Galactose-1-phosphate uridyltransferase deficiency: A literature review of the putative mechanisms of short and long-term complications and allelic variants.

Galactosemia type 1 is an autosomal recessive disorder of galactose metabolism, determined by a deficiency in the enzyme galactose-1-phosphate uridyltransferase (GALT). GALT deficiency is classified as severe or variant depending on biochemical phenotype, genotype and potential to develop acute and long-term complications. Neonatal symptoms usually resolve after galactose-restricted diet; however, some patients, despite the diet, can develop long-term complications, in particular when the GALT enzyme activity results absent or severely decreased. The mechanisms of acute and long-term complications are still discussed and several hypotheses are presented in the literature like enzymatic inhibition, osmotic stress, endoplasmic reticulum stress, oxidative stress, defects of glycosylation or epigenetic modification. This review summarizes the current knowledge of galactosemia, in particular the putative mechanisms of neonatal and long-term complications and the molecular genetics of GALT deficiency.

Comparison of X-chromosome inactivation in Duchenne muscle/myocardium-manifesting carriers, non-manifesting carriers and related daughters.

Female carriers of Duchenne muscular dystrophy (DMD) are usually asymptomatic. However, 2.5-7.8% of them may present muscle symptoms and cardiomyopathy, attributed to a reduced production of dystrophin, probably because of skewed patterns of X-chromosome inactivation (XCI). To evaluate the role of XCI in symptomatic (at muscle or heart level) and asymptomatic DMD carriers, 44 subjects were selected from our database (12 manifesting, 21 non-manifesting, 11 healthy females), and XCI pattern determined in the lymphocytes by the androgen receptor methylation-based assay. The results showed that DMD-manifesting carriers had a preferential inactivation of the X-chromosome carrying the normal allele, while non-manifesting carriers and healthy females showed a random XCI pattern. Moreover, when comparing muscle with heart manifesting carriers, the former group showed a higher degree of skewing. No concordance in XCI was found between mothers and daughters, when symptomatic/asymptomatic mother-daughter pairs were analyzed. The results confirm that DMD clinical manifestations in carriers are associated with non-random patterns of X inactivation.

Autosomal recessive limb-girdle muscular dystrophy, LGMD2F, is caused by a mutation in the delta-sarcoglycan gene.

Limb-girdle muscular dystrophies (LGMD) are a heterogeneous group of inherited neuromuscular disorders characterized by proximal muscular weakness of the pelvic and shoulder girdles and a variable progression with symptoms, ranging from very severe to mild. One autosomal dominant (LGMD1A, at chromosome 5q22.3-31.3) (ref. 3) and five autosomal recessive (AR) loci responsible for this phenotype have been identified: LGMD2A at 15q (ref. 4); LGMD2B at 2p (ref. 5), LGMD2C at 13q (ref. 6), LGMD2D at 17q (ref. 7) and LGMD2E at 4q (refs 8,9). In the muscle membrane, dystrophin associates with several proteins and glycoproteins organized in two main subcomplexes: the dystroglycan (DG) and sarcoglycan (SG) complexes. The genes for LGMD2C, LGMD2D and LGMD2E code for proteins of the SG complex. We recently mapped a sixth AR form of LGMD, LGMD2F, to chromosome 5q33-34 in two Brazilian families. In the same chromosomal interval we also mapped the delta SG gene, encoding a novel 35-kD component of the sarcoglycan (SG) complex. We now show that a homozygous mutation in the delta SG gene (a single nucleotide deletion that alters its reading frame) is the cause of LGMD2F.

Italian validation of INQoL, a quality of life questionnaire for adults with muscle diseases.

