Human ageing can be characterized by a profile of circulating microRNAs (miRNAs), which are potentially predictors of biological age. They can be used as a biomarker of risk for age-related inflammatory outcomes, and senescent endothelial cells (ECs) have emerged as a possible source of circulating miRNAs. In this paper, a panel of four circulating miRNAs including miR-146a-5p, miR-126-3p, miR-21-5p, and miR-181a-5p, involved in several pathways related to inflammation, and ECs senescence that seem to be characteristic of the healthy ageing phenotype. The circulating levels of these miRNAs were determined in 78 healthy subjects aged between 22 to 111 years. Contextually, extracellular miR-146a-5p, miR-126-3p, miR-21-5p, and miR-181a-5p levels were measured in human ECs in vitro model, undergoing senescence. We found that the levels of the four miRNAs, using ex vivo and in vitro models, progressively increase with age, apart from ultra-centenarians that showed levels comparable to those measured in young individuals. Our results contribute to the development of knowledge regarding the identification of miRNAs as biomarkers of successful and unsuccessful ageing. Indeed, they might have diagnostic/prognostic relevance for age-related diseases.

Anderson–Fabry disease is a lysosomal storage disorder caused by mutations in the GLA gene, which encodes the enzyme α-galactosidase A. The GLA gene is located on the X-chromosome, causing an X-linked pathology: due to lyonization, female patients usually manifest a variable symptomatology, ranging from asymptomatic to severe phenotypes. The confirmation of the clinical diagnosis of Fabry disease, achieved by measuring α-galactosidase A activity, which is usually the first test used, shows differences between male and female patients. This assay is reliable in male patients with causative mutations in the GLA gene, in whom the enzymatic activity is lower than normal values; on the other hand, in female Fabry patients, the enzymatic activity is extremely variable between normal and pathological values. These fluctuations are also found in female patients’ blood levels of globotriaosylsphingosine (LysoGb3) for the same reason. In this paper, we present a retrospective study conducted in our laboratories on 827 Fabry patients with causative mutations in the GLA gene. Our results show that 100% of male patients had α-galactosidase A activity below the reference value, while more than 70% of female patients had normal values. It can also be observed that almost half of the female patients with pathogenic mutations in the GLA gene showed normal values of LysoGb3 in blood. Furthermore, in women, blood LysoGb3 values can vary over time, as we show in a clinical case presented in this paper. Both these tests could lead to missed diagnoses of Fabry disease in female patients, so the analysis of the GLA gene represents the main diagnostic test for Fabry disease in women to date.

Fabry disease (FD) is a lysosomal storage disorder (LSD) characterized by lysosomal accumulation of glycosphingolipids in a wide variety of cytotypes, including endothelial cells (ECs). FD patients experience a significantly reduced life expectancy compared to the general population; therefore, the association with a premature aging process would be plausible. To assess this hypothesis, miR-126-3p, a senescence-associated microRNA (SA-miRNAs), was considered as an aging biomarker. The levels of miR-126-3p contained in small extracellular vesicles (sEVs), with about 130 nm of diameter, were measured in FD patients and healthy subjects divided into age classes, in vitro, in human umbilical vein endothelial cells (HUVECs) “young” and undergoing replicative senescence, through a quantitative polymerase chain reaction (qPCR) approach. We confirmed that, in vivo, circulating miR-126 levels physiologically increase with age. In vitro, miR-126 augments in HUVECs underwent replicative senescence. We observed that FD patients are characterized by higher miR-126-3p levels in sEVs, compared to age-matched healthy subjects. We also explored, in vitro, the effect on ECs of glycosphingolipids that are typically accumulated in FD patients. We observed that FD storage substances induced in HUVECs premature senescence and increased of miR-126-3p levels. This study reinforces the hypothesis that FD may aggravate the normal aging process.

