Identificazione e caratterizzazione di nuovi marcatori epigenetici coinvolti nella patogenesi dell'osteoporosi

Osteoporosis (OP) is a multifactorial disorder in which environmental factors along with genetic variants and epigenetic mechanisms are implicated. Long non-coding RNAs (lncRNAs) have a crucial role in regulating many important biological processes in bone metabolism and OP aetiology, including inflammation. We focused on 84 lncRNAs regulating the expression of pro-inflammatory and anti-inflammatory genes and miRNAs, to identify lncRNAs mis-regulated in osteoblast primary cultures derived from OP patients and healthy controls. Six lncRNAs resulted significantly downregulated in OP patients: CEP83-AS1, RP11-84C13.1, CTC- 487M23.5, GAS5, NCBP2-AS2, and SDCBP2-AS1. Among them, the lncRNA GAS5 has shown a five-fold downregulation of OP primary osteoblasts, thus indicating a possible pivotal role in OP development. Therefore, we decided to analyze GAS5 lncRNA expression level in serum from our study cohort and we found a significantly increased level in OP patients, with a statistically higher significance in fractured OP individuals. Genetic analysis of GAS5 rs145204276 polymorphism revealed that the deletion allele (Del) was correlated with a higher expression of GAS5 in OP patients, confirming previous studies which indicate rs145204276 Del allele as a modulator of GAS5 transcription activity. Our results suggest circulating GAS5 as a putative biomarker for the diagnosis and prognosis of OP and fragility fractures. In silico analysis was also performed to predict the interaction of GAS5 lncRNA with its targets. This led to identify PTX3 as downstream target of GAS5 lncRNA, with a mediated interaction through miR-21-5p, a regulator of OP pathogenesis and known sponge target of GAS5. PTX3 is generating great interest given the recent discovery of its involvement in bone metabolism. For this reason, we also performed serum PTX3 analysis revealing a statistically significant increase of circulating levels in OP patients. It is therefore possible that circulating PTX3 increase is a downstream effect, mediated by known systemic downregulation of miR-21-5p, caused by the upregulation of GAS5 lncRNA in OP patients. We speculate that an increase in circulating GAS5 in OP patients could be explained by its induction by a proapoptotic signal to accumulate in exosomes, leading to a reduced expression level within osteoblasts. Altogether, these data open a new regulatory mechanism of gene expression in bone homeostasis and could direct the development of future therapeutic approaches.