Knockdown of CDC20 Promotes Adipogenesis of Bone Marrow-derived Stem Cells by Modulating β-catenin

Background: Bone is a rigid organ that provides physical protection and support to vital organs of the body. Bone loss disorders are commonly associated with increased bone marrow adipose tissue. Bone marrow mesenchymal stromal/stem cells (BMSCs) are multipotent progenitors that can differentiate into osteoblasts, adipocytes, and chondrocytes. Cell division cycle 20 (CDC20) is a co-activator of anaphase promoting complex/cyclosome (APC/C), and is required for ubiquitin ligase activity. Our previous study showed that CDC20 promoted the osteogenic commitment of BMSCs and Cdc20 conditional knockout mice suggested a decline in bone mass. In this study, we found that knockdown of CDC20 promoted adipogenic differentiation of BMSCs by modulating β-catenin, which suggested a link between adipogenesis and osteogenesis.

Methods: Lentivirus containing a CDC20 shRNA was used for CDC20 knockdown in human BMSCs (hBMSCs). Primary mouse BMSCs (mBMSCs) were isolated from Cdc20 and Sp7-Cre;Cdc20 mice. Adipogenesis was examined using quantitative real-time reverse transcription PCR (qRT-PCR) and western blotting analysis of adipogenic regulators, Oil Red O staining, and transplantation into nude mice. CDC20 knockout efficiency was determined through immunochemistry, qRT-PCR, and western blotting of bone marrow. Accumulation of adiposity was measured through histology and staining of bone sections. Exploration of the molecular mechanism was determined through western blotting, Oil Red O staining, and qRT-PCR.

Results: CDC20 expression in hBMSCs was significantly decreased during adipogenic differentiation. CDC20 knockdown enhanced hBMSC adipogenic differentiation in vitro. CDC20-knockdown hBMSCs showed more adipose tissue-like constructs upon hematoxylin and eosin (H&E) and Oil Red O staining. Sp7-Cre;Cdc20 mice presented increased adipocytes in their bone marrow compared with the control mice. mBMSCs from Sp7-Cre;Cdc20 mice showed upregulated adipogenic differentiation. Knockdown of CDC20 led to decreased β-catenin levels, and a β-catenin pathway activator (lithium chloride) abolished the role of CDC20 in BMSC adipogenic differentiation.

Conclusions: Our findings showed that CDC20 knockdown enhanced adipogenesis of hBMSC and mBMSCs adipogenesis in vitro and in vivo. CDC20 regulates both adipogenesis and osteogenesis of BMSCs, and might lead to the development of new therapeutic targets for "fatty bone" and osteoporosis.