Mucopolysaccharidosis type II: identification of 30 novel mutations among Latin American patients.

In this study, 103 unrelated South-American patients with mucopolysaccharidosis type II (MPS II) were investigated aiming at the identification of iduronate-2-sulfatase (IDS) disease causing mutations and the possibility of some insights on the genotype-phenotype correlation The strategy used for genotyping involved the identification of the previously reported inversion/disruption of the IDS gene by PCR and screening for other mutations by PCR/SSCP. The exons with altered mobility on SSCP were sequenced, as well as all the exons of patients with no SSCP alteration. By using this strategy, we were able to find the pathogenic mutation in all patients. Alterations such as inversion/disruption and partial/total deletions of the IDS gene were found in 20/103 (19%) patients. Small insertions/deletions/indels (<22 bp) and point mutations were identified in 83/103 (88%) patients, including 30 novel mutations; except for a higher frequency of small duplications in relation to small deletions, the frequencies of major and minor alterations found in our sample are in accordance with those described in the literature.

Associations between parenting behavior and anxiety in a rodent model and a clinical sample: relationship to peripheral BDNF levels.

Adverse early-life environment is associated with anxiety-like behaviors and disorders. Brain-derived neurotrophic factor (BDNF) is sensitive to this environment and could be a marker of underlying brain changes. We aimed at evaluating the development of anxiety-like behaviors in a rat model of early adversity, as well as the possible association with BDNF levels. Similar associations were investigated in a sample of adolescent humans. For the rat study, Wistar rat litters were divided into: early-life stress (ELS, limited access to nesting material) and control groups. Maternal behavior was observed from days 1 to 9 of life and, as adults, rats were subjected to behavioral testing and BDNF measurements in plasma, hippocampus, amygdala and periaqueductal gray. For the human study, 129 adolescents were evaluated for anxiety symptoms and perceived parental care. Serum BDNF levels and the Val66Met polymorphism of the BDNF gene were investigated. We found that ELS dams showed more pure contact, that is, contact with low care and high control, toward pups, and their adult offspring demonstrated higher anxiety-like behaviors and plasma BDNF. Also the pure contact correlated positively with adult peripheral BDNF. Similarly in humans, there was a positive correlation between maternal overprotection and serum BDNF only in Met carriers. We also found negative correlations between maternal warmth and separation anxiety, social phobia and school phobia. Finally, our translational approach revealed that ELS, mediated through variations in maternal care, is associated with anxiety in both rats and humans and increased peripheral BDNF may be marking these phenomena.

Report of a Large Brazilian Family With a Very Attenuated Form of Hunter Syndrome (MPS II).

Hunter syndrome, or Mucopolysaccharidosis type II (MPS II), is a rare X-linked recessive disorder caused by a deficiency of the lysosomal enzyme iduronate-2-sulfatase (IDS). The phenotypic spectrum varies from severe to attenuated clinical forms. We report a large Brazilian family with 16 affected individuals exhibiting a very attenuated form of MPS II. Fourteen female carriers were also identified. Twelve affected male patients, whose ages ranged from 1 to 35 years, were examined. Molecular analysis showed a novel missense mutation (p.A77D) in the IDS gene, confirming the diagnosis. Nine of the family members presented some degree of heart damage, though only the proband became symptomatic and required heart transplantation. One 19-year-old adult and 1-year-old twin boys each had a normal echocardiogram. Short stature was found in two adults while macrocephaly was found in one; the remaining adults had anthropometric measures within normal range. All affected adults had normal cognitive development and were able to perform normal daily activities, except one who had mild learning disability. Two patients died due to natural causes beyond 70 years of age. The female carriers did not present any signs of disease. In this large family with a mild form of MPS II and variable degree of clinical manifestations, it is noteworthy that several affected individuals have remained asymptomatic even at advanced age and even without enzyme replacement therapy.

Clinical and biochemical studies in mucopolysaccharidosis type II carriers.

The aim of the study was to characterize clinically and biochemically mucopolysaccharidosis type II (MPS II) heterozygotes. Fifty-two women at risk to be a carrier, with a mean age of 34.1 years (range 16-57 years), were evaluated through pedigree analysis, medical history, physical examination, measurement of iduronate sulfatase (IDS) activities in plasma and in leukocytes, quantification of glycosaminoglycans (GAGs) in urine, and analysis of the IDS gene. Eligibility criteria for the study also included being 16 years of age or older and being enrolled in a genetic counselling programme. The pedigree and DNA analyses allowed the identification of 40/52 carriers and 12/52 non-carriers. All women evaluated were clinically healthy, and their levels of urinary GAGs were within normal limits. Median plasma and leukocyte IDS activities found among carriers were significantly lower than the values found for non-carriers; there was, however, an overlap between carriers' and non-carriers' values. Our data suggests that MPS II carriers show lower plasma and leukocyte IDS activities but that this reduction is generally associated neither with changes in levels of urinary GAGs nor with the occurrence of clinical manifestations.

