PDF(Pubmed)
Abstract：
UNASSIGNED: Stem cell therapy for regenerative medicine has been sincerely investigated, but not still popular although some clinical trials show hopeful results. This therapy is suggested to be a representative candidate such as bone defect due to the accident, iatrogenic resection oncological tumor, congenital disease, and severe periodontitis in oral region. Recently, the Bio-3D printer \"Regenova®\" has been introduced as an innovative three-dimensional culture system, equipped scaffold-free bio-assembling techniques without any biomaterials. Therefore, we expected a mount of bone defect could be repaired by the structure established from this Bio-3D printer using osteogenic potential stem cells.
UNASSIGNED: The gingival tissue (1x1 mm) was removed from the distal part of the lower wisdom tooth of the patients who agreed our study. Human Gingival Mesenchymal Stem Cells (hGMSCs) were isolated from this tissue and cultured, since we confirmed the characteristics such as facile isolation and accelerated proliferation, further, strong potential of osteogenic-differentiation. Spheroids were formed using hGMSC in 96-well plates designed for low cell adhesion. The size of the spheroids was measured, and fluorescent immunostaining was employed to verify the expression of stem cell and apoptosis marker, and extracellular matrix. Following four weeks of bone differentiation, μCT imaging was performed. Calcification was confirmed by alizarin red and von Kossa staining. Fluorescent immunostaining was utilized to assess the expression of markers indicative of advanced bone differentiation.
UNASSIGNED: We have established and confirmed the spheroids (∼600 μm in diameter) constructed from human GMSCs (hGMSCs) still maintain stem cell potentials and osteogenic differentiation abilities from the results that CD73 and not CD34 were expressed as stem cell positive and negative marker, respectively. These spheroids were pilled up like cylindal shape to the \"Kenzan\" platform of Bio-3D printer and cultured for 7days. The cylindal structure originated from compound spheroids were tried to differentiate into bone four weeks with osteogenic induction medium. The calcification of bio-3D printed bone-like structures was confirmed by alizarin red and Von Kossa staining. In addition, μCT analysis revealed that the HU (Hounsfield Unit) of the calcified structures was almost identical to that of trabecular bone. Immunofluorescent staining detected osteocalcin expression, a late-stage bone differentiation marker.
UNASSIGNED: For the first time, we have achieved the construction of a scaffold-free, bone-like luminal structure through the assembly of spheroids comprised of this hGMSCs. This success is sure to be close to the induction of clinical application against regenerative medicine especially for bone defect disease.
UNASSIGNED: The gingival tissue (1x1 mm) was removed from the distal part of the lower wisdom tooth of the patients who agreed our study. Human Gingival Mesenchymal Stem Cells (hGMSCs) were isolated from this tissue and cultured, since we confirmed the characteristics such as facile isolation and accelerated proliferation, further, strong potential of osteogenic-differentiation. Spheroids were formed using hGMSC in 96-well plates designed for low cell adhesion. The size of the spheroids was measured, and fluorescent immunostaining was employed to verify the expression of stem cell and apoptosis marker, and extracellular matrix. Following four weeks of bone differentiation, μCT imaging was performed. Calcification was confirmed by alizarin red and von Kossa staining. Fluorescent immunostaining was utilized to assess the expression of markers indicative of advanced bone differentiation.
UNASSIGNED: We have established and confirmed the spheroids (∼600 μm in diameter) constructed from human GMSCs (hGMSCs) still maintain stem cell potentials and osteogenic differentiation abilities from the results that CD73 and not CD34 were expressed as stem cell positive and negative marker, respectively. These spheroids were pilled up like cylindal shape to the \"Kenzan\" platform of Bio-3D printer and cultured for 7days. The cylindal structure originated from compound spheroids were tried to differentiate into bone four weeks with osteogenic induction medium. The calcification of bio-3D printed bone-like structures was confirmed by alizarin red and Von Kossa staining. In addition, μCT analysis revealed that the HU (Hounsfield Unit) of the calcified structures was almost identical to that of trabecular bone. Immunofluorescent staining detected osteocalcin expression, a late-stage bone differentiation marker.
