21-hydroxylase Gene Research Articles

Functional characterisation of the H365Y mutation of the 21-hydroxylase gene in congenital adrenal hyperplasia

The study subject was a 13 day-old boy admitted to hospital, with weight loss since birth. He presented with the vomiting and hypotension that are classical features of congenital adrenal hyperplasia (CAH). The most common type of CAH is an autosomal recessive disorder caused by mutations in the 21-hydroxylase (CYP21A2) gene. To examine the CYP21A2 gene, gene-specific PCR was carried out, followed by sequencing. The baby was shown to be a compound heterozygote H365Y/R356W for two CYP21A2 gene mutations each inherited from a different parent. One of the mutations has not previously been functionally characterised. The mutations were reconstructed in an expression plasmid and characterised in vitro after transient transfection into human embryonic kidney (HEK293T) and hepatoblastoma (C3A) cell lines followed by measurement of enzyme activity. The CYP21A2 H365Y mutant exhibited minimal 21-hydroxylase activity to convert 17-hydroxyprogesterone to 11-deoxycortisol or progesterone to 11-deoxycorticosterone. Western immunoblotting indicated that the H365Y enzyme was produced in more variable amounts than wild type; in particular, the H365Y mutant protein may be unstable and/or subject to a more rapid degradation by the human proteosome as well as catalytically inefficient. The double mutant genotype with a severe mutation on each allele is compatible with the clinical presentation.

Chimeric CYP21A1P/ CYP21A2 genes identified in Czech patients with congenital adrenal hyperplasia

Congenital adrenal hyperplasia (CAH) comprises a group of autosomal recessive disorders caused by an enzymatic deficiency which impairs the biosynthesis of cortisol and, in the majority of severe cases, also the biosynthesis of aldosterone. Approximately 95% of all CAH cases are caused by mutations in the steroid 21-hydroxylase gene ( CYP21A2). The CYP21A2 gene and its inactive pseudogene ( CYP21A1P) are located within the HLA class III region of the major histocompatibility complex (MHC) locus on chromosome 6p21.3. In this study, we describe chimeric CYP21A1P/ CYP21A2 genes detected in our patients with 21-hydroxylase deficiency (21OHD). Chimeric CYP21A1P/ CYP21A2 genes were present in 171 out of 508 mutated CYP21A2 alleles (33.8%). We detected four types of chimeric CYP21A1P/ CYP21A2 genes: three of them have been described previously as CH-1, CH-3, CH-4, and one type is novel. The novel chimeric gene, termed CH-7, was detected in 21.4% of the mutant alleles. Possible causes of CYP21A1P/ CYP21A2 formation are associated with 1) high recombination rate in the MHC locus, 2) high recombination rate between highly homologous genes and pseudogenes in the CYP21 gene area, and 3) the existence of chi-like sequences and repetitive minisatellite consensus sequences in CYP21A2 and CYP21A1P which play a role in promoting genetic recombination.

Autopsy and genetic diagnosis of 21-hydroxylase deficiency with bilateral testicular tumors in a case under no medication for over one year

The autopsy findings of an adult patient with 21-hydroxylase deficiency are presented. Genetic analysis of the 21-hydroxylase gene ( CYP21A2) was performed for accurate diagnosis of congenital adrenal hyperplasia (CAH), and bilateral testicular tumors were characterized. We report a 29-year-old Japanese man who was diagnosed with CAH (21-hydroxylase deficiency) in infancy and had continued steroid therapy until the age of 28. However, for more than one year, he had not been treated for CAH and was found dead. In the medico-legal autopsy findings, both adrenal glands were enlarged, and hypertrophy of adrenal cortices and bilateral testicular tumors positive for melan-A were observed. Genomic DNA was prepared from cervical lymph nodes collected during autopsy, and CYP21A2 was PCR amplified and sequenced directly using newly designed primers. From the morphological findings, the bilateral testicular tumors were considered to be adrenogenital syndrome (TTAGS). Through the whole sequence of CYP21A2, the intron 2 splice mutation 656A to 656G was found. TTAGS were thought to be adrenal rests enlarged by ACTH stimulus. From the autopsy findings and the result of genetic analysis, he was diagnosed with the salt-wasting form of 21-hydroxylase deficiency and his cause of death was presumed to be heart failure based on abnormal electrolytes.

