Molecular Diagnostics of Copper-Transporting Protein Mutations Allows Early Onset Individual Therapy of Menkes Disease.

Menkes disease is a severe X-linked recessive disorder caused by a defect in the ATP7A gene, which encodes a membrane copper-transporting ATPase. Deficient activity of the ATP7A protein results in decreased intestinal absorption of copper, low copper level in serum and defective distribution of copper in tissues. The clinical symptoms are caused by decreased activities of copper-dependent enzymes and include neurodegeneration, connective tissue disorders, arterial changes and hair abnormalities. Without therapy, the disease is fatal in early infancy. Rapid diagnosis of Menkes disease and early start of copper therapy is critical for the effectiveness of treatment. We report a molecular biology-based strategy that allows early diagnosis of copper transport defects and implementation of individual therapies before the full development of pathological symptoms. Low serum copper and decreased activity of copperdependent mitochondrial cytochrome c oxidase in isolated platelets found in three patients indicated a possibility of functional defects in copper-transporting proteins, especially in the ATPA7 protein, a copper- transporting P-type ATPase. Rapid mutational screening of the ATP7A gene using high-resolution melting analysis of DNA indicated presence of mutations in the patients. Molecular investigation for mutations in the ATP7A gene revealed three nonsense mutations: c.2170C>T (p.Gln724Ter); c.3745G>T (p.Glu1249Ter); and c.3862C>T (p.Gln1288Ter). The mutation c.3745G>T (p.Glu1249Ter) has not been identified previously. Molecular analysis of the ATOX1 gene as a possible modulating factor of Menkes disease did not reveal presence of pathogenic mutations. Molecular diagnostics allowed early onset of individual therapies, adequate genetic counselling and prenatal diagnosis in the affected families.

Prediction of coronary vessel involvement on the basis of atherosclerosis risk factor analysis.

The prediction of coronary vessel involvement by means of noninvasive tests is one of the fundamental objectives of preventive cardiology. This review describes the current possibilities of coronary vessel involvement prediction by means of ultrasonographic examination of carotid arteries, analysis of polymorphisms in the genes encoding enzymes responsible for production of nitric oxide and carbon monoxide and assessment of levels of certain proinflammatory cytokines. In the presented work these noninvasive markers are correlated with the extent of coronary vessel involvement as assessed by coronary angiography, intravascular ultrasound and virtual histology (Fig. 5, Ref. 40).

Stress perception and (GT)n repeat polymorphism in haem oxygenase 1 promoter are both risk factors in development of eating disorders.

Haem oxygenase 1 (HO-1) plays a pivotal role in metabolic stress protecting cells in dependence on reactive oxygen species. This study investigated a potential gene environment interaction between the (GT)n repeat HO1 polymorphism and the stress perception in patients with eating disorder and in controls. Stress perception and (GT)n polymorphism were measured in 127 patients with eating disorders and in 78 healthy controls using Stress and Coping Inventory and genotyping. Based on the inventory, overall, specific and weighted stress scores were defined. Clinical stress score was generated according to the patient's history and interviews. According to our hypothesis, 1) all stress scores describing subjective stress perception were significantly higher in patients compared to controls (P ≤ 0.001; P ≤ 0.002; P ≤ 0.001), 2) the L/L genotype of GT promoter repeats (L < 25 GT repeats, S < 25 GT repeats) in the patients was associated with higher overall (P ≤ 0.001), specific (P ≤ 0.010) and weighted stress score (P ≤ 0.005) compared to the L/S variant, and 3) Pearson's correlation of clinical versus objective stress scores showed not very tight relationship (0.198; 0.287; 0.224, respectively). We assume potential risk of the L allele of HO1 promoter polymorphism for the stress response and contribution of the subjective stress perception together with the L/L genotype to the development of eating disorder. Decreased HO1 expression in the presence of L/L genotype plus more intensive stress perception in the patients can lead to secondary stress, with increasing severity of the symptoms and aggravation of the disease.

Microsatellite polymorphism in haem oxygenase 1 gene promoter in multiple sclerosis.

Previous studies suggested that increased activity of haem oxygenase 1 may ameliorate autoimmune neuroinflammation in experimental models of multiple sclerosis. This increased activity is associated with an augmented number of GT repeats (≥ 25) within the HMOX1 gene promoter. Here we examined 338 patients with multiple sclerosis to determine the influence of their HMOX1 gene promoter (GT)n polymorphism and other individual characteristics on the course of the disease. The patients were divided into those with "rapid" or "delayed" course, based on reaching expanded disability status scale step 4 within nine years of disease onset, and the correlations between the disease course and the investigated characteristics were sought using logistic regression analysis. No statistically significant effect of HMOX1 gene promoter (GT)n polymorphism on the rate of disability progression was found (P = 0.9). This was confirmed by Cox regression analysis, which did not find any difference in the cumulative risk of reaching expanded disability status scale step 4 between the patients with long and short HMOX1 gene promoter (P = 0.7). In contrast, covariates significantly associated with the faster disability progression were: progressive course of multiple sclerosis, shorter duration of disease-modifying treatment and older age at disease onset (P ≤ 0.04). The observed absence of effect of the HMOX1 promoter (GT)n polymorphism could be attributed to its known dualistic role in the pathogenesis of autoimmune disorders. As a secondary outcome, we have seen that disease-modifying drugs have the potential to delay disability progression in patients with multiple sclerosis.

Genetic variants in haem oxygenase-1 and endothelial nitric oxide synthase influence the extent and evolution of coronary artery atherosclerosis.

The genetic basis for atherosclerosis development and progression is poorly characterized. We aimed to assess the relationship between endothelial nitric oxide synthase (ENOS) 894 G/T, haem oxygenase-1 (HO1) dinucleotide-length promoter polymorphisms and coronary artery atherosclerotic invol vement and its changes during statin therapy. Coronary angiography, intravascular ultrasound (IVUS), IVUS-derived virtual histology (VH) and genetic polymorphism analysis were performed at study entry. Patients were randomized 1:1 to standard or aggressive hypolipidaemic treatment, and a follow-up evaluation was performed after twelve months. Plaque magnitude was significantly higher in carriers of HO1 risk variants when compared with carriers of the protective variants (< 25 GT repeats). Similarly, the total coronary atherosclerotic burden was significantly greater in HO1 risk variant carriers than in HO1 protective variant carriers. Both parameters did not differ with respect to the ENOS genotype. A higher prevalence of thin-cap fibroatheroma (TCFA) in HO1 risk variant carriers was observed, compared with the HO1 protective variant carriers. The prevalence of TCFA was not influenced by the ENOS genotype. Baseline plaque composition did not differ significantly with respect to both polymorphisms. Significant interactions between plaque composition changes and ENOS and HO1 genotypes were observed during statin treatment. In conclusion, the protective HO1 promoter polymorphism correlates with a lower coronary artery plaque burden, whereas the protective ENOS 894 G/T polymorphism seems to favourably influence changes of coronary artery plaque composition during statin therapy, but has no significant correlation to the magnitude of coronary atherosclerosis.