Background: Gaucher disease is a lysosomal storage disorder caused by functional glucocerebrosidase enzyme deficiency. Hepatosplenomegaly and hematological complications are found in both Gaucher disease and Acid Sphingomyelinase Deficiency, which is caused by acid sphingomyelinase dysfunction. The possible overlap in clinical presentation can cause diagnostic errors in differential diagnosis. For this reason, in patients with an initial clinical suspicion of Gaucher disease, we aimed to carry out a parallel screening of acid sphingomyelinase and glucocerebrosidase. Methods: Peripheral blood samples of 627 patients were collected, and enzymatic activity analysis was performed on both glucocerebrosidase and acid sphingomyelinase. The specific gene was studied in samples with null or reduced enzymatic activity. Specific molecular biomarkers helped to achieve the correct diagnosis. Results: In 98.7% of patients, normal values of glucocerebrosidase activity excluded Gaucher disease. In 8 of 627 patients (1.3%), the glucocerebrosidase enzymatic activity assay was below the normal range, so genetic GBA1 analysis confirmed the enzymatic defect. Three patients (0.5%) had normal glucocerebrosidase activity, so they were not affected by Gaucher disease, and showed decreased acid sphingomyelinase activity. SMPD1 gene mutations responsible for Acid Sphingomyelinase Deficiency were found. The levels of specific biomarkers found in these patients further strengthened the genetic data. Conclusions: Our results suggest that in the presence of typical signs and symptoms of Gaucher disease, Acid Sphingomyelinase Deficiency should be considered. For this reason, the presence of hepatosplenomegaly, thrombocytopenia, leukocytopenia, and anemia should alert clinicians to analyze both enzymes by a combined screening. Today, enzyme replacement therapy is available for the treatment of both pathologies; therefore, prompt diagnosis is essential for patients to start accurate treatment and to avoid diagnostic delay.

Anderson–Fabry disease is a hereditary, progressive, multisystemic lysosomal storage disorder caused by a functional deficiency of the enzyme α-galactosidase A (α-GalA). This defect is due to mutations in the GLA gene, located in the long arm of the X chromosome (Xq21-22). Functional deficiency of the α-GalA enzyme leads to reduced degradation and accumulation of its substrates, predominantly globotriaosylceramide (Gb3), which accumulate in the lysosomes of numerous cell types, giving rise to the symptomatology. Clinical diagnosis can still be difficult today due to the peculiarities of the disease, which presents with clinical manifestations that overlap with those of other pathologies and a wide possibility of differential diagnoses, which lead to missed diagnoses, misdiagnosis, or a diagnostic delay. Patients with clinical suspicion of Fabry disease undergo a diagnostic workup that includes an evaluation of α-GALA enzyme activity, genetic analysis of the GLA gene, and the measurement of blood Lyso-Gb3, a soluble derivative of Gb3. In this paper, we describe four novel mutations identified in the GLA gene which are associated with absent or reduced α-GalA activity, pathological accumulation of the specific substrate, and characteristic clinical manifestations of Fabry disease. We identified two mutations (c.583insGAATA and p.Y207X) that result in the formation of a premature translation stop codon, resulting in a truncated protein and thus a completely non-functional enzyme. The other two identified gene alterations (p.G261C and c.786G>T, which determine p.W262C) are missense mutations that cause reduced α-GALA activity, the accumulation of blood Lyso-Gb3, and symptoms consistent with Fabry disease, and therefore may be associated with this disorder. The identification of these new mutations in patients with symptoms attributable to Fabry disease increases the molecular knowledge of the GLA gene and provides important support to the clinician, for a more accurate and timely diagnosis of the pathology.

Studying a patient with Pompe disease (PD) is like opening Pandora’s box. The specialist is faced with numerous clinical features similar to those of several diseases, and very often the symptoms are well hidden and none is associated with this rare disease. In recent years, scientific interest in this disease has been growing more and more, but still no symptom is recognized as key to a correct diagnosis of it, nor is there any specific disease marker to date. New diagnostic/therapeutic proposals on disease allow for the diffusion of knowledge of this pathology for timely diagnosis of the patient. Due to unawareness and difficulty in diagnosis, many adults with PD are diagnosed with great delay. In this article, we report and discuss current knowledge of PD and provide new data from work conducted on a cohort of 2934 Italian subjects recruited in recent years. A genetic analysis of the GAA gene was performed on patients with significant clinical signs and pathological enzyme activity to define the genetic profile of subjects. This identified 39 symptomatic PD subjects with low acid alpha-glucosidase enzyme activity and the presence of two causative mutations in GAA gene regions. Furthermore, 22 subjects with genetic variants of uncertain significance (GVUS) were identified.