Table 1

Author	Structure (sample)	Topography	Type	Evaluation	Result
Gittens et al. 2012 [24]	PT-smooth SLA-sand blast acid etched NMPT-nano mod NMSLA-nano mod s.blast etch	~5 nm - Not indicated ~20 nm - Not indicated (Only average roughness indicated from graph)	In vitro MG63 Human osteoblast-like	- DNA assay (cell amount) Evaluated after confluence At least x 2 times	Cell number, which decreases as cells transition from a proliferative to a more mature state, was lower for MG63s on the microrough surfaces compared to the microsmooth control, with the lowest levels on the combined microrough SLA and nanostructured NMSLA surfaces. P < 0.05
Gittens et al. 2012 [25]	sTiAlV - microsmooth rTiAlV - microrough NMsTiAlV-nanomod NMrTiAlV-nanomod	- some micro scale - some sub micro scale - 73 nm - 61 nm	In vitro primery HOBs	- Histology (Z1 Coulter particle counter) Evaluated after confluence At least x 2 times	Osteoblast cell number, which decreases in differentiated cells due to a transcriptionally-restricted transition between proliferation and differentiation, was lower on the microrough surfaces, with the lowest levels on the combined microrough and nanostructured NMrTiAlV, biggest on sTiAlV. P < 0.05
Gittens et al. 2011 [26] (manually selected)	- PT - SLA - NMPT (45 m) protuberance - NMPT (90 m) - NMPT (180 m) coarser str. - NMSLA	- 0.43 µm roughness - 14 ± 6 nm roughness - 40 to 200 nm - 40 to 360 nm - 500 to 1000 nm - 18 ± 3 nm roughness	In vitro MG63 cell	- DNA assay Evaluated at confluence At least x 2 times	The number of MG63 osteoblast from DNA measurements for the NMPT, SLA and NMSLA samples were lower than for the PT. This reduction in cells paralleled an increase in mean nanoscale roughness (NMPT vs. PT) and the microscale roughness (SLA and NMSLA vs. PT). P < 0.05
Tetè et al. 2011 [31]	- Sandblasted - Blast+acidetched - Full contact coverage (galvanotactic anodizing process in a phosphate-sulfate)	- 10 to 20 µm particles - Micro and nanopores 2 to 150 nm - Circular pores 10 to 700 nm (Mostly roughness explained)	In vitro hDPaSCs OB like cells	- Histology - SEM (prolif) 20 or 7 days	After 7 days, culturing onto FCC titanium coating was possible to evaluate a higher number of cells growing on the titanium surface, distributed around more samples areas, and the typical net morphology tended to form a confluent layer. After 20 days, SEM analysis of FCC coating showed a great amount of cell proliferation.
Zuo et al. 2013 [33]	- machined Ti plates (Ti-m) - polished Ti plates (Ti-p) - DBD Ti plate (Ti-tr)	- Ti-m less particles ~1 μm pores - Ti-p very sparse round nanoparticles ~1 μm pores - spherical nanoparticles 50 to 125 nm	In vitro MC3T3-E1 Pre OB	- Histology 24, 48, and 72 h - Hemocytometer x 3 times	Initially dielectric barrier discharge (DBD) modification significantly enhanced cell adhesion, spread, and proliferation of preosteoblasts with no negative effects on cell differentiation. At 72 h, there was no remarkable difference between three groups. P < 0.05 and P < 0.01
Tsukimura et al 2011 [34]	- Acid etched TI - Acid etched+TiO deposits - Acid etched+TiO deposits+UV treated	- Microscale - 100 nm, 300 nm, 500 nm - 100 nm, 300 nm, 500 nm + UV	In vitro Rat BMCs	- Histology - WST-1 (density) - BrdU proliferation assay Day 3	Increase in the proliferative activity of cells on the nanonodular surfaces both before and after UV treatment, with that on the 300 nm nanonodules being the greatest. P < 0.05
Kubo et al. 2009 [35]	- A Micropits - B micropits+nanonoduls - C micropits+nanonoduls - D micropits+nanonoduls	- 0.5 to 1.5 μm - 100 nm - 300 nm - 500 nm	In vitro Rat OB	- WST-1 6, 24 h 2, 5 days - BrdU marker	Cell density measured at culture days of 2 and 5 was substantially greater on the surfaces with nanonodules. The result of the BrdU incorporation per cell at day 2 confirmed the increased proliferation on the nanonodular surfaces, with the greatest one on the 300 nm nanonodules. P < 0.01
Han et al. 2011 [36]	- Ti6Al4V Smooth - Ti6Al4V porous	- Micro scale scratches - 10 to 20 nm grains	In vitro neonatal rat calvaria OB	- Histology - MTT assay 1, 3, 7, 10 and 14 days	The growth curves showed that the osteoblasts on nanophase Ti6Al4V substrate appeared to have a not only higher but also longer growth phase for cell proliferation than those cultured on any of other surface. P < 0.01
Yu et al. 2010 [37]	- Smooth-Ti - TN unannealed - Annealed 450 °C - Annealed 550 °C	- Not indicated ~80 nm ~80 nm ~80 nm	In vitro MC3T3-E1 Mouse pre OB	- MTT assay 24, 48 and 72 h	The proliferation of osteoblast cultured on anatase or anatase/rutile nanotube layersd showed significantly higher than smooth layer and amorphous nanotube layers, which means the crystal structure of nanotube layers can over-ride the chemistry effect and plays a main role in cell proliferation and mineralization. P < 0.05
