Genetic aspects of adolescent idiopathic scoliosis in a family with multiple affected members: a research article

Abstract Background The etiology of idiopathic scoliosis remains unknown and different factors have been suggested as causal. Hereditary factors can also determine the etiology of the disease; however, the pattern of inheritance remains unknown. Autosomal dominant, X-linked and multifactorial patterns of inheritances have been reported. Other studies have suggested possible chromosome regions related to the etiology of idiopathic scoliosis. We report the genetic aspects of and investigate chromosome regions for adolescent idiopathic scoliosis in a Brazilian family. Methods Evaluation of 57 family members, distributed over 4 generations of a Brazilian family, with 9 carriers of adolescent idiopathic scoliosis. The proband presented a scoliotic curve of 75 degrees, as determined by the Cobb method. Genomic DNA from family members was genotyped. Results Locating a chromosome region linked to adolescent idiopathic scoliosis was not possible in the family studied. Conclusion While it was not possible to determine a chromosome region responsible for adolescent idiopathic scoliosis by investigation of genetic linkage using microsatellites markers during analysis of four generations of a Brazilian family with multiple affected members, analysis including other types of genomic variations, like single nucleotide polymorphisms (SNPs) could contribute to the continuity of this study.

Patient with toxoplasmosis and glucose-6-phosphate dehydrogenase deficiency: a case report

Abstract Introduction Toxoplasmosis, a zoonotic protozoal disease caused by toxoplasma gondii, is prevalent throughout the world, affecting a large proportion of persons who usually have no symptoms. Glucose 6 phosphate dehydrogenase deficiency, an X-linked inherited disorder, is present in over 400 million people world wide. It is more common in tropical and subtropical countries and is one of the important causes of hemolytic anemia. Case presentation This case report relates the occurrence of the two diseases simultaneously in a child of five years old. Conclusion Patients with glucose-6-phosphate dehydrogenase deficiency are more susceptible to toxoplasmosis and this case report, reinforce the findings of this propensity and alert us for such possibility, what it is important, therefore, the treatment of toxoplasmosis can cause serious hemolysis in these patients.

Morbus Hunter : Neutralisationskapazität von Antikörpern unter Enzymersatztherapie und deren klinischen Auswirkungen