Metabolic disorders in newly diagnosed young adult female patients with simple virilizing 21-hydroxylase deficiency

Classical congenital adrenal hyperplasia (CAH) is characterized by the defects in cortisol and aldosterone secretion, and accompanied with adrenal hyperandrogenism. It is likely that the impaired adrenocortical function and intermittent treatment-related hypercortisolism may predispose patients to the development of metabolic syndrome in adulthood. Our aim was to assess the impact of hyperandrogenism on metabolic profiles in CAH women without glucocorticosteroid treatment. We evaluated the clinical characteristics and metabolic profiles in 30 untreated Chinese female adults with simple virilizing congenital adrenal hyperplasia (SV-CAH). Mutation analysis was performed by sequencing the entire 21-hydroxylase gene (CYP21A2). As compared with the controls, CAH patients had higher BMI (BMI, 21.5±2.1 vs. 20.0±1.8 kg/m2, P<0.05), higher 2 h post-load plasma glucose levels (6. 35±1.74 vs. 5. 35±1.17 mmol/l, P<0.05), higher serum triglycerides (TG) (1.12±0.64 vs. 0.63±0.15 mmol/l, P<0.01), and lower high-density lipoprotein cholesterol (HDL-c) (1.30±0.39 vs. 1.67±0.29 mmol/l, P<0.01). Moreover, CAH patients had higher fasting insulin and homeostasis model assessment of insulin resistance (HOMA-IR) (1.81±0.99 vs. 1.24±0.50, P<0.05), while ΔIns30/ΔGlu30 showed no statistically significant difference in two groups. In addition, a marked reduction of serum adiponectin levels were observed in CAH patients (7.0±3.3 vs. 13.2±4.8 μg/ml, P<0.001), however, serum CRP levels were not different between patients and the controls. Further regression analysis showed that higher serum testosterone concentrations were associated with metabolic disorder indexes and reduction of serum adiponectin. Our study demonstrates that untreated CAH patients are prone to have metabolic disorders in association with elevated serum testosterone levels and reduced insulin insensitivity.

Identification of CYP21A2 mutant alleles in Czech patients with 21-hydroxylase deficiency

Congenital adrenal hyperplasia (CAH) is comprised of a group of autosomal recessive disorders caused by an enzymatic deficiency which impairs the biosynthesis of cortisol and, in most of the severe cases, also the biosynthesis of aldosterone. Approximately 90-95% of all the CAH cases are due to mutations in the steroid 21-hydroxylase gene (CYP21A2). In this study, the molecular genetic analysis of CYP21A2 was performed in 267 Czech probands suspected of 21-hydroxylase deficiency (21OHD). 21OHD was confirmed in 241 probands (2 mutations were detected). In 26 probands, a mutation was found only in 1 CYP21A2 allele. A set of 30 different mutant alleles was determined. We describe i) mutated CYP21A2 alleles carrying novel point mutations (p.Thr168Asn, p.Ser169X and p.Pro386Arg), ii) mutated CYP21A2 alleles carrying the novel chimeric gene designated as CH-7, which was detected in 21.4% of the mutant alleles, iii) an unusual genotype with a combination of the CYP21A2 duplication, 2 point mutations and the CYP21A2 large-scale gene conversion on the second allele, and (iv) a detailed analysis of the chimeric CYP21A1P/CYP21A2 genes. In conclusion, our genotyping approach allowed for the accurate identification of the CYP21A2 gene mutations in 21OHD patients and their families and provided some useful information on diagnosis and genetic counselling.

Correlation between genotype and hormonal levels in heterozygous mutation carriers and non-carriers of 21-hydroxylase deficiency.