Mucopolysaccharidosis type VI: Identification of novel mutations on the arylsulphatase B gene in South American patients.

Mucopolysaccharidosis type VI (Maroteaux-Lamy syndrome, MPS VI) is an autosomal recessive disorder caused by deficiency of N-acetylgalactosamine-4-sulphatase (ARSB),which leads to the lysosomal accumulation and excretion of dermatan sulphate (DS). In this study, 13 unrelated MPS VI patients (12 Brazilian and 1 Chilean) were investigated regarding the identification of the ARSB gene mutations using PCR, SSCP and sequencing. The exons with altered mobility on SSCP were sequenced, as well as all the exons of patients with no SSCP alteration. Seven novel mutations were identified: D59N, L72R, Q88H, P93S, R197X, 1279delA and c.1143-8T > G. The previously reported mutations 1533del23, R315Q and 427delG were found in six, three and two alleles respectively. The other mutations already reported, S384N and G144R, were found in only one allele. In addition, three polymorphisms previously described (V358M, V376M and P397P) were detected in the patients analysed. Our findings are in agreement with the literature confirming the great genetic heterogeneity associated with MPS VI.

In vivo evidence of brain galactitol accumulation in an infant with galactosemia and encephalopathy.

In a newborn infant with galactose-1-phosphate uridyltransferase deficiency and encephalopathy, brain magnetic resonance imaging revealed cytotoxic edema in white matter. Using in vivo proton magnetic resonance spectroscopy, we detected approximately 8 mmol galactitol per kilogram of brain tissue, an amount potentially relevant to the pathogenesis of brain edema.

Gossypol blood levels and inhibition of spermatogenesis in men taking gossypol as a contraceptive. A multicenter, international, dose-finding study.

The safety and efficacy of gossypol continues to be controversial. The aim of this study was to evaluate gossypol as a contraceptive pill for men at doses lower than those previously prescribed and in men from various ethnic origin. A total of 151 men from Brazil, Nigeria, Kenya, and China were divided into two groups. Both groups received 15 mg gossypol/day for 12 or 16 weeks to reach spermatogenesis suppression. Subjects were then randomized to either 7.5 or 10 mg/day for 40 weeks. In addition, 51 men were enrolled as a control group. In all, 81 subjects attained spermatogenesis suppression. Only one man discontinued treatment because of tiredness. Potassium levels fluctuated within the normal range. FSH increased consistently. Testicular volume decreased, but after discontinuation, values returned to levels not statistically different from admission. Of 19 subjects on the 7.5 mg/day dose group, 12 recovered sperm counts >20 million/mL within 12 months of discontinuing gossypol. In the 10 mg/day group, sperm counts recovered in only 10 of 24 subjects. Eight of the 43 patients remained azoospermic 1 year after stopping gossypol. All men diagnosed with varicocele failed to reverse spermatogenesis suppression. Gossypol blood levels indicated that sperm suppression occurs independently of concentration, whereas spermatogenesis recovery appears to be concentration-dependent. Gossypol may become a medical alternative to surgical vasectomy when the delay in onset of infertility is acceptable. When taken for 1 year, gossypol causes no reduction in sexual desire or frequency of intercourse. The possibility of reversal, occurring in 51% of the men on this regimen within 1 year after stopping gossypol, is an advantage of this compound as compared with surgical sterilization in many parts of the world.

PIP: The safety and efficacy of gossypol continues to be controversial. The aim of this study was to evaluate gossypol as a contraceptive pill for men at doses lower than those previously prescribed and in men from various ethnic origin. A total of 151 men from Brazil, Nigeria, Kenya, and China were divided into two groups. Both groups received 15 mg gossypol/day for 12 or 16 weeks to reach spermatogenesis suppression. Subjects were then randomized to either 7.5 or 10 mg/day for 40 weeks. In addition, 51 men were enrolled as a control group. In all, 81 subjects attained spermatogenesis suppression. Only 1 man discontinued treatment because of tiredness. Potassium levels fluctuated within the normal range. FSH increased consistently. Testicular volume decreased, but after discontinuation, values returned to levels not statistically different from admission. Of 19 subjects in the 7.5 mg/day dose group, 12 recovered sperm counts higher than 20 million/ml within 12 months of discontinuing gossypol. In the 10 mg/day group, sperm counts recovered in only 10 of 24 subjects. 8 of the 43 patients remained azoospermic 1 year after stopping gossypol. All men diagnosed with varicocele failed to reverse spermatogenesis suppression. Gossypol blood levels indicated that sperm suppression occurs independently of concentration, whereas spermatogenesis recovery appears to be concentration-dependent. Gossypol may become a medical alternative to surgical vasectomy when the delay in onset of infertility is acceptable. When taken for 1 year, gossypol causes no reduction in sexual desire or frequency of intercourse. The possibility of reversal, occurring in 51% of the men on this regimen within 1 year after stopping gossypol, is an advantage of this compound as compared with surgical sterilization in many parts of the world.

