粗化处理对新型骨植入钛合金的生物相容性影响

目的研究国内新研制的骨植入钛合金TZS进行喷砂并酸蚀处理对成骨细胞生物学行为的影响。方法采用SD大鼠体外原代培养方法培养成骨细胞,并将细胞分别接种于新型钛合金（ 光滑表面）及表面处理的材料表面,建立体外共同培养模型。采用MTT比色法检测第3天成骨细胞的增殖率,扫描电镜（ SEM）观察细胞形态学变化,培养第5天进行细胞碱性磷酸酶功能活性检测。结果喷砂与酸蚀处理组表面的成骨细胞,在3 d时的细胞增殖率与光滑组比较有统计学差异（ P<0.05）;5 d时的处理组碱性磷酸酶活性检测值与光滑组比较无统计学差异（ P>0.05）;SEM观察成骨细胞在喷砂与酸蚀组表面具有典型形态特征,伸展良好,具有丰富的板状、丝状伪足,优于光滑组。结论喷砂并酸蚀处理的新型钛合金在体外实验表现为不同程度的促进成骨细胞增殖及功能活性表达。     

The effect of titanium surface roughening on protein absorption, cell attachment, and cell spreading

PURPOSE: The purpose of this study was to compare properties of roughened and polished titanium with respect to their ability to attach to cells and bind to protein as well as their cell spreading behavior. MATERIALS AND METHODS: Three different titanium surface treatments were compared for their ability to support cell attachment and spreading: sandblasted and acid-etched, resorbable blast media, and machine-polished titanium. The surface of the materials was characterized for surface roughness, surface energy, and surface chemistry. Osteoblast-like MG-63 cells were tested for in vitro attachment and spreading in the presence of serum proteins. Cell attachment was assessed by direct counting, dye binding, and microculture titanium assays. Cell spreading was determined by measuring area/cell in phalloidin-AlexaFluor 488 stained cells. Absorption of bovine serum albumin was determined by assay. RESULTS: Scanning electron micrography and x-ray diffractometry confirmed increased surface roughness of the roughened materials. All 3 materials had similar albumin binding kinetics. Three different methods confirmed that roughened surfaces enhance early cell attachment to titanium in the presence of serum. Cells spread better on smoother machined surfaces than on the roughened surfaces. CONCLUSION: Roughened titanium surfaces exhibited better early cell attachment than smooth surfaces in the presence of serum. The cells attached to roughened titanium were less spread than those attached to machined titanium. Although albumin binding was not different for roughened surfaces, it is possible that roughened surfaces preferentially bound to serum adhesive proteins to promote early cell attachment.     

The effects of Mg-ion implantation and sandblasting on Porphyromonas gingivalis attachment

Objectives: The purpose of this study was to evaluate the effect of titanium surface treatment on Porphyromonas gingivalis bacterial attachment. Materials and methods: Titanium disks of 15 mm in diameter and 1 mm in thickness (n=40) were subjected to mechanical grinding, or sandblasting. Magnesium (Mg) ions were implanted onto the titanium surface using a plasma source ion implantation method. The structure, chemistry, and surface morphologies of the titanium surfaces were analyzed using scanning electron microscopy (SEM), X‐ray photoelectron spectroscopy and Auger electron spectroscopy. Surface roughness was measured using a laser profilometer. Half of the titanium disks in each group were dipped in saliva for 24 h. All of the titanium specimens were rinsed with distilled water. A P. gingivalis strain was cultured in anaerobic conditions at 37°C for 72 h, and all titanium specimens were dipped in the bacterial suspension at 37°C for 24 h. Specimens were examined at × 3000 magnification using a SEM. The number of bacteria in each of 10 separate fields was determined by directly counting the number of bacterial colonies that adhered to each specimen. The mean values were calculated afterward. The resulting data were analyzed to assess the significance of observed differences based on the method of the surface treatment, ion implantation, and saliva dipping. Results: The amount of P. gingivalis attached to the sandblasted specimens was greater than that on the ground specimens (P<0.001). Moreover, surfaces with Mg‐ion implantation had more attachments than nonimplanted surfaces (P<0.001). Saliva dipping acted synergistically with surface roughness and chemical composition of the specimens. Conclusions: Chemically modified surface increase the attachment of a major periodontopathic bacterium, P. gingivalis. To cite this article:
Kim ML, Jeong CM, Jeon YC, Byon ES, Jeong YS, Cho LR. The effects of Mg‐ion implantation and sandblasting on Porphyromonas gingivalis attachment.
Clin. Oral Impl. Res. 23 , 2012; 245–252.
doi: 10.1111/j.1600‐0501.2010.02138.x     

