Martin J Y, Schwartz Z, Hummert T W, Schraub D M, Simpson J, Lankford J, Dean D D, Cochran D L, Boyan B D
Wilford Hall Medical Center, Lackland AFB, Texas, USA.
J Biomed Mater Res. 1995 Mar;29(3):389-401. doi: 10.1002/jbm.820290314.
The effect of surface roughness on osteoblast proliferation, differentiation, and protein synthesis was examined. Human osteoblast-like cells (MG63) were cultured on titanium (Ti) disks that had been prepared by one of five different treatment regimens. All disks were pretreated with hydrofluroic acid-nitric acid and washed (PT). PT disks were also: washed, and then electropolished (EP); fine sandblasted, etched with HCl and H2SO4, and washed (FA); coarse sandblasted, etched with HCl and H2SO4, and washed (CA); or Ti plasma-sprayed (TPS). Standard tissue culture plastic was used as a control. Surface topography and profile were evaluated by brightfield and darkfield microscopy, cold field emission scanning electron microscopy, and laser confocal microscopy, while chemical composition was mapped using energy dispersion X-ray analysis and elemental distribution determined using Auger electron spectroscopy. The effect of surface roughness on the cells was evaluated by measuring cell number, [3H]thymidine incorporation into DNA, alkaline phosphatase specific activity, [3H]uridine incorporation into RNA, [3H]proline incorporation into collagenase digestible protein (CDP) and noncollagenase-digestible protein (NCP), and [35S]sulfate incorporation into proteoglycan. Based on surface analysis, the five different Ti surfaces were ranked in order of smoothest to roughest: EP, PT, FA, CA, and TPS. A TiO2 layer was found on all surfaces that ranged in thickness from 100 A in the smoothest group to 300 A in the roughest. When compared to confluent cultures of cells on plastic, the number of cells was reduced on the TPS surfaces and increased on the EP surfaces, while the number of cells on the other surfaces was equivalent to plastic. [3H]Thymidine incorporation was inversely related to surface roughness. Alkaline phosphatase specific activity in isolated cells was found to decrease with increasing surface roughness, except for those cells cultured on CA. In contrast, enzyme activity in the cell layer was only decreased in cultures grown on FA- and TPS-treated surfaces. A direct correlation between surface roughness and RNA and CDP production was found. Surface roughness had no apparent effect on NCP production. Proteoglycan synthesis by the cells was inhibited on all the surfaces studied, with the largest inhibition observed in the CA and EP groups. These results demonstrate that surface roughness alters osteoblast proliferation, differentiation, and matrix production in vitro. The results also suggest that implant surface roughness may play a role in determining phenotypic expression of cells in vivo.
研究了表面粗糙度对成骨细胞增殖、分化和蛋白质合成的影响。将人成骨样细胞（MG63）培养在通过五种不同处理方案之一制备的钛（Ti）盘上。所有盘均先用氢氟酸 - 硝酸预处理并洗涤（PT）。PT盘还进行了以下处理：洗涤后再进行电解抛光（EP）；细喷砂处理，用HCl和H2SO4蚀刻，然后洗涤（FA）；粗喷砂处理，用HCl和H2SO4蚀刻，然后洗涤（CA）；或钛等离子喷涂（TPS）。使用标准组织培养塑料作为对照。通过明场和暗场显微镜、冷场发射扫描电子显微镜和激光共聚焦显微镜评估表面形貌和轮廓，同时使用能量色散X射线分析绘制化学成分图，并使用俄歇电子能谱确定元素分布。通过测量细胞数量、[3H]胸苷掺入DNA、碱性磷酸酶比活性、[3H]尿苷掺入RNA、[3H]脯氨酸掺入胶原酶可消化蛋白（CDP）和非胶原酶可消化蛋白（NCP）以及[35S]硫酸盐掺入蛋白聚糖来评估表面粗糙度对细胞的影响。基于表面分析，将五种不同的Ti表面按从最光滑到最粗糙的顺序排列：EP、PT、FA、CA和TPS。在所有表面上均发现了一层TiO2，其厚度范围从最光滑组的100埃到最粗糙组的300埃。与塑料上的汇合细胞培养物相比，TPS表面上的细胞数量减少，EP表面上的细胞数量增加，而其他表面上的细胞数量与塑料相当。[3H]胸苷掺入与表面粗糙度呈负相关。发现分离细胞中的碱性磷酸酶比活性随表面粗糙度增加而降低，但在CA上培养的细胞除外。相反，细胞层中的酶活性仅在FA和TPS处理的表面上生长的培养物中降低。发现表面粗糙度与RNA和CDP产生之间存在直接相关性。表面粗糙度对NCP产生没有明显影响。在所研究的所有表面上，细胞的蛋白聚糖合成均受到抑制，在CA和EP组中观察到的抑制作用最大。这些结果表明，表面粗糙度在体外改变成骨细胞的增殖、分化和基质产生。结果还表明，植入物表面粗糙度可能在决定体内细胞的表型表达中起作用。
