[Initial osteoblast functions on a type of near β-type titanium alloys surfaces modified by the double glow plasma nitriding technology].

Objective: To evaluate the adhesion, proliferation, alkaline phosphatase (ALP) activity and the expression of osteogenesis-related genes and osteoprotegerin (OPG)/receptor activator of NF-κB ligand (RANKL) of osteoblast-like cells on a type of near β-type titanium alloys (Ti-5Zr-3Sn-5Mo-15Nb, TLM) surfaces modified by the double glow plasma nitriding technology, and to investigate the effect of the modified surfaces on the initial functions of osteoblast-like cells. Methods: The surfaces of TLM were modified by the double glow plasma nitriding technology. TLM surfaces without modification were used as control. Cell morphology was observed with scanning electron microscopy (SEM). Methyl thiazolyl tetrazolium (MTT) method was used to measure cell proliferation. Cell ALP activity was evaluated by using reagent kits. The mRNA expression of Runt-related transcription factor-2 (RUNX2), typeⅠcollagen alpha 1 chain (COLⅠ α1) and OPG/RANKL were examined by quantitative real-time PCR(qRT-PCR). Results: Four hour following cell alture, cells on modified surfaces extend filopodia and intercellular junction was tight. Three days later, cell proliferation (0.277±0.007) was significantly higher than that in control group (0.249±0.004) (P<0.01). After two weeks, ALP activity on TLM modified layer (173.6±1.89) was significantly higher than that on unmodified TLM (162.6±2.4) (P<0.01). The mRNA expression of osteoblast marker RUNX2, COLⅠα1 were stronger than that in control group (P<0.05). The expression of OPG mRNA was higher than that in control group (P<0.01), and RANKL mRNA expression was significantly lower than that in control group (P<0.05). Conclusions: The TLM surface modified by the double glow plasma nitriding technology has a positive effect on osteoblasts initial adhesion, proliferation and differentiation, and it can also improve expression of OPG mRNA and has an inhibitory effect on RANKL mRNA expression of osteoblasts.目的： 研究近β钛合金（Ti-5Zr-3Sn-5Mo-15Nb，TLM）渗氮改性层对成骨细胞早期功能的影响，为探索口腔种植新材料提供依据。 方法： 圆片形TLM试样经抛光、清洗后均分为两组（每组20个），渗氮组表面采用双层辉光离子渗氮技术进行渗氮处理，另一组不处理作为对照组。将小鼠颅顶前骨细胞亚克隆14接种于试样表面，扫描电镜观察培养4 h后细胞的黏附形态；甲基噻唑基四唑法检测3、5、7 d后细胞增殖情况；检测7、14 d后细胞碱性磷酸酶（alkaline phosphatase，ALP）活性；荧光实时定量PCR（quantitative real-time PCR，qRT-PCR）检测细胞Runt相关转录因子2（Runt-related transcription factor-2，RUNX2）、Ⅰ型胶原α1链、护骨因子、NF-κB受体激活蛋白配体（receptor activator of NF-κB ligand，RANKL）mRNA的表达情况。 结果： 接种4 h，渗氮组细胞伸展良好，有细丝状伪足伸出，细胞间连接紧密；培养3 d，渗氮组细胞增殖A值（0.277±0.007）显著高于对照组（0.249±0.004）（P<0.01）；培养14 d，渗氮组ALP活性（173.6±1.9）显著高于对照组（162.6±2.4）（P<0.01）；渗氮组RUNX2、Ⅰ型胶原α1链、护骨因子mRNA相对表达量均显著高于对照组（P<0.05），RANKL mRNA相对表达量显著低于对照组（P<0.05）。 结论： 近β钛合金表面采用双层辉光离子渗氮改性后，改性层可促进成骨细胞早期黏附、增殖，利于成骨细胞分化，并提高细胞护骨因子mRNA表达量，抑制RANKL mRNA表达。.