TiO_2喷砂酸蚀处理对钛片表面氧化膜及成骨细胞生长影响的研究

目的研究纯钛钛片经喷砂及喷砂酸蚀处理后,表面氧化膜金相结构和化学成分的变化及对成骨细胞黏附和生长特性的影响。方法将直径为15 mm、厚度为1 mm的纯钛钛片分4组进行表面处理：1）机械打磨组（S0）;2）喷砂组（SB）;3）喷砂酸蚀1组（SLA1）;4）喷砂酸蚀2组（SLA2）。采用电子探针分析仪及X射线衍射仪检测4组钛片表面氧化膜的厚度、化学成分以及金相结构,扫描电镜观察其表面微观形态。而后将成骨细胞培养于4组钛片表面,采用MTT法分析比较4组钛片表面对成骨细胞黏附率以及增殖率的影响。结果与S0组相比,SB、SLA1、SLA2组的粗糙度明显增大（P<0.05）。SB、SLA1、SLA2组间表面平均粗糙度差异无统计学意义（P>0.05）。酸蚀处理使喷砂形成的氧化膜变薄,密度减低,且结构发生改变：原有的金红石型TiO2峰消失,锐钛矿型TiO2减少。在表面平均粗糙度相同条件下,SB组钛片表面氧化膜均匀致密,有利于成骨细胞早期的黏附和增殖。结论喷砂和喷砂酸蚀处理均增加了钛片表面的粗糙度,有利于成骨细胞的黏附和增殖,但酸蚀使TiO2喷砂表面的氧化膜层变薄,在平均粗糙度不变的情况下,单纯喷砂表面成骨细胞的黏附和增殖优于喷砂酸蚀处理表面。     

Determining optimal surface roughness of TiO(2) blasted titanium implant material for attachment, proliferation and differentiation of cells derived from human mandibular alveolar bone

In the complex process of bone formation at the implant-tissue interface, implant surface roughness is an important factor modulating osteoblastic function. In this study, primary cultures of osteoblast-like cells, derived from human mandibular bone, were used. The aim was to examine the effect of varying surface roughness of titanium implant material on cellular attachment, proliferation and differentiation. A recognized method of increasing surface roughness and enlarging the surface area of titanium implants is by blasting with titanium dioxide particles: the four specimen types in the study comprised surfaces which were machine-turned only, or blasted after turning, with 63-90 microm, 106-180 microm, or 180-300 microm TiO(2) particles, respectively. The specimens were analyzed by scanning electron microscopy and confocal laser scanning. The turned samples had the smoothest surfaces: average height deviation (S(a)) of 0.20 microm. The roughest were those blasted with 180-300 microm particles, S(a) value 1.38 microm. Blasting with intermediate particle sizes yielded S(a) values of 0.72 microm and 1.30 microm, respectively. Cell profile areas were measured using a semiautomatic interactive image analyzer. Figures were expressed as percentage of attachment. DNA synthesis was estimated by measuring the amount of [(3)H]-thymidine incorporation into trichloroacetic acid (TCA) insoluble cell precipitates. The specific activity of alkaline phosphatase was assayed using p-nitrophenylphosphate as a substrate. The ability of the cells to synthesize osteocalcin was investigated in serum-free culture medium using the ELSA-OST-NAT immunoradiometric kit. After 3 h of culture, the percentage of cellular attachment did not differ significantly between specimens blasted with 180-300 micromparticles and the turned specimens. All blasted surfaces showed significantly higher [(3)H]-thymidine incorporation than the turned surfaces (P<0.05), with the highest on the surfaces blasted with 180-300 microm particles. Osteocalcin synthesis by the cells in response to stimulation by 1,25(OH)2D3, was also significantly greater (P<0.05) on the surfaces blasted with TiO(2) particles. However, analysis of alkaline phosphatase activity disclosed no significant differences among the four surface modifications. It is concluded that in this cellular model, the proliferation and differentiation of cells derived from human mandibular bone is enhanced by surface roughness of the titanium implant. However, increasing the size of the blasting particles to 300 microm does not further increase the initial attachment of the cells compared to turned surfaces and those blasted with 63-90 microm particles.     