UNASSIGNED: For the first time, we have achieved the construction of a scaffold-free, bone-like luminal structure through the assembly of spheroids comprised of this hGMSCs. This success is sure to be close to the induction of clinical application against regenerative medicine especially for bone defect disease.
摘要：
■用于再生医学的干细胞疗法已经得到了真诚的研究，但仍然不受欢迎，尽管一些临床试验显示出希望的结果。该疗法被认为是由于事故导致的骨缺损等代表性候选，医源性肿瘤切除，先天性疾病,和口腔区域的严重牙周炎。最近,Bio-3D打印机“Regenova®”作为一种创新的三维文化系统被引入，配备无支架生物组装技术，无需任何生物材料。因此,我们预计，通过使用成骨潜能干细胞从Bio-3D打印机建立的结构，可以修复大量的骨缺损。
■从同意我们研究的患者的下智齿远端部分去除牙龈组织（1x1mm）。从该组织中分离人牙龈间充质干细胞(hGMSCs)并进行培养,由于我们证实了容易分离和加速增殖等特征，进一步,成骨分化的强大潜力。使用hGMSC在设计用于低细胞粘附的96孔板中形成球状体。测量球体的大小，用荧光免疫染色验证干细胞和凋亡标志物的表达,和细胞外基质。骨分化四周后，进行μCT成像。通过茜素红和vonKossa染色证实钙化。利用荧光免疫染色来评估指示晚期骨分化的标志物的表达。
■我们已经建立并证实了由人GMSCs（hGMSCs）构建的球体（直径约600μm）仍然保持干细胞潜能和成骨分化能力，结果CD73而不是CD34表达为干细胞阳性和阴性标记。分别。这些球体像圆柱形一样堆积到Bio-3D打印机的“Kenzan”平台上，并培养7天。尝试使用成骨诱导培养基将源自复合球体的圆柱形结构分化为骨骼4周。通过茜素红和VonKossa染色证实了生物3D打印的骨样结构的钙化。此外,μCT分析显示，钙化结构的HU（Hounsfield单位）几乎与小梁骨相同。免疫荧光染色检测骨钙蛋白的表达，晚期骨分化标志物。
■第一次，我们已经实现了无脚手架的建设，通过由这种hGMSC组成的球状体的组装形成骨样管腔结构。这一成功肯定会接近诱导针对再生医学的临床应用，特别是针对骨缺损疾病。
■从同意我们研究的患者的下智齿远端部分去除牙龈组织（1x1mm）。从该组织中分离人牙龈间充质干细胞(hGMSCs)并进行培养,由于我们证实了容易分离和加速增殖等特征，进一步,成骨分化的强大潜力。使用hGMSC在设计用于低细胞粘附的96孔板中形成球状体。测量球体的大小，用荧光免疫染色验证干细胞和凋亡标志物的表达,和细胞外基质。骨分化四周后，进行μCT成像。通过茜素红和vonKossa染色证实钙化。利用荧光免疫染色来评估指示晚期骨分化的标志物的表达。
■我们已经建立并证实了由人GMSCs（hGMSCs）构建的球体（直径约600μm）仍然保持干细胞潜能和成骨分化能力，结果CD73而不是CD34表达为干细胞阳性和阴性标记。分别。这些球体像圆柱形一样堆积到Bio-3D打印机的“Kenzan”平台上，并培养7天。尝试使用成骨诱导培养基将源自复合球体的圆柱形结构分化为骨骼4周。通过茜素红和VonKossa染色证实了生物3D打印的骨样结构的钙化。此外,μCT分析显示，钙化结构的HU（Hounsfield单位）几乎与小梁骨相同。免疫荧光染色检测骨钙蛋白的表达，晚期骨分化标志物。
■第一次，我们已经实现了无脚手架的建设，通过由这种hGMSC组成的球状体的组装形成骨样管腔结构。这一成功肯定会接近诱导针对再生医学的临床应用，特别是针对骨缺损疾病。
