Neonatal screening for congenital adrenal hyperplasia in North-Eastern Italy: a report three years into the program

AIMS: To evaluate the incidence of congenital adrenal hyperplasia (CAH) in the Northern Italian population and the efficiency of the North-Eastern Italy screening program. To adjust cut-off levels for 17-hydroxyprogesterone (17-OHP) in relation to gestational age and birth weight, comparing the benefits in terms of reduction of recall rates with the two approaches and ultimately choosing the better of the two. SUBJECTS AND METHODS: Since September 2001, blood samples from neonates born in North-Eastern Italy have been screened with a fluoroimmunoassay method for 17-OHP determination (DELFIA). A preliminary cut-off level of > or = 30 nmol/l was set both for term and preterm newborns. The values of 17-OHP were analysed using statistical methods in relation to gestational age and birth weight in order to modify the cut-off on the basis of our data. RESULTS: After 33 months of screening we screened 128,282 newborns and detected 6 affected babies. During the first 8 months of screening among the recalled babies, 89.6 and 78.1% were preterm and low-birth-weight newborns, respectively, with a recall rate of 2.59% for premature neonates and of 4.94% for babies with birth weights < 2,500 g. We chose a new cut-off value of 50 nmol/l for preterm newborns only and, after 4 months, the recall rate was reduced to 0.83% for these infants and to 1.83% for low-birth-weight infants. CONCLUSION: After 33 months of screening for CAH in North-Eastern Italy, we report an incidence of 1:21,380. In 5 out of 6 affected babies, the diagnosis was established only after a positive screening test, which prevented a severe salt-wasting crisis in these babies. The cut-off level related to gestational age led to a significant reduction in the number of false-positives among preterm babies.We therefore intend to continue with the screening program for CAH in North-Eastern Italy, keeping a gestational-age-related cut-off in the hope that our data may encourage a national screening program for CAH.     

Growth patterns and final height in congenital adrenal hyperplasia due to classical 21-hydroxylase deficiency. Results of a multicenter study

BACKGROUND: Longitudinal growth and bone age (BA) development are the most important clinical parameters for monitoring adequate glucocorticoid replacement in children with congenital adrenal hyperplasia (CAH). AIM OF THE STUDY: To analyze the growth pattern of patients treated for CAH of the salt wasting (SW) and simple virilizing (SV) clinical forms; to evaluate final height as compared to reference data and individual target height; to evaluate the course of BA development. PATIENTS AND METHODS: A large database of 598 patients with CAH was created in 5 Central European countries and growth data of 341 treated patients with 21-hydroxylase deficiency were analyzed retrospectively. The patients were of Caucasian origin. Centiles were constructed in a cross-sectional manner and an additional longitudinal analysis was performed in order to evaluate the pubertal growth spurt by applying particular statistical methods (Preece-Baines model). RESULTS: The growth of SW CAH patients was impaired in infancy and early childhood (0-3 years of age), but followed normal patterns in childhood until puberty. In contrast, children with SV CAH had normal patterns of growth in infancy and early childhood and were considerably taller than healthy references during childhood. In the longitudinal study, peak height velocity in both boys and girls was normal, but it occurred at an earlier age than in the standard population. The final height of patients with CAH was reduced in comparison to both the reference and the individual target height. No correlations were found between final height and age at the start of the therapy in SV patients or between final height and year of birth. BA was advanced in both types of CAH, but more accelerated in SV patients. CONCLUSION: Characteristic growth patterns for treated SV and SW CAH children were identified, with a normal pubertal growth spurt and reduced final height being observed.     

