Proliferation and differentiation of MC3T3-E1 cells on acid-alkali treated porous titanium

Abstract

Objective To evaluate the proliferation and differentiation of MC3T3-E1 cells on acid-alkali treated porous titanium. Methods Porous titanium samples were fabricated by powder metallurgy process, then immersed in a mixed acid and NaOH solution in different periods to get acid-alkali-treated porous Ti (AAPT) . 73 AAPT samples, 41 non-treated porous Ti (NTPT) samples and 30 non-treated commercial dense titanium (NTDT) samples were made. The surface morphology was observed under a scanning electron microscopy. Protein adhesion was quantitatively analyzed by BCA method. The optimum treating time of alkali-treated in acid-alkali treatment was determined. For NTPT and AAPT, the contact angle and surface chemical structure were examined by an optical contact angle measuring device and a Raman spectrometer. CCK-8 and real-time fluorescence quantitative polymerase chain reaction were carried out to measure the proliferative activity and the gene expression of alkaline phosphatase (ALP) and osteocalcin (OCN) , respectively. The level of significance was determined by independent sample t test and an One-Way ANOVA followed by LSD-t test for a multiple comparison procedure. Results The scanning electron micrographs showed the nano structure of 12 h alkali treatment (AA12PT) was most distinct and obvious. And it had the highest protein adsorption (0.367 mg/ml) (F= 400.835, P<0.05) . So AA12PT was chosen in the subsequent experiments. The contact angle on the surfaces of NTPT (93.7°) exhibited a statistically significantly higher in comparison to the contact angle of AA12PT (22.08°) (F= 1.194, P<0.001) . The Raman spectroscopy indicated the presence of sodium hydrogen titanate on the surface of AA12PT. MC3T3-E1 cells got better proliferation rate on NTDT than AA12PT (F3 d= 245.517, F5 d= 102.442, F7 d= 119.047, P<0.001) at day 3, 5 and 7. And AA12PT enhanced cell proliferation significantly than NTPT (P3 d= 0.005, P5 d= 0.038, P7 d= 0.010) . At day 7, the gene expression of ALP and OCN of AA12PT and NTPT was higher than that of NTDT (FALP= 13.698, PALP-AA12PT<0.001, PALP-NTPT= 0.002; FOCN= 175.859, POCN<0.001) . At day 14, the gene expression of ALP and OCN of NTDT was higher than that of NTPT (FALP= 33.691, PALP= 0.045; FOCN= 42.789, POCN= 0.044) . Conclusion Acid-alkali treated porous titanium can significantly improve the amount of protein adsorption and promote proliferation and differentiation of osteoblasts. Key words: Titanium, Porousity; Osteoblasts; Protein adsorption; Alkaline phosphatase; Osteocalcin