Congenital adrenal hyperplasia, both in its classic (CCAH) and non-classic form (NCAH), is a morbid condition sustained by the absent or reduced function of one of the enzymes involved in cortisol biosynthesis - mainly 21 hydroxylase - associated with different levels of clinical androgenization. In a wide group of relatives of patients affected by CCAH and NCAH (no.=222) and healthy volunteers (no.=30), a clinical, hormonal and genetic evaluation was performed in order to differentiate between the condition of heterozygous mutation carrier and non-carrier of any among 21-hydroxylase gene (CYP21) mutations. This study shows that clinical presentation and basal 17α-hydroxyprogesterone (17α-OHP) are not able to differentiate between heterozygous carriers and non-carriers, whereas 17α-OHP value after ACTH bolus is significantly different between heterozygous carriers and non-carriers: p<0.001 with a cut-off value of 3 ng/ml (90% sensitivity and 74,3% specificity). Moreover, our data indicate that 17α-OHP response to ACTH may be a useful tool to select subjects for genetic analysis.

Reversal of Primary Male Infertility and Testicular Adrenal Rest Tumors in Salt-Wasting Congenital Adrenal Hyperplasia

A26-yr old male with a salt-wasting form of congenital adrenal hyperplasia presented with a 4-yr history of primary infertility. He had done well on deflazacort (range 6–18 mg/d) and fludrocortisone [0.05 mg twice a day (bid)] until late adolescence when he had several Addison’s crises due to decreased therapeutic adherence. He was lost to follow-up for several years before infertility work-up. LH and FSH were undetectable, whereas serum testosterone level was 37.4 nmol/liter normal (N): 8–26 , ACTH 501 ng/liter (N: 10 – 60), and 17-hydroxyprogesterone 1047 nmol/liter (N: 1.8 –9.2). Computed tomography scan showed massive bilateral adrenal hyperplasia (Fig. 1), and testicular ultrasonography multiple bilateral testicular adrenal rest tumors (TARTs). Semen analysis showed azoospermia. Congenital adrenal hyperplasia was shown to be due to a missense mutation in exon 8 of the 21hydroxylase gene. In response to dexamethasone (0.5 mg bid) and fludrocortisone (0.05 mg bid), ACTH and 17-hydroxyprogesterone levels fell (8 ng/liter and 2.5nmol/liter, respectively), whereas LH (4.2 U/liter) and FSH (9.3 U/liter) rose, followed by spontaneous conception. The patient’s wife delivered a healthy term male baby 12 months after the change of replacement therapy. However, repeat semen analysis and paternity confirmation were not performed. Repeat imaging studies after 13 months of treatment disclosed dramatic reduction of adrenal hyperplasia and the disappearance of TARTs. These data demonstrate the rapid reversibility of TARTs using dexamethasone as first described by Cunnah et al. (1) without side effects in contrast to previous reports (2). The chronological association of TART disappearance with pregnancy suggests that mechanical obstruction on seminal tubules might have caused this patient’s infertility, although gonadal axis suppression may also have contributed to infertility, as discussed recently (3, 4).

Refractory Acne and 21-Hydroxylase Deficiency in a Selected Group of Female Patients

Background: Excessive androgen production, suspected in women when acne is accompanied by hirsutism and menstrual irregularities, may be due to congenital adrenal hyperplasia. This inherited disorder of cortisol biosynthesis is caused in more than 90–95% of all cases by 21-hydroxylase deficiency (21-OHD). The steroid 21-hydroxylase gene (CYP21) has a high degree of variability. Objective: This study was conducted to evaluate CYP21 gene mutations in a selected group of women with papulopustular and comedonal acne refractory to treatment, irregular menses and hirsutism. Methods: 30 out of 61 women enrolled underwent pelvic ultrasound examination and hormonal screening. In 9 patients with a polycystic ovary and hormonal pattern of adrenal hyperandrogenism a significant elevation of adrenocorticotropic hormone (ACTH) stimulated 17-hydroxyprogesterone was detected. These women positive in the ACTH stimulation test were submitted to CYP21 gene analysis. Results: Genetic testing revealed several different point mutations and demonstrated that a cohort of patients resistant to acne therapy can be carriers or affected by non-classical 21-OHD (late onset). Conclusion: Persistent acne can be the unique presenting sign of non-classical 21-OHD. Evaluation of CYP21 gene mutations may identify female carriers or patients for genetic counselling.