Zhao et al. 2010 [38]	- Smooth - acid-etched - etch/anod 5 V - etch/anod 20 V	- Not indicated (smooth) - Micropits ~15 nm NT ~80 nm NT	In vitro PRCOB	- MTT assay 1, 4 and 7 days	After 7 days cell number on the acid-etched/20 V anodized surface is observed to be slightly higher. Addition of nanotubes to microstructured surface enhances osteoblast behaviors with nearly all the cell functions retained or promoted. P < 0.05
Zhao et al. 2011 [40]	Polished 5 V anodized 20 V anodized	- Not indicated ~25 nm nanonet textur ~80 nm nanotubular texture	In vitro PRCO (Primery rat calvarial ob)	- Histology - SEM - MTT assay 30, 60, 120 m 1, 4, 7 days	No significant difference in ad cell numbers on the 5 V, 20 V and polished is observed after 30, 60, 120 min and 1, 4, and 7 days except cell proliferation on the 5 V anodized surface is a little lower than on the other two Ti surfaces at days 1 and 4 P < 0.01
Xia et al. 2012 [41]	- Nanotubes - Micropores - Flat (control)	~100 nm ~ 10 to 20 μm - Shallow pits grooves	In vitro MG63 cells In vivo (rabit)	- Histology - MTT assay 1, 4, 7 days x times n = 9	More osteoblasts aggregated on the surface of TiO₂ nanotube. This result was in accordance with the increased cellular proliferation on the TiO₂ nanotubes observed in the In vitro study. Results showed increased proliferation on TiO₂ nanotube than on microporous or polished Ti plates. P < 0.05
Brammer et al. 2009 [42]	A-Ti B-TiO₂ nanotubes B-TiO₂ nanotubes C-TiO₂ nanotubes D-TiO₂ nanotubes	- Not indicated - 30 nm - 50 nm - 70 nm - 100 nm	In vitro MC3T3-E1	- Histology 2, 12, 24, 24 h 7 days - MTT 24, 48 h	The number of adhered cells on the smallest 30 nm diameter nanotubes was notably higher than all the other sizes of nanotubes, but the cells started to be more elongated on nanotube diameters above 70 nm. P < 0.05
Zhang et al. 2012 [43] (manually selected)	- Ti-control - Ti-6h small size sawtooth nanonetwork - Ti-24h large size sawtooth nanonetwork	- Not indicated ~10 nm, 100 to 200 nm distance ~30 nm, 200 to 300 nm distance	In vitro Rat BMMSCs	- Histology(prolif) (laser scanning microscope) 1, 4, 7 days x 3 times	The large nano-sawtooth structur approximately 30 nm produced the largest cell responses, including adhesion, proliferation, and differentiation properties. P < 0.05 and P < 0.01
Rani et al. 2012 [44]	Nanotube (NT) Nanoscaffold (NS) Nanoleaf (NL) Nanoneedles (NN) polished Ti (contr)	- Diameter 60 to 80 nm - roughness 166 nm - roughness 228 nm - roughness 940 nm - Not indicated	In vitro pHOB In vivo (rats)	- Histology - SEM - alamarBlueTM assay (prolif) 3, 5, 7 days	On days 5 and 7, the proliferation rate was higher on the nanoleafy surface amongst all. These results correlate with the enhanced protein adsorption on nanoleafy samples. cells grown on NN surfaces showed a significant reduction in proliferation, despite high protein adsorption. P < 0.05
Zhuang et al. 2014 [45]	- Smooth - SLA - micro, macro pits - SB-AH1 - nanoneedles - SB-AH2 - nanoporous	- 0.4 ± 0.05 µm rough - 1.97 ± 0.19 µm rough - 0.94 ± 0.04 µm rough - 1.31 ± 0.06 µm rough (only roughness is indicated)	In vitro MC3T3-E1 In vivo Rabits	- CCK-8(WST8) 1, 3, 5, 7 days - Micro-CT 4 and 8 weeks	Proliferation assay showed increased proliferation for SB-AH1 and SB-AH2 when SB-AH2 statistically better (P < 0.05). I Vivo study showed higher bone volume on SB-AH1 and SB-AH2 as well.
Ross et al. 2013 [57]	A polished B bead blasted C anod-sulfuric D anod+hydrofl anod E anod+hydrof etche	- striations 1 to 2 μm - grain structure - rounded feature 2 μm - pore size of 1 to 2 μm - numerous nanometer features	In vitro HOBs	- Histology - Auto T4 Cellometer 1, 4, and 7 days x 3 times	Anodization of Ti6Al4V using sulfuric acid followed by hydrofluoric acid with microporous surface 1 to 2 μm in diameter, and this promoted osteoblast densities D > E P < 0.01
Yu et al. 2014 [58]	- Smooth - Nano-foveolae	- Not indicated - 10 to 20 μm grains with 80 nm nano-foveolae structures.	In vitro MC3T3-E1 Mice pre OB	- MTT 24, 72 h	The proliferation rate of preosteoblasts was statistically similar at 24 h and statistically lower on nano-foveolae structures at 72 h. P < 0.05
Hori et al. 2010 [62]	- Smooth (machined) - Micropits - TiO₂ micro-nano-hybrid	- Not indicated - 0.5 to 1.5 μm -198.5 ± 22.3 nm (nanonoduls)	In vitro Rat OB	- WST-1 6, 24 h 2, 5 days - BrdU marker	The addition of nanonodules to the micropits, increased the number of cells two to three times to a level even greater than on the machined surface at 5th day. The result of the BrdU incorporation per cell at day 2 confirmed that proliferative activity of osteoblasts was impaired on the micropitted surface, whereas the proliferation on the micro-nano-hybrid surface was raised to a level equivalent to smooth surface. P < 0.05