BACKGROUND AND PURPOSE: A quality of life (QoL) questionnaire for neuromuscular diseases was recently constructed and validated in the United Kingdom in a sample of adult patients with a variety of muscle disorders. Preliminary results suggested it could be a more relevant and practical measure of QoL in muscle diseases than generic health measures of QoL. The purpose of our work was: (i) To validate INQoL in Italy on a larger sample of adult patients with muscle diseases (ii) to compare INQoL to SF-36. METHODS: We have translated into Italian and applied language adaptations to the original UK INQoL version. We studied 1092 patients with different muscle disorders and performed (i) test-retest reliability (n = 80); (ii) psychometric (n = 345), known-group (n = 1092), external criterion (n = 70), and concurrent validity with SF-36 (n = 183). RESULTS: We have translated and formally validated the Italian version of INQoL confirming and extending results obtained in the United Kingdom. In addition to good results in terms of reliability, known-group and criterion validity, a comparison with the SF-36 scales showed a stronger association between INQoL total index and SF-36 physical (r = -0.72) than mental (r = -0.38) summary health indexes. When considering comparable domains of INQoL and SF-36 with respect to an objective measure of muscle strength assessment (MMRC), regression analysis showed a stronger correlation using INQoL rather than SF-36 scores. CONCLUSIONS: INQoL is recommended to assess QoL in muscle diseases because of its ability to capture physical limitations that are specifically relevant to the muscle condition.

Risk of arrhythmia in type I myotonic dystrophy: the role of clinical and genetic variables.

OBJECTIVE: To examine the association between the presence of arrhythmia in type 1 myotonic dystrophy (DM1) and clinical-genetic variables, evaluating their role as predictors of the risk of arrhythmia. METHODS: 245 patients with genetically proven DM1 underwent clinical and non-invasive cardiological evaluation. Severity of muscular involvement was assessed according to the 5 point Muscular Disability Rating Score (MDRS). Data were analysed by univariate and multivariate models. RESULTS: 245 patients were examined and cardiac arrhythmias were found in 63 subjects, 40 of whom required a device implant. Statistical analyses revealed that men had more than double the risk of developing arrhythmias compared with women (p = 0.018). Addition of each year of age caused an increased risk of arrhythmia equal to 3% (p = 0.030). Subjects with MDRS 5 had a risk of arrhythmia 12 times higher than patients with MDRS 1-2 (p<0.001). Although all of these variables were significantly associated with cardiac rhythm dysfunction, they had a low sensitivity for the prediction of arrhythmic risk CONCLUSION: Male sex, age and muscular disability were strongly associated with the development of arrhythmia in DM1. However, all of these variables were weak predictors of arrhythmic risk. These results suggest that other factors may be involved in the development of cardiac conduction abnormalities in DM1.

Usefulness of heart rate variability as a predictor of sudden cardiac death in muscular dystrophies.

Sudden cardiac death, or cardiac arrest, is a major health problem, causing about 166,200 deaths each year among adults in the United States. It may be caused by almost all known heart diseases. Most cardiac arrests occur when the diseased heart begins to exhibit rapid and/or chaotic activity, such as ventricular tachycardia or fibrillation. Some are due to extreme slowing of the heart. All these events are called life-threatening arrhythmias. Arrhythmogenic cardiomyopathy is a frequent feature in several muscular dystrophies with a potential risk of cardiac sudden death. Among the measures able to predict the propensity to develop life-threatening arrhythmias, heart rate variability is an accepted non invasive measurement of cardiac autonomic modulation. The use of heart rate variability to measure the extent of changes in autonomic nervous system is an established risk stratification procedure in different diseases. In fact numerous studies have demonstrated the positive prognostic power of altered heart rate variability values to predict all-cause mortality, cardiac events, sudden cardiac death and heart transplantation. Usefulness of heart rate variability as a predictor of sudden cardiac death in muscular dystrophies has been reviewed.

Analysis of Single Nucleotide Polymorphisms (SNPs) of the small-conductance calcium activated potassium channel (SK3) gene as genetic modifier of the cardiac phenotype in myotonic dystrophy type 1 patients.

Myotonic dystrophy type 1 (DM1) is the most frequently inherited neuromuscular disease in adults. It is a multisystemic disorder with major cardiac involvement most commonly represented by first-degree atrioventricular heart block (AVB), followed by different degrees of bundle-branch and intraventricular blocks In search for candidate genes, modifiers of the AVB phenotype in DM1, the expression of the small-conductance calcium activated potassium channel (SK3) gene was analysed in muscle biopsies from DM1 patients. The association between SK3 polymorphisms and the AVB phenotype was then studied analyzing 40 DM1 patients with AVB and 40 age-matched DM1 affected individuals with no ECG abnormalities. [CTG]n repeat length and cardiac clinical picture were also assessed for correlation. QRT-PCR experiments showed an over-expression of the SK3 transcript in DM1 muscle biopsies compared to healthy controls. However, no statistical association between the AVB phenotype and either the [CTG]n expansion length or the presence of specific SNPs in the SK3 gene were detected. These findings suggest that modifier genes, other than SK3, should be identified in order to explain the cardiac phenotypic variability among DM1 patients.