Intron 2 Splice Mutation at CYP21 Gene in Patients with Congenital Adrenal Hyperplasia in the Republic of Macedonia

Intron 2 Splice Mutation at CYP21 Gene in Patients with Congenital Adrenal Hyperplasia in the Republic of MacedoniaCongenital adrenal hyperplasia (CAH) is an autosomal recessive disorder. In 90-95% of cases it results from mutations in the gene for 21-hydroxylase (CYP21, also termed CYP21A2 and P450c21). The IVS-II-656 (C/A&gt;G) mutation leaves ~2.0% enzyme activity, and comprises 25% of the classic CYP21 deficiency alleles and 51% of alleles in the salt-wasting form.We performed direct molecular diagnosis of the IVS-II mutation in 41 Macedonian patients with different clinical forms of CAH and 55 of their healthy parents and siblings from 37 unrelated families, using the differential polymerase chain reaction/amplification created restriction site method (PCR/ACRS). The IVS-II mutation was detected in 41.5% patients (29.3% were homozygotes and 12.2% were heterozygotes). All homozygotes had a severe classical CAH phenotype (of which 91.7% were salt-wasting and 8.3% were simple virilizing). Three of the heterozygotes had a salt-wasting (SW) phenotype and were compound heterozygotes. The IVS-II mutation was also found in 30.9% of the family members (18.2% were homozygous and 12.7% were heterozygous) and none had any clinical manifestation. The frequency of the IVS-II mutation (41.5%) in these subjects was similar to that reported elsewhere.

A p.P30L Mutation at the CYP21A2 Gene in Macedonian Patients with Nonclassical Congenital Adrenal Hyperplasia

A p.P30L Mutation at the CYP21A2 Gene in Macedonian Patients with Nonclassical Congenital Adrenal HyperplasiaNonclassical congenital adrenal hyperplasia (NCAH) is an autosomal recessive imbalance in cortisol synthesis with adrenal androgen excess. Although rarely recognized in infants, it may cause premature adrenarche and pubarche, virilization in young women and variable symptoms in young men. It is commonly caused by mutations in CYP21A2, the gene for steroid 21-hydroxylase. Patients with the p.P30L allele tend to have pronounced evidence of androgen excess but are categorized as nonclassical. We used direct molecular detection of the p.P30L mutation in CYP21A2 in 11 Macedonian NCAH patients and in 17 members of their families using polymerase chain reaction/amplification created restriction site (PCR/ACRS) analysis and digestion with restriction enzymes. The p.P30L mutation was found in a homozygous state in seven (63.6%) and in a heterozygous state in four (36.4%) patients. Of the latter, one was also heterozygous for the p.Q318X mutation. The p.P30L mutation was found in a heterozygous state in 10 (58.8%) and in a homozygous state in one (5.9%) of the family members. These findings support a role of the p.P30L mutation in NCAH.

Congenital adrenal hyperplasia and the function of adrenal medulla.

