Visfatin-mediated increase of matrix mineralization and reduced collagen type I expression could contribute to bone fragility. Visfatin is involved in impaired bone remodeling at the adipose tissue/bone interface through induction of proinflammatory factors and dysregulated MMP/TIMP balance during MSC differentiation.

Systemic sclerosis (SSc) is a rare multi-organ autoimmune disease characterized by progressive skin fibrosis. Inflammation, type 2 immunity, and fibrogenic processes are involved in disease development and may be affected by sphingolipids. However, details about early-stage pathophysiological mechanisms and implicated mediators remain elusive. The sphingolipid sphingosine-1-phosphate (S1P) is elevated in the sera of SSc patients, and its receptor S1P5 is expressed in skin tissue. Nevertheless, almost nothing is known about the dermatological contribution of S1P5 to inflammatory and pro-fibrotic processes leading to the pathological changes seen in SSc. In this study, we observed a novel effect of S1P5 on the inflammatory processes during low-dose bleomycin (BLM)-induced fibrogenesis in murine skin. By comparing 2-week-treated skin areas of wild-type (WT) and S1P5-deficient mice, we found that S1P5 is important for the transcriptional upregulation of the Th2 characteristic transcription factor GATA-3 under treatment-induced inflammatory conditions, while T-bet (Th1) and FoxP3 (Treg) mRNA expression was regulated independently of S1P5. Additionally, treatment caused a regulation of S1P receptor 1 and S1P receptor 3 mRNA as well as a regulation of long-chain ceramide profiles, which both differ significantly between the genotypes. Despite S1P5-dependent differences regarding inflammatory processes, similar macroscopic evidence of fibrosis was detected in the skin histology of WT and S1P5-deficient mice after 4 weeks of subcutaneous BLM treatment. However, at the earlier 2-week point in time, the mRNA data of pro-collagen type 1 and SMAD7 indicate a pro-fibrotic S1P5 contribution in the applied SSc mouse model. In conclusion, we propose that S1P5 plays a role as a novel modulator during the early phase of BLM-caused fibrogenesis in murine skin. An immediate relationship between dermal S1P5 expression and fibrotic processes leading to skin alterations, such as formative for SSc pathogenesis, is indicated but should be studied more profound in further investigations. Therefore, this study is an initial step in understanding the role of S1P5-mediated effects during early stages of fibrogenesis, which may encourage the ongoing search for new therapeutic options for SSc patients.

Background The Delta/Notch like EGF-related receptor (DNER) is a single-pass transmembrane protein with characteristic EGF-like repeats in the extracellular domain, similar to those of the Notch receptor and its ligand Delta. DNER is an activator of the Notch signaling pathway, which plays a major role in cell fate determination and differentiation. In our previous study, an overexpression of DNER was observed in lesional areas of human osteoarthritic articular cartilage when compared to unaffected zones of the same tissue. Objectives Based on these findings we analyzed the role of DNER in the chondrogenic differentiation of human mesenchymal stem cells (hMSC). Methods HMSCs were nucleofected with an overexpression vector for DNER and a control vector. Chondrogenic differentiation of the hMSCs was performed in a pellet mass culture system for 28 days. The culture medium was supplemented with transforming growth factor beta 3 (TGF-β3) and dexamethasone. Overexpression of DNER and the effect on specific chondrogenic markers like aggrecan, collagen type 1 and 2, Sox9, collagen type 10 were analyzed with immunohistochemical staining and semi-quantitative real-time PCR. Chondrogenesis of the hMSCs was monitored using alcian blue staining. Results Alcian blue staining increased over time assuming chondrogenic differentiation of the hMSCs. Histologically, an overexpression of DNER was detected 7 days after nucleofection and remained stable during chondrogenesis, whereas no staining of DNER could be detected in the mock-transfected cells (ctr cells). Collagen 1 expression was higher in the ctr cells at the beginning of chondrogenesis compared to the pDNER-nucleofected cells (pDNER cells) and decreased in both over time. Collagen 2 was slightly expressed from day 21 in the ctr and pDNER cells. Sox9 was greater expressed in the ctr cells at the beginning of chondrogenesis and decreased in a time-dependent manner. In pDNER cells the Sox9 expression increased at day 14 and remained stable over time. The collagen 10 level is constantly higher in the pDNER nucleofected cells compared to the ctr cells. In both, the expression increased in a time-dependent manner. Aggrecan expression increased during chondrogenesis in the ctr and pDNER cells but the expression was higher in the pDNER nucleofected cells on day 28. The semi-quantitative real-time PCR data confirmed the overexpression of DNER 2 days after nucleofection and the effect, that collagen 1 expression is downregulated and collagen 2 expression upregulated in the pDNER transfected cells compared to control until day 14. Conclusions Since DNER overexpression led to a downregulation of collagen type 1 in the hMSCs, while the expression of collagen type 2 and aggrecan were upregulated, our results support the idea that DNER might positively influence chondrogenesis and exert a reparative role during OA pathophysiology. This promotion might be regulated via an intermediate overexpression of collagen type 10. Disclosure of Interest None declared DOI 10.1136/annrheu...

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