Fertility and sexual activity in patients with Triple A syndrome.

Objective: Triple A syndrome, caused by autosomal recessively inherited mutations in the AAAS gene is characterized by alacrima, achalasia, adrenal insufficiency, and neurological impairment. To the best of our knowledge, no patients of both sexes have been reported to have offspring. Our aim was to assess the causes of infertility in male patients with this multisystemic syndrome, and to present a female patient that spontaneously conceived a child. Design: Cross-sectional study. Methods: Six males aged 19-48 years were included. Gonadotropins, testosterone, DHEAS, androstenedione, inhibin B, anti-Mullerian hormone measurements and testicular ultrasound were performed. Results: All six male patients had impaired general health and neurological symptoms including erectile and ejaculatory dysfunction. None of them had an offspring. The only demonstrated cause of infertility in our male patients was erectile and ejaculatory dysfunction which precludes sexual intercourse. Our patients had normal libido but were sexually abstinent. Except for low adrenal androgen levels, the concentrations of all measured hormones as well as testicular ultrasound were normal which may indicate the possibility of spermatogenesis in male patients with triple A syndrome. Little is known about fertility in female patients, but based on our observations spontaneous pregnancies seem to be possible. Conclusion: Our results contribute to still scarce knowledge on fertility in patients with Triple A syndrome and as well represents a foundation for further research on causes of infertility and possible treatment options.

A single-cell transcriptional landscape of immune cells shows disease-specific changes of T cell and macrophage populations in human achalasia.

Achalasia is a rare motility disorder of the esophagus caused by the gradual degeneration of myenteric neurons. Immune-mediated ganglionitis has been proposed to underlie the loss of myenteric neurons. Here, we measure the immune cell transcriptional profile of paired lower esophageal sphincter (LES) tissue and blood samples in achalasia and controls using single-cell RNA sequencing (scRNA-seq). In achalasia, we identify a pattern of expanded immune cells and a specific transcriptional phenotype, especially in LES tissue. We show C1QC+ macrophages and tissue-resident memory T cells (TRM), especially ZNF683+ CD8+ TRM and XCL1+ CD4+ TRM, are significantly expanded and localized surrounding the myenteric plexus in the LES tissue of achalasia. C1QC+ macrophages are transcriptionally similar to microglia of the central nervous system and have a neurodegenerative dysfunctional phenotype in achalasia. TRM also expresses transcripts of dysregulated immune responses in achalasia. Moreover, inflammation increases with disease progression since immune cells are more activated in type I compared with type II achalasia. Thus, we profile the immune cell transcriptional landscape and identify C1QC+ macrophages and TRM as disease-associated immune cell subsets in achalasia.

Don't forget Allgrove syndrome in adult patients as a bulbar-ALS mimicker.

INTRODUCTION: Allgrove syndrome is a genetic disorder characterized by a multisystem involvement manifesting mainly in childhood with esophageal achalasia, adrenal insufficiency, and alacrima. Associated neurological manifestations are frequent in patients with late-onset forms and include peripheral, central, and autonomic dysfunction. The definitive diagnosis remains genetic, but neurological symptoms/signs could be a relevant clue for the diagnosis. DISCUSSION: This syndrome is rare, but it is not impossible for it to occur in adults, so all neurologists must be alert. Moreover, in this regard, neurological symptoms can sometimes be very similar to those of motor neuron disease patients, so that, although rare, Allgrove syndrome may also enter into the differential diagnosis with the bulbar variant of amyotrophic lateral sclerosis. Nevertheless, attention to extra-neurological symptoms must remain high as these play an equally important role in reaching the diagnosis. CASE REPORT: Here we present the case of a patient with some peculiarities that are onset at an advanced age, genetic confirmation of the diagnosis, and prominent neurological involvement, which also opens the differential diagnosis to amyotrophic lateral sclerosis.

SCARB1 downregulation in adrenal insufficiency with Allgrove syndrome.