The mucosal attachment to titanium implants with different surface characteristics: an experimental study in dogs

BACKGROUND: Findings from in vitro studies have indicated that the orientation and proliferation of cells on titanium surfaces may be influenced by the topography of the surface on which they are grown. It may be argued, therefore, that differences may occur in the mucosal attachment to titanium implants with different surface roughness. AIM: The present experiment was performed to study the composition of the soft tissue barrier that formed to implants prepared with well-defined smooth or rough surfaces. MATERIAL AND METHODS: Five beagle dogs were used. Four implants made of c.p. titanium were placed in the right edentulous mandibular premolar region. After 3 months, two different types of abutments were connected: one experimental (OA) with a dual, thermal acid-etched surface ('Osseotite'), and one regular (RA) abutment with a 'turned' surface. At the end of a 6-month period during which proper plaque control had been maintained, biopsies including the implant and the surrounding soft and hard tissues were obtained, decalcified and processed for light and electron microscopy. A confocal He-Ne laser profilometer was used to study the surface topography of the abutments. RESULTS: The attachment between the peri-implant mucosa and titanium abutments with either a turned (RA; 'smooth') or acid-etched (OA; 'rough') surface was similar from both a quantitative and a qualitative aspect. The attachment comprised a barrier epithelium and a zone of connective tissue attachment of similar dimension at RA and OA. It was further observed that the 'inner' zone of the connective tissue attachment at both types of abutment was composed of about 30-33% fibroblasts and 63-66% collagen. CONCLUSION: It was demonstrated that the soft tissue attachment that formed to implants made of c.p. titanium was not influenced by the roughness of the titanium surface.     

喷砂酸蚀与双重酸蚀钛表面对成骨细胞行为影响的对比研究

目的 对比研究喷砂酸蚀与双重酸蚀钛表面对成骨细胞生物学行为的影响。 方法 在纯钛试件表面分别进行喷砂酸蚀与双重酸蚀处理。以光滑钛表面（Ti）为对照组,喷砂酸蚀钛表面（Ti-SLA）、双重酸蚀钛表面（Ti-DA）为实验组,通过扫描电镜（SEM）、表面接触角测试、X射线光电子能谱（XPS）观察分析三种钛表面的微形貌、润湿性和元素组成。将MC3T3-E1成骨细胞接种于三组试件表面,研究不同钛表面对成骨细胞生物学行为的影响。 结果 SEM观察显示Ti-DA组试件表面形成了较Ti-SLA组更均匀细密的微米级凹坑结构;各组试件的表面接触角无显著差异;XPS分析显示Ti-SLA组试件表面有微量铝元素残留;成骨细胞在三组试件表面的粘附、增殖无显著差异,而Ti-DA组显著促进成骨细胞的分化。 结论 与喷砂酸蚀钛表面相比,双重酸蚀钛表面的微米级凹坑结构更均匀细密,且无铝元素残留,能更有效地促进成骨细胞分化。     

Surface properties and biocompatibility of acid-etched titanium

The purpose of this study was to evaluate the effects of acid-etched titanium on the biological responses of osteoblast-like MC3T3-E1 cells. Four types of treatments (polishing, sandblasting, concentrated H2SO4 etching, and concentrated H2SO4 etching with vacuum firing) were carried out on the surfaces of commercially pure titanium (cpTi) disks. MC3T3-E1 cells were then cultured on the treated cpTi surfaces. Through surface roughness measurement and SEM analysis, it was found that the acid-etched surfaces showed higher roughness values than the sandblasted ones. Scanning electron microscope analysis showed that the cells on the disks treated with acid-etching and acid-etching with vacuum firing spread as well as the sandblasted ones. There were no significant differences in cell proliferation and collagen production on cpTi among the four different surface treatments. Based on the results of this study, it was concluded that etching with concentrated sulfuric acid was a simple and effective way to roughen the surface of titanium without compromising its biocompatibility.     