Morbus Hunter, eine lysosomale Speicherkrankheit, ist eine seltene, progrediente, x-chromosomal vererbte Stoffwechselkrankheit, die durch ein Defizit an Iduronat-2-sulfatase (IDS) hervorgerufen wird. Als Folge daraus erfolgt kein Abbau von Heparan- und Dermatansulfat und die Glykosaminoglykane reichern sich in de Lysosomen der Zelle an. M. Hunter ist eine Multisystemerkrankung und weist ein breites klinisches Spektrum mit interindividuell unterschiedlichem Krankheitsbeginn, Ausprägungen und Progression der Symptome auf. Seit 2007 besteht die Therapieoption einer Enzymersatztherapie (ERT) mit Elaprase®. Einige Patienten entwickeln Antikörper gegen das substituierte Enzym, welche partiell neutralisierende Eigenschaften besitzen. Ziel dieser Untersuchung war es zu klären, ob die Neutralisationskapazität der gebildeten Antikörper mittels einer Bestimmung im Mischserum festgestellt werden kann und ob persistierende Antikörper mit Neutralisationskapazität zu einer Einschränkung der Wirksamkeit der Enzymersatztherapie führen. Es sollte weiterhin untersucht werden, ob sich mittels Messung der neuronenspezifischen Enolase (NSE) und S-100 Rückschlüsse auf eine neuropathische Beteiligung ziehen lassen, da bis jetzt noch keine klinische oder biochemische Messmethode existiert, die für M. Hunter-Patienten eine verlässliche Vorhersage für eine neuropathische Beteiligung bietet. 30 Patienten wurden in die retrospektive/prospektive Kohortenstudie eingeschlossen. Bei der Bestimmung der IDS-Aktivität im Mischserum mit einem gesunden Menschen zeigten fünf der Patienten (17%) in zwölf Mischseren eine um ≥ 40% reduzierte Aktivität. Zwei (7%) der 30 untersuchten Patienten wurden mit dieser Methode als positiv für persistierende neutralisierende Antikörper identifiziert. Zum gleichen Ergebnis bezüglich der persistierenden neutralisierenden Antikörper führten die Anti-Elaprase®-Immunglobulin-Bestimmungen unter Berücksichtigung des Bestimmungszeitpunkts, die bei Shire Pharmaceuticals durchgeführt wurden. Die Untersuchungsergebnisse lassen den Schluss zu, dass die gebildeten Antikörper auch intraindividuell unterschiedlich sind. Zudem interagieren sie mit den verschiedensten Epitopen des Enzyms der ERT und besitzen nicht alle neutralisierende Eigenschaften. Aufgrund der heterogenen Zusammensetzung folgt die Hemmung der Enzymaktivität vermutlich keiner eindeutigen Kinetik. Anti-Elaprase®-Immunglobulin G spielt für die Neutralisationskapazität jedoch eine wichtige Rolle. Die Auswertung und Beurteilung der Einschränkung der Wirksamkeit der Therapie hervorgerufen durch die Antikörper mit Neutralisationskapazität gestaltete sich kompliziert. Im Ergebnis zeigte sich, dass sich die beiden Patienten mit persistierenden neutralisierenden Antikörpern in der Entwicklung der klinischen Parameter interindividuell stark unterschieden. Um einen Zusammenhang zwischen klinischem Verlauf und Antikörperbildung gegen die ERT zu finden, müssen in einem größeren Patientenkollektiv mehr Patienten mit persistierenden neutralisierenden Antikörpern identifiziert werden und der Einfluss der Antikörper untersucht werden. Die Untersuchung der NSE und S-100 ergab, dass weder die Konzentration der NSE noch der S-100 Rückschlüsse auf die neuropathische Beteiligung des Patienten zulässt.

Cross-sectional observational study of 208 patients with non-classical urea cycle disorders

Urea cycle disorders (UCDs) are inherited disorders of ammonia detoxification often regarded as mainly of relevance to pediatricians. Based on an increasing number of case studies it has become obvious that a significant number of UCD patients are affected by their disease in a non-classical way: presenting outside the newborn period, following a mild course, presenting with unusual clinical features, or asymptomatic patients with only biochemical signs of a UCD. These patients are surviving into adolescence and adulthood, rendering this group of diseases clinically relevant to adult physicians as well as pediatricians. In preparation for an international workshop we collected data on all patients with non-classical UCDs treated by the participants in 20 European metabolic centres. Information was collected on a cohort of 208 patients 50% of which were ≥ 16 years old. The largest subgroup (121 patients) had X-linked ornithine transcarbamylase deficiency (OTCD) of whom 83 were female and 29% of these were asymptomatic. In index patients, there was a mean delay from first symptoms to diagnosis of 1.6 years. Cognitive impairment was present in 36% of all patients including female OTCD patients (in 31%) and those 41 patients identified presymptomatically following positive newborn screening (in 12%). In conclusion, UCD patients with non-classical clinical presentations require the interest and care of adult physicians and have a high risk of neurological complications. To improve the outcome of UCDs, a greater awareness by health professionals of the importance of hyperammonemia and UCDs, and ultimately avoidance of the still long delay to correctly diagnose the patients, is crucial.