BACKGROUND: Allgrove disease is a rare genetic syndrome characterized by adrenal insufficiency, alacrimia, achalasia and complex neurological involvement. Allgrove disease is due to recessive mutations in the AAAS gene, which encodes for the nucleoporin Aladin, implicated in the nucleocytoplasmic transport. The adrenal insufficiency has been suggested to rely on adrenal gland-ACTH resistance. However, the link between the molecular pathology affecting the nucleoporin Aladin and the glucocorticoid deficiency is still unknown. RESULTS: By analyzing postmortem patient's adrenal gland, we identified a downregulation of Aladin transcript and protein. We found a downregulation of Scavenger receptor class B-1 (SCARB1), a key component of the steroidogenic pathway, and SCARB1 regulatory miRNAs (mir125a, mir455) in patient's tissues. With the hypothesis of an impairment in the nucleocytoplasmic transport of the SCARB1 transcription enhancer cyclic AMP-dependent protein kinase (PKA), we detected a reduction of nuclear Phospho-PKA and a cytoplasmic mislocalization in patient's samples. CONCLUSIONS: These results shed a light on the possible mechanisms linking ACTH resistance, SCARB1 impairment, and defective nucleocytoplasmic transport.

GWAS using low-pass whole genome sequence reveals a novel locus in canine congenital idiopathic megaesophagus.

Congenital idiopathic megaesophagus (CIM) is a gastrointestinal disorder of dogs wherein the esophagus is dilated and swallowing activity is reduced, causing regurgitation of ingesta. Affected individuals experience weight loss and malnourishment and are at risk for aspiration pneumonia, intussusception, and euthanasia. Great Danes have among the highest incidences of CIM across dog breeds, suggesting a genetic predisposition. We generated low-pass sequencing data for 83 Great Danes and used variant calls to impute missing whole genome single-nucleotide variants (SNVs) for each individual based on haplotypes phased from 624 high-coverage dog genomes, including 21 Great Danes. We validated the utility of our imputed data set for genome-wide association studies (GWASs) by mapping loci known to underlie coat phenotypes with simple and complex inheritance patterns. We conducted a GWAS for CIM with 2,010,300 SNVs, identifying a novel locus on canine chromosome 1 (P-val = 2.76 × 10-10). Associated SNVs are intergenic or intronic and are found in two clusters across a 1.7-Mb region. Inspection of coding regions in high-coverage genomes from affected Great Danes did not reveal candidate causal variants, suggesting that regulatory variants underlie CIM. Further studies are necessary to assess the role of these non-coding variants.

The clinical and laboratory features of patients with triple A syndrome: a single-center experience in Turkey.

AIM: Triple-A Syndrome (TAS) is a rare autosomal recessive disorder characterized by adrenal insufficiency, achalasia, and alacrimia. This disorder is caused by mutations in the AAAS gene. The aim of this study is to discuss the clinical, laboratory and molecular genetic analysis results of 12 patients with TAS. METHOD: We evaluated 12 patients from 8 families. Clinical and laboratory data were retrospectively collected from the medical records of the patients in the database for the period 2015-2020. All exons and exon-intron junctions of the AAAS gene were evaluated by next-generation sequencing method. Detected variants were classified according to American Collage of Medical Genetics criteria. RESULTS: Alacrimia was found in all patients (100%); achalasia was found in 10 patients (83.3%) and adrenal insufficiency was found in 10 patients (83.3%). In addition, hyperreflexia(6/12), learning disability(5/12), hypernasal speech(5/12), muscle weakness(8/12), delayed walking(7/12), delayed speech(6/12), excessive sweating(7/12), optic atrophy(1/12), epilepsy(1/12), palmoplantar hyperkeratosis(5/12), multiple dental caries(9/12), atrophy of the thenar/hypothenar muscles(4/12) and short stature(4/12) were detected. The DHEA-S levels were measured in 10 patients and were found to be low in 8 of them. In all patients, the sodium and potassium levels were found to be normal. AAAS gene sequencing revealed four previously reported c.1066_1067del (p.Leu356fs*8), c.1432 C > T (p.Arg478*), c.688 C > T (p.Arg230*), and c.1368_1372del (p.Gln456fs*38) variants and two novel homozygous c.1250-1 G > A and c.398_399 + 2del variants in the AAAS gene. CONCLUSION: We detected two novel variants in the AAAS gene. While the classic triad is present in 66.7% of the cases, neurological dysfunction, skin and dental pathologies also occur quite frequently. The earliest and most common finding of TAS is alacrimia. Therefore, adrenal insufficiency should be investigated in all patients with alacrimia and if necessary, genetic analysis should be performed for TAS. In addition, TAS should be followed up with a multidisciplinary approach since it involves many systems.

