High Frequency of Copy Number Variations and Sequence Variants at CYP21A2 Locus: Implication for the Genetic Diagnosis of 21-Hydroxylase Deficiency

BACKGROUND: The systematic study of the human genome indicates that the inter-individual variability is greater than expected and it is not only related to sequence polymorphisms but also to gene copy number variants (CNVs). Congenital Adrenal Hyperplasia due to 21-hydroxylase deficiency (21OHD) is the most common autosomal recessive disorder with a carrier frequency of 1:25 to 1:10. The gene that encodes 21-hydroxylase enzyme, CYP21A2, is considered to be one of the most polymorphic human genes. Copy number variations, such as deletions, which are severe mutations common in 21OHD patients, or gene duplications, which have been reported as rare events, have also been described. The correct characterization of 21OHD alleles is important for disease carrier detection and genetic counselling METHODOLOGY AND FINDINGS: CYP21A2 genotyping by sequencing has been performed in a random sample of the Spanish population, where 144 individuals recruited from university students and employees of the hospital were studied. The frequency of CYP21A2 mutated alleles in our sample was 15.3% (77.3% were mild mutations, 9% were severe mutations and 13.6% were novel variants). Gene dosage assessment was also performed when CYP21A2 gene duplication was suspected. This analysis showed that 7% of individuals bore a chromosome with a duplicated CYP21A2 gene, where one of the copies was mutated. CONCLUSIONS: As far as we know, the present study has shown the highest frequency of 21OHD carriers reported by a genotyping analysis. In addition, a high frequency of alleles with CYP21A2 duplications, which could be misinterpreted as 21OHD alleles, was found. Moreover, a high frequency of novel genetic variations with an unknown effect on 21-hydroxylase activity was also found. The high frequency of gene duplications, as well as novel variations, should be considered since they have an important involvement in carrier testing and genetic counseling

C4B null alleles are not associated with genetic polymorphisms in the adjacent gene CYP21A2 in autism

<p>Abstract</p> <p>Background</p> <p>Research indicates that the etiology of autism has a strong genetic component, yet so far the search for genes that contribute to the disorder, including several whole genome scans, has led to few consistent findings. However, three studies indicate that the complement <it>C4B </it>gene null allele (i.e. the missing or nonfunctional <it>C4B </it>gene) is significantly more frequent in individuals with autism. Due to the close proximity of the <it>CYP21A2 </it>gene to the <it>C4B </it>locus (3 kb) it was decided to examine samples from autistic subjects, including many with known <it>C4B </it>null alleles for common <it>CYP21A2 </it>mutations.</p> <p>Methods</p> <p>Samples from subjects diagnosed with autism and non-autistic controls (controls) previously typed for <it>C4B </it>null alleles were studied. Allele specific polymerase chain reaction (PCR) methods were used to determine 8 of the most common <it>CYP21A2 </it>genetic mutations, known to completely or partially inhibit 21-hydroxylase, the enzyme encoded by the <it>CYP21A2 </it>gene.</p> <p>Results</p> <p>Although the combined autism and control study subjects had 50 <it>C4B </it>null alleles only 15 <it>CYP21A2 </it>mutations were detected in over 2250 genotypes. Eight mutations were detected in the autistic samples and 7 in the controls. The frequency of <it>CYP21A2 </it>mutations was similar between the autism and control samples. Only one individual (autistic) carried a chromosome containing both <it>C4B </it>null allele and <it>CYP21A2 </it>mutations.</p

Common Genetic Variants of the Human Steroid 21-Hydroxylase Gene (CYP21A2) Are Related to Differences in Circulating Hormone Levels

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Hungarian Scientific Research Fund (OTKA, PD100648 (AP)) Technology Innovation Fund, National Developmental Agency (KTIA-AIK-2012-12-1-0010). AP is the recipient of a “Lendület” grant from the Hungarian Academy of Sciences

Glutathione enhances fibroblast collagen contraction and protects keratinocytes from apoptosis in hyperglycaemic culture

Background  Cutaneous wound healing is relatively slow in patients with diabetes. Objectives  To test the hypothesis that this defect in healing of wounds in patients with diabetes results from dysfunction of skin fibroblasts and epidermal keratinocytes and that this dysfunction is related to disrupted intracellular glutathione (GSH) homeostasis. Methods  We investigated the effects of esterified GSH on the contraction of fibroblasts in a fibroblast-populated collagen lattice and on keratinocyte apoptosis. Results  High glucose medium (hyperglycaemia) reduced the contraction ability of fibroblasts ( P <  0·05). The normalization of glucose medium concentrations for hyperglycaemic fibroblasts did not restore the contraction capacity. The percentage of apoptotic keratinocytes was statistically higher in hyperglycaemic cells ( P <  0·05). GSH media concentrations ranging from 0·1 to 100 µmol L −1 restored the ability of hyperglycaemic fibroblasts to contract the gels in a concentration-dependent manner. Primary human keratinocytes grown in hyperglycaemic medium were more susceptible to apoptosis, and treatment with esterified GSH rescued the keratinocytes from apoptosis. Conclusions  These data suggest that intracellular GSH can normalize skin cell functions disrupted by in vitro cell growth under hyperglycaemic conditions.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/74552/1/j.1365-2133.2004.06329.x.pd

Mutational spectrum of steroid 21-hydroxylase and the genotype-phenotype association in Middle European patients with congenital adrenal hyperplasia

OBJECTIVE: To analyze the mutational spectrum of steroid 21-hydroxylase (CYP21) and the genotype- phenotype correlation in patients with congenital adrenal hyperplasia (CAH) registered in the Middle European Society for Pediatric Endocrinology CAH database, and to design a reliable and rational approach for CYP21 mutation detection in Middle European populations. DESIGN AND METHODS: Molecular analysis of the CYP21 gene was performed in 432 CAH patients and 298 family members. Low-resolution genotyping was performed to detect the eight most common point mutations. High-resolution genotyping, including Southern blotting and sequencing was performed to detect CYP21 gene deletions, conversions, point mutations or other sequence changes. RESULTS: CYP21 gene deletion and In2 and Ile172Asn mutation accounted for 72.7% of the affected alleles in the whole study group. A good genotype-phenotype correlation was observed, with the exception of Ile172Asn and Pro30Leu mutations. In 37% of patients low resolution genotyping could not identify the causative mutation or distinguish homozygosity from hemizygosity. Using high-resolution genotyping, the causative mutations could be identified in 341 out of 348 analyzed patients. A novel mutation Gln315Stop was found in one simple virilising CAH (SV-CAH) patient from Austria. In the remaining seven patients polymorphisms were identified as the leading sequence alteration. The presence of elevated basal and ACTH-stimulated 17-hydroxyprogesterone, premature pubarche, advanced bone age and clitoral hypertrophy directly implicated Asn493Ser polymorphism in the manifestation of nonclassical- (NC) and even SV-CAH. CONCLUSIONS: By genotyping for the most common point mutations, CYP21 gene deletion/conversion and the 8 bp deletion in exon 3, it should be possible to identify the mutation in 94-99% of the diseased alleles in any investigated Middle European population. In patients with a mild form of the disease and no detectable mutation CYP21 gene polymorphisms should be considered as a plausible disease-causing mutation