DNAI1 mutations explain only 2% of primary ciliary dykinesia

BACKGROUND: Primary ciliary dyskinesia (PCD) is a rare recessive hereditary disorder characterized by dysmotility to immotility of ciliated and flagellated structures. Its main symptoms are respiratory, caused by defective ciliary beating in the epithelium of the upper airways (nose, bronchi and paranasal sinuses). Impairing the drainage of inhaled microorganisms and particles leads to recurrent infections and pulmonary complications. To date, 5 genes encoding 3 dynein protein arm subunits (DNAI1, DNAH5 and DNAH11), the kinase TXNDC3 and the X-linked RPGR have been found to be mutated in PCD. OBJECTIVES: We proposed to determine the impact of the DNAI1 gene on a cohort of unrelated PCD patients (n = 104) recruited without any phenotypic preselection. METHODS: We used denaturing high-performance liquid chromatography and sequencing to screen for mutations in the coding and splicing site sequences of the gene DNAI1. RESULTS: Three mutations were identified: a novel missense variant (p.Glu174Lys) was found in 1 patient and 2 previously reported variants were identified (p.Trp568Ser in 1 patient and IVS1+2_3insT in 3 patients). Overall, mutations on both alleles of gene DNAI1 were identified in only 2% of our clinically heterogeneous cohort of patients. CONCLUSION: We conclude that DNAI1 gene mutation is not a common cause of PCD, and that major or several additional disease gene(s) still remain to be identified before a sensitive molecular diagnostic test can be developed for PCD.

Periodontal disease progression in subjects with orofacial clefts over a 25-year follow-up period

AIMS: To assess rates of periodontal disease progression in subjects with cleft lip, alveolus and palate (CLAP) over a 25-year period without regular maintenance care in a specialist setting and to compare those with those of subjects without alveolar clefts, i.e. cleft lip (CL) or cleft palate (CP). MATERIAL AND METHODS: Ten subjects with CLAP and 10 subjects with CL/CP were examined in 1979, 1987, 1993 and 2004. Probing pocket depth (PPD), clinical attachment level (CAL), bleeding on probing (BoP) and plaque control record (PCR) scores were recorded in all 20 subjects. RESULTS: High plaque and BoP scores were recorded at all examinations in both groups. Over 25 years, a statistically significant loss of mean full-mouth CAL of 1.52 +/- 0.12 mm (SD) and 1.66 +/- 0.15 mm occurred in the CLAP and CL/CP group respectively (p<0.05). A statistically significant increase (p<0.05) in mean full-mouth PPD of 0.35 +/- 0.12 mm was observed in the CL/CP group, whereas only a trend for a mean full-mouth increase in PPD of 0.09 +/- 0.11 mm was observed in the CLAP group. In subjects with CLAP, a statistically significant increase (p<0.05) in PPD of 0.92 +/- 1.13 mm at cleft sites was observed compared with that of 0.17 +/- 0.76 mm at control sites. With respect to CAL, the loss at the corresponding sites amounted to 2.71 +/- 1.46 and to 2.27 +/- 1.62 mm, respectively (p=0.36). CONCLUSIONS: When stringent and well-defined supportive periodontal therapy was not provided, subjects with orofacial clefts were at high risk for periodontal disease progression. Over 25 years, alveolar cleft sites tended to have more periodontal tissue destruction compared with control sites.

The story of RP10: A positional cloning approach to the identification of an autosomal dominant retinitis pigmentosa gene

