Birth Size in Neonates with Congenital Adrenal Hyperplasia due to 21-hydroxylase Deficiency

Objective: Classic congenital adrenal hyperplasia (CAH) secondary to 21-hydroxylase deficiency is characterized by increased prenatal adrenal androgen secretion. There are a small number of reports in the literature showing higher birth weight and length in CAH newborns. Methods: We analyzed birth weight and length data of 116 German newborns (48 boys, 68 girls) with classic CAH who were born during the period from 1990 to 2017. All children have been followed or are currently treated as outpatients in our clinic. All children were born at term. The mothers were healthy and their pregnancies were uneventful. The diagnosis of CAH was confirmed by molecular analyses of the CYP21A2 gene. Birth data were calculated as standard deviation (SD) scores according to German reference values. Results: Weight and length in male CAH newborns (mean ± SD) (3601±576 g; 52.4±2.85 cm) were significantly higher than in female CAH newborns (3347±442 g; 51.2±2.55 cm), but male-female differences in the CAH cohort were lost when the data were converted into SD scores. The birth sizes of the CAH newborns did not differ from the reference group. The birth sizes also did not differ between the different CAH genotypes. Maternal age, mode of delivery and maternal parity had no influence on birth size. Conclusion: Our data show that prenatal hyperandrogenism does not affect fetal growth.

Adrenal crisis in a 14-year-old boy 12 years after hematopoietic stem cell transplantation.

We report on a boy of Albanian descent with the history of juvenile myelomonocytic leukemia (JMML). JMML was diagnosed at the age of 17 months and treated by hematopoietic stem cell transplantation (HSCT). At the age of 14.3 years, about 12 years after HSCT, he was hospitalized with an adrenal crisis. Hormone findings were consistent with primary adrenal insufficiency. Autoimmune adrenalitis was confirmed by positive autoantibodies against 21-hydroxylase and adrenal tissue. Since autoimmune Hashimoto thyroiditis was already known from the age of 9 years, we assume that both diseases are part of the spectrum of autoimmune polyglandular syndrome (APS) type 2. APS type 2 is a rare endocrine disease characterized by Addison's disease along with autoimmune thyroid disease and/or type 1 diabetes. LEARNING POINTS: Endocrine sequelae after hematopoietic stem cell transplantation (HSCT) are common and can develop over a long period.Primary adrenal insufficiency after HSCT is absolutely rare.The combination of adrenal autoimmune disease and Hashimoto thyroiditis is consistent with autoimmune polyglandular syndrome type 2.

Short-term adverse effects of testosterone used for priming in prepubertal boys before growth hormone stimulation test.

BACKGROUND: Despite the fact that priming with sex steroids in prepubertal children before growth hormone (GH) provocative tests is recommended, there is an ongoing controversial discussion about the appropriate age of the children, the drug used for priming, the dose and the period between priming and the GH test. Interestingly, there is no discussion on the safety of this procedure. To date, only little data have been available on the possible side effects of priming with testosterone. METHODS: We analyzed the outcome in 188 short-statured prepubertal boys who had been primed with testosterone enanthate (n=136: 50 mg; n=51: 125 mg, and accidentally one boy with 250 mg) 7 days prior to the GH test. Serum testosterone levels were measured on the day of the GH test in 99 boys. RESULTS: Overall, only five boys developed adverse side effects. Two boys (dose 125 mg) showed severe low-flow priapism and had to undergo decompression of the corpora cavernosa. One boy suffered from self-limiting priapism and testicular pain (dose 50 mg). Two patients reported testicular pain (each dose 50 mg). The single patient with 250 mg testosterone did not show any adverse effects. The total side effect rate was 2.7%. The serum testosterone levels of the boys with side effects were not different from the testosterone levels of the boys without any side effects. CONCLUSIONS: Parents and patients should be informed about the possible side effects of priming with testosterone such as priapism and testicular pain. However, the overall side effect rate is low. We found no correlation between the outcome and the testosterone dose used and/or the level of serum testosterone.

Mutant Hoxd13 induces extra digits in a mouse model of synpolydactyly directly and by decreasing retinoic acid synthesis.

Individuals with the birth defect synpolydactyly (SPD) have 1 or more digit duplicated and 2 or more digits fused together. One form of SPD is caused by polyalanine expansions in homeobox d13 (Hoxd13). Here we have used the naturally occurring mouse mutant that has the same mutation, the SPD homolog (Spdh) allele, and a similar phenotype, to investigate the molecular pathogenesis of SPD. A transgenic approach and crossing experiments showed that the Spdh allele is a combination of loss and gain of function. Here we identify retinaldehyde dehydrogenase 2 (Raldh2), the rate-limiting enzyme for retinoic acid (RA) synthesis in the limb, as a direct Hoxd13 target and show decreased RA production in limbs from Spdh/Spdh mice. Intrauterine treatment with RA restored pentadactyly in Spdh/Spdh mice. We further show that RA and WT Hoxd13 suppress chondrogenesis in mesenchymal progenitor cells, whereas Hoxd13 encoded by Spdh promotes cartilage formation in primary cells isolated from Spdh/Spdh limbs, and that this was associated with increased expression of Sox6/9. Increased Sox9 expression and ectopic cartilage formation in the interdigital mesenchyme of limbs from Spdh/Spdh mice suggest uncontrolled differentiation of these cells into the chondrocytic lineage. Thus, we propose that mutated Hoxd13 causes polydactyly in SPD by inducing extraneous interdigital chondrogenesis, both directly and indirectly, via a reduction in RA levels.

