ncRNA jako predyktory powstawania skostnień heterotopowych

The aim of the study
Heterotopic ossification (HO) is a dysregulation of skeletal muscle homeostasis and regeneration that leads to the
formation of mature bone in untypical locations. The direct molecular mechanism leading to the formation of HO
has not yet been described. Importantly, HO formation is related to the state of hypoxia and inflammation, and is
likely related to disturbances in BMP (bone morphogenic protein) and TGFβ (transforming growth factor β) signaling
pathways. However, other factors, such as TLR (toll-like receptor), that play a key role in inducing an innate immune
response, could be involved. Importantly, the knowledge about the role of ncRNA (non-coding RNA), such as miRNA
(microRNA) and lncRNA (long non-coding RNA), in the formation of HO is limited. The miRNA and lncRNA are
molecules that affect the expression of target mRNAs. Thus, identification of miRNA and lncRNA is important for
understanding the mechanism of HO formation. Our preliminary data (high-throughput sequencing) showed
changes in mRNA and lncRNA expression in muscles of patients suffering from HO after hip arthroplasty compared
to healthy donors. On the basis of bioinformatic analysis, we selected miRNA and lncRNA molecules that could play
a role in the HO formation. We hypothesized that the formation of HO is related to disturbances in the expression
of miRNAs or lncRNAs. We believe that these molecules may affect the expression of BMPs, TGFβ, TLR or other
proteins involved in signaling that impairs myogenesis and induces osteogenesis. Thus, the aim of this project is
to verify the role of selected molecules in the HO formation. To do so, human mesenchymal stromal cells or human
myoblasts will be transfected with selected miRNA and lncRNA molecules and an analysis of their differentiation
into chondroblasts or osteoblasts, cultured under hypoxic, normoxic or inflammatory conditions in vitro, will be
performed. Furthermore, the transfected human cells will be transplanted into SCID mice muscles or
subcutaneously, what will allow functional in vivo verification of the obtained data. The new information on the
molecules involved in the formation of HO will allow us to validate their role in HO and follow their level in patients.
Thus, the level of selected miRNAs, lncRNAs, and mRNA of proteins involved in osteogenesis (such as BMPs, TGFβ,
TLR) will be analyzed in the tissue samples of healthy donors, patients who underwent hip arthroplasty, and patients
suffering from HO. Identifying miRNAs and lncRNAs would be of great importance to understand the mechanism of
HO formation. In addition, the identification of such molecules would be crucial to evaluate them as predictors of
HO formation. Such molecules could be used as markers to monitor the ossification formation and possibly
implement the appropriate treatment. Furthermore, these molecules could become a potential targets for therapy.