# Potential Marker Genes for Predicting Adipogenic Differentiation of Mesenchymal Stromal Cells
## 4 Discussion  
In our previous study, we selected 95 genes as candidate MSC marker genes based on microarray analysis data [10]. Although the 95 genes may not be enough to identify useful markers, and other genes may serve as more reliable markers, these 95 genes can be a promising starting point for further analysis. In this study, we correlated mRNA expression levels in MSCs before induction with GPDH activities after adipogenic differentiation in order to identify marker genes for MSCs with high adipogenic potential. Twenty-two out of the 95 genes showed the expression profiles significantly correlated with GPDH activities. Expression levels of 11 out of the 22 genes in ilium MSCs, which show high differentiation potential, were more than three times higher than those in jaw MSCs, which have only limited potential. We selected the 11 genes, ITGA5, MCAM, GPR37, ACLY, DNCII, P4 HA2, LIF, CDKN2D, CD74, AURKB, and SLC2A1, as potential markers. Although none of these genes accurately distinguished ilium MSCs from jaw MSCs, combined 3D analysis of CDKN2D, ITGA5, and CD74 mRNA expression allowed us to completely separate the two types of MSCs. Thus, the 3D analysis can provide an effective strategy to select MSCs with high adipogenic potential. All 11 genes identified in the present study are involved in cell growth and/or cell cycle regulation [14; 15; 16; 17; 18; 19; 20; 21; 22; 23; 24]. ITGA5 codes the important adhesion molecule involved in adipogenesis [14], and ACLY codes a key enzyme in fat synthesis [17; 25]. MCAM (CD146) regulates the proliferation and differentiation of MSCs [15] and serves as a cell surface marker for predicting the potential of MSCs to differentiate into chondrocytes [8]. Although our results suggest that ITGA5 plays a role in the differentiation of MSCs at the early stage of adipogenesis, the exact function of these genes in adipogenesis remains to be investigated in future studies. The differentiation potential and direction of MSCs seems to differ depending on their origins, although MSCs can be obtained from various tissues, including the bone marrow, adipose tissue, synovium, and dental pulp. [PERSON] et al. [5] found that MSCs derived from alveolar bone marrow have poor adipogenic or chondrogenic differentiation potential, but high osteogenic differentiation  
Figure 3: Effects of the anti-ITGA5 antibody on adipogenic differentiation of ilium MSCs. (**a**) MSCs were treated with the anti-ITGA5 or control IgG for 21 days during adipogenic induction and stained with oil red O (scale bar, 70 \(\upmu\)m). (**b**) Oil droplet area was quantified by using ImageJ software. Data are presented as mean percent area \(\pm\) standard error (\(n=3\)). Student’s \(t\)-test, **\(p<0.01\)**. potential similar to ilium MSCs. On the other hand, MSCs from synovium are superior to ilium MSCs in both adipogenesis and chondrogenesis [26]. In this study, we also found bone marrow MSCs obtained from the jaw to have a limited capacity to differentiate into adipocytes as compared with bone marrow MSCs from the ilium, although jaw MSCs have a high ability to become osteoblasts. Furthermore, [PERSON] et al. [27] demonstrated that MSCs from adipose tissues have a higher adipogenic ability than MSCs from bone marrow. In contrast, bone marrow MSCs showed a higher ability to differentiate into osteoblasts and chondrocytes than adipose MSCs. However, differentiation levels determined by oil red O staining varied widely among donors of MSCs. In addition, MSCs obtained from adipose tissues have a reduced proliferative capacity [28; 29]. Since MSCs from the ilium bone marrow have a high proliferative and multipotent capacity to differentiate into various types of cells, we speculate that ilium bone marrow MSCs have a high adipogenic differentiation potential, and can serve as materials for soft tissue engineering, although their differentiation potential varies among cell lines. In most previous studies, preadipocytes have been used for tissue engineering of soft connective tissues [30].