R. Carmena

Large rearrangements of the LDL receptor gene and lipid profile in a FH Spanish population

Background Familial hypercholesterolemia (FH) is an autosomal dominant disease caused by mutations in the low‐density lipoprotein receptor (LDLR) gene. To date, there has not been a systematic survey of the frequency of gross mutations in the LDLR gene in the Spanish population. The objective of our study was to investigate large rearrangements in the Spanish FH population and the relation between the kind of large rearrangement and the phenotype in carrier families.

Circulating mononuclear cells nuclear factor-kappa B activity, plasma xanthine oxidase, and low grade inflammatory markers in adult patients with familial hypercholesterolaemia

Eur J Clin Invest 2010; 40 (2): 89–94

Misclassification of subjects with insulin resistance and associated cardiovascular risk factors by homeostasis model assessment index. Utility of a postprandial method based on oral glucose tolerance test.

Different methods are available for assessing insulin sensitivity in the fasting state. However, insulin resistance (IR) is initially a postprandial disturbance; and usually, when basal (fasting) disturbance appears, the process has been in progress for some time. Our aim was to investigate if a postprandial measurement, performing an oral glucose tolerance test (OGTT), is more sensitive than fasting values. We wished to identify early IR states in healthy, nonobese individuals and ascertain if this situation was associated with other cardiovascular risk factors. A total of 90 nonobese, nondiabetic, and nonsmoker individuals were studied. They were divided into 3 groups according to IR state--group 1: non-IR--homeostasis model assessment of IR (HOMA(IR)) and insulin sensitivity index of Matsuda-De Fronzo (ISI-Mat) were normal (HOMA(IR) <3.2 and ISI-Mat >4.0); group 2: with IR post-OGTT (ISI-Mat ≤4.0 and HOMA(IR) <3.2); and group 3: subjects with IR in basal conditions (HOMA(IR) ≥3.2). An intravenous glucose tolerance test to compare both indices was also performed. In 14.4% of subjects, the fasting HOMA(IR) values failed to identify IR (false-negative results). The ISI-Mat values were better correlated than HOMA(IR) (r = 0.875, P = .0001 and r = -0.631, P = .0001, respectively) with insulin sensitivity index obtained with intravenous glucose tolerance test. Subjects with IR had higher prevalence of a cluster of cardiovascular risk factors than non-IR subjects. These data show that that a significant percentage of subjects were misclassified with HOMA(IR). Early identification of IR by OGTT was associated with other cardiovascular risk factors. The OGTT is a simple method that could be applied to accurately identify IR subjects in the general population.

Increased oxidative stress levels and normal antioxidant enzyme activity in circulating mononuclear cells from patients of familial hypercholesterolemia

Familial hypercholesterolemia (FH) is a clinical condition with high risk for developing atherosclerosis. Increased oxidative stress (OS) and FH have been related to atherosclerosis, but no data are available on levels of OS and antioxidant enzyme activity in circulating mononuclear cells (CMCs) from FH patients. Circulating mononuclear cells are important mediators in atherosclerosis development, and chronically increased blood OS present in FH can induce modification in CMC activity. The objective of the study was to analyze the OS levels in CMCs from FH patients and controls. We have selected 30 nonrelated FH index patients and 30 normoglycemic and normocholesterolemic controls matched by age, sex, body mass index, abdominal circumference, and homeostasis model assessment index. Production of free radicals was analyzed by measurement of xanthine oxidase activity in plasma, reduced and oxidized glutathione (GSH and GSSG, respectively), and malonyldialdehyde in levels CMCs. Antioxidant status was analyzed by measuring antioxidant enzyme activity as superoxide dismutase, catalase, and glutathione peroxidase. We have found that FH patients showed significantly higher xanthine oxidase and malonyldialdehyde enzyme activities, as well as increased GSSG and lower GSH values resulting in a higher GSSG/GSH ratio. These data indicate a higher free radical production in plasma and increased OS levels in CMCs from patients than from controls. No significant differences were found in superoxide dismutase, catalase, and glutathione peroxidase activities between both groups. These data show an important alteration of OS regulation in FH and the absence of antioxidant response in CMCs mediated by some of the major antioxidant enzymes.

Reduced penetrance of autosomal dominant hypercholesterolemia in a high percentage of families: importance of genetic testing in the entire family.