Retinitis pigmentosa (RP) is an inherited retinal degenerative disease that is the leading cause of inherited blindness worldwide. Characteristic features of the disease include night blindness, progressive loss of visual fields, and deposition of pigment on the retina in a bone spicule-like pattern. RP is marked by extreme genetic heterogeneity with at least 19 autosomal dominant, autosomal recessive and X-linked loci identified. RP10, which maps to chromosome 7q, was the fifth autosomal dominant RP locus identified, and accounts for the early-onset disease in two independent families. Extensive linkage and haplotype analyses have been performed in these two families which have allowed the assignment of the disease locus to a 5-cM region on chromosome 7q31.3. In collaboration with Dr. Eric Green (National Center for Human Genome Research, National Institutes of Health), a well-characterized physical map of the region was constructed which includes YAC, BAC and cosmid coverage. The entire RP10 critical region resides within a 9-Mb well-characterized YAC contig. These physical maps not only provided the resources to undertake the CAIGES (cDNA amplification for identification of genomic expressed sequences) procedure for identification of retinal candidate genes within the critical region, but also identified a number of candidate genes, including transducin-$\gamma$ and blue cone pigment genes. All candidate genes examined were excluded. In addition, a number of ESTs were mapped within the critical region. EST20241, which was isolated from an eye library, corresponded to the 3$\sp\prime$ region of the ADP-ribosylation factor (ARF) 5 gene. ARF5, with its role in vesicle transport and possible participation in the regulation of the visual transduction pathway, became an extremely interesting candidate gene. Using a primer walking approach, the entire 3.2 kb genomic sequence of the ARF5 gene was generated and developed intronic primers to screen for coding region mutations in affected family members. No mutations were found in the ARF5 gene, however, a number of additional ESTs have been mapped to the critical region, and, as the large-scale sequencing projects get underway, megabases of raw sequence data from the RP10 region are becoming available. These resources will hasten the isolation and characterization of the RP10 gene. ^

Esx1 is an X-chromosome-imprinted regulator of placental formation and fetal growth

Placental formation and genomic imprinting are two important features of embryonic development in placental mammals. Genetic studies have demonstrated that imprinted genes play a prominent role in regulating placental formation. In marsupials, mice and humans, the paternally derived X chromosome is preferentially inactivated in the placental tissues of female embryos. This special form of genomic imprinting may have evolved under the same selective forces as autosomal imprinted genes. This chromosomal imprinting phenomenon predicts the existence of maternally expressed X-linked genes that regulate placental development.^ In this study, an X-linked homeobox gene, designated Esx1 has been isolated. During embryogenesis, Esx1 was expressed in a subset of placental tissues and regulates formation of the chorioallantoic placenta. Esx1 acted as an imprinted gene. Heterozygous female mice that inherit an Esx1-null allele from their father developed normally. However, heterozygous females that inherit the Esx1 mutation from their mother were born 20% smaller than normal and had an identical phenotype to hemizygous mutant males and homozygous mutant females. Surprisingly, although Esx1 mutant embryos were initially comparable in size to wild-type controls at 13.5 days post coitum (E13.5) their placentas were significantly larger (51% heavier than controls). Defects in the morphogenesis of the labyrinthine layer were observed as early as E11.5. Subsequently, vascularization abnormalities developed at the maternal-fetal interface, causing fetal growth retardation. These results identify Esx1 as the first essential X-chromosome-imprinted regulator of placental development that influences fetal growth and may have important implications in understanding human placental insufficiency syndromes such as intrauterine growth retardation (IUGR). ^

Differential spatial and structural organization of the X chromosome underlies dosage compensation in C. elegans.

The adjustment of X-linked gene expression to the X chromosome copy number (dosage compensation [DC]) has been widely studied as a model of chromosome-wide gene regulation. In Caenorhabditis elegans, DC is achieved by twofold down-regulation of gene expression from both Xs in hermaphrodites. We show that in males, the single X chromosome interacts with nuclear pore proteins, while in hermaphrodites, the DC complex (DCC) impairs this interaction and alters X localization. Our results put forward a structural model of DC in which X-specific sequences locate the X chromosome in transcriptionally active domains in males, while the DCC prevents this in hermaphrodites.