The most common form of congenital adrenal hyperplasia (CAH) is the deficiency of steroid 21-hydroxylase which results from deletion or mutation of the cytochrome P450 21-hydroxylase gene. The low level of glucocorticoids and in some cases low level of mineralocorticoids has an important pathophysiological influence on the axis of the hypothalamo-pituitary-adrenal cortex. Using determination of plasmatic metanephrine, normetanephrine and chromogranin A, we wanted to investigate the structure and function of adrenal medulla in patients with CAH, because adrenocortical and adrenomedullary systems are intimately linked anatomically and functionally. Levels of plasmatic metanephrine, normetanephrine and chromogranin A were measured in our group of 37 patients (age range: 5-45years, 18 females and 19 males) with the classic salt-wasting form of CAH. The reference range was 73% for metanephrine (<10ng/L, 83.3% females, 63.2% males) and 59.5% for metanephrine (<15ng/L, 72.2% females, 47.4% males). The concentration of plasmatic nephrines in the first quartile reference range was achieved in the case of metanephrine in all patients (<23ng/L), and in the case of normetanephrine in 86.5% of patients (<42.5ng/L). The level in chromogranin A was normal in all patients. No significant differences were found in plasmatic concentrations of nephrines and chromogranin A between males and females with CAH, nor was there a significant correlation with genetic results (severe or moderate salt-wasting form of CAH). Impaired secretion of glucocorticoids in patients with CAH leads to the structural changes in adrenal medulla which are expressed by low production of metanephrine, and to a lesser extent, normetanephrine.

Mutations of the Steroid 21-Hydroxylase gene among Filipino Patients with Congenital Adrenal Hyperplasia

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Mutational Characterization of Steroid 21-Hydroxylase Gene in Portuguese patients with Congenital Adrenal Hyperplasia

Congenital adrenal hyperplasia (CAH) due to steroid 21-hydroxylase deficiency is a common inherited disorder of adrenal hormone biosynthesis due to mutations in the 21-hydroxylase gene, CYP21A2. Genotyping for ten of the most frequent mutations was performed in 84 Portuguese CAH patients: 10 salt-wasters, 6 simple-virilizers and 68 non-classical patients. The patients were diagnosed by a level of 17-hydroxyprogesterone above 10 ng/ml either in basal conditions or after an ACTH 0,25 mg IV Test. A variety of genotyping techniques were used to detect these ten mutations. CYP21A2 mutations were detected in 91.7% (77/84) of the patients. The frequency of alleles carrying two or more CYP21A2 mutations (9.5% - 16/168) is higher than in other populations. The most frequent mutations identified in our population were V281L (41.7%) and deletions/conversions involving the promoter region of the CYP21A2 gene (28.3%). A decreased frequency of IVS2-12C/A>G mutation (5.6%) was the most characteristic feature of our population. This study allow the characterization of the mutational spectrum of CAH patients, mainly non-classical CAH, with 21-hydroxylase deficiency from Portugal showing specific genetic features of this population which reveals differences with worldwide countries.

Síndrome suprarrenogenital congénito virilizante con mutación de novo I172N: estudio de un nuevo caso

The classical form of congenital adrenal hyperplasia is the result of mutations in the 21-hydroxylase gene ( CYP21A2). Most deficient alleles carry pre-existing mutations in the CYP21PA homologue pseudogene, located in tandem. Mutant alleles are inherited from carrier parents, and de novo mutations during gametogenesis or foetal development are exceptional. The present paper describes a de novo mutation occurring at the maternal allele (I172N) of a patient with a classical form of 21-hydroxylase deficiency, whose father was heterozygous for R356W. The mother did not carry the mutation. Microsatellite analyses confirmed a correct allelic segregation. The I172N mutation (in compound heterozygosity with a null mutation) gives rise to a virilizing phenotype not associated with salt-wasting.

Functional analysis of two rare CYP21A2 mutations detected in Italian patients with a mildest form of congenital adrenal hyperplasia

More than 90% of all cases of congenital adrenal hyperplasia (CAH) result from steroid 21-hydroxylase gene (CYP21A2) mutations. Most of these mutations result from intergenic recombinations between CYP21A2 and closely linked CYP21A1P pseudogene. Rare mutations not generated by gene conversion account for 5-10% of 21-hydroxylase deficiency alleles. Functional analysis of two novel CYP21A2 missense mutations (p.R224W and p.D407N) was performed. Our study was composed of two Italian patients suffering from a very mild form of nonclassic CAH (NC-CAH). To assay the enzymatic activity of mutants, the in vitro analysis was performed in transiently transfected COS-1 cells. The residual activities obtained for p.R224W and p.D407N mutants allow their classification as NC-CAH mutations. These results correlate with the rate of severity of the patients' disease. In this paper, we report two novel CYP21A2 mutations in two Italian individuals affected by 21-hydroxylase deficiency. Based on the functional in vitro analysis we can classify these mutations as NC-CAH variants.