Cytokine release by osteoblast-like cells cultured on implant discs of varying alloy compositions

OBJECTIVE: The aims of this study were (i). to assess the morphological features of osteo-blast-like, osteosarcoma cells (cell line SaOS-2) cultured on implant surfaces of varying alloys and (ii). to evaluate the biological activity of these cells, following their adhesion onto these surfaces. MATERIALS AND METHODS : SaOS-2 cells (6 x 104) were grown on titanium discs (diameter 30 mm), each with a surface of differing composition and roughness (commercially pure titanium, titanium-aluminium-vanadium alloy, oxide-blasted titanium and Astra-Tech special treatment titanium; the alloys are directly comparable with those used to construct implants). The cells were grown for time periods of 1, 3, 5 and 7 days, the media were collected and the cells were fixed with 2.5% glutaraldehyde. The media were then assayed (using enzyme-linked immunosorbant assay) for the levels of interleukin (IL)-1, interleukin-6, interleukin-18 and osteoprotegerin (OPG) produced by the cells. The discs, with the cells fixed on them, were viewed under scanning electron microscopy (SEM, x 2.0 k) to evaluate cell morphology. RESULTS: Following attachment, the cells changed their morphology and released local factors known to activate osteoclasts. Commercially pure titanium stimulated the cells the most and titanium-aluminium-vanadium alloy the least. All implant materials stimulated production of IL-1, IL-6, IL-18 above that produced by cells grown on Petri dishes (polystyrene). The titanium-aluminium-vanadium alloy allowed cell attachment but levels of IL-1 in this medium were significantly lower (31.5 +/- 5.2 pg/ml on same day) than cultures with pure titanium (201.8 +/- 11.5 pg/ml at day 5). The same pattern was observed with the IL-6, IL-18, and OPG with polystyrene appearing to stimulate most production of OPG. Titanium-aluminium-vanadium produced the least biological response.     

成骨细胞在微弧氧化处理后纯钛表面的附着、增殖及ALP活性

目的 :对表面微弧氧化 (microarcoxidation ,MAO)处理后的纯钛材料进行成骨细胞生物相容性检测 ,评价改进的MAO工艺应用于钛植入材料表面处理的可能性。方法 :纯钛材料经过 2种MAO处理后 (MAO 1和MAO 2工艺 ) ,采用MC 3T3细胞系对不同时间点成骨细胞在材料表面的附着率、生长增殖情况以及ALP活性进行检测 ,以未经处理光滑纯钛表面作为对照 ,以SPSS对实验结果进行统计分析。结果 :早期 (0 .5h、1h)细胞附着率差异有统计学意义 ,MAO 1组 >MAO 2组 >纯钛组 ;2h后 ,MAO 1组与MAO 2组无差异 ,2组细胞附着率都显著高于纯钛组。细胞增殖及ALP活性测试中 ,MAO 1处理组在各时间点都显著高于另外 2组。结论 :MAO处理后的纯钛对成骨细胞的生物相容性优于未处理组 ,改进的MAO 1处理工艺较一般工艺可以更有效提高成骨细胞的早期粘附、增殖及ALP活性。     

Influence of surface treatments developed for oral implants on the physical and biological properties of titanium. (I) Surface characterization.

We present an investigation of the physico‐chemical surface properties of commercially pure titanium coverslips which were submitted to various treatments designed to optimize their topography in view of application in oral implantology. The surface microroughness, chemical composition and water wettability were analyzed on titanium coverslips prepared by mechanical polishing, acid attack in HCl/HZSOI, after mechanical polishing or sandblasting, and titanium plasma‐spray. The chemical composition has been measured by Auger electron spectroscopy. The treatments have no major influence on the surface chemical composition and all the samples display a composition approaching that of TiOZ with minor amounts of carbon. sulfur, silicon and calcium as impurities. The roughness has been measured by scanning force microscopy on an area of 20 μm x 20 μm on each sample. Polished titanium is smooth (peak‐to‐valley roughness 81 nm). whereas the acid‐attacked surfaces exhibit a micro‐roughness in the pm range (2100 nm for polished and acid attacked; 3600 nm for sandblasted and acid attacked) which is quite reproducible over large areas of the sample. The acid attacked samples present a subsurface layer which contains hydrogen below the native passivating oxide layer. Water wettability measurement shows that all surfaces are hydrophobic with a slightly higher contact angle for the acid attacked surfaces. The different treatments analyzed in this study essentially influence the surface roughness by preserving the chemical composition and the wettability properties of titanium native oxide surface layer.     