Positional candidate cloning of the RP10 form of retinitis pigmentosa

Retinitis pigmentosa (RP) is a genetically heterogeneous group of retinal degenerations that affects over one million people worldwide. To date, 11 autosomal dominant, 13 autosomal recessive, and 5 X-linked forms of retinitis pigmentosa have been identified through linkage analysis, but the disease-causing genes and mutations have been found for only half of these loci. My research uses a positional candidate cloning approach to identify the gene and mutations responsible for one type of autosomal dominant retinitis pigmentosa, RP10. The premise is that identifying the genes and mutations responsible for disease will provide insight into disease mechanisms and provide treatment options. Previous research mapped the RP10 locus to a 5cM region on chromosome 7q31 between markers D7S686 and D7S530. Linkage and fine-point haplotype analysis was used to reduce and refine the RP10 disease interval to a 4cM region located between D7S2471 and a new marker located 45,000bp telomeric of D7S461. In order to identify genes located in the RP10 interval, an extensive EST map was created of this region. Five EST clusters from this map were analyzed to determine if mutations in these genes cause the RP10 form of retinitis pigmentosa. The genomic structure of a known metabotrophic glutamate receptor, GRMS8, was determined first. DNA sequencing of GRM8 in RP10 family members did not identify any disease-causing mutations. Four other EST clusters (A170, A173, A189, and A258) were characterized and determined to be part of the same gene, UBNL1 (ubinuclein-like 1). The full-length mRNA sequence and genomic structure of UBNL1 was determined and then screened in patients. No disease-causing mutations were identified in any of the RP10 family members tested. Recent data made available with the release of the public and Celera genome assemblies indicates that UBNL1 is outside of the RP10 disease region. Despite this complication, characterization of UBNL1 is still important in the understanding of normal visual processes and it is possible that mutations in UBNL1 could cause other forms of retinopathy. The EST map and list of RP10 candidates will continue to aid others in the search for the RP10 gene and mutations. ^

Resistance to apoptosis caused by PIG-A gene mutations in paroxysmal nocturnal hemoglobinuria

Paroxysmal nocturnal hemoglobinuria (PNH) is a clonal hematopoietic stem cell disorder resulting from mutations in an X-linked gene, PIG-A, that encodes an enzyme required for the first step in the biosynthesis of glycosylphosphatidylinositol (GPI) anchors. PIG-A mutations result in absent or decreased cell surface expression of all GPI-anchored proteins. Although many of the clinical manifestations (e.g., hemolytic anemia) of the disease can be explained by a deficiency of GPI-anchored complement regulatory proteins such as CD59 and CD55, it is unclear why the PNH clone dominates hematopoiesis and why it is prone to evolve into acute leukemia. We found that PIG-A mutations confer a survival advantage by making cells relatively resistant to apoptotic death. When placed in serum-free medium, granulocytes and affected CD34+ (CD59−) cells from PNH patients survived longer than their normal counterparts. PNH cells were also relatively resistant to apoptosis induced by ionizing irradiation. Replacement of the normal PIG-A gene in PNH cell lines reversed the cellular resistance to apoptosis. Inhibited apoptosis resulting from PIG-A mutations appears to be the principle mechanism by which PNH cells maintain a growth advantage over normal progenitors and could play a role in the propensity of this disease to transform into more aggressive hematologic disorders. These data also suggest that GPI anchors are important in regulating apoptosis.

Evidence of evolutionary up-regulation of the single active X chromosome in mammals based on Clc4 expression levels in Mus spretus and Mus musculus

Previous studies have shown that the chloride channel gene Clc4 is X-linked and subject to X inactivation in Mus spretus, but that the same gene is autosomal in laboratory strains of mice. This exception to the conservation of linkage of the X chromosome in one of two interfertile mouse species was exploited to compare expression of Clc4 from the X chromosome to that from the autosome. Clc4 was found to be highly expressed in brain tissues of both mouse species. Quantitative analyses of species-specific expression of Clc4 in brain tissues from mice resulting from M. spretus × laboratory strain crosses, demonstrate that each autosomal locus has half the level of Clc4 expression as compared with the single active X-linked locus. In contrast expression of another chloride channel gene, Clc3, which is autosomal in both mouse species is equal between alleles in F1 animals. There is no evidence of imprinting of the Clc4 autosomal locus. These results are consistent with Ohno’s hypothesis of an evolutionary requirement for a higher expression of genes on the single active X chromosome to maintain balance with autosomal gene expression [Ohno, S. (1967) Sex Chromosomes and Sex-Linked Genes (Springer, Berlin)].