Prevalence and Characteristics of Adrenal Tumors and Myelolipomas in Congenital Adrenal Hyperplasia: A Systematic Review and Meta-Analysis

Objective: The prevalence of adrenal tumors in congenital adrenal hyperplasia (CAH) is uncertain. Our objective was to estimate the prevalence and characteristics of adrenal tumors and myelolipoma in CAH, and investigate clinical features of this population.Methods: We carried out systematic searches in Medline Ovid and Embase for articles published until January, 2020. Studies with confirmed CAH, biochemically and/or genetically, were included. The two authors independently extracted data from each study.Results: Six cohort studies were included in the prevalence calculation. In addition, 32 case reports on adrenal myelolipomas and CAH were included. The prevalence of adrenal tumors in CAH was 29.3%. When only studies with genetically verified cytochrome P450, Family 21, subfamily A, polypeptide 2 gene (CYP21A2) mutations were included the prevalence was 23.6%. The prevalence of myelolipoma in CAH was 7.4% (verified CYP21A2 mutations 8.6%). The proportion of myelolipoma in the adrenal tumors was 25.4% (genetically verified 36.6%). The median (range) age at tumor diagnosis was 36.0 (12 to 60) years and there were more tumors in males than in females (37.9% versus 22.1%; P<.05). In patients with myelolipomas, 93.5% had an undiagnosed or poorly managed CAH.Conclusion: Patients with CAH had a high prevalence of adrenal tumors, particularly myelolipomas. Those with myelolipomas had a high frequency of late-diagnosed or poorly controlled CAH. Adrenal imaging may be considered in patients with CAH, especially if abdominal pain is present.     

Masculinized finger length patterns in human males and females with congenital adrenal hyperplasia

The ratio of the length of the second digit (2D) to the length of the fourth digit (4D) is greater in women than in men. Since androgens are involved in most somatic sex differences and since the sexual dimorphism in 2D:4D is stable from 2 years of age in humans, it was hypothesized that finger length pattern development might be affected by early androgen exposure. Human females with congenital adrenal hyperplasia (CAH) are exposed prenatally to higher than normal levels of adrenal androgens, providing an opportunity to test the effects of early androgen exposure on digit ratios. The 2D:4D was calculated for females with CAH, females without CAH, males with CAH, and males without CAH. Females with CAH had a significantly smaller 2D:4D on the right hand than did females without CAH. Males with CAH had a significantly smaller 2D:4D on the left hand than did males without CAH. A subset of six males with CAH had a significantly smaller 2D:4D on both hands compared with their male relatives without CAH. These results are consistent with the idea that prenatal androgen exposure reduces the 2D:4D and plays a role in the establishment of the sex difference in human finger length patterns. Finger lengths may therefore offer a retrospective marker of perinatal androgen exposure in humans.     

How many deaths can be prevented by newborn screening for congenital adrenal hyperplasia?

BACKGROUND/AIMS: Congenital adrenal hyperplasia (CAH) is increasingly being included in newborn screening programs. Screening can prevent neonatal mortality in children with salt-wasting CAH, but the number of deaths prevented is not known. Cost-effectiveness analyses of screening require estimates of the probability of mortality in CAH. METHODS: We reviewed the literature to identify cohort studies of children with CAH ascertained clinically in the absence of screening. We abstracted the numbers of infant deaths attributable to CAH. We also addressed sex ratios among children with clinically detected CAH and the contribution of ascertainment bias to unbalanced ratios. RESULTS: The evidence suggests a probability of infant death due to adrenal crises in salt-wasting CAH of 4% or less in contemporary advanced economies without screening for CAH. This is lower than previous estimates, although the rate of mortality could be considerably higher in populations with limited clinical awareness or access. CONCLUSION: Although screening for CAH is conducted in a number of countries, further research is still needed to provide reliable estimates on the numbers of prevented deaths, along with evidence-based assessments of the potential benefits, harms, and costs of screening.     

Prevalence of intranuclear particles in liver pathologies

An ultrastructural study of the prevalence of electron dense 23-27 nm intranuclear particles was carried out on liver biopsies from patients with NANB chronic active hepatitis (CAH), Delta + CAH, HBsAg + CAH, nonviral liver pathologies and in one healthy volunteer. The particles were classified according to aggregation pattern and were found to be correlated with NANB CAH and Delta + CAH. No particles were observed in nonviral liver pathologies. A close antigenic relationship has been shown between the cytoplasmic alterations observed in NANB and delta hepatitis in chimpanzees. Our data indicate that there is a structural similarity between the intranuclear particles seen in both Delta and NANB hepatitis, thus reinforcing the hypothesis that the NANB and Delta agents are closely related.     