Extensive scanning of the calpain-3 gene broadens the spectrum of LGMD2A phenotypes.

BACKGROUND: The limb girdle muscular dystrophies (LGMD) are a heterogeneous group of Mendelian disorders highlighted by weakness of the pelvic and shoulder girdle muscles. Seventeen autosomal loci have been so far identified and genetic tests are mandatory to distinguish among the forms. Mutations at the calpain 3 locus (CAPN3) cause LGMD type 2A. OBJECTIVE: To obtain unbiased information on the consequences of CAPN3 mutations. PATIENTS: 530 subjects with different grades of symptoms and 300 controls. METHODS: High throughput denaturing HPLC analysis of DNA pools. RESULTS: 141 LGMD2A cases were identified, carrying 82 different CAPN3 mutations (45 novel), along with 18 novel polymorphisms/variants. Females had a more favourable course than males. In 94% of the more severely affected patient group, the defect was also discovered in the second allele. This proves the sensitivity of the approach. CAPN3 mutations were found in 35.1% of classical LGMD phenotypes. Mutations were also found in 18.4% of atypical patients and in 12.6% of subjects with high serum creatine kinase levels. CONCLUSIONS: A non-invasive and cost-effective strategy, based on the high throughput denaturing HPLC analysis of DNA pools, was used to obtain unbiased information on the consequences of CAPN3 mutations in the largest genetic study ever undertaken. This broadens the spectrum of LGMD2A phenotypes and sets the carrier frequency at 1:103.

Cardiac treatment in neuro-muscular diseases.

The authors have been treating heart involvement in muscle dystrophy since 1978. However, this study aimed to define recent therapeutic protocols, evaluating the results of cardiac treatment, performed between 1st February 2004 and 31st July, 2006. In this period, 100 Becker, 136 Duchenne, 44 Limb-girdle and 116 Steinert patients were treated. In that same period, a large group of MD patients refusing cardiac therapy have also been followed. All patients had previously been classified in the appropriate stage of cardiomyopathy and examined at least twice every year and even every week if presenting heart failure. The results show the usefulness of the recent protocols of treatment of cardiac involvement in muscle dystrophy patients.

The lack of the Celf2a splicing factor converts a Duchenne genotype into a Becker phenotype.

Substitutions, deletions and duplications in the dystrophin gene lead to either the severe Duchenne muscular dystrophy (DMD) or mild Becker muscular dystrophy depending on whether out-of-frame or in-frame transcripts are produced. We identified a DMD case (GSΔ44) where the correlation between genotype and phenotype is not respected, even if carrying a typical Duchenne mutation (exon 44 deletion) a Becker-like phenotype was observed. Here we report that in this patient, partial restoration of an in-frame transcript occurs by natural skipping of exon 45 and that this is due to the lack of Celf2a, a splicing factor that interacts with exon 45 in the dystrophin pre-mRNA. Several experiments are presented that demonstrate the central role of Celf2a in controlling exon 45 splicing; our data point to this factor as a potential target for the improvement of those DMD therapeutic treatments, which requires exon 45 skipping.

Implicación ocular en pacientes con distrofia miotónica / Eye involvement in patients with myotonic dystrophy

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Paroxysmal atrial fibrillation in myotonic dystrophy type 1 patients: P wave duration and dispersion analysis.

