Cutaneous silent periods in patients with Fabry disease

We assessed the cutaneous silent period (CSP) in 24 patients with Fabry disease with small-fiber sensory neuropathy and 12 normal subjects to test the hypothesis that small-diameter afferents are responsible for producing the CSP. Sensory nerve conduction studies and quantitative sensory testing for cold and vibration detection thresholds were also measured. Overall, Fabry patients had impaired thermal, but not vibration, detection thresholds, with greatest impairment in the feet. In the upper extremity, CSP latencies, duration, and suppression of electromyographic activity (EMG) did not differ. In the lower extremity, patients had reduced suppression of EMG during the CSP compared to normal controls. CSP durations exhibited a bimodal distribution in patients, including a subset of seven patients with durations shorter than all controls. This subset had profound loss of thermal sensation in the feet, but this was also true of some patients who had normal CSPs. Patients with shortened CSPs had modestly elevated vibration thresholds and reduced sensory potentials in comparison to patients with normal CSPs. Reduced CSPs in Fabry patients are associated with, but not entirely explained by, the severity of small-fiber neuropathy as measured by quantitative sensory testing. The possibility that large-diameter fibers provide a minor contribution to producing the CSP should be considered.     

Effect of vitamin D receptor (VDR) genotypes on the risk for osteoporosis in type 1 Gaucher disease

The almost universal finding of osteopenia in Gaucher disease and the unpredictable advent of avascular necrosis and/or pathological fractures are clinically relevant issues that are not necessarily reversed by specific enzyme replacement therapy (ERT). However, genetic/epigenetic factors may explain variance for bone mineral density (BMD) in the normal population. Among the candidate genes, vitamin D receptor (VDR) polymorphisms have been correlated with BMD at the lumbar spine (LS) and femoral neck (FN) and/or fracture risk in various populations. Helsinki Committee approval was received. DNA samples from 15 healthy individuals and 83 non-neuronopathic adult patients with recent BMD data at LS and FN were tested for VDR polymorphisms FokI, ApaI, TaqI, and BsmI. Chi-square, Fisher’s exact, analysis of variance (ANOVA), Scheffe post hoc, and non-parametric Kruskal-Wallis ANOVA were applied as appropriate. P values <0.05 was considered statistically significant. There were no significant differences between patients and controls in frequency of any of polymorphic genotypes. The ApaI and BsmI genotypes were correlated with the N370S/N370S Gaucher genotype (p = 0.029, and p = 0.061, respectively). There was a correlation between the BsmI genotype and skeletal involvement (p = 0.018) and a trend to significance with Z scores at LS (p = 0.084). There was a statistically significant correlation between the Taq1 variants and BMD at the FN (p = 0.030) with the TT variant associated with lower BMD. As in other populations, VDR polymorphic genotype may be an independently sorting modifier in the prediction of BMD and bone involvement in Gaucher disease.     

Single-sperm analysis for haplotype construction of de-novo paternal mutations: application to PGD for neurofibromatosis type 1

BACKGROUND: Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder caused by mutations in the neurofibromin gene. Approximately, 50% of cases are caused by de-novo mutations. Even when the NF1 mutation is known, accuracy of PGD is highly enhanced by simultaneous analysis of linked markers. In a childless couple referred to PGD, the male carried a de-novo mutation, precluding the possibility of typing relatives to establish the mutation-associated haplotype. We developed a single-sperm haplotype analysis strategy to establish the haplotype linked to the NF1 mutation. METHODS: Spermatozoa from freshly ejaculated semen were used as a substrate for multiplex PCR on single sperm. RESULTS: In addition to the NF1 mutation, six informative polymorphic markers flanking the NF1 gene (D17S1294, D17S1849, D17S841, D17S975, NF1TG2 and NF1AC5) were linked to individual alleles in single sperm from the affected male. CONCLUSIONS: Single-sperm analysis established the haplotypes of both mutant and wild-type NF1 alleles and enabled the implementation of a PGD protocol using polymorphic marker analysis. This method is generally applicable to PGD for any disease in which the haplotype of paternal mutations cannot be determined by typing relatives.     