Analysis of the spectra of mutational damage of the 21-hydroxylase gene in patients with adreno-genital syndrome

The spectrum of mutations in the steroid 21-hydroxylase gene (CYP21B) and the frequency of 11 mutations among 66 patients with different forms of congenital adrenal hyperplasia (CAH) were analyzed by means of PCR amplification. Each of the CAH forms was characterized by specific spectrum of diagnostically important mutations. The salt-losing (SL) form of the disease was most frequently associated with gene deletion (39%) and the 668-13C-G mutation in the second intron (23.5%), whereas the majority of simple virilizing (SV) CAH cases were associated with the 1172N mutation in exon 4 (22%), gene deletion (16.5%), and the 668-13C-G mutation (16.5%). Mutations in the steroid 21-hydroxylase gene were detected in 70% of the chromosomes from the patients with the SL and SV forms of CAH, and only in 1.3% of the chromosomes from the patients with the nonclassic (NC) form. A total of 78 mutant chromosomes from the NC CAH patients were examined, and only one case of a gene deletion in the heterozygous state was revealed. In the individuals examined, the V281L and P30L mutations described in the NC CAH patients from other populations were not detected. This result can be explained either by the fact that NC CAH cases in Russia are associated with other major mutations, or by difficult clinical diagnosis questionable CAH cases.     

Congenital Adrenal Hyperplasia Due to 21-Hydroxylase Deficiency Presenting as Adrenal Incidentaloma: A Systematic Review and Meta-Analysis

Objective: Adrenal incidentalomas (AIs) may be due to congenital adrenal hyperplasia (CAH) due to homozygous CYP21A2 mutations, or perhaps from heterozygous carrier status. It is unclear if genetic or biochemical testing of CYP21A2 status in AI is justified, despite its potential for avoiding adrenal crises in those referred for adrenalectomy.Methods: We systematically searched PubMed/MEDLINE for articles published up to October 19, 2015 containing all terms associated with adrenal tumors and CAH. Meta-analyses were used to estimate the CAH or carrier prevalence in AI and assess clinical factors that may guide testing.Results: Thirty-six publications were included. Of AI patients biochemically screened for CAH, 58/990 (5.9%) were diagnosed with CAH. Genetic screening of all AIs revealed only 2/252 (0.8%) with clear CAH. The carrier prevalence was 10.2% (36/352). The rate of 0.8% (8/1,000) genetically confirmed CAH is higher than the 1/15,000 affected by classic CAH or 1/1,000 by nonclassic CAH in the Caucasian population. The rate of heterozygous CYP21A2 mutation frequency is similar to those in reported in population studies. Levels of both basal and stimulated 17-hydroxyprogesterone positively correlated with AI diameter. Although bilateral incidentalomata were frequent in CAH, their presence did not predict CYP21A2 status.Conclusion: The presence of an AI does not increase the probability of detection of CAH or CYP21A2 carrier status to the extent routine genetic testing is justified. Screening with 17-hydroxyprogesterone levels appears to lack specificity in the setting of an AI. CYP21A2 mutation analysis is probably the only reliable method for CAH diagnosis in AIs.Abbreviations:ACC = adrenocortical carcinomaACTH = adrenocorticotropic hormoneAI = adrenal incidentalomaCAH = congenital adrenal hyperplasiaNCAH = nonclassic congenital adrenal hyperplasia17OHP = 17-hydroxyprogesteroneSV = simple virilizing     

A new DNA diagnostic system for the detection of human <Emphasis Type="Italic">CYP21</Emphasis> gene mutations associated with adrenal cortex hyperplasia

