Yanagi Tsukasa, Kajiya Hiroshi, Fujisaki Seiichi, Maeshiba Munehisa, Yanagi-S Ayako, Yamamoto-M Nana, Kakura Kae, Kido Hirofumi, Ohno Jun
Oral Medicine Research Center, Fukuoka Dental College, 2-15-1Tamura, Fukuoka, Japan.
Department of Oral Rehabilitation, Fukuoka Dental College, 2-15-1Tamura, Fukuoka, Japan.
Regen Ther. 2021 Nov 11;18:472-479. doi: 10.1016/j.reth.2021.10.004. eCollection 2021 Dec.
Mesenchymal stromal/stem cells (MSCs) are multipotent, self-renewing cells that are extensively used in tissue engineering. Dedifferentiated fat (DFAT) cells are derived from adipose tissues and are similar to MSCs. Three-dimensional (3D) spheroid cultures comprising MSCs mimic the biological microenvironment more accurately than two-dimensional cultures; however, it remains unclear whether DFAT cells in 3D spheroids possess high osteogenerative ability. Furthermore, it is unclear whether DFAT cells from 3D spheroids transplanted into calvarial bone defects are as effective as those from two-dimensional (2D) monolayers in promoting bone regeneration.
We compared the osteogenic potential of rat DFAT cells cultured under osteogenic conditions in 3D spheroids with that in 2D monolayers. Furthermore, to elucidate the ability of 3D spheroid DFAT cells to promote bone healing, we examined the osteogenic potential of transplanting DFAT cells from 3D spheroids or 2D monolayers into a rat calvarial defect model.
Osteoblast differentiation stimulated by bone morphogenetic protein-2 (BMP-2) or osteogenesis-inducing medium upregulated osteogenesis-related molecules in 3D spheroid DFAT cells compared with 2D monolayer DFAT cells. BMP-2 activated phosphorylation in the canonical Smad 1/5 pathways in 3D spheroid DFAT cells but phosphorylated ERK1/2 and Smad2 in 2D monolayer DFAT cells. Regardless of osteogenic stimulation, the transplantation of 3D DFAT spheroid cells into rat calvarial defects promoted new bone formation at a greater extent than that of 2D DFAT cells.
Compared with 2D DFAT cells, 3D DFAT spheroid cells promote osteoblast differentiation and new bone formation via canonical Smad 1/5 signaling pathways. These results indicate that transplantation of DFAT cells from 3D spheroids, but not 2D monolayers, accelerates bone healing.
间充质基质/干细胞（MSCs）是多能的自我更新细胞，广泛应用于组织工程。去分化脂肪（DFAT）细胞来源于脂肪组织，与MSCs相似。与二维培养相比，包含MSCs的三维（3D）球体培养能更准确地模拟生物微环境；然而，3D球体中的DFAT细胞是否具有高成骨能力仍不清楚。此外，尚不清楚移植到颅骨缺损处的3D球体DFAT细胞在促进骨再生方面是否与二维（2D）单层DFAT细胞一样有效。
我们比较了在成骨条件下培养的大鼠DFAT细胞在3D球体和2D单层中的成骨潜力。此外，为了阐明3D球体DFAT细胞促进骨愈合的能力，我们检测了将3D球体或2D单层的DFAT细胞移植到大鼠颅骨缺损模型中的成骨潜力。
与2D单层DFAT细胞相比，骨形态发生蛋白-2（BMP-2）或成骨诱导培养基刺激的成骨细胞分化上调了3D球体DFAT细胞中成骨相关分子。BMP-2激活了3D球体DFAT细胞中经典Smad 1/5信号通路的磷酸化，但在2D单层DFAT细胞中磷酸化了ERK1/2和Smad2。无论成骨刺激如何，将3D DFAT球体细胞移植到大鼠颅骨缺损处比2D DFAT细胞更能促进新骨形成。
与2D DFAT细胞相比，3D DFAT球体细胞通过经典Smad 1/5信号通路促进成骨细胞分化和新骨形成。这些结果表明，移植3D球体而非2D单层的DFAT细胞可加速骨愈合。
