Alicia Chan

T1 Mapping With Cardiovascular MRI Is Highly Sensitive for Fabry Disease Independent of Hypertrophy and Sex

Background— Fabry disease (FD) is an X-linked disorder of lysosomal metabolism affecting multiple organs with cardiac disease being the leading cause of death. Current imaging evaluations of the heart are suboptimal. The goals of the current study are to evaluate the potential of quantitative T1 mapping with cardiovascular MRI as a disease-specific imaging biomarker. Methods and Results— A total of 31 patients with FD, 23 healthy controls, and 21 subjects with concentric remodeling or hypertrophy underwent cardiovascular MRI to measure left ventricular (LV) morphology, function, delayed enhancement, as well as myocardial T1 values, and derived parameters (extracellular volume). All subjects had LV ejection fraction >50% and similar volumes. FD and concentric remodeling or hypertrophy had similarly increased mass, wall thickness, and mass/volume as compared with controls. A total of 16 of 31 FD subjects and 10 of 21 concentric remodeling or hypertrophy subjects had LV hypertrophy. Noncontrast myocardial T1 values were substantially lower in FD as compared with controls and concentric remodeling or hypertrophy (1070±50, 1177±27, and 1207±33 ms, respectively; P<0.001), but extracellular volume was similar in all groups (21.7±2.4%, 22.2±3.1%, and 21.8±3.9%, respectively). Single-voxel NMR spectroscopy in 4 FD and 4 healthy control subjects showed a significant negative linear relationship between lipid content and noncontrast T1 values (r=−0.9; P=0.002). Female subjects had lower LV mass and wall thickness, longer myocardial T1 values and larger extracellular volume suggesting a key sex difference in cardiac remodeling. Conclusions— Reduced noncontrast myocardial T1 values are the most sensitive and specific cardiovascular MRI parameter in patients with FD irrespective of sex and LV morphology and function.

T1 Mapping with CMR Is Highly Sensitive for Fabry Disease Independent of Hypertrophy and Gender

Background— Fabry disease (FD) is an X-linked disorder of lysosomal metabolism affecting multiple organs with cardiac disease being the leading cause of death. Current imaging evaluations of the heart are suboptimal. The goals of the current study are to evaluate the potential of quantitative T1 mapping with cardiovascular MRI as a disease-specific imaging biomarker. Methods and Results— A total of 31 patients with FD, 23 healthy controls, and 21 subjects with concentric remodeling or hypertrophy underwent cardiovascular MRI to measure left ventricular (LV) morphology, function, delayed enhancement, as well as myocardial T1 values, and derived parameters (extracellular volume). All subjects had LV ejection fraction >50% and similar volumes. FD and concentric remodeling or hypertrophy had similarly increased mass, wall thickness, and mass/volume as compared with controls. A total of 16 of 31 FD subjects and 10 of 21 concentric remodeling or hypertrophy subjects had LV hypertrophy. Noncontrast myocardial T1 values were substantially lower in FD as compared with controls and concentric remodeling or hypertrophy (1070±50, 1177±27, and 1207±33 ms, respectively; P<0.001), but extracellular volume was similar in all groups (21.7±2.4%, 22.2±3.1%, and 21.8±3.9%, respectively). Single-voxel NMR spectroscopy in 4 FD and 4 healthy control subjects showed a significant negative linear relationship between lipid content and noncontrast T1 values (r=−0.9; P=0.002). Female subjects had lower LV mass and wall thickness, longer myocardial T1 values and larger extracellular volume suggesting a key sex difference in cardiac remodeling. Conclusions— Reduced noncontrast myocardial T1 values are the most sensitive and specific cardiovascular MRI parameter in patients with FD irrespective of sex and LV morphology and function.

The paradox of the carnitine palmitoyltransferase type Ia P479L variant in Canadian Aboriginal populations

Investigation of seven patients from three families suspected of a fatty acid oxidation defect showed mean CPT-I enzyme activity of 5.9+/-4.9 percent of normal controls. The families, two Inuit, one First Nation, live in areas of Canada geographically very distant from each other. The CPT1 and CPT2 genes were fully sequenced in 5 of the patients. All were homozygous for the same P479L mutation in a highly conserved region of the CPT1 gene. Two patients from the first family were also homozygous for the CPT2 F352C polymorphism in the CPT2 gene. Genotyping the patients and their family members confirmed that all seven patients were homozygous for the P479L variant allele in the CPT1 gene, as were 27 of 32 family members. Three of the seven patients and two cousins had hypoketotic hypoglycemia attributable to CPT-Ia deficiency, but adults homozygous for the variant denied hypoglycemia. We screened 422 consecutive newborns from the region of one of the Inuit families for this variant; 294 were homozygous, 103 heterozygous, and only 25 homozygous normal; thus the frequency of this variant allele is 0.81. There was an infant death in one family and at least 10 more deaths in those infants (7 homozygous, 3 heterozygous) consecutively tested for the mutation at birth. Thus there is an astonishingly high frequency of CPT1 P479L variant and, judging from the enzyme analysis in the seven patients, also CPT-I deficiency in the areas of Canada inhabited by these families. Despite the deficiency of CPT-Ia which is the major rate-limiting enzyme for long chain fatty acid oxidation, clinical effects, with few exceptions, were slight or absent. One clue to explaining this paradox is that, judging from the fatty acid oxidation studies in whole blood and fibroblasts, the low residual activity of CPT-Ia is sufficient to allow a reasonable flux through the mitochondrial oxidation system. It is likely that the P479L variant is of ancient origin and presumably its preservation must have conveyed some advantage.