Preimplantation genetic diagnosis (PGD) for a treatable disorder: Gaucher disease type 1 as a model

BackgroundPreimplantation genetic diagnosis (PGD) is a technique that enables identification of unaffected embryos prior to in vitro fertilization (IVF) transfer in couples at risk for a Mendelian disorder. Most cases involve severe genetic diseases with neurological features and/or major malformations. We present two couples in which PGD was performed for prevention of type 1 Gaucher disease, a non-neuronopathic, non-lethal disorder.Materials and methodsWe developed a multiplex fluorescent PCR protocol, simultaneously amplifying the familial mutations and eight closely spaced, highly polymorphic informative microsatellite markers surrounding the gene, to be used for PGD analysis.ResultsCouple #1 mother was homozygous for the N370S mutation and the father carried the 84GG mutation; their first daughter receives specific Gaucher therapy. One PGD cycle resulted in seven embryos of which four had the paternal wild type allele; two were transferred resulting in a healthy baby boy born at term. Couple #2, each a carrier (N370S and R359Q), whose first-born child had died (age 5 years) of Gaucher disease, underwent 7 PGD cycles. Only one cycle resulted in a clinical pregnancy but a miscarriage was followed at 10 weeks.ConclusionsPGD is an effective and accurate method for preventing Gaucher disease type I in carrier couples. Since this disease is treatable, special ethical considerations and careful selection of couples should be performed.     

Nonprecipitous changes upon withdrawal from imiglucerase for Gaucher disease because of a shortage in supply

Because of an unpredicted and unavoidable shortage in global supplies of imiglucerase for Gaucher disease, we collected clinical and laboratory data from patients who were evaluated ≤ 6 months before drug withdrawal and then retested before/during reinstatement of therapy. Hemoglobin, platelet counts, and liver and spleen index volumes by ultrasound as well as chitotriosidase were evaluated as percent change over time and relative to dose-years of exposure to enzyme therapy. Deterioration was seen in all four clinical parameters and in chitotriosidase activity in most patients and even among patients who stopped for only 3 months. No patient consistently showed nondeterioration in all five parameters. Platelet counts were most sensitive to therapy withdrawal. There was no overt correlation between percent change in these parameters and dose-years of therapy. Although we no longer think that drug vacations should be considered as a form of maintenance for patients requiring enzyme therapy for Gaucher disease, but if necessary—for compelling personal reasons—one may be reassured that a short-term drug interruption is not likely to lead to irreversible complications or return to baseline values of critical clinical parameters.     

Successful polar body-based preimplantation genetic diagnosis for achondroplasia

Achondroplasia, the most common form of dwarfism, is a candidate for preimplantation genetic diagnosis (PGD) because a single mutation accounts for almost all cases. Multiplex fluorescent assay including the common G380R mutation in the FGFR3 gene and eight close polymorphic markers was developed. First and second polar bodies (PB) were used for PGD analysis. An affected woman was treated with routine long-protocol ovarian stimulation and puncture. In the first PGD cycle, out of four fertilized oocytes, PB analysis revealed two mutant oocytes, one with total amplification failure of the maternal allele and one with inconclusive results. In the second PGD cycle, 14 oocytes were retrieved following a higher FSH dose and by performing oocyte retrieval and by placing the patient in the anti-Trendelenburg position using abdominal pressure to allow all follicles to be drained. Following PB analysis, two embryos containing the wild-type FGFR3 allele were transferred. This led to an uncomplicated pregnancy and delivery by Caesarean section at week 38 of a healthy boy, carrying the FGFR3 wild-type maternal allele. In conclusion, oocyte retrieval, while difficult in patients with achondroplasia, can be successfully performed. PB analysis is a reliable and sensitive method for PGD for maternal achondroplasia.