RNA-sequencing reveals molecular and regional differences in the esophageal mucosa of achalasia patients.

Achalasia is an esophageal motility disorder characterized by the functional loss of myenteric plexus ganglion cells in the distal esophagus and lower esophageal sphincter. Histological changes have been reported in the esophageal mucosa of achalasia, suggesting its involvement in disease pathogenesis. Despite recent advances in diagnosis, our understanding of achalasia pathogenesis at the molecular level is very limited and gene expression profiling has not been performed. We performed bulk RNA-sequencing on esophageal mucosa from 14 achalasia and 8 healthy subjects. 65 differentially expressed genes (DEGs) were found in the distal esophageal mucosa of achalasia subjects and 120 DEGs were identified in proximal esophagus. Gene expression analysis identified genes common or exclusive to proximal and distal esophagus, highlighting regional differences in the disease. Enrichment of signaling pathways related to cytokine response and viral defense were observed. Increased infiltration of CD45+ intraepithelial leukocytes were seen in the mucosa of 38 achalasia patients compared to 12 controls. Novel insights into the molecular changes occurring in achalasia were generated in this transcriptomic study. Some gene changes observed in the mucosa of achalasia may be associated with esophagitis. Differences in DEGs between distal and proximal esophagus highlight the importance of better understanding regional differences in achalasia.

[A case of triple A syndrome with c.463C>T mutation in the AAAS gene].

A 47-year-old woman was admitted to our hospital for scrutiny of limb weakness and orthostatic hypotension that had progressed from childhood. She had been treated for alacrima and esophageal achalasia from childhood. On admission, she had hyperreflexia of upper and lower extremities, distal predominant muscle atrophy in the lower extremities, decreased sensation of the distal extremities, and autonomic neuropathy. Her blood test results ruled out adrenal insufficiency, but Schirmer's test was positive. Given the lacrimation symptoms, esophageal achalasia, and neuropathy, the patient was diagnosed with triple A syndrome in whom a c.463C>T mutation (p.R155C) was found in the AAAS gene by genetic testing. Triple A syndrome is an autosomal recessive inherited disease caused by mutations in the AAAS gene. Genetic testing of the AAAS gene should be considered in patients with one or two of main symptoms of triple A syndrome.

Recurrent pulmonary infection leads to the diagnosis of triple A syndrome: a case report.

INTRODUCTION: Triple A syndrome is a very uncommon disease marked by a triad of adrenocorticotrophic hormone (ACTH)-resistant features: adrenal insufficiency, alacrimia, and achalasia. It presents in several clinical forms with undetermined incidence and shows an autosomal pattern of inheritance. It is caused by a variety of mutations in the AAAS genes which encode a protein of unknown function called ALADIN. Diagnosis depends on clinical manifestations, laboratory test results, imaging and endoscopic findings, and Schirmer's test. The treatment includes artificial tears, glucocorticoid replacement therapy, and treatment of achalasia. CASE PRESENTATION: A 12-year-old Syrian girl was referred to Damascus University's Children's Hospital for recurrent pulmonary infection. Her mother had noted an absence of tears when crying since birth, diffused pigmentations since birth, especially on the cheeks and genitals, recurrent vomiting of both solid and liquid foods, and recurrent exacerbations of bronchitis and recurrent pneumonia. ACTH and blood cortisol levels indicated an adrenal insufficiency, chest computed tomography and barium swallow test results indicated achalasia, tear break-up time as well as eye examination indicated alacrimia, which led to the diagnosis of triple A syndrome. Treatment included Heller cardiomyotomy, artificial tears, and hydrocortisone (15-30 mg/m2), as well as continuous observation of ACTH levels. DISCUSSION: Triple A syndrome (which is characterized by the triad of achalasia, alacrima, adrenal insufficiency) is a rare multisystem disease. It has a genetic background and is potentially fatal. This syndrome is often misdiagnosed, especially in regions where it is expected to have a high prevalence rate (regions with documented cases and high rate of consanguinous marriage), This study is the first documentation of triple A syndrome in Syria, a country where consanguineous marriage is common. This syndrome should be kept in mind when a child presents with one or more of its characteristic features.