Congenital Adrenal Hyperplasia (CAH) is one of the most widespread severe autosomal recessive hereditary diseases. CAH is caused by the impaired biosynthesis of the key human hormones cortisol and aldosterone and is accompanied by the excess synthesis of androgens. Over 90% of CAH cases are caused by a deficiency of the steroid 21-hydrohylase (P450c21). The degree of damage in this enzyme is responsible for the severity of the clinical manifestation of CAH from potentially lethal to mild symptoms. Various mutations of the gene encoding this enzyme are the main source of the reduced activity of the steroid-21-hydrolase. The location of the highly homological pseudogene CYP21P in close proximity to the functional gene impedes the DNA diagnostics of CAH. To detect the eight most frequent CYP21 gene mutations associated with CAH, we developed a new real-time PCR-based system of DNA diagnostics using new allele-specific primers and TaqMan probes for the analyzed mutations. The method was primarily tested on artificial DNA templates, where the analyzed mutations were introduced by site-directed mutagenesis. Then, it was tested on DNA samples from 43 patients with clinical and biochemical manifestations of CAH; seven patients were used as a control. Two mutant alleles were detected in two different individuals: the nonsense Q318X and the missense V281L mutations.     

HLA Typing of Patients with 21-Hydroxylase Deficiency in Iranian Children with Congenital Adrenal Hyperplasia

Congenital adrenal hyperplasia (CAH) is a group of potentially life-threatening disorders, most often caused by deficiency of steroid 21-hydroxylase. Children with ambiguous genitalia, hermaphroditism, or signs and symptoms of CAH admitted to Children's Medical Center were enrolled in the survey, and 101 patients were found. Karyotyping, clinical examination, and paraclinical tests were done. HLA typing was done in patients with proven classical CAH and their parents. HLA antigens were typed in children with CAH-type 21-hydroxylase deficiency. The antigen frequencies were compared with those of the control population. The studies revealed that two HLA antigens, HLA-B18 and HLA-B21, showed a significant increase in frequency. The calculated relative risk value was high, distinguishing the population of patients and their parents. The relative risk among patients was 11.82 for HLA-B18 and 1.75 for HLA-B21 antigens. There was no relationship between HLA-DR antigens and CAH. Studies on the correlation between HLA and CAH indicate an association with HLA-B18 and HLA-B21 antigens, and they can be used as genetic markers of the disorder in the Iranian population, if they are restricted to Iranian patients.     

Analysis of the association of HLA-DQ1 alleles with mutation of the 21-hydroxylase gene in patients with congenital adrenal hyperplasia

In 96 patients with congenital adrenal hyperplasia (CAH) and 50 healthy donors from northwestern Russia the distribution of the HLA-DQA1 alleles and the mutation spectrum and frequency at the CYP21B gene were examined. In the patients with nonclassical (NC) CAH, the distribution of the HLA-DQA1 polymorphic alleles was similar to that in the population sample. In the patients with the salt-wasting form of the disease a statistically significant decrease of the *0401 or *501 major allele frequency was observed. The prevalence of certain HLA-DQA1 genotypes, namely, HLA5, HLA3, and HLA4, was observed in the patients with the NC, salt-wasting (SW), and simple virilizing CAH, respectively. Each clinical group was characterized by a specific spectrum of clinically valuable mutations. An association between the CYP21B mutations most frequently found in case of SW and SV CAH (delB, I2splice, and I172N) and certain HLA-DQA1 alleles was demonstrated. The necessity of more precise clinical diagnostics of the NC CAH cases along with detailed examination of this group for determination of the major mutations typical of the NC CAH cases from northwestern Russia is discussed.     

Plasma insulin-like growth factor I levels during treatment of congenital adrenal hyperplasia

We tested the hypotheses that the levels of insulin-like growth factor I (IGF-I) reflect the degree of control during the treatment of patients with congenital adrenal hyperplasia (CAH), and that suppressed IGF-I levels during puberty contribute to decreased adult height. We measured serial IGF-I levels in 26 CAH patients followed over 6 months to 5 years. The control of CAH was variable, with several cases of overtreatment in infancy, and frequent undertreatment in childhood and adolescence. IGF-I levels were quite variable and inconsistent with the control of CAH. Our results suggest that IGF-I levels are not helpful in monitoring CAH patients; IGF-I does not seem to play a role in the growth failure during CAH therapy.     