Uridine diphosphoglucose content of human erythrocytes: assessment by conversion to uridine diphosphoglucuronate.

To settle the ongoing controversy regarding differential uridine diphosphoglucose (UDPG) and uridine diphosphogalactose (UDPGal) content of erythrocytes, which may be important in evaluating the metabolic abnormality in patients with galactosemia, we derived a combined enzymatic-high-performance liquid chromatography (HPLC) assay. Uridine diphosphoglucuronate (UDPGA), the unique product of UDPG dehydrogenase activity, was separated and quantified by HPLC in extracts of human erythrocytes. The quantity of UDPGA produced in cell filtrates incubated with the enzyme corresponds to the amount of UDPG directly determined by HPLC. The amount of UDPGA produced was independent of the enzyme purity or activity used. On the other hand, the amounts of UDPG estimated by fluorometric measurement of the production of reduced nicotinamide adenine dinucleotide varied with the enzyme purity and activity. The combined enzymatic-HPLC method confirms the direct determinations of UDPG content of normal erythrocytes. The results indicate that, under appropriate conditions, the fluorometric-based assay will give accurate estimates of UDPG, but the direct HPLC method yields consistent and correct UDPG and UDPGal determinations.

Intellectual outcome in children with maple syrup urine disease.

We report a controlled study of intellectual outcome in 16 children with maple syrup urine disease (MSUD) that compares the outcome of MSUD diagnosed after symptoms became apparent with that of MSUD diagnosed prospectively and in unaffected siblings and parents. The mean IQ (+/- SD) score in the children with classic MSUD was 78 +/- 24; however, there were two discrete groups: one with normal IQ (greater than 84) whose MSUD had been diagnosed at a mean age of 3.5 days and a second group, with IQ below normal, whose MSUD was diagnosed at a mean of 10 days of age. Affected children treated presymptomatically had higher IQ scores than their affected siblings treated when their disease was symptomatic. Multiple regression analysis indicated that the important influences on IQ were age at the time of diagnosis and long-term metabolic control; control at the time of testing also might have affected performance. The mean score of unaffected siblings was 92 +/- 5 and the mean parental IQ was 83 +/- 9. The mean IQ scores of children with variant MSUD, 97 +/- 4, was similar to that of their parents, 103 +/- 6. This study was not longitudinal and thus could not identify subtle developmental learning problems. We conclude that early and meticulous treatment of MSUD can result in intellectually normal children.

Prospective study of the pituitary-ovarian function after tubal sterilization by the Pomeroy or Uchida techniques.

Surgical sterilization has become one of the most widely used methods of fertility regulation in the world. A common concern is the not yet clarified possible disturbance of the ovarian function resulting from the surgical procedure. A prospective study was carried out to contribute toward answering this question. Twenty subjects were enrolled in a longitudinal study of ovarian endocrine function before, and 2 and 6 months after, tubal ligation. The Pomeroy technique, which may compromise ovarian circulation, and the Uchida technique, which is free of this risk, were used randomly. Eight women who underwent to the Pomeroy technique and 9 who underwent the Uchida technique completed the protocol. Luteinizing hormone, follicle-stimulating hormone, estradiol, and progesterone were measured daily from days 10 through 18 of the cycle and every other day until the onset of the next menses. No change in any of the hormones studied was observed, with the exception of a significant increase in progesterone 2 months after tubal ligation by the Uchida technique.

Effect of uridine on hepatic galactose-1-phosphate uridyltransferase.