A rare cause of infantile achalasia: GMPPA-congenital disorder of glycosylation with two novel compound heterozygous variants.

Achalasia is rare in the pediatric population and should prompt clinicians to consider genetic disorders associated with this condition. While AAA syndrome (also known as Allgrove or Triple A syndrome) is commonly considered, GMPPA-congenital disorder of glycosylation (CDG) should also be in the differential diagnosis. We report a 9-month-old female born to nonconsanguineous parents with achalasia and alacrima found to have two novel compound heterozygous variants in the GMPPA gene associated with GMPPA-CDG. This rare disorder is commonly associated with developmental delay and intellectual disability. We discuss management of this disorder including the importance of confirming a genetic diagnosis and summarize reported cases.

Functional validation of a novel AAAS variant in an atypical presentation of Allgrove syndrome.

BACKGROUND: Achalasia-addisonianism-alacrima syndrome, frequently referred to as Allgrove syndrome or Triple A syndrome, is a multisystem disorder resulting from homozygous or compound heterozygous pathogenic variants in the gene encoding aladin (AAAS). Aladin is a member of the WD-repeat family of proteins and is a component of the nuclear pore complex. It is thought to be involved in nuclear import and export of molecules. Here, we describe an individual with a paternally inherited truncating variant and a maternally inherited, novel missense variant in AAAS presenting with alacrima, achalasia, anejaculation, optic atrophy, muscle weakness, dysarthria, and autonomic dysfunction. METHODS: Whole-exome sequencing was performed in the proband, sister, and parents. Variants were confirmed by Sanger sequencing. The localization of aladin to the nuclear pore was assessed in cells expressing the patient variant. RESULTS: Functional testing of the maternally inherited variant, p.(Arg270Pro), demonstrated decreased localization of aladin to the nuclear pore in cells expressing the variant, indicating a deleterious effect. Follow-up testing in the proband's affected sister revealed that she also inherited the biallelic AAAS variants. CONCLUSIONS: Review of the patient's clinical, pathological, and genetic findings resulted in a diagnosis of Triple A syndrome.

AAMR syndrome in a 22-month-old and literature review.

PURPOSE: Alacrima is characterized by severely decreased or deficient tear production. It can be associated with systemic findings; most commonly Triple-A Syndrome with alacrima, achalasia, and adrenal dysfunction. METHODS: A case report and review of the literature. RESULTS: A 22-month-old boy with cognitive delay presented with a lack of tearing. MRI brain and orbits showed a bilateral absence of lacrimal glands. Biochemical testing revealed normal adrenal function. Genetic testing showed no abnormalities in the AAAS gene. Mutations in the AAAS gene are associated with Triple-A syndrome. Whole-exome sequencing did reveal compound heterozygosity for the GMPPA gene, and mutations in the GMPPA gene have been associated with AAMR syndrome with alacrima, achalasia, and mental retardation. A literature review revealed all AAMR syndrome patients with consanguineous parents. This is the first reported case of AAMR syndrome in non-consanguineous parents. CONCLUSIONS: In children older than 6 months old with decreased or deficient tear production, systemic work-up should include radiographic, biochemical, and genetic testing. Conditions associated with alacrima include familial dysautonomia, anhidrotic ectodermal dysplasia, Triple-A syndrome, and AAMR syndrome. One key clinical difference between the two syndromes is that patients with normal adrenal function have AAMR syndrome rather than Triple-A syndrome. After a child has a diagnosis with alacrima, these patients should be evaluated closely by a pediatric multidisciplinary team. Treatment for alacrima depends on the severity of ocular symptoms and ranges from the use of lubricant tears and ointment to moisture chambers.

Downexpression of miR-200c-3p Contributes to Achalasia Disease by Targeting the PRKG1 Gene.