OBJECTIVE: Myotonic dystrophy type 1 (MD1) is characterized by cardiac involvement, in about 80% of case, that predominantly affects the conduction system. Aim of our study was to evaluate the P-wave duration and dispersion (PD) in MD1 patients underwent pacemaker implantation with conserved systolic and diastolic function. PATIENTS AND METHODS: We enrolled 60 MD1 patients (age 51.3 ± 5 years; 11 females) underwent dual chamber pacemaker implantation for various grade of atrioventricular (AV) block. Sixty sex-and age matched non-MD1 subjects were recruited as controls. P-wave duration and dispersion were carefully measured using 12-lead electrocardiogram. RESULTS: Compared with healthy control group, MD1 patients presented increased maximum P wave duration (106.4 ± 20.9 vs 65.9 ± 8.2 ms, p = 0.03) and PD values (40.1 ± 11 vs 27.1 ± 4.2 ms, p = 0.003). No statistically significant difference was found in minimum P wave duration (69.7 ± 11.8 vs 65.4 ± 8.1 ms, p = 0.4). The MD1 patients with paroxysmal atrial fibrillation, compared with MD1 patients without evidence of atrial fibrillation, presented increased maximum P wave duration (108.1 ± 10.4 vs 78.1 ± 7.9 ms, p = 0.001) and PD values (41.1 ± 8.5 vs 33.2 ± 4.2 ms, p = 0.003). Minimum P wave duration (68.4 ± 8.2 vs 67.1 ± 4.9 ms, p = 0.5) didn't differ between the two groups. CONCLUSIONS: Our data showed a significantly increased P wave duration and dispersion in MD1 patients compared with age and sex-matched healthy controls. We showed a statistically significant increase in PD and P max in MD1 patients subgroup with AF compared to MD1 patients with no arrhythmias.

Molecular diagnosis in LGMD2A: mutation analysis or protein testing?

Limb girdle muscular dystrophy (LGMD) type 2A (LGMD2A) is caused by mutations in the CAPN3 gene encoding for calpain-3, a muscle specific protease. While a large number of CAPN3 gene mutations have already been described in calpainopathy patients, the diagnosis has recently shifted from molecular genetics towards biochemical assay of defective protein. However, an estimate of sensitivity and specificity of protein analysis remains to be established. Thus, we first correlated protein and molecular data in our large LGMD2A patient population. By a preliminary immunoblot screening for calpain-3 protein of 548 unclassified patients with various phenotypes (LGMD, myopathy, or elevated levels of serum creatine kinase [hyperCKemia]), we selected 208 cases for CAPN3 gene mutation analysis: 69 had protein deficiency and 139 had normal expression. Mutation search was conducted using SSCP, denaturing high performance liquid chromatography (DHPLC), amplification refractory mutation system (ARMS-PCR), and direct sequencing methods. We identified 58 LGMD2A mutant patients: 46 (80%) had a variable degree of protein deficiency and 12 (20%) had normal amount of calpain-3. We calculated that the probability of having LGMD2A is very high (84%) when patients show a complete calpain-3 deficiency and progressively decreases with the amount of protein; this new data offers an important tool for genetic counseling when only protein data are available. A total of 37 different CAPN3 gene mutations were detected, 10 of which are novel. In our population, 87% of mutant alleles were concentrated in seven exons (exons 1, 4, 5, 8, 10, 11, and 21) and 61% correspond to only eight mutations, indicating the regions where future molecular analysis could be restricted. This study reports the largest collection of LGMD2A patients so far in which both protein and gene mutations were obtained to draw genotype-protein-phenotype correlations and provide insights into a critical protein domain.

Mutation of dystrophin gene and cardiomyopathy.

The correlations between the type of gene mutation and the cardiac clinical picture were examined in 284 patients with dystrophinopathy (200 Duchenne and 84 Becker). The subjects with normal heart showed deletions including exons 48-49 in 21.4% DMD and in 25% BMD, and other deletions in 35.7% DMD and 25% BMD; vice versa the cases with severe cardiac involvement showed deletions including 48-49 in 38.8% DMD and 37.5% BMD and other deletions in 32.9% DMD and 20% BMD. The age of death was 18 years in DMD patients with deletions including 48-49 whereas the age was about 22 in the cases with other deletions. The differences were statistically significant.

Deflazacort in Duchenne muscular dystrophy: a comparison of two different protocols.