Large-scale mapping of mutations affecting zebrafish development.

BACKGROUND: Large-scale mutagenesis screens in the zebrafish employing the mutagen ENU have isolated several hundred mutant loci that represent putative developmental control genes. In order to realize the potential of such screens, systematic genetic mapping of the mutations is necessary. Here we report on a large-scale effort to map the mutations generated in mutagenesis screening at the Max Planck Institute for Developmental Biology by genome scanning with microsatellite markers. RESULTS: We have selected a set of microsatellite markers and developed methods and scoring criteria suitable for efficient, high-throughput genome scanning. We have used these methods to successfully obtain a rough map position for 319 mutant loci from the Tübingen I mutagenesis screen and subsequent screening of the mutant collection. For 277 of these the corresponding gene is not yet identified. Mapping was successful for 80 % of the tested loci. By comparing 21 mutation and gene positions of cloned mutations we have validated the correctness of our linkage group assignments and estimated the standard error of our map positions to be approximately 6 cM. CONCLUSION: By obtaining rough map positions for over 300 zebrafish loci with developmental phenotypes, we have generated a dataset that will be useful not only for cloning of the affected genes, but also to suggest allelism of mutations with similar phenotypes that will be identified in future screens. Furthermore this work validates the usefulness of our methodology for rapid, systematic and inexpensive microsatellite mapping of zebrafish mutations.

The other trinucleotide repeat: polyalanine expansion disorders.

Expansions of trinucleotide repeats encoding polyalanine tracts have been recognized as the cause of several diseases, predominantly congenital malformation syndromes. To date, nine genes with alanine tract expansions have been described. With the exception of PABPN1, which codes for a poly(A)-binding protein, all these genes encode transcription factors that play important roles during development. Recent in vitro and in vivo findings indicate that expansions of polyalanine tracts beyond a certain threshold result in protein misfolding, aggregation and subsequent degradation. Polyalanine tracts are relatively common in the genome and occur most frequently in transcription factors and other proteins with nuclear localization. The molecular role of alanine tracts is unknown, but their strong evolutionary conservation suggests the existence of potent functional or structural constraints.

A molecular pathogenesis for transcription factor associated poly-alanine tract expansions.

Poly-alanine (Ala) tract expansions in transcription factors have been shown to be associated with human birth defects such as malformations of the brain, the digits, and other structures. Expansions of a poly-Ala tract from 15 to 22 (+7)-29 (+14) Ala in Hoxd13, for example, result in the limb malformation synpolydactyly in humans and in mice [synpolydactyly homolog (spdh)]. Here, we show that an increase of the Ala repeat above a certain length (22 Ala) is associated with a shift in the localization of Hoxd13 from nuclear to cytoplasmic, where it forms large amorphous aggregates. We observed similar aggregates for expansion mutations in SOX3, RUNX2 and HOXA13, pointing to a common mechanism. Cytoplasmic aggregation of mutant Hoxd13 protein is influenced by the length of the repeat, the level of expression and the efficacy of degradation by the proteasome. Heat shock proteins Hsp70 and Hsp40 co-localize with the aggregates and activation of the chaperone system by geldanamycin leads to a reduction of aggregate formation. Furthermore, recombinant mutant Hoxd13 protein forms aggregates in vitro demonstrating spontaneous misfolding of the protein. We analyzed the mouse mutant spdh, which harbors a +7 Ala expansion in Hoxd13 similar to the human synpolydactyly mutations, as an in vivo model and were able to show a reduction of mutant Hoxd13 and, in contrast to wt Hoxd13, a primarily cytoplasmic localization of the protein. Our results provide evidence that poly-Ala repeat expansions in transcription factors result in misfolding, degradation and cytoplasmic aggregation of the mutant proteins.

The synpolydactyly homolog (spdh) mutation in the mouse -- a defect in patterning and growth of limb cartilage elements.

We have investigated the recessive mouse mutant synpolydactyly homolog (spdh) as a model for human synpolydactyly (SPD). As in human SPD, the spdh phenotype consists of central polydactyly, syndactyly and brachydactyly and is caused by the expansion of a polyalanine encoding repeat in the 5' region of the Hoxd13 gene. We performed a detailed phenotypic and functional analysis of spdh/spdh embryos using skeletal preparations, histology, in situ hybridization, BrdU labeling of proliferating cells, and in vitro expression studies. The absence of normal phalangeal joints and the misexpression of genes involved in joint formation demonstrate a role for Hox-genes in joint patterning. The spdh mutation results in abnormal limb pattering, defective chondrocyte differentiation, and in a drastic reduction in proliferation. Abnormal chondrocyte differentiation and proliferation persisted after birth and correlated with the expression of the mutant Hoxd13 and other Hox-genes during late-embryonic and postnatal growth.