatal events, peculiar of CIAS1-mutated patients. Seven patients carried CIAS1 missense mutation, localized within the nucleotide binding domain of cryopyrin. Four previously described mutations and three new heterozygous CIAS1 missense mutations were identified. ASC gene, encoding for a direct interactor of cryopyrin, was not mutated in Italian CINCA patients. Finally, we reported the efficacy and safety of anti-IL1 therapy (Anakinra) in seven patients with a particularly severe CINCA phenotype. CONCLUSION: Despite some common signs-used as syndrome hallmarks-we observed a high variability in symptoms, genetic results and outcomes in Italian CINCA patients. In contrast with other authors, we cannot find out any correlation between mutations in CIAS1 and CINCA severity, but we underlined the concomitance of perinatal events and mental retardation only in CIAS1 mutated subjects. Finally, we confirmed the efficacy of Anakinra treatment, both in CIAS1-mutated and non-mutated patients.

Preservation of clonal heterogeneity of the Pneumocystis carinii-specific CD4 T cell repertoire in HIV infected, asymptomatic individuals.

The loss of CD4 lymphocytes in HIV disease associates with opportunistic infections. Since diverse CD4 T cell clones respond to an opportunistic pathogen, we asked whether CD4 depletion deletes selected clones in the repertoire (vertical depletion) or it affects all clones by reducing the cell number in each progeny without affecting the overall number of clones (horizontal depletion). Understanding this point may help explain the mode of CD4 depletion and the mode of immunoreconstitution after therapy. Therefore we examined the CD4 T cell repertoire specific for Pneumocystis carinii, a relevant opportunistic pathogen in AIDS, in HIV-infected, asymptomatic individuals. We identified two patients of 36 asymptomatics for lack of proliferation to P. carinii, suggesting selective depletion of specific CD4 cells. To investigate clonal heterogeneity of P. carinii-responsive CD4 lymphocytes, specific CD4 T cell lines were generated and studied by TCR BV gene family usage and CDR3 length analysis (spectratyping). Clonal heterogeneity was similar in antigen-specific CD4 lines generated from P. carinii non-responding HIV seropositives and from controls. Thus, despite undetectable response to the pathogen, residual specific cells probably prevent overt infection and, when expanded in vitro, exhibit a clonal diversity similar to normal controls. These findings suggest a horizontal, rather than vertical, depletion in these asymptomatic patients.

Characterization of the response to myelin basic protein in a non human primate model for multiple sclerosis.

The common marmoset Callithrix jacchus (C. jacchus) is an outbred species characterized by a naturally occurring bone marrow chimerism and susceptibility to a form of experimental autoimmune encephalomyelitis (EAE) resembling multiple sclerosis (MS). T cell clones specific for the myelin antigen, myelin basic protein (MBP), can be derived from both naive and immunized marmosets and can adoptively transfer EAE to compatible chimeric siblings. Here, we demonstrate that several different antigenic determinants of MBP are recognized by these encephalitogenic T cell clones. Furthermore, PCR-based analysis of TCR Vbeta families does not show the preferential usage of any gene segment. Characterization of third complementarity determining regions (CDR3) fails to demonstrate a recurring motif characteristic of the T cell immune response to MBP in this species. Nevertheless, brief amino acid motifs are shared among marmoset clones and CDR3 sequences from MS samples. These data suggest that, due to its outbred condition, the C. jacchus marmoset mounts a diverse pathogenic response to MBP. However, the findings that certain CDR3 sequences are identically expressed in different animals, or by different T cell clones, suggest that MBP-specific T cell populations may be clonally expanded following chronic antigenic stimulation in vivo.

Dynamics of the reactivity to MBP in multiple sclerosis.

Though many lines of evidence support the importance of myelin basic protein (MBP) in the pathogenesis of experimental autoimmune encephalomyelitis (EAE), its role in multiple sclerosis (MS) is still debated as well as the significance of epitope spreading in disease progression. We characterised the response to MBP in eight MS subjects and three of these were followed over time. In one case, the follow up lasted over a 6-year period. Clonal expansion, clonal persistence and epitope spreading against other MBP determinants was detected irrespective of disease course. In one patient we identified a novel T-cell receptor variable gene (BV28S2) which may be involved in the selection of MBP determinants, as suggested by experiments performed in the presence of mismatched antigen presenting cells (APC) between two subjects compatible for HLA-DR2 subtype but differing for the epitope recognised. Our findings do not sustain a role for the response to MBP effecting on clinical course and suggest that a novel TCR gene may be involved in the recognition of unusual self antigens.

Congenital hypoplastic anaemia in a patient with a new multiple congenital anomalies-mental retardation syndrome.

We report on a girl with congenital hypoplastic anaemia, "coarse" face, generalized hypertrichosis with scalp hypotrichosis, short fifth finger, hypoplastic toenails, and mental retardation. A sister of the proposita, who died at the age of 1 year, had severe congenital anaemia, hypoplastic fingernails, low birth weight, failure to thrive, and repeated upper respiratory tract infections. Based on family history, we suspect that hypoplastic anaemia and the same multiple congenital anomalies-mental retardation syndrome (MCA/MR) were also present in this sister. To the best of our knowledge, this patient represents the first report of congenital hypoplastic anaemia and such a complex MCA/MR syndrome, probably inherited as an autosomal recessive trait.

Identification of LMX1B gene point mutations in italian patients affected with Nail-Patella syndrome.

Nail-Patella syndrome, or osteo-onychodysplasia, is an autosomal dominant disorder characterized by nail dysplasia, absent or hypoplastic patellae, iliac horns and nephropathy. Previous studies have demonstrated linkage of the Nail-Patella locus with polymorphic markers on human chromosome 9q34. Recently, point mutations in the LMX1B gene have been identified in Nail-Patella patients and in families with recurrence of Nail-Patella syndrome and open-angle glaucoma. We describe here the identification of additional point mutations in the LMX1B gene in a set of Italian patients affected with Nail-Patella syndrome: two deletions of 1 and 2 bp causing a frameshift in two sporadic patients and nonsense mutations in two familial and one sporadic cases have been identified. All the mutations affect the homeodomain of the LMX1B protein and could cause the Nail-Patella syndrome through a loss of function as well as a dominant negative effect. Haplotype analysis in the two familial cases carrying the same stop codon mutation suggests the presence of a founder effect. Finally, analysis of cDNA clones obtained from human fetal kidney has revealed the existence of two different transcripts of LMX1B gene likely due to an alternative splicing.