Inflammation-mediated age-dependent effects of casein kinase 2-interacting protein-1 on osteogenesis in mesenchymal stem cells

Background: The casein kinase 2-interacting protein-1 (CKIP-1) is important in the development of osteoblasts and cardiomyocytes. However, the effects of CKIP-1 on osteoblast precursor mesenchymal stem cells (MSCs) remain unclear. This study aimed to determine whether CKIP-1 affects osteogenic differentiation in MSCs and explore the relationship ofCKIP-1and inflammation. Methods: Bone marrow MSCs of CKIP-1 wild type (WT) and knockout (KO) mice were cultivatedin vitro. Cell phenotype was analyzed by flow cytometry, colony formation was detected to study the proliferative ability. Osteogenic and adipogenic induction were performed. The osteogenic ability was explored by alizarin red staining, alkaline phosphatase (ALP) staining and ALP activity detection. Quantitative real-time polymerase chain reaction (qRT-PCR) was carried out to determine the mRNA expression levels of osteoblast marker genes. The adipogenic ability was detected by oil red O staining. Content of the bone was analyzed to observe the differences of bone imaging parameters including trabecular bone volume/tissue volume (BV/TV), bone surface area fraction/trabecular BV, trabecular number (Tb.N), and trabecular spacing (Tb.sp). Interleukin (IL)-1 beta was injected on WT mice of 2 months old and 18 months old, respectively. Difference inCKIP-1expression was detected by RT-PCR and western blot. The relationship betweenCKIP-1and inflammation was explored by RT-PCR and western blot. Results: ALP assays, alizarin red staining, and qRT-PCR showed that MSCs derived fromCKIP-1KO mice exhibited a stronger capability for osteogenesis. Micro-computed tomography detection showed that among 18-month-old mice,CKIP-1KO mice presented significantly higher bone mass compared with WT mice (P = 0.02). No significant difference was observed in 2-month-old mice.In vivodata showed that expression ofCKIP-1was higher in the bone marrow of aging mice than in young mice (4.3-fold increase at the mRNA level,P = 0.04). Finally, the expression levels ofCKIP-1in bone marrow (3.2-fold increase at the mRNA level,P = 0.03) and cultured MSCs were up-regulated on chronic inflammatory stimulation by IL-1 beta. Conclusions: CKIP-1is responsible for negative regulation of MSC osteogenesis with age-dependent effects. Increasing levels of inflammation with aging may be the primary factor responsible for higher expression levels of CKIP-1 but may not necessarily affect MSC aging.