BACKGROUND Autosomal dominant hypercholesterolemias (ADHs) are characterised by increased plasma levels of total and LDL cholesterol, predisposing to premature atherosclerosis. ADHs comprise several diseases with undistinguishable phenotype, caused by mutations in different genes: LDLR, APOB and PCSK9. Genetic studies are usually performed in patients with altered cholesterol levels. However, some persons carrying pathogenic mutations are normocholesterolemic and there are no further studies about this subject. We have studied the frequency of families and individuals carrying ADH mutations who do not present the disease in Spanish population. METHODS We have analysed genes known to cause ADH by direct sequencing in 24 ADH families (215 members). Functional effect of some LDLR gene mutations was assessed by transfecting cultured cells with plasmids. RESULTS Six families with mutations presented 7 mutation carriers who did not show ADH phenotype: 30% of ADH families presented normocholesterolemic individuals, and 7% of carriers of pathogenic mutations did not show ADH phenotype. We have analysed the effect of some of these mutations and they are responsible for impaired LDL receptor function. We have excluded mutations in APOB and PCSK9 genes that could reduce LDLc levels. CONCLUSIONS An important percentage of ADH families presented individuals who do not show an ADH phenotype, but who are able to transmit the pathogenic mutation to their offspring. Genetic study of all subjects in ADH families should be performed in order to identify normocholesterolemic carriers that allow the detection of mutations in their descendants and the prevention of the disease consequences.

Plasma homocysteine levels are associated with ulceration of the foot in patients with type 2 diabetes mellitus

To examine the association of biochemical markers of risk (plasma Hcy, microalbuminuria, lipoprotein (a)(Lp(a)) and diabetic dyslipidaemia) with the prevalence of diabetic foot ulceration in type 2 diabetic patients.

A New PCSK9 Gene Promoter Variant Affects Gene Expression and Causes Autosomal Dominant Hypercholesterolemia

CONTEXT Autosomal dominant hypercholesterolemia (ADH) is a genetic disorder characterized by increased low-density lipoprotein (LDL)-cholesterol levels, leading to high risk of premature cardiovascular disease. More than 900 mutations in LDL receptor, six in APOB and 10 in PCSK9 have been identified as a cause of the disease in different populations. All known mutations in PCSK9 causing hypercholesterolemia produce an increase in the enzymatic activity of this protease. Up to now, there are data about the implication of PCSK9 in ADH in a low number of populations, not including a Spanish population. OBJECTIVE The objective of the study was to study the prevalence of PCSK9 mutations in ADH Spanish population. PARTICIPANTS We screened PCSK9 gene in 42 independent ADH patients in whom mutations in LDL receptor and APOB genes had been excluded. RESULTS None of the known mutations causing ADH was detected in our sample, but we found two variations in the promoter region that could cause ADH, c.-288G>A and c.-332C>A (each in one proband). The analysis of the effect of these two variations on the transcription activity of the PCSK9 promoter showed that c.-288G>A did not modify the transcription, whereas c.-332C>A variant caused a 2.5-fold increase when compared with the wild-type sequence, either with or without lovastatin. CONCLUSIONS PCSK9 is a rare cause of ADH in Spanish population and, up to what we know, none of the previously described mutations has been detected. We have identified a new mutation that could cause ADH by increasing the transcription of PCSK9.

Association of selected ABC gene family single nucleotide polymorphisms with postprandial lipoproteins: Results from the population-based Hortega study

The aim of the study was to determine the influence of twenty single nucleotide polymorphisms (SNPs) of the ABCA1, ABCG1, ABCG5 and ABCG8 genes on the plasmatic concentrations of total cholesterol (TC), HDL and LDL cholesterol (HDLc, LDLc) in the postprandial state with a representative Spanish Caucasian population (1473 individuals, 50.0% women, ages ranging 21-85 years). In men, subjects with the AA genotype of the ABCA1 rs2230806 (R219K) polymorphism were associated with increased plasma LDLc levels, while the ABCA1 haplotype, which included the rs2230806 A allele, was associated with higher TC and LDLc plasma concentrations. In women, significant relationships were found between rs1893590 polymorphisms (ABCG1 gene) and HDLc plasma concentrations (subjects with the AA genotype had lower HDLc levels). For the ABCG8 gene, the rs4148211 polymorphism was associated with higher plasma TC and LDLc concentrations in the total population. Moreover, an ABCG5-G8 haplotype, which included the rs6544718 T allele, was associated with higher HDLc plasma concentrations in women. In conclusion, different SNPs of the ABCA1, ABCG1 and ABCG5-ABCG8 genes were associated, some under gender-specific analysis, with variations in the plasma lipid levels under postprandial conditions in a representative Spanish population.

