ECTS Abstracts (2015) 1 P153

Osteogenic differentiation of human mesenchymal stromal cell relies on autocrine/paracrine leptin activity/action

Bram van der Eerden, Marijke Schreuders-Koedam, Yolande de Lege, Solveig Staurland, Jeroen van de Peppel & Johannes van Leeuwen

Erasmus MC, Internal Medicine, Rotterdam, The Netherlands.

Leptin is an important molecule linking energy to bone metabolism. Although the central and peripheral effects have been extensively studied, the role of locally produced leptin by bone itself has not yet been fully explored. Therefore, we assessed the role of endogenous leptin on osteogenic differentiation of human mesenchymal stromal cells (MSC). Leptin and leptin receptor (LEPR) mRNA were expressed in both MSC-derived osteoblasts and adipocytes but the level of leptin in osteoblasts was 10-fold higher compared to adipocytes. This was confirmed at the protein level, using ELISA. Immunocytochemically, leptin was located throughout the cytoplasm. Next, we assessed whether leptin produced by osteoblasts had an autocrine/paracrine effect on osteoblast differentiation by either blocking the binding to LEPR, using a leptin neutralising antibody (nAb), or by using short hairpin RNA (shRNA) against the LEPR. Compared with untreated osteoblasts, mineralisation was strongly reduced following nAb treatment (-80% at day 14, -40% at day 17 of culture). This was corroborated by inhibition (60-100%) of mineralisation by 3 different LEPR shRNAs. Osteoblast marker genes were unaffected in the first 10 days of culture compared with controls by nAb treatment. Interestingly, while the expression of these genes started to decrease in the control condition after 10 days at the onset of mineralisation, neutralising leptin led to a persistent high expression: collagen I (+300% and +400% at days 14 and 17), alkaline phosphatase (+150% at days 14 and 17) and RUNX2 (+300% at day 17) compared with control. In conclusion, osteoblast maturation and mineralisation require endogenously produced leptin, thereby adding complexity to the role of leptin in bone metabolism. We hypothesise that leptin signalling plays a role in transition to the mineralisation phase and that lack of leptin signalling prevents the necessary downregulation of osteoblast differentiation genes and thereby inhibits the differentiation and delays mineralisation.

Disclosure: The authors declared no competing interests.

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