Genetic and Clinical Heterogeneity in eIF2B-Related Disorder

Eukaryotic initiation factor 2B (eIF2B)-related disorders are heritable white matter disorders with a variable clinical phenotype (including vanishing white matter disease and ovarioleukodystrophy) and an equally heterogeneous genotype. We report 9 novel mutations in the EIF2B genes in our subject population, increasing the number of known mutations to more than 120. Using homology modeling, we have analyzed the impact of novel mutations on the 5 subunits of the eIF2B protein. Although recurrent mutations have been found at CpG dinucleotides in the EIF2B genes, the high incidence of private or low frequency mutations increases the challenge of providing rapid genetic confirmation of this disorder, and limits the application of EIF2B screening in cases of undiagnosed leukodystrophy.

Mild Elevation of N-Acetylaspartic Acid and Macrocephaly: Diagnostic Problem

Patients with slightly increased excretion of N-acetylaspartic acid in urine, together with macrocephaly, present a dignostic dilemma for Canavans disease. We describe a 13-year-old male patient with macrocephaly, mild developmental delay, increased signal intensity in the basal ganglia bilaterally, partial cortical blindness, and retinitis pigmentosa. Although the clinical course and magnetic resonance imaging findings did not resemble typical Canavans disease, N-acetylaspartic acid excretion in the patients urine was slightly elevated, 99.90 ± 4.00 μg/mg creatinine, whereas the normal control range was < 83 μg/mg creatinine. Cultured skin fibroblasts from the patient showed no aspartoacylase activity. Cloning of genomic DNA isolated from the patients fibroblasts showed an intronic mutation, specifically deletion of -2A and -3C at the acceptor site of exon 3 and disrupting the normal splicing of the gene. A second mutation was found in exon 6, 863 A→G in aspartoacylase complementary DNA, causing a tyrosine-to-cysteine (Y288C) amino acid substitution. Expression of the mutation on exon 6 showed normal aspartoacylase activity. These data suggest that expression of the mutation may help to understand the enzyme defect in a patient with slightly increased N-acetylaspartic acid excretion. (J Child Neurol 2003;18:809—812).

Longitudinal observations of serum heparin cofactor II-thrombin complex in treated Mucopolysaccharidosis I and II patients.

Monitoring of therapeutic response in mucopolysaccharidosis (MPS) patients is problematic as most biomarkers are specific for either disease complications or specific organ system involvement. Recent studies have indicated that serum heparin-cofactor II-thrombin complex (HCII-T) may serve as an important biomarker in the group of MPSs where dermatan sulphate is stored. This complex forms when blood coagulates in the presence of glycosaminoglycans (GAGs) where the ultimate amount of HCII-T that forms reflects the concentration of circulating GAGs. We have studied serum HCII-T levels in 9 MPS I and 11 MPS II treated patients and have compared values to studies of urinary GAGs. In severe MPS I patients treated with either transplantation or enzyme replacement therapy (ERT), serum HCII-T levels never reach the range of normal despite normalization of uGAGs in some patients. Some attenuated MPS I patients have normalization of HCII-T but require a protracted exposure time relative to the drop in urinary GAGs. Treated MPS II patients show a clear correlation of serum HCII-T levels with the presence of antibodies to Idursulfase, with antibody positive patients showing an early drop in HCII-T levels with eventual increases in levels often to levels above those seen at baseline. This is contrasted by a robust and persistent drop in uGAGs. Antibody negative MPS II patients show a drop in HCII-T levels on treatment but levels never normalize despite normalization of uGAGs. This study highlights the utility and biologic relevance of serum HCII-T levels in monitoring therapy in these disorders.

Mitochondriome and cholangiocellular carcinoma

Cholangiocellular carcinoma (CCA) of the liver was the target of more interest, recently, due mainly to its increased incidence and possible association to new environmental factors. Somatic mitochondrial DNA (mtDNA) mutations have been found in several cancers. Some of these malignancies contain changes of mtDNA, which are not or, very rarely, found in the mtDNA databases. In terms of evolutionary genetics and oncology, these data are extremely interesting and may be considered a sign of poor fitness, which may conduct in some way to different cellular processes, including carcinogenesis. MitoChip analysis is a strong tool for investigations in experimental oncology and was carried out on three CCA cell lines (HuCCT1, Huh-28 and OZ) with different outcome in human and a Papova-immortalized normal hepatocyte cell line (THLE-3). Real time quantitative PCR, western blot analysis, transmission electron microscopy, confocal laser microscopy, and metabolic assays including L-Lactate and NAD+/NADH assays were meticulously used to identify mtDNA copy number, oxidative phosphorylation (OXPHOS) content, ultrastructural morphology, mitochondrial membrane potential (ΔΨm), and differential composition of metabolites, respectively. Among 102 mtDNA changes observed in the CCA cell lines, 28 were non-synonymous coding region alterations resulting in an amino acid change. Thirty-eight were synonymous and 30 involved ribosomal RNA (rRNA) and transfer RNA (tRNA) regions. We found three new heteroplasmic mutations in two CCA cell lines (HuCCT1 and Huh-28). Interestingly, mtDNA copy number was decreased in all three CCA cell lines, while complexes I and III were decreased with depolarization of mitochondria. L-Lactate and NAD+/NADH assays were increased in all three CCA cell lines. MtDNA alterations seem to be a common event in CCA. This is the first study using MitoChip analysis with comprehensive metabolic studies in CCA cell lines potentially creating a platform for future studies on the interactions between normal and neoplastic cells.