Achalasia is an esophageal smooth muscle motility disorder with unknown pathogenesis. Taking into account our previous results on the downexpression of miR-200c-3p in tissues of patients with achalasia correlated with an increased expression of PRKG1, SULF1, and SYDE1 genes, our aim was to explore the unknown biological interaction between these genes and human miR-200c-3p and if this relation could unravel their functional role in the etiology of achalasia. To search for putative miR-200c-3p binding sites in the 3'-UTR of PRKG1, SULF1 and SYDE1, a bioinformatics tool was used. To test whether PRKG1, SULF1, and SYDE1 are targeted by miR-200c-3p, a dual-luciferase reporter assay and quantitative PCR on HEK293 and fibroblast cell lines were performed. To explore the biological correlation between PRKG1 and miR-200c-3p, an immunoblot analysis was carried out. The overexpression of miR-200c-3p reduced the luciferase activity in cells transfected with a luciferase reporter containing a fragment of the 3'-UTR regions of PRKG1, SULF1, and SYDE1 which included the miR-200c-3p seed sequence. The deletion of the miR-200c-3p seed sequence from the 3'-UTR fragments abrogated this reduction. A negative correlation between miR-200c-3p and PRKG1, SULF1, and SYDE1 expression levels was observed. Finally, a reduction of the endogenous level of PRKG1 in cells overexpressing miR-200c-3p was detected. Our study provides, for the first time, functional evidence about the PRKG1 gene as a direct target and SULF1 and SYDE1 as potential indirect substrates of miR-200c-3p and suggests the involvement of NO/cGMP/PKG signaling in the pathogenesis of achalasia.

Allgrove syndrome with amyotrophy.

Allgrove syndrome is an autosomal recessive disease mostly caused by mutations in the AAAS gene. It has variable clinical features but its cardinal features comprise the triad of achalasia, alacrimia and adrenal insufficiency. It typically develops during the first decade of life, but some cases have second and third decades onset. We describe a 25-year-old woman with Allgrove syndrome who had progressive amyotrophy, achalasia, dry eyes and adrenal insufficiency since childhood. Awareness of its neurological manifestations and multisystem features helps to shorten the time for diagnosis and allow appropriate symptomatic treatment.

Achalasia and acromegaly: Co-incidence of these diseases or a new syndrome?

BACKGROUND: Acromegaly is a disorder associated with hypersecretion of growth hormone, most usually caused by a pituitary adenoma. Dysmotility of the gastrointestinal tract has been reported in acromegalic patients. Achalasia is a disorder characterized by aperistalsis of the oesophagus with incomplete lower oesophageal sphincter relaxation and whose aetiology remains unknown. Mutations in some genes have previously been associated with the development of acromegaly or achalasia. The study aims were to analyse mutations in selected genes in a woman having both of these diseases, to identify their aetiological factors, and to suggest explanations for the co-incidence of acromegaly and achalasia. METHODS AND RESULTS: A female patient with acromegaly, achalasia, and a multinodular thyroid gland with hyperplastic colloid nodules underwent successful treatment of achalasia via laparoscopic Heller myotomy, a thyroidectomy was performed, and the pituitary macroadenoma was surgically excised via transnasal endoscopic extirpation. Germline DNA from the leukocytes was analysed by sequencing methods for a panel of genes. No pathogenic mutation in AAAS, AIP, MEN1, CDKN1B, PRKAR1A, SDHB, GPR101, and GNAS genes was found in germline DNA. The somatic mutation c.601C>T/p.R201C in the GNAS gene was identified in DNA extracted from a tissue sample of the pituitary macroadenoma. CONCLUSIONS: We here describe the first case report to our knowledge of a patient with both acromegaly and achalasia. Association of acromegaly and soft muscle tissue hypertrophy may contribute to achalasia's development. If one of these diagnoses is determined, the other also should be considered along with increased risk of oesophageal and colorectal malignancy.

Biallelic CAV1 null variants induce congenital generalized lipodystrophy with achalasia.