Serum osteocalcin and insulin-like growth factor I levels in children with congenital adrenal hyperplasia

Patients with the virilizing forms of congenital adrenal hyperplasia (CAH) need a life-long glucocorticoid replacement therapy and also an additional mineralocorticoid replacement in cases with the salt-wasting form of the disease. Glucocorticoids are reported to decrease the serum osteocalcin levels and to inhibit the effects of insulin-like growth factor I (IGF-I). To collect data on the age related patterns of osteocalcin and IGF-I production in patients with CAH, measurements of these compounds have been carried out in a considerably large sample of treated CAH patients and control subjects in childhood and adolescence. Data of 62 patients between 0. 3-19 years of age were compared to the data of 188 control children. Osteocalcin and IGF-I were determined by radioimmunoassay. A lower than normal level of serum osteocalcin was found in both male and female patients at chronological ages above 11.6 and 9.6 years, respectively. Furthermore, no pubertal osteocalcin peak could be seen when data were evaluated according to the bone age. Serum IGF-I levels were higher in male CAH patients at the chronological age of 0.3-15.5 years and in female patients at the chronological age of 4. 6-9.5 years. In pubertal years serum IGF-I concentrations were lower in CAH patients when data were evaluated according to the bone age. We conclude that serum osteocalcin is decreased during and after puberty in CAH patients on replacement doses of glucocorticoids. Normal to elevated serum levels of IGF-I in treated CAH cases suggest that the shorter final height of these patients may not be due to the decreased activity in the growth hormoneIGF-I axis, but rather to the advanced bone maturation and the premature epiphyseal fusion.     

Psychosexual Development of Women with Congenital Adrenal Hyperplasia

Women with congenital adrenal hyperplasia (CAH) (N= 31) and their unaffected sisters or female cousins (N= 15) participated in a study of psychosexual development. All participants were ≥18 years of age (mean age, 25 years; range, 18–40). Comparisons were also made between the CAH women with the salt-wasting (SW) form of the disorder and those with simple virilization (SV). A psychosexual assessment protocol examined six variables: (1) sex assignment at birth (probands only); (2) recalled sex-typed behavior during childhood; (3) gender identity and gender role identification in adulthood; (4) relationship status; (5) sexual orientation in fantasy; and (6) sexual orientation in behavior. Salt-wasting status and sex assignment at birth were also ascertained for the CAH women who either refused to participate in the study (N= 10) or could not be traced (N= 13). Compared to the controls, the women with CAH recalled more cross-gender role behavior and less comfort with their sense of “femininity” during childhood. The two groups did not differ in degree of gender dysphoria in adulthood, although the probands showed more cross-gender role identification. Three of the nonparticipant probands were living, as adults, in the male social role (2 reared from birth as boys and 1 who changed from the female to the male social role during adolescence). The CAH women and the controls did not differ in relationship status (married/cohabiting vs. single). The CAH women had lower rates of exclusive heterosexual fantasy and fewer sexual experiences with men than the controls; however, the CAH women did not have more sexual experiences with women than the controls. Comparisons between the SW and SV revealed several differences: the SW were less likely to be assigned to the female sex at birth, recalled more cross-gender role behavior during childhood, were less likely to be married or cohabiting, and had lower rates of sexual experiences with men. The results were discussed in relation to the effects of prenatal androgens on psychosexual differentiation.     

Working memory performance is reduced in children with congenital adrenal hyperplasia

Individuals with classic congenital adrenal hyperplasia (CAH) experience impaired glucocorticoid production and are treated postnatally with glucocorticoids. Prior research with animals and other human populations indicates that glucocorticoids can influence memory, particularly working memory. We tested the hypothesis that children with CAH would show reduced working memory. Children in the United Kingdom, aged 7–11 years, with classical CAH (31 girls, 26 boys) were compared to their unaffected relatives (30 girls, 20 boys) on a test of working memory, the Digit Span test. Vocabulary was also assessed to measure verbal intelligence for control purposes. Children with CAH showed reduced working memory performance compared to controls, on both components of the Digit Span test: p = .008 for Digit Span Forward, and p = .027 for Digit Span Backward, and on a composite score, p = .004. These differences were of moderate size (d = .53 to .70). Similar differences were also seen in a subset of 23 matched pairs of children with CAH and their relatives (d = .78 to .92). There were no group differences on Vocabulary. Glucocorticoid abnormality, including treatment effects, could be responsible for the reduced Digit Span performance in children with CAH. Other factors related to CAH, such as salt-wasting crises, could also be involved. Additional research is needed to identify the cause of the memory reduction, which will help to determine if more rapid diagnosis or more precise glucocorticoid treatment would help prevent memory reduction. Educational interventions might also be considered for children with CAH.     

