Background Systemic Sclerosis (SSc) is characterized by vascular alteration with a progressive loss of capillaries resulting in insufficient blood flow and chronic peripheral hypoxia, associated to a failure of reparative angiogenesis, followed by progressive fibrosis of skin and internal organs[1]. An impaired production of angiogenic molecules is involved in this dysfunctional angiogenesis [2]. Tie2 is a trans-membrane receptor, exclusively expressed on the surface of endothelial cell (EC). Tie2 modulates the vessels quiescence, or alternatively, induces angiogenesis, via the interaction with its ligands, Ang1 and Ang2. Of note, activation of endothelial Tie2 signalling, by Ang1, which is produces by perivascular cells, enhances ECs barrier integrity and endothelial-pericyte interaction. On the contrary, Ang2, produced by ECs, normally acts as an Ang1 antagonist [3].

Objectives The aim of the present study was to assess if an impaired cross-talk between ECs and perivascular mesenchymal stem cells (MSCs), during SSc, may affect the normal interaction among Ang1, Ang2 and Tie2 thus contributing to the impaired angiogenesis. In this work, bone marrow derived MSCs were used as pericytes surrogate, considering that, perivascular cells share surface markers and differentiative ability with bone marrow MSCs, and MSCs express pericyte markers and cooperate with endothelial cells to form a vascular network, supporting the concept that pericytes are members of the adult multipotent MSCs family [4].

Methods We investigated Ang1, Ang2 and their receptor performing co-cultures with ECs and bone marrow MSCs obtained from patients and healthy controls (HC). After 48 hours, cells were sorted and analysed for molecular assays. Furthermore, we investigated, by ELISA assay, the proteins released in the supernatants. Finally, we silenced Ang-1 expression in HC-MSCs by Ang1-siRNA.

Results At molecular level, SSc-MSCs, cultured alone, expressed lower amount of Ang1 when compared to HC-MSCs. After co-culture, a significant decreased of Ang1 mRNA levels was observed in the SSc-MSCs/SSc-ECs. On the contrary, SSc-ECs expressed higher levels of Ang2 and Tie2 in each co-culture condition, when compared to the expressions of cells cultured alone. The WB and ELISA assays mirrored the results observed in gene expression. HC-MSCs transfected with Ang1-siRNA lacked the ability to support the formation of tube like structure.

Conclusions In this work we provided evidence that an imbalance of Ang1/Ang2 molecules and a decreased expression of their receptor, Tie2, during ECs-perivascular MSCs interplay, may modulate vessel stability, and vascular tube formation, thus contributing to the angiogenic alteration observed during SSc.

References

1. Cipriani P et al. Autoimmun Rev 2011,10:641-646.

2. Cipriani P et al. Arthritis Res Ther 2013,16:442.

3. Fukuhara S et al. Histol Histopathol 2010,25:387–396.

4. Cipriani P et a. Angiogenesis 2013,16:595-607.

Disclosure of Interest None declared