Intracellular density is a novel indicator of differentiation stages of murine osteoblast lineage cells

Abstract

Osteoblasts are primary bone‐making cells originating from mesenchymal stem cells (MSCs) in the bone marrow. The differentiation of MSCs to mature osteoblasts involves an intermediate stage called preosteoblasts, but the details of this process remain unclear. This study focused on the intracellular density of immature osteoblast lineage cells and hypothesized that the density might vary during differentiation and might be associated with the differentiation stages of osteoblast lineage cells. This study aimed to clarify the relationship between intracellular density and differentiation stages using density gradient centrifugation. Primary murine bone marrow stromal cell cultures were prepared in an osteogenic induction medium, and cells were separated into three fractions (low, intermediate, and high‐density). The high‐density fraction showed elevated expression of osteoblast differentiation markers (Sp7, Col1a1, Spp1, and Bglap) and low expression of MSC surface markers (Sca‐1, CD73, CD105, and CD106). In contrast, the low‐density fraction showed a high expression of MSC surface markers. These results indicated that intracellular density increased during differentiation from preosteoblasts to committed osteoblasts. Intracellular density may be a novel indicator for osteoblast differentiation stages. Density gradient centrifugation is a novel technique to study the process by which preosteoblasts transform into bone‐forming cells.

Regulon active landscape reveals cell development and functional state changes of human primary osteoblasts in vivo

“…During osteogenesis in mice, the expression of BM-MSC-related markers Lepr [ 33 ] and Vcam1 [ 34 ] gradually decreased over pseudotime. Further, the expression of osteoblast-related markers Runx2 [ 8 ] and Bglap [ 35 ] gradually increased as osteogenesis progressed (Fig. 12 M).…”

Single-cell RNA sequencing reveals in vivo osteoimmunology interactions between the immune and skeletal systems

“…For different subclusters of OBC, cell trajectory inference analysis results showed that OBC-2 was the late-stage cell compared with early-stage subcluster OBC-1 ( Figures 1C, D ). OBC-1 also highly expressed BMSCs markers such as LEPR ( 38 ) and VCAM1 ( 39 ); while OBC-2 highly expressed osteogenic markers such as RUNX2, BGLAP and COL1A1 ( 40 , 41 ) ( Figure 1E ). A branch heatmap ( Figure 1F ) of the branch point 2 in Figures 1C , 2D further showed a tendency toward up-regulation of ossification related genes ( Figure 1G ) in cell fate 1 (late-stage cell fate, left branch after branch point 2 in Figure 1D ).…”