Uridine sugar nucleotides are important intermediates in galactose metabolism and may play a role in the long-term galactose toxicity in human galactose-1-phosphate uridyltransferase deficiency galactosemia. Since administration of uridine, a precursor of uridine nucleotides, has been considered as a therapeutic measure, we have investigated the effects of this compound on the activity of rat hepatic transferase. Uridine has been found to be an inhibitor of the enzyme in in vitro studies and to cause an increase in galactose-1-phosphate in liver perfused with galactose which is consistent with physiologic inhibition of the enzyme. Uridine is a partial linear competitive inhibitor of UDPglucose and an uncompetitive inhibitor of galactose-1-phosphate. These findings suggest caution should be applied in giving the compound to subjects with genetically limited transferase activity because of the possibility of inhibiting the small amount of residual enzyme.

Cystinuria in dogs: comparison of the cystinuric component of the Fanconi syndrome in basenji dogs to isolated cystinuria.

Two animal models for cystinuria have been examined: the Basenji dog with Fanconi syndrome and cystine stone-forming dogs of various breeds. Brush-border membranes were isolated from these animals and uptake of D-glucose and L-cystine was characterized. Experiments with isolated brush-border vesicles from Basenji dogs with cystinuria as a component of the Fanconi syndrome showed diminished sodium-dependent D-glucose uptake but no decrease in L-cystine uptake even though the cystine defect in vivo was as high as 94% (ie, 6% reabsorption). In contrast, brush-border vesicles isolated from the kidney of a cystine stone-forming dog (Welsh Corgi) with a cystine defect of only 16% (ie, 84% reabsorption) had decreased uptake of cystine compared to values found for Beagle and Basenji vesicles. Thus, cystinuria found in Basenji dogs with the Fanconi syndrome differs from that in classic stone-forming cystinuric dogs. The alteration responsible for the cystinuria of Basenji dogs with Fanconi syndrome does not appear to have a membrane locus and may reflect altered energetics for transport, which are not detected in isolated vesicles. The cystine defect in cystinuric stone-forming dogs does appear to be reflected in the isolated membrane.

Holocarboxylase synthetase deficiency: 9-year follow-up of a patient on chronic biotin therapy and a review of the literature.

We report on the long-term medical and neurodevelopmental follow-up of a patient with the rare and potentially lethal disease, holocarboxylase synthetase deficiency. He was originally treated prenatally with biotin megatherapy and for 9 years with 6 mg/day since his only episode of fulminant acidosis at 3 months of age. While growth and general health have been normal, the patient has exhibited signs of minimal brain dysfunction. However, evaluation of unaffected siblings suggests that this may be unrelated to his metabolic disease. A review of the literature and recommendations for optimal treatment are provided.

Isovaleric acidemia: medical and neurodevelopmental effects of long-term therapy.

Nine patients with isovaleric acidemia were treated with a low-protein diet and supplemental glycine for up to 10 years. Carnitine was added to the therapy in four patients. Overall, the treatment was well tolerated, resulting in no significant side effects other than persistent hyperglycinemia. Normal growth was observed in all patients. Of four patients with the chronic phenotype, three, whose treatment was delayed beyond the first year of life, are mentally retarded. Two of five patients with the acute phenotype are retarded. The outcome in these two was complicated in one by neonatal intraventricular hemorrhage and in the other by therapeutic noncompliance. In our patients, only those who were treated successfully from early infancy and had no complications did not develop mental retardation. After initiation of therapy, there was a significant decrease in ketoacidotic attacks requiring hospitalization. Glycine is indicated for the treatment of acute ketoacidosis in these patients; none of the catastrophically ill newborn who received glycine died. The aim of treatment is to reduce the isovaleric acid burden to a minimum. Therapy consisting of leucine restriction with supplemental glycine and carniline should be started as soon as possible after birth.

Angiosarcoma presenting as diffuse alveolar hemorrhage.

Angiosarcoma has been reported as occurring in both postirradiation and postradical mastectomy patients. Described is a patient, postmastectomy and irradiation, with alveolar hemorrhage secondary to angiosarcoma. Angiosarcoma, primary or metastatic to lung, should be included in the differential diagnosis of diffuse alveolar hemorrhage in this patient population.

Holocarboxylase synthetase deficiency: a biotin-responsive organic acidemia.

The clinical and biochemical features of an infant affected by holocarboxylase synthetase deficiency are presented. The patient was the sibling of the deceased child in whose cultured skin fibroblasts the precise enzymatic disorder was first determined. This fact permitted administration of specific therapy in the form of oral biotin, resulting in immediate improvement from impending respiratory failure and shock. The clinical response to biotin was accompanied by recovery of the biochemical mechanisms known to be biotin-dependent, as manifested by disappearance of intermediates in urine and blood. The variability of biotin responsiveness and the diversity of clinical presentation in the patients originally thought to have a deficiency of beta methylcrotonylCoA carboxylase, a biotin-dependent enzyme, raises the question of a separate, specific apocarboxylase defect.