A new CYP21A1P/CYP21A2chimeric gene identified in an Italian woman suffering from classical congenital adrenal hyperplasia form

BackgroundMore than 90% of Congenital Adrenal Hyperplasia (CAH) cases are associated with mutations in the 21-hydroxylase gene (CYP21A2) in the HLA class III area on the short arm of chromosome 6p21.3. In this region, a 30 kb deletion produces a non functional chimeric gene with its 5' and 3' ends corresponding to CYP21A1P pseudogene and CYP21A2, respectively. To date, five different CYP21A1P/CYP21A2 chimeric genes have been found and characterized in recent studies. In this paper, we describe a new CYP21A1P/CYP21A2 chimera (CH-6) found in an Italian CAH patient.MethodsSouthern blot analysis and CYP21A2 sequencing were performed on the patient. In addition, in order to isolate the new CH-6 chimeric gene, two different strategies were used.ResultsThe CYP21A2 sequencing analysis showed that the patient was homozygote for the g.655C/A>G mutation and heterozygote for the p.P30L missense mutation. In addition, the promoter sequence revealed the presence, in heterozygosis, of 13 SNPs generally produced by microconversion events between gene and pseudogene. Southern blot analysis showed that the woman was heterozygote for the classic 30-kb deletion producing a new CYP21A1P/CYP21A2 chimeric gene (CH-6). The hybrid junction site was located between the end of intron 2 pseudogene, after the g.656C/A>G mutation, and the beginning of exon 3, before the 8 bp deletion. Consequently, CH-6 carries three mutations: the weak pseudogene promoter region, the p.P30L and the g.655C/A>G splice mutation.ConclusionWe describe a new CYP21A1P/CYP21A2 chimera (CH-6), associated with the HLA-B15, DR13 haplotype, in a young Italian CAH patient.

Expressional dynamics of minisatellite 33.15 tagged spermatozoal transcriptome in Bubalus bubalis

BackgroundTranscriptionally quiescent spermatozoa have been established to be a repository of mRNA coding for several functionally essential cellular proteins. This entourage of mRNA is envisaged to be involved in post-fertilization and early embryogenesis. Minisatellites tagged with mRNA transcripts have been implicated with gene organization, regulation and function. However, the organization and expression of the minisatellite tagged transcript diversity, particularly in spermatozoa, remains unclear.ResultsIn the present study, we identified and characterized 12 mRNA transcripts from the spermatozoa of water buffalo Bubalus bubalis employing minisatellite associated sequence amplification (MASA) and a consensus sequence of 33.15 repeat loci. Of these 33.15 tagged transcripts, only one was found to be homologous to Bovine steroid 21-hydroxylase (P-450-c21) gene. Other ten transcripts showed significant similarity with various mRNAs or chromosomal contigs across the species. The remaining one construed to be novel since this was unreported in the database (NCBI GenBank). All these uncharacterized and known transcripts showed highest expression in testis and spermatozoa compared to that in somatic tissues and ovary. Of these 12 mRNA transcripts, 4 showed differential expression in the forebrain and hindbrain of buffalo. Moreover, genes corresponding to all the 33.15 tagged spermatozoal transcripts were found to be conserved across 13 other species analyzed.ConclusionOur results show MASA as an important tool to capture mRNA transcript diversity tagged with minisatellites in the spermatozoa. Comprehensive characterization of these transcripts is envisaged to augment our understanding on the genes involved in testicular functions and sustenance of a viable paternal genome during pre- and post- fertilization events and early stages of development. Prospects of this approach in genome analysis in general and comparative genomics in particular are highlighted.