Adrenal Myelolipomas in Patients with Congenital Adrenal Hyperplasia: Review of the Literature and A Case Report

ObjectiveTo review the association between congenital adrenal hyperplasia (CAH) and adrenal myelolipomas and report a case of bilateral, giant adrenal myelolipomas in a patient with untreated CAH due to 21-hydroxylase deficiency.MethodsWe describe the patient’s clinical presentation, imaging findings, and laboratory test results and review the relevant English-language literature concerning patients with both CAH and myelolipomas.ResultsA 45-year-old man with untreated CAH due to 21-hydroxylase deficiency presented with increasing abdominal girth and abdominal pain. Computed tomography of the abdomen demonstrated very low-density adrenal masses (22 × 11 cm on the left side and 6 × 5.5-cm on the right side) consistent with adrenal myelolipomas. The left adrenal myelolipoma was resected (24.4 × 19.0 × 9.5 cm; 2557 g). The mass was composed of mature adipose tissue with areas of hematopoietic cells of myeloid, erythroid, and megakaryocytic cell lines. Islands of adrenal cortical cells were scattered between the adipose and hematopoietic tissue. Including the present case, we identified 31 patients with both CAH and myelolipomas who have been described in the English-language literature. The details of these cases were reviewed.ConclusionsPersons with CAH may be at increased risk of developing adrenal myelolipomas, particularly if their CAH is poorly controlled. How and whether chronic exposure of the adrenal glands to high corticotropin levels increases the risk of developing myelolipomas remains a matter of speculation. (Endocr Pract. 2011;17:441-447)     

Sex differences in children's free drawings: a study on girls with congenital adrenal hyperplasia

Sex differences are recognized in children's free drawings with respect to motifs, colors, figure compositions, and expression. Boys tend to draw mobile objects and mechanical objects with dark or cold colors and often use bird's-eye-view composition when they draw pictures, whereas girls like to draw human motifs (especially girls and women), flowers, and butterflies with light and warm colors and tend to arrange motifs in a row on the ground. Analyses were made on the drawings of girls with congenital adrenal hyperplasia (CAH) and unaffected boys and girls, using masculine and feminine indexes. Sex difference in masculine and feminine indexes was clear in the drawings by unaffected boys and girls. Their drawings do not or mostly do not contain characteristics typical of the opposite sex. Compared with those of unaffected girls, the pictures of CAH girls more strongly showed masculine characteristics. The feminine index for the pictures of CAH girls was significantly lower than that for unaffected girls, while the masculine index for CAH girls was significantly higher than that for unaffected girls. Furthermore, the masculine index for CAH girls was not significantly different from that of unaffected boys. These results suggest that androgen exposure during fetal life may contribute to shaping masculine characteristics in children's free drawings.     

Genetic linkage studies between congenital adrenal hyperplasia and the HLA blood group system

Seventeen families with one or two children suffering from congenital adrenal hyperplasia (CAH) were not only typed for their HLA-A,B, and D antigens but also tested biochemically forCAH heterozygosity after ACTH stimulation. The lod score analysis showed a close genetic linkage betweenCAH andHLA, indicating that theC-21-hydroxylase deficiency gene(s) causing CAH in the homozygous deficient state are-located in close proximity to theHLA complex on chromosome 6 with an estimated recombination fraction of 0 to 5%. HLA typing in 21 unrelated CAH patients revealed a statistically significant association to the HLA-B5 antigen with a relative risk value of 5.8. There was a significant correlation (P=0.0025) between theHLA segregation data and theCAH heterozygosity test results in relatives of CAH patients although a few ‘false negative’ results in theCAH heterozygosity test were observed. Thus, the combination of HLA typing and this biochemical test at present provides the most precise approach for detecting CAH carriers in families of CAH patients.