We compare the long-term benefits and side effects of deflazacort using two treatment protocols from Naples (N) and Toronto (T). Boys with Duchenne muscular dystrophy between the ages of 8 and 15 years and who had four or more years of deflazacort treatment were reviewed. Diagnostic criteria included males with proximal muscle weakness evident before 5 years, increased serum creatine kinase and genetic testing and/or a muscle biopsy consistent with Duchenne muscular dystrophy. Thirty-seven boys were treated with protocol-N using deflazacort at a dose of 0.6 mg/kg per day for the first 20 days of the month and no deflazacort for the remainder of the month. Boys with osteoporosis received daily vitamin D and calcium. Deflazacort treatment started between 4 and 8 years of age. Thirty-two were treated with protocol-T using deflazacort at a dose of 0.9 mg/kg per day, plus daily vitamin D and calcium. Treatment started between 6 and 8 years of age. All boys were monitored every 4-6 months. The results were compared with age-matched controls in the two groups (19 for protocol-N and 30 for protocol-T). For the boys treated with protocol-N, 97% were ambulatory at 9 years (control, 22%), 35% at 12 years (control, 0%), 25% at 15 years (control, 0%). For the 32 boys treated with protocol-T, 100% were ambulatory at 9 years (control, 48%), 83% at 12 years (control, 0%) and 77% at 15 years (control, 0%). No aids or leg braces were used for ambulation. In boys 13 years and older, a scoliosis of >20 degrees developed in 30% of the boys on protocol-N, 16% on protocol-T and 90% of controls. For protocol-N, no cataracts were observed while in protocol-T, 30% of boys had asymptomatic cataracts that required no treatment. Fractures occurred in 19% (control 16%) of boys on protocol-N and 16% (control, 20%) of boys on protocol-T. This report illustrates: (a) the importance of collaborative studies in developing treatment protocols in Duchenne muscular dystrophy and (b) the long-term beneficial effects of deflazacort treatment in both protocols. However, the protocol-T seems to be more effective and frequently is associated with asymptomatic cataracts.

Discordant clinical outcome in type III spinal muscular atrophy sibships showing the same deletion pattern.

Three type III spinal muscular atrophy (SMA) families are described in which the same deletion pattern for SMN gene and flanking loci is apparent in both affected and unaffected siblings. Deletions extending to include the NAIP gene are reported in one sibship. All three individuals in which SMN and/or NAIP deletions were detected showed the same haplotypes for SMA linked microsatellite markers as their affected sibs. The three index cases had a SMA III with early onset (1.5-2 yr) and became chairbound at the age 4, 5 and 20 yr. The three haploidentical sibs were given a clinical severity score. One of them showed no sign of the disease at the age of 4 yr and was considered "unaffected"; a 35-yr-old female, who had no symptoms but showed tongue fasciculations and hand tremor was considered "asymptomatic"; a 34-yr-old female, who had mild muscular weakness since the age of 24, was rated "mild". These observations demonstrate the presence of a continuum of clinical variability within SMA III families. These data suggest that, in these three families at least, the SMA phenotype is caused or influenced by another gene(s) additional to SMN.

Evaluation of cardiac and respiratory involvement in sarcoglycanopathies.

Sarcoglycanopathies constitute a subgroup of limb-girdle recessive muscular dystrophies due to defects in sarcoglycan complex that comprises five distinct transmembrane proteins called alpha-, beta-, gamma-, delta-and epsilon-sarcoglycans. As it is well known that sarcoglycans are expressed both in heart and in skeletal muscles and a complete deficiency in delta-sarcoglycan is the cause of the Syrian hamster BIO.14 cardiomyopathy, we studied cardiac and respiratory involvement in 20 patients with sarcoglycanopathies by clinical, electrocardiographic, echocardiographic, scintigraphic and spirometric assessments. A normal heart function was found in 31.3% of all patients; a preclinical cardiomyopathy in 43.7%; an arrhythmogenic cardiomyopathy in 6.3% and initial signs of dilated cardiomyopathy in 18.7%. In one patient the data were examined retrospectively. No correlation was found between cardiac and skeletal muscle involvement. With reference to the type of sarcoglycanopathy, signs of hypoxic myocardial damage occurred in beta-, gamma- and delta-sarcoglycanopathies, while initial signs of a dilated cardiomyopathy in gamma- and delta-sarcoglycanopathies were found. A normal respiratory function was observed in 23.5% of all patients, a mild impairment in 35.4%, a moderate impairment in 29.4%, and a severe impairment in 11.7%.