Increased levels of plasma lysosomal enzymes in patients with Lowe syndrome

Lowe syndrome is an X-linked disorder that has a complex phenotype that includes progressive renal failure and blindness. The disease is caused by mutations in an inositol polyphosphate 5-phosphatase designated OCRL. It has been shown that the OCRL protein is found on the surface of lysosomes and that a renal tubular cell line deficient in OCRL accumulated substrate phosphatidylinositol 4,5-bisphosphate. Because this lipid is required for vesicle trafficking from lysosomes, we postulate that there is a defect in lysosomal enzyme trafficking in patients with Lowe syndrome that leads to increased extracellular lysosomal enzymes and might lead to tissue damage and contribute to the pathogenesis of the disease. We have measured seven lysosomal enzymes in the plasma of 15 patients with Lowe syndrome and 15 age-matched male controls. We find a 1.6- to 2.0-fold increase in all of the enzymes measured. When the data was analyzed by quintiles of activity for all of the enzymes, we found that 95% of values in the lowest quintile come from normal subjects whereas in the highest quintile 85% of the values are from patients with Lowe syndrome. The increased enzyme levels are not attributable to renal insufficiency because there was no difference in enzyme activity in the four patients with the highest creatinine levels compared with the six patients with the lowest creatinine values.

A first-generation X-inactivation profile of the human X chromosome

In females, most genes on the X chromosome are generally assumed to be transcriptionally silenced on the inactive X as a result of X inactivation. However, particularly in humans, an increasing number of genes are known to “escape” X inactivation and are expressed from both the active (Xa) and inactive (Xi) X chromosomes; such genes reflect different molecular and epigenetic responses to X inactivation and are candidates for phenotypes associated with X aneuploidy. To identify genes that escape X inactivation and to generate a first-generation X-inactivation profile of the X, we have evaluated the expression of 224 X-linked genes and expressed sequence tags by reverse-transcription–PCR analysis of a panel of multiple independent mouse/human somatic cell hybrids containing a normal human Xi but no Xa. The resulting survey yields an initial X-inactivation profile that is estimated to represent ≈10% of all X-linked transcripts. Of the 224 transcripts tested here, 34 (three of which are pseudoautosomal) were expressed in as many as nine Xi hybrids and thus appear to escape inactivation. The genes that escape inactivation are distributed nonrandomly along the X; 31 of 34 such transcripts map to Xp, implying that the two arms of the X are epigenetically and/or evolutionarily distinct and suggesting that genetic imbalance of Xp may be more severe clinically than imbalance of Xq. A complete X-inactivation profile will provide information relevant to clinical genetics and genetic counseling and should yield insight into the genomic and epigenetic organization of the X chromosome.

Direct interaction of the Wiskott-Aldrich syndrome protein with the GTPase Cdc42.

Wiskott-Aldrich syndrome (WAS) is an X-linked immunodeficiency disorder with the most severe pathology in the T lymphocytes and platelets. The disease arises from mutations in the gene encoding the WAS protein. T lymphocytes of affected males with WAS exhibit a severe disturbance of the actin cytoskeleton, suggesting that the WAS protein could regulate its organization. We show here that WAS protein interacts with a member of the Rho family of GTPases, Cdc42. This interaction, which is guanosine 5'-triphosphate (GTP)-dependent, was detected in cell lysates, in transient transfections and with purified recombinant proteins. A weaker interaction was also detected with Rac1 using WAS protein from cell lysates. It was also found that different mutant WAS proteins from three affected males retained their ability to interact with Cdc42 and that the level of expression of the WAS protein in these mutants was only 2-5% of normal. Taken together these data suggest that the WAS protein might function as a signal transduction adaptor downstream of Cdc42, and in affected males, the cytoskeletal abnormalities may result from a defect in Cdc42 signaling.