Hereditary Haemorrhagic Telangiectasia: mutation analysis, new variant characterization and study of circulating microRNAs in a rare disease

The topic of my PhD project is the Hereditary Haemorrhagic Telangiectasia (HHT), also known as Rendu-Osler-Weber syndrome, a rare genetic disease with an incidence of about 1:5000, inherited as an autosomal dominant trait. Up to date, mutations in four genes (ENG, ACVRL1, MADH4 and GDF2) have been described in HHT, however, ENG and ACVRL1 mutations account for the 85% of patients. All these genes encode for proteins belonging to the TGF-β/BMPs signalling pathway and are involved in the regulation of angiogenesis. HHT is clinically diagnosed if a patient presents at least three out of four criteria, known as “Curaҫao criteria”: (i) spontaneous and recurrent epistaxis; (ii) telangiectases at characteristic sites, as lips, oral cavity, nose, fingertips and gastrointestinal mucosa; (iii) arteriovenous malformations at characteristic sites, as liver, lungs and central nervous system, and (iv) family history, i.e. a first degree relative with a diagnosis of HHT according to the same criteria. The aim of this thesis is to further investigate the genetic background of HHT in order to better understand the underlying molecular mechanisms and the genotype-phenotype correlations. In particular, my work involved the mutation analyses of ENG and ACVRL1 coding exons and the study of circulating miRNAs in a group of HHT patients. Mutation analyses were performed in new index cases with a HHT clinical diagnosis and their relatives, to identify the disease-causing mutation. Then, the results collected were added to data previously obtained by the Medical Genetics Laboratory (headed by Prof. C. Danesino) and used to perform a descriptive study of the HHT Italian Population, carried out in collaboration with Prof. C. Sabbà, from the University of Bari. Moreover, during my PhD I attended a three-months period in the laboratory of Prof. C. Bernabeu, in Madrid, in order to study the putative pathogenic effect of a particular ENG variant (c.1852+42 C>T) found in a HHT family. The study of circulating miRNAs was performed in collaboration with Prof. M. Denti (University of Trento). We collected plasma samples from 15 people (5 controls; 5 patients with ENG mutation and 5 patients with ACVRL1 mutation) and evaluated the expression level of 752 human miRNAs to identify those misregulated in patients compared to controls. Firstly, we analysed results grouping the patients according to the mutated gene and we identified about 20 misregulated miRNAs. Then, we evaluated if some miRNAs were differentially expressed in patients according to their clinical symptoms. In conclusion, the obtained results increased the knowledge on the molecular pathogenesis, and shed a light on the role exploited by miRNAs in this disorder, thus suggesting new genotype-phenotype correlations, although a validation of the results in a bigger population is mandatory.