Mizoguchi Takahiro, Ueno Koji, Takeuchi Yuriko, Samura Makoto, Suzuki Ryo, Murata Tomoaki, Hosoyama Tohru, Morikage Noriyasu, Hamano Kimikazu
Departments of Surgery and Clinical Sciences, Graduate School of Medicine, Yamaguchi University, Yamaguchi, Japan.
Institute of Laboratory Animals, Yamaguchi University, Yamaguchi, Japan.
Cell Physiol Biochem. 2018;47(1):201-211. doi: 10.1159/000489767. Epub 2018 May 10.
BACKGROUND/AIMS: We have developed a mixed-cell sheet consisting of autologous fibroblasts and peripheral blood mononuclear cells with a high potency for angiogenesis and wound healing against refractory cutaneous ulcers in mouse and rabbit models. To increase the effectiveness of the mixed sheet, we developed a multilayered mixed sheet.
We assessed the therapeutic effects of multilayered sheets on cutaneous ulcers in mice. Growth factors and chemokines were assessed by enzyme-linked immunosorbent assay. Angiogenesis and fibroblast migration were measured by using tube formation and migration assays. Wound healing rate of cutaneous ulcers was evaluated in mice with diabetes mellitus.
The concentration of secreted vascular endothelial growth factor, hepatocyte growth factor, transforming growth factor, C-X-C motif chemokine ligand (CXCL)-1, and CXCL-2 in multilayered sheets was much higher than that in single-layered mixed-cell sheets (single-layered sheets) and multilayered sheets of fibroblasts alone (fibroblast sheets). The supernatant in multilayered sheets enhanced angiogenic potency and fibroblast migration compared with single-layered and fibroblast sheets in an in vitro experiment. The wound healing rate in the multilayered sheet-treated group was higher compared with the no-treatment group (control) at the early stage of healing. Moreover, both vessel lumen area and microvessel density in tissues treated with multilayered sheets were significantly increased compared with tissues in the control group.
Multilayered sheets promoted wound healing and microvascular angiogenesis in the skin by supplying growth factors and cytokines. Accordingly, our data suggest that multilayered sheets may be a promising therapeutic material for refractory cutaneous ulcers.
背景/目的：我们已经研发出一种由自体成纤维细胞和外周血单核细胞组成的混合细胞片，在小鼠和兔模型中，其对难治性皮肤溃疡具有高效的血管生成和伤口愈合能力。为了提高混合细胞片的有效性，我们开发了一种多层混合细胞片。
我们评估了多层细胞片对小鼠皮肤溃疡的治疗效果。通过酶联免疫吸附测定法评估生长因子和趋化因子。使用管腔形成和迁移测定法测量血管生成和成纤维细胞迁移。在糖尿病小鼠中评估皮肤溃疡的伤口愈合率。
多层细胞片中分泌的血管内皮生长因子、肝细胞生长因子、转化生长因子、C-X-C基序趋化因子配体（CXCL）-1和CXCL-2的浓度远高于单层混合细胞片（单层细胞片）和仅成纤维细胞的多层细胞片（成纤维细胞片）。在体外实验中，与单层细胞片和成纤维细胞片相比，多层细胞片的上清液增强了血管生成能力和成纤维细胞迁移。在愈合早期，多层细胞片治疗组的伤口愈合率高于未治疗组（对照组）。此外，与对照组组织相比，多层细胞片治疗组织的血管腔面积和微血管密度均显著增加。
多层细胞片通过提供生长因子和细胞因子促进皮肤伤口愈合和微血管生成。因此，我们的数据表明多层细胞片可能是治疗难治性皮肤溃疡的一种有前景的治疗材料。