Plasma homocysteine levels are independently associated with the severity of peripheral polyneuropathy in type 2 diabetic subjects

Peripheral polyneuropathy (PN) is a frequent complication of diabetes. However, mechanisms underlying the development of PN are multifactorial and not well understood. Our aim was to examine the association of plasma homocysteine (Hcy) with the prevalence and grade of peripheral PN in patients with type 2 diabetes (T2DM). We studied a cohort of 196 subjects with T2DM classified according to the grade of PN (Neuropathy Disability Score, NDS). Subjects with the highest grade of PN were older and had significantly increased levels of creatinine, microalbuminuria, HbA1c, and plasma Hcy compared to the other two groups. The differences in plasma Hcy values were maintained after correcting for confounding factors. Plasma Hcy, HbA1c, duration of diabetes, and age were predictors of the grade of PN. In conclusion, for each increase of 1 µmol in plasma Hcy there was a 23% increase of the risk of diabetic PN evaluated by NDS. Moreover, the grade of PN was predicted by plasma Hcy and HbA1c values, age and duration of diabetes. Further prospective studies should be conducted to confirm the association of plasma Hcy levels with the grade of PN in subjects with T2DM.

Dietary polyunsaturated fatty acids may increase plasma LDL-cholesterol and plasma cholesterol concentrations in carriers of an ABCG1 gene single nucleotide polymorphism: Study in two Spanish populations

BACKGROUND ABCG1 mediates cellular cholesterol transport, but there is very little known about the influence of ABCG1 polymorphisms on human plasma lipoprotein cholesterol concentrations or on the interactions of these polymorphisms with diet. OBJECTIVE Our objective was to investigate whether interactions between PUFA intake and ABCG1 polymorphisms modulate associations with plasma total cholesterol (TC), LDL- and HDL-cholesterol in two Spanish populations. METHODS We grounded our investigation on two general population-based studies: the Hortega study (population A) and the Pizarra study (population B). Participants included 1178 individuals (50.0% women, age range 21-85 years) and 763 individuals (66% women, age range 23-73 years) from populations A and B, respectively, without lipid lowering drugs. Subjects were genotyped for ABCG1 variants. Biochemical measurements were taken by standard procedures. Dietary intakes were estimated with a validated questionnaire. RESULTS In population A, the A allele homozygotes of SNP rs4148102 had higher TC and LDLc concentrations in subjects on a high PUFA diet than did the carriers of the G allele (242.1 ± 38.9 vs. 198.0 ± 36.0mg/dL, p = 0.003, and 149.8 ± 37.9 vs. 111.4 ± 32.1mg/dL, p = 0.005, respectively), and significant gene-diet interactions were observed (p=0.020 and p = 0.013, respectively). In population B, similar differences in TC and LDLc concentrations were also found in association with this SNP under a high PUFA diet (253.2±24.9 vs. 197.7 ± 39.9 mg/dL, p = 0.009, and 171.8 ± 20.5 vs. 120.4 ± 34.2mg/dL, p = 0.004, respectively), but the gene-diet interactions observed were not significant (p = 0.379 and p = 0.422, respectively). In the pooled populations, differences in the TC and LDLc concentrations increased (246.8 ± 32.9 vs. 198.0 ± 37.5, p = 6 × 10(-5), and 159.0±32.6 vs. 114.3 ± 33.1, p = 3 × 10(-5), respectively), and significant gene-diet interactions were maintained (p = 0.006 and p = 0.003, respectively). CONCLUSION In two Spanish populations, the ABCG1 polymorphism rs4148102 was associated with variations in plasma lipoprotein cholesterol concentrations in subjects with high PUFA intakes. Carriers of the AA genotype consuming high PUFA diet showed higher plasma LDLc concentrations.