Characterization of sterilized CP titanium implant surfaces

Surface analysis techniques and in vitro biologic assays were used to characterize sterilized commercially pure titanium surfaces. Significant surface alterations were observed following sterilization treatments. These alterations led to decreased fibroblast cell attachment and altered cellular spreading phenomena compared to nonsterilized control surfaces.     

Short-term plasma-cleaning treatments enhance in vitro osteoblast attachment to titanium.

The purpose of this research was to characterize the in vitro cellular behavior of osteoblast-like cells on titanium surfaces prepared with argon plasma-cleaning (PC) treatments for various lengths of time. The highest levels of cell attachment were observed for surfaces which had been plasma-treated for one min. Surface analyses indicated that although PC treatments dramatically improved surface wettability, the presence of inorganic contaminants was observed with longer treatment times and may have interfered with cell attachment. Further work is suggested to investigate the longer-term phenotypic expression of osteoblasts when grown on implant surfaces.     

Attachment of human marrow stromal cells to titanium surfaces

The attachment of human bone marrow stromal cells to titanium alloy (Ti6Al4V) surfaces was investigated. Titanium disks were polished and modified by surface roughening and by passivation in nitric add. Cell attachment to titanium surfaces and tissue culture plastic (TCP) was determined by tetrazolium bromide (MTT) assay at 2, 6, 24, and 48 hours after seeding. Cell proliferation was determined by thymidine incorporation. Attachment on titanium surfaces was 75.6% to 94.9% of attachment on TCP control. The difference between cell attachment on the TCP compared with smooth or rough titanium was statistically significant (P < .05). However, no statistically significant difference was found between attachment to TCP and passivated titanium. Cell proliferation on titanium surfaces after 24 hours was approximately 70% of proliferation on TCP. There was a statistically significant difference (P < .05) between proliferation on tissue culture and smooth and passivated titanium but not on rough titanium. These results indicate that titanium provides a surface that is conducive to cell attachment and that passivating titanium improves cell attachment, approaching levels seen with TCP, a surface specifically developed to enhance cell attachment. Increasing surface roughness results in improved cell proliferation on titanium.     

Effect of heat-treated titanium surfaces on protein adsorption and osteoblast precursor cell initial attachment

The clinical success of dental implants is governed in part by surface properties of implants and their interactions with the surrounding tissues. The objective of this study was to investigate the effect of heat-treated titanium surfaces on protein adsorption and osteoblast precursor cell attachment in vitro. Passivated titanium samples used in this study were either non heat treated or heat treated at 750 degrees C for 90 minutes. It was observed that the contact angle on heat-treated titanium surfaces was statistically lower compared with the non-heat-treated titanium surfaces. The non-heat-treated titanium surface was also observed to be amorphous oxide, whereas heat treatment of titanium resulted in the conversion of amorphous oxide to crystalline anatase oxide. No significant difference in albumin and fibronectin adsorption was observed between the heat-treated and non-heat-treated titanium surfaces. In addition, no significant difference in initial cell attachment was observed between the two groups. It was concluded that heat treatment of titanium resulted in significantly more hydrophilic surfaces compared to non-heat-treated titanium surfaces. However, differences in oxide crystallinity and wettability were not observed to affect protein adsorption and initial osteoblast precursor cell attachment.     

钛表面羟磷灰石/壳聚糖-转化生长因子-β1缓释微球复合涂层的制备及其对成骨细胞黏附与增殖的影响

目的 探讨在钛表面制备羟磷灰石（HA）/壳聚糖（CS）-转化生长因子-β1（TGF-β1）缓释微球复合涂层及其对成骨细胞黏附与增殖的影响。方法 通过物理-化学-生物改性联合方式制备HA/CS-TGF-β1 缓释微球复合涂层。运用扫描电镜（SEM）、X射线衍射仪（XRD）、傅里叶变换红外光谱（FTIR）等分析涂层的形貌和物相;使用CCK-8及免疫荧光检测其对成骨细胞的细胞毒性及细胞黏附和增殖的影响。结果 成功制备HA/CS-TGF-β1缓释微球复合涂层,该涂层制备具有超亲水性,体外释药稳定且时间长,成骨细胞在此复合涂层的钛片上生长无抑制,且对细胞的黏附与增殖具有促进作用。结论 HA/CS-TGF-β1缓释微球复合涂层在体外对于成骨细胞的黏附与早期增殖有明显的促进作用,具有良好的运用前景。     