OBJECTIVE: CAV1 encodes caveolin-1, a major protein of plasma membrane microdomains called caveolae, involved in several signaling pathways. Caveolin-1 is also located at the adipocyte lipid droplet. Heterozygous pathogenic variants of CAV1 induce rare heterogeneous disorders including pulmonary arterial hypertension and neonatal progeroid syndrome. Only one patient was previously reported with a CAV1 homozygous pathogenic variant, associated with congenital generalized lipodystrophy (CGL3). We aimed to further delineate genetic transmission, clinical, metabolic, and cellular characteristics of CGL3. DESIGN/METHODS: In a large consanguineous kindred referred for CGL, we performed next-generation sequencing, as well as clinical, imagery, and metabolic investigations. We studied skin fibroblasts from the index case and the previously reported patient with CGL3. RESULTS: Four patients, aged 8 months to 18 years, carried a new homozygous p.(His79Glnfs*3) CAV1 variant. They all displayed generalized lipodystrophy since infancy, insulin resistance, low HDL-cholesterol, and/or high triglycerides, but no pulmonary hypertension. Two patients also presented at the age of 15 and 18 years with dysphagia due to achalasia, and one patient had retinitis pigmentosa. Heterozygous parents and relatives (n = 9) were asymptomatic, without any metabolic abnormality. Patients' fibroblasts showed a complete loss of caveolae and no protein expression of caveolin-1 and its caveolin-2 and cavin-1 partners. Patients' fibroblasts also displayed insulin resistance, increased oxidative stress, and premature senescence. CONCLUSIONS: The CAV1 null variant investigated herein leads to an autosomal recessive congenital lipodystrophy syndrome. Loss of caveolin-1 and/or caveolae induces specific manifestations including achalasia which requires specific management. Overlapping phenotypic traits between the different CAV1-related diseases require further studies.

Whole-exome sequencing reveals common and rare variants in immunologic and neurological genes implicated in achalasia.

Idiopathic achalasia (IA) is a severe motility disorder characterized by neuronal degeneration in the myenteric plexus, but the etiology remains largely unknown. We performed whole-exome sequencing (WES) in 100 IA-affected individuals and 313 non-IA control subjects and validated the results in 230 IA-affected individuals and 1,760 non-IA control subjects. Common missense variants rs1705003 (CUTA, GenBank: NC_000006.11:g.33385953A>G) and rs1126511 (HLA-DPB1, GenBank: NC_000006.11:g.33048466G>T) at 6p21.32 were reproducibly associated with increased risk of IA (rs1126511: OR = 1.83, p = 2.34 × 10-9; rs1705003: OR = 2.37, p = 3.21 × 10-7), meeting exome-wide significance. Both variants can affect the expression of their target genes at the transcript level. An array-based association analysis in 280 affected individuals and 1,121 control subjects determined the same signal at 6p21.32. Further conditional analyses supported that the two missense variants identified in WES-based association study were potential causal variants of IA. For rare variants, the top genes identified by gene-based analysis were significantly enriched in nerve and muscle phenotypic genes in the mouse. Moreover, the functional rare variants in these genes tended to cooccur in IA-affected individuals. In an independent cohort, we successfully validated three rare variants (CREB5, GenBank: NC_000007.13:g.28848865G>T; ESYT3, GenBank: NC_000003.11:g.138183253C>T; and LPIN1, GenBank: NC_000002.11:g.11925128A>G) which heightens the risk of developing IA. Our study identified and validated two common variants and three rare variants associated with IA in immunologic and neurological genes, providing new insight into the etiology of IA.

Frequency and clinical characteristics of special types of achalasia in Japan: A large-scale, multicenter database study.

BACKGROUND AND AIM: Achalasia is a rare disease, with an incidence of one in 100 000. Genetic factors and autoimmune involvement have been reported in its etiology, and their involvement is strongly suspected, especially in patients with familial achalasia and those with comorbid hereditary or autoimmune diseases. However, these special types of achalasia are rare, and their frequency and clinical characteristics remain unclear. METHODS: This retrospective, multicenter cohort study included Japanese patients with a diagnosis of achalasia, treated between 2010 and 2019 across six tertiary centers in Japan. The frequency and clinical characteristics of special types of achalasia, namely, familial achalasia, achalasia with a comorbid hereditary disease, and achalasia with a comorbid autoimmune disease, were retrospectively investigated using a large-scale multicenter database. RESULTS: During the study period, 1115 patients were treated for achalasia at six tertiary centers. Familial achalasia, achalasia with a comorbid hereditary disease, and achalasia with a comorbid autoimmune disease occurred in 7 (0.63%), 11 (0.99%), and 27 (2.4%) patients, respectively. Familial achalasia had a slightly younger age of onset (37.6 ± 12.1 years old) and a higher incidence in male patients (six patients; 85.7%). Down's syndrome was the most common hereditary comorbidity, and thyroid disease was the most common autoimmune comorbidity. CONCLUSIONS: We clarified the frequency and clinical characteristics of special types of achalasia. Although special types of achalasia are rare, these comorbidities should be considered when treating patients with achalasia.