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

doi:10.3877/cma.j.issn.1674-1366.2016.01.006

Abstract

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 (F_{3 d}= 245.517, F_{5 d}= 102.442, F_{7 d}= 119.047, P<0.001) at day 3, 5 and 7. And AA₁₂PT enhanced cell proliferation significantly than NTPT (P_{3 d}= 0.005, P_{5 d}= 0.038, P_{7 d}= 0.010) . At day 7, the gene expression of ALP and OCN of AA₁₂PT and NTPT was higher than that of NTDT (F_ALP= 13.698, P_ALP-AA12PT<0.001, P_ALP-NTPT= 0.002; F_OCN= 175.859, P_OCN<0.001) . At day 14, the gene expression of ALP and OCN of NTDT was higher than that of NTPT (F_ALP= 33.691, P_ALP= 0.045; F_OCN= 42.789, P_OCN= 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

Xuelian Li, Shuai Liu, Yutao Jian, Yuezhong Meng, Ke Zhao. Proliferation and differentiation of MC3T3-E1 cells on acid-alkali treated porous titanium[J]. Chinese Journal of Stomatological Research(Electronic Edition), 2016, 10(01): 30-37.

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Figures/Tables 9

References 21

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基因	引物序列
ALP	正向：5′?AACCCAGACACAAGCATTCG?3′
?	反向：5′?GCCTTTGAGGTTTTTGGTCT?3′
OCN	正向：5′?TTCTGCTCACTCTGCTGACG?3′
?	反向：5′?ACCACTCCAGCACAACTCCA?3′

基因	引物序列
ALP	正向：5′?AACCCAGACACAAGCATTCG?3′
?	反向：5′?GCCTTTGAGGTTTTTGGTCT?3′
OCN	正向：5′?TTCTGCTCACTCTGCTGACG?3′
?	反向：5′?ACCACTCCAGCACAACTCCA?3′

组别	3d	5d	7d
NTDT组	0.43 ± 0.02	0.53 ± 0.05	0.65 ± 0.04
NTPT组	0.20 ± 0.02	0.27 ± 0.02	0.33 ± 0.01
AA₁₂PT组	0.23 ± 0.02	0.31 ± 0.02	0.40 ± 0.05

组别	3d	5d	7d
NTDT组	0.43 ± 0.02	0.53 ± 0.05	0.65 ± 0.04
NTPT组	0.20 ± 0.02	0.27 ± 0.02	0.33 ± 0.01
AA₁₂PT组	0.23 ± 0.02	0.31 ± 0.02	0.40 ± 0.05

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