磁控溅射ZrSiN/ZrO2复合梯度涂层纯钛表面对钛瓷结合强度的影响

目的:探讨磁控溅射ZrSiN/ZrO2复合梯度涂层作为钛瓷结合中间层对钛瓷结合强度的影响.方法:将50个钛试件分为2组(n=25),实验组在钛基底表面用磁控溅射的方法制备ZrSiN/ZrO2复合梯度涂层,对照组试件表面采用常规喷砂处理.2组试件各随机抽取10个进行粗糙度和表面能检测,各取10个用钛专用瓷粉烧结并采用三点弯曲试验进行钛瓷结合强度测试,用SEM和EDS对2组试件瓷剥脱面和钛瓷结合横截面进行观察、对比、分析.结果:ZrSiN/ZrO2复合梯度涂层组的粗糙度低(P＜0.05),接触角大(P＜0.05);三点弯曲结合强度试验表明实验组钛瓷结合强度高(P＜0.05);瓷剥脱面和钛瓷结合横截面的SEM和EDS分析显示,各元素原子百分比含量及线性分布情况均提示实验组试件表面瓷残留量多于对照组.结论:磁控溅射ZrSiN/ZrO2复合梯度涂层能够控制钛金属表面过度氧化,有效缓冲钛瓷结合热应力,提升钛瓷结合强度.     

Surface modifications of titanium implant material with excimer laser for more effective osseointegration

The biointegration of dental and orthopaedic implants depends mainly on the morphology and physical-chemical properties of their surfaces. Accordingly, the development of the desired microstructure is a relevant requirement in the bulk manufacture. Besides the widely used sandblasting plus acid etching and plasma-spray coating techniques, the laser surface modification method offers a plausible alternative. In order to analyze the influence of the laser treatment, the surfaces of titanium samples were exposed to excimer laser irradiation. The aim of this study was to develop surfaces that provide optimal conditions for bone-implant contact, bone growth, formation and maintenance of gingival attachment. For this purpose, holes were ablated on the surface of samples by nanosecond (18 ns, ArF) and also sub-picosecond (0,5 ps, KrF) laser pulses. Using pulses of ns length, due to melt ejection, crown-like protrusions were formed at the border of the holes, which made them sensitive to mechanical effects. To avoid these undesirable crown-like structures ultrashort KrF excimer laser pulses were successfully applied. On the other hand, titanium samples were laser-polished in favour of formation and connection of healthy soft tissues. Irradiation by a series of nanosecond laser pulses resulted in an effective smoothening as detected by atomic force microscopy (AFM). By inhibiting plaque accumulation this favours formation of gingival attachment. X-ray photoelectron spectroscopy (XPS) studies showed that laser treatment, in addition to micro-structural and morphological modification, results in decreasing of surface contamination and thickening of the oxide layer. X-ray diffraction (XRD) analysis revealed that the original alpha-titanium crystalline structure of the laser-polished titanium surface was not altered by the irradiation.     

Effect of chemically modified titanium surfaces on protein adsorption and osteoblast precursor cell behavior

PURPOSE: To investigate the effects of different chemically modified titanium surfaces on protein adsorption and the osteoblastic differentiation of human embryonic palatal mesenchymal (HEPM) cells. MATERIALS AND METHODS: Three different surfaces were evaluated. The first, a machined surface (Ti-M), was considered a control. The second surface was acid etched (Ti-AE). The third surface was prepared by exposing the Ti-AE samples to sodium hydroxide (NaOH) solution (Ti-AAE). The surface characteristics of chemically modified titanium were investigated by means of scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and profilometry. To evaluate the production of biomarkers, commercial kits were utilized. RESULTS: Surface composition and morphology affected the kinetics of protein adsorption. Ti-AE surfaces manifested a greater affinity for fibronectin adsorption compared to Ti-M or Ti-AAE surfaces. It was observed that Ti-AE and Ti-AAE surfaces promoted significantly greater cell attachment compared to Ti-M surfaces. Statistically significant differences were also observed in the expression of alkaline phosphatase (ALP) activity, osteocalcin, and osteopontin on all 3 titanium surfaces. ALP activity and osteocalcin production up to day 12 suggested that differentiation of the cells into osteoblasts had occurred and that cells were expressing a bone-forming phenotype. CONCLUSIONS: It was thus concluded from this study that surface morphology and composition play a critical role in enhancing HEPM cell proliferation and differentiation into osteoblast cells.