Abstract
BackgroundAdipose-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.MethodsHuman 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.ResultsWe 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.ConclusionsWe 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.
Highlights
Adipose-derived stem cells (ASCs) that show multidifferentiation and anti-immune rejection capacities have been widely used in plastic and reconstructive surgery
We investigated the changes in microtubules and the linker of the nucleoskeleton and cytoskeleton (LINC) complex, and we examined its regulation by Sad1/UNC-84 2 (SUN2) during adipogenesis in Human adipose-derived stem cell (hASC) to demonstrate the interaction between microtubules and the LINC complex and the possible mechanisms of adipogenic differentiation of ASCs
For adipogenic and osteogenic differentiation, Oil Red O and Alizarin Red staining were performed as described previously [16, 17]. hASCs between three and six passages were used in the experiments, and all experiments were repeated at least three times using hASCs from three donors. hASCs were cultured in growth medium consisting of high-glucose Dulbecco’s modified Eagle’s medium (DMEM; Gibco), 10% fetal bovine serum (FBS; Gibco), and 1% penicillin/streptomycin (Gibco)
Summary
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. The role of mechanical force in the determination of lineage fate is still unclear. The regulation of stem cell differentiation by soluble factors is well characterized, the role of mechanical force in the determination of lineage fate is still unclear. Physical cues play an important role in the determination of lineage fate in stem cells. Mechanical and biophysical interactions between cells and their surrounding environment regulate essential processes, such as growth, survival [3], and differentiation [4]. The giant isoforms of Nesprin-1 and Nesprin-2 interact with actin via their N-terminal actin-binding domain, whereas other isoforms can associate with microtubule motors [9, 10]
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