Linkage analysis of the C4A/ C4B copy number variation and polymorphisms of the adjacent steroid 21-hydroxylase gene in a healthy population

Genes encoding the steroid 21-hydroxylase ( CYP21A2) and the complement component C4 proteins ( C4A and C4B) are located in the MHC region in a strongly linked structure named RCCX module. Previous studies found that carriers of C4B gene deficiency ( C4B*Q0) have higher risk for cardiovascular diseases. A potential explanation is that lacking the C4B gene may result in altered function of the neighboring CYP21A2 gene. Therefore we sequenced the CYP21A2 gene in 96 healthy individuals to identify polymorphisms and to characterize their linkage pattern. Fifty-three variations were detected including a new one which alters the TATA-box of the gene. Only three known mutations (V281L, Q318X and R479L) associated with congenital adrenal hyperplasia, were found in 7, 2 and 1 subjects, respectively. Linkage analysis revealed that some variations exhibit strong correlation with the C4 copy number polymorphism and constituents of the MHC III region. Rare alleles of three polymorphisms were identified as components of the 8.1 ancestral haplotype. Haplotyping and family study confirmed that the variant alleles of two intronic SNPs were constituents of haplotype blocks lacking the C4B gene. These results suggest that variations of CYP21A2 gene can be involved in disease associations of the 8.1 haplotype and the C4B*Q0 genotype.

Transient hyper-17-hydroxyprogesteronemia: a clinical subgroup of patients diagnosed at neonatal screening for congenital adrenal hyperplasia

Neonatal screening for congenital adrenal hyperplasia (CAH) is characterized by a high false-positive rate, mainly among preterm and low birth weight infants. The aims of this study were to describe a subgroup of infants with transient serum hyper-17-hydroxyprogesteronemia (hyper-17-OHPemia) and to compare them with false positive and affected by 21-hydroxylase deficiency newborns. We retrospectively analyzed the clinical data of all newborns positive at CAH neonatal screening, who were referred to our hospital to confirm the diagnosis from 2002 to 2006. They were submitted to clinical investigations and blood tests to evaluate 17-hydroxyprogesterone (17-OHP), renin, and electrolyte levels. CAH-unaffected newborns with increased serum 17-OHP were submitted to strict follow-up monitoring, which included an ACTH-stimulating test and genetic analysis of the 21-hydroxylase gene, until serum 17-OHP decreased. Thirty-seven newborns with gestational ages ranging from 33 to 40 weeks were studied. Eight infants (three male and five female) were affected by CAH (serum 17-OHP: 277.5 (210-921) nmol/l), 14 (ten male and four female) were false positives (17-OHP: 3.75 (0.3-8.4) nmol/l), and 15 (ten male and five female) showed a serum hyper-17-OHPemia (17-OHP: 15.9 (9.9-33) nmol/l). No mutations of the 21-hydroxylase gene were found in infants with hyper-17-OHPemia and their serum 17-OHP levels were normalized by the third month of life. We identified a population of infants with transient serum hyper-17-OHPemia, and no clinical signs of disease or 21-hydroxylase gene mutations. No further investigations are necessary after birth in these newborns if 17-OHP levels decrease, other confirmatory tests such as ACTH-stimulation test or genotyping analysis are necessary only if symptoms appear.

Funcriohal analysis of novel mutations P459H and R483W in 21-hydroxylase gene

Mutations P459H and R483W detected in CYP21A2 gene in two Chinese patients with simple virilizing 21-hydroxylase deficiency were studied.Plasmid vectors containing P459H and R483W were constructed and transfected into COS-7 cells.The converting rate of progesterone to 11-desoxycortisone was calculated.P459H reduce 21-hydroxylase activity to 6.8%,while the residual enzyme activity of R483W was only 2.9%. Key words: Congenital adrenal hyperplasia; CYP2 1 gene; Mutation; Enzyme assay