ABSTRACT

Purpose: Adipose-derived stem cells (ADSCs) are ideal for cell-based therapies to support bone regeneration. It is vital to understand the critical genes and molecular mechanisms involved in the functional regulation of ADSCs for enhancing bone regeneration. In the present study, we investigated the Gremlin 1 (GREM1) effect on ADSCs osteogenic differentiation and senescence.

Materials and methods: The in vitro ADSCs osteogenic differentiation potential was evaluated by determining alkaline phosphatase (ALP) activity, mineralization ability, and the expression of osteogenic markers. Cell senescence is determined by SA-β-gal staining, telomerase assay, and the expression of aging markers.

Results: GREM1 overexpression in ADSCs reduced ALP activity and mineralization, inhibited the expression of osteogenic related genes OCN, OPN, DSPP, DMP1, and BSP, and key transcription factors, RUNX2 and OSX. GREM1 knockdown in ADSCs enhanced ALP activity and mineralization, promoted the expression of OCN, OPN, DSPP, DMP1, BSP, RUNX2, and OSX. GREM1 overexpression in ADSCs reduced the percent SA-β-Gal positive cells, P16 and P53 expressions, and increased telomerase activity. GREM1 knockdown in ADSCs increased the percentage of SA-β-Gal positive cells, P16 and P53 expressions, and reduced telomerase activity. Furthermore, GREM1 reduced the mRNA expression levels of BMP2, BMP6, and BMP7.

Conclusions: In summary, our findings suggested that GREM1 inhibited ADSCs senescence and osteogenic differentiation and antagonized BMP transcription.

摘要

目的：脂肪干细胞 (ADSC) 是支持骨再生的细胞疗法的理想选择。了解参与 ADSC 功能调节以增强骨再生的关键基因和分子机制至关重要。在本研究中，我们研究了 Gremlin 1 (GREM1) 对 ADSCs 成骨分化和衰老的影响。

材料和方法：通过测定碱性磷酸酶（ALP）活性、矿化能力和成骨标志物的表达来评估体外ADSCs成骨分化潜能。细胞衰老通过 SA-β-gal 染色、端粒酶测定和衰老标志物的表达来确定。

结果： ADSCs中GREM1过表达降低了ALP活性和矿化，抑制了成骨相关基因OCN、OPN、DSPP、DMP1和BSP以及关键转录因子RUNX2和OSX的表达。ADSCs 中的 GREM1 敲低增强了 ALP 活性和矿化，促进了OCN、OPN、DSPP、DMP1、BSP、RUNX2和OSX的表达。ADSCs 中 GREM1 过表达降低了 SA-β-Gal 阳性细胞百分比、P16和P53表达，并增加了端粒酶活性。ADSC 中的 GREM1 敲低增加了 SA-β-Gal 阳性细胞、P16和P53的百分比表达，并降低端粒酶活性。此外，GREM1 降低了 BMP2、BMP6 和 BMP7 的 mRNA 表达水平。

结论：综上所述，我们的研究结果表明 GREM1 抑制 ADSCs 衰老和成骨分化并拮抗 BMP 转录。