Effect of concentrated growth factors on proliferation and differentiation of human gingival fibroblasts

doi:10.3969/j.issn.2095-4344.1768

Abstract

Abstract:

BACKGROUND: In the blood, there are a large number of growth factors and platelet components with regeneration and repair functions, which can promote the regeneration of soft and hard tissues.
OBJECTIVE: To study the effect of concentrated growth factor extract on the proliferation and osteogenic differentiation of human gingival fibroblasts and to explore the clinical application of concentrated growth factors in oral implantology.
METHODS: Patients admitted at the Department of Stomatology, People’s Hospital of Longhua, Shenzhen were selected. (1) In vitro experiment: Concentrated growth factors were extracted by centrifugation, and human gingival fibroblasts were isolated and cultured. The cells were divided into four groups and cultured in complete culture media containing 0 (control), 25%, 50% and 100% concentrated growth factor extract). (2) In vivo experiment: The affected teeth were extracted followed by dental implantation. Concentrated growth factor was placed to induce the rapid healing of soft and hard tissues. Six months later, two-stage implantation was performed. The modified envelope flap was used to increase the thickness of the labial gingiva and the amount of soft tissue in the proximal and distal areas for 3 months to obtain soft tissue contours in coordination with the adjacent teeth.
RESULTS AND CONCLUSION: (1) In vitro experiment results: Compared with the control group, the proliferation of human gingival fibroblasts was significantly increased by 50% and 100% concentrated growth factor extract on the 3^rd day of culture. On the 14^th day of culture, compared with the control group, the activity of alkaline phosphatase, the molecular weight of mineralization and the expression of osteoblast-associated genes RUNX2 and osteocalcin were significantly increased in the 100% concentrated growth factor extract group. (2) In vivo experiment results: Concentrated growth factor extract could promote bone regeneration around oral implants. There was no obvious retraction at the gingival margin, and the gingival papilla filled the space of the teeth. The bone mass around the implant increased significantly at 14 months in relative to 3 months after implantation. To conclude, concentrated growth factor extract can promote the proliferation and osteogenic differentiation of human gingival fibroblasts and the peri-implant regeneration of soft and hard tissue.

Key words: concentrated growth factor, concentrated growth factor extract, platelets, human gingival fibroblasts, proliferation, differentiation, oral implantation, tissue construction, soft tissue regeneration, bone tissue regeneration

CLC Number:

R452
R783

Li Xiaoju, Song Guangbao, Yang Jianzhen, Wei Bo, Wu Bin. Effect of concentrated growth factors on proliferation and differentiation of human gingival fibroblasts[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(21): 3342-3348.

Figures/Tables 2

2.1 受试者数量分析参与试验的患者均进入结果分析，中途无退出者。 2.2 各组CCK-8法检测人牙龈成纤维细胞增殖变化实验第1天，100%浓缩生长因子萃取液组与对照组相比，人牙龈成纤维细胞增殖的效果显著(P < 0.05)。在实验第2，3天，与对照组相比，50%浓缩生长因子萃取液组增殖的效果显著(P < 0.05)，100%浓缩生长因子萃取液组数值显著增加(P < 0.01)；而25%浓缩生长因子萃取液组与对照组相比，人牙龈成纤维细胞增殖的促进效果不显著(P > 0.05)，见图2。 2.3 各组碱性磷酸酶活性变化经过7，14天的培养，与对照组相比，100%浓缩生长因子萃取液组碱性磷酸酶活性显著升高(P < 0.01)，同时50%浓缩生长因子萃取液组在这2个时间点与对照组相比碱性磷酸酶活性均升高(P < 0.05)，见图3。 2.4 成骨诱导培养基培养在干预培养21 d后，两组细胞在显微镜下均可见无定形物质(显微镜：IX73，Olympus Corporation，Tokyo，Japan)。并且经过茜素红染色后，这些无定形物质均呈深红色结节状，形态多种多样，并且100%浓缩生长因子萃取液组比对照组形成更多的钙结节，见图4，这进一步说明浓缩生长因子萃取液能够促进人牙龈成纤维细胞的成骨分化。 2.5 各组成骨相关基因表达在干预培养第3，7，14天，对成骨相关基因RUNX2和骨钙素的表达进行RT-PCR检测，结果见图5，6。在干预培养第3，7，14天，与对照组相比，25%与50%浓缩生长因子萃取液组中RUNX2表达量升高(P < 0.05)；100%浓缩生长因子萃取液组中RUNX2表达量显著增高(P < 0.01)。在干预培养第7，14天，与对照组相比，25%，50%浓缩生长因子萃取液组中骨钙素表达量增加(P < 0.05)，100%浓缩生长因子萃取液组中骨钙素表达量有显著增高(P < 0.01)。实验结果表明浓缩生长因子萃取液能够显著上调成骨相关基因RUNX2和骨钙素的表达。 2.6 口腔种植形态上颌左侧中切牙即刻种植并同期置入浓缩生长因子膜后，软组织愈合良好，通过暂冠塑形获得比较满意的穿龈轮廓，戴入永久冠后牙龈轮廓和形态得到较好的维持，龈缘无明显退缩，龈乳头获得较好的充盈。X射线影像片检查显示种植体形成了良好的骨整合，术后14个月与术后3个月对比，种植体周围骨量增加，见图7。"

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