Department of Anatomy, Guangdong Provincial Key Laboratory of Medicine and Biomechanics, School of Basic Medical Sciences, Southern Medical University, Guangzhou, 510515, China.
Departments of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, China.
Stem Cell Res Ther. 2018 May 2;9(1):125. doi: 10.1186/s13287-018-0836-y.
Adipose-derived stem cells (ASCs) that show multidifferentiation and anti-immune rejection capacities have been widely used in plastic and reconstructive surgery. Previous studies have indicated that mechanical and biophysical interactions between cells and their surrounding environment regulate essential processes, such as growth, survival, and differentiation, and the cytoskeleton system plays an important role in the mechanotransduction. However, the role of mechanical force in the determination of lineage fate is still unclear.
Human ASCs (hASCs) were obtained from three different donors by liposuction. Adipogenesis and osteogenesis were determined by Oil Red O and Alizarin Red staining, respectively. The mRNA levels of the cytoskeleton system, PPARγ, and C/EBPα were determined by real-time polymerase chain reaction (RT-PCR). The level of cytoskeleton, PPARγ, and C/EBPα protein levels were measured by Western blotting. The morphology of the cytoskeleton system during adipogenesis was observed with confocal microscopy. hASCs were transfected with a SUN2-specific shRNA to knockdown sun2, and a nontargeting shRNA was used as a control.
We found that disrupting the physiological balance between the cytoskeleton and the linker of the nucleoskeleton and cytoskeleton (LINC) complex (especially SUN2) could impact the adipogenesis of hASCs in vitro. Microtubule (MT) depolymerization with nocodazole (which interferes with the polymerization of MTs) increased the expression of SUN2 and PPARγ, while taxol (an inhibitor of MT disassembly) showed the opposite results. Meanwhile, hASCs with sun2 knockdown overexpressed MTs and decreased PPARγ expression, thereby inhibiting the adipogenesis. Furthermore, knockdown of sun2 changed the structure of perinuclear MTs.
We demonstrated the presence of cross-talk between MT and SUN2, and this cross-talk plays a critical role in the rebalance of the mechanical environment and is involved in the regulation of PPARγ transport during adipogenic differentiation of hASCs.
脂肪来源的干细胞（ASCs）具有多向分化和抗免疫排斥能力，已广泛应用于整形和重建外科。先前的研究表明，细胞与其周围环境之间的机械和生物物理相互作用调节着重要的过程，如生长、存活和分化，细胞骨架系统在机械转导中起着重要作用。然而，机械力在决定谱系命运中的作用仍不清楚。
通过脂肪抽吸从 3 位不同供体中获得人脂肪干细胞（hASCs）。通过油红 O 和茜素红染色分别确定脂肪生成和成骨。通过实时聚合酶链反应（RT-PCR）测定细胞骨架系统、PPARγ 和 C/EBPα 的 mRNA 水平。通过 Western blot 测定细胞骨架、PPARγ 和 C/EBPα 蛋白水平。用共聚焦显微镜观察细胞骨架系统在脂肪生成过程中的形态。用 SUN2 特异性 shRNA 转染 hASCs 以敲低 sun2，并用非靶向 shRNA 作为对照。
我们发现，破坏细胞骨架与核骨架和细胞骨架连接体（LINC）复合物（特别是 SUN2）之间的生理平衡会影响 hASCs 的体外脂肪生成。用长春新碱（干扰微管聚合）去聚合微管会增加 SUN2 和 PPARγ 的表达，而紫杉醇（微管解聚抑制剂）则相反。同时，SUN2 敲低的 hASCs 过度表达微管并降低 PPARγ 表达，从而抑制脂肪生成。此外，SUN2 敲低改变了核周微管的结构。
我们证明了 MT 和 SUN2 之间存在交叉对话，这种对话在机械环境的再平衡中起着关键作用，并参与调节 hASCs 脂肪生成分化过程中 PPARγ 的运输。
