J. Mater. Sci. Technol. ›› 2022, Vol. 97: 201-212.DOI: 10.1016/j.jmst.2021.04.056
• Research Article • Previous Articles Next Articles
Yuan Zhanga, Shan Fua, Lei Yanga,b, Gaowu Qina,b, Erlin Zhanga,b,*(
)
Received:2021-03-15
Revised:2021-04-06
Accepted:2021-04-17
Published:2021-07-02
Online:2021-07-02
Contact:
Erlin Zhang
About author:* E-mail address: zhangel@atm.neu.edu.cn (E. Zhang).Yuan Zhang, Shan Fu, Lei Yang, Gaowu Qin, Erlin Zhang. A nano-structured TiO2/CuO/Cu2O coating on Ti-Cu alloy with dual function of antibacterial ability and osteogenic activity[J]. J. Mater. Sci. Technol., 2022, 97: 201-212.
| Gene | Forward primer sequence (5′-3′) | Reverse primer sequence (5′-3′) |
|---|---|---|
| ALP | CCAGAAAGACACCTTGACTGTGG | TCTTGTCCGTGTCGCTCACCAT |
| RUNX-2 | TTGCCCTCATCCTTCACTCC | GGCTCCTCCCTTCTCAACCT |
| OCN | ACCATCTTTCTGCTCACTCTGCT | CCTTATTGCCCTCCTGCTTG |
| COL-I | GACATGTTCAGCTTTGTGGACCTC | GGGACCCTTAGGCCATTGTGTA |
| GAPDH | TGTGTCCGTCGTGGATCTGA | TTGCTGTTGAAGTCGCAGGAG |
Table 1 Primer sequences used for quantitative RT-PCR analysis.
| Gene | Forward primer sequence (5′-3′) | Reverse primer sequence (5′-3′) |
|---|---|---|
| ALP | CCAGAAAGACACCTTGACTGTGG | TCTTGTCCGTGTCGCTCACCAT |
| RUNX-2 | TTGCCCTCATCCTTCACTCC | GGCTCCTCCCTTCTCAACCT |
| OCN | ACCATCTTTCTGCTCACTCTGCT | CCTTATTGCCCTCCTGCTTG |
| COL-I | GACATGTTCAGCTTTGTGGACCTC | GGGACCCTTAGGCCATTGTGTA |
| GAPDH | TGTGTCCGTCGTGGATCTGA | TTGCTGTTGAAGTCGCAGGAG |
Fig. 1. Surface morphology of different samples observed by SEM. (a-d) Low-magnification SEM images of cp-Ti, cp-Ti-AH, Ti-5Cu and Ti-5Cu-AH, respectively; (e-h) high-magnification SEM images of cp-Ti, cp-Ti-AH, Ti-5Cu and Ti-5Cu-AH, respectively. Red rectangles mark the EDS analysis positions.
| Samples | O (wt.%) | Na (wt.%) | Ti (wt.%) | Cu (wt.%) |
|---|---|---|---|---|
| cp-Ti | <0.01 | - | >99.99 | - |
| cp-Ti-AH | 35.31 | <0.01 | 64.69 | - |
| Ti-5Cu | <0.01 | - | 94.99 | 5.01 |
| Ti-5Cu-AH | 34.33 | <0.01 | 62.21 | 3.46 |
Table 2 EDS analysis results of four kinds of specimens.
| Samples | O (wt.%) | Na (wt.%) | Ti (wt.%) | Cu (wt.%) |
|---|---|---|---|---|
| cp-Ti | <0.01 | - | >99.99 | - |
| cp-Ti-AH | 35.31 | <0.01 | 64.69 | - |
| Ti-5Cu | <0.01 | - | 94.99 | 5.01 |
| Ti-5Cu-AH | 34.33 | <0.01 | 62.21 | 3.46 |
Fig. 2. XPS patterns of different samples. (a) XPS full spectra for four kinds of specimens; (b) high-resolution spectra of Ti 2p for four kinds of specimens; (c) high-resolution spectra of Cu 2p for Ti-5Cu sample; (d) high-resolution spectra of Cu 2p for Ti-5Cu-AH sample.
Fig. 3. XRD patterns of four kinds of samples. (a) XRD patterns of 2θ from 20° to 80°; (b) XRD patterns of 2θ from 22° to 30°.
Fig. 4. Roughness and water contact angles of four kinds sample. (a) Surface roughness; (b) water contact angle (* p < 0.05).
| Sample | Ti-5Cu | Ti-5Cu-AH |
|---|---|---|
| Cu ion concentration (μg/L) | 76.64 ± 16.79 | 1676.15 ± 217.49 |
Table 3 Cu ions release of Ti-5Cu and Ti-5Cu-AH after immersed in cell culture medium for 24 h.
| Sample | Ti-5Cu | Ti-5Cu-AH |
|---|---|---|
| Cu ion concentration (μg/L) | 76.64 ± 16.79 | 1676.15 ± 217.49 |
Fig. 5. Antibacterial property of materials. (a-d) Bacterial colony of S. aureus cultivated on cp-Ti, cp-Ti-AH, Ti-5Cu and Ti-5Cu-AH, respectively; (e) antibacterial rates of four kinds of specimens (* p < 0.05).
Fig. 6. SEM images of S. aureus on four kinds of specimens after 12 h and 24 h culture. Red arrows mark the dead bacteria, and the white arrow marks the sunken cell wall of bacteria.
Fig. 7. Live/dead staining of S. aureus on four kinds of specimens after cultured for 12 h and 24 h.
Fig. 8. Initial adhesion and spreading of cells on four kinds of samples stained using Rhodamine-phalloidin/DAPI after culture for 2 h, 4 h and 12 h.
Fig. 9. Proliferation of cells on four kinds of specimens after 1 day, 3 days and 5 days incubation: (a) stained with Rhodamine-phalloidin/DAPI; (b) viability of cells evaluated by MTT assay. (c) Viability of MC3T3-E1 cells exposed to Cu2+ with doses of 0, 300, 3000 and 30,000 μg/L by MTT assay after 1 day and 3 days incubation (* p < 0.05).
Fig. 10. Osteogenic differentiation of MC3T3-E1 cells on four kinds of specimens. ALP activities assay of cells on four kinds of samples after 4 days and 7 days incubation: (a) stained by BCIP/NBT kit and (b) quantitative assay ALP activity. ECM mineralized nodule assay of MC3T3-E1 cells on four kinds of specimens after 28 days culture: (c) images of mineralized nodules stained by Alizarin Red S and (d) quantitative assay of ECM mineralized nodules. Osteogenesis-related gene expressions of cells after 7 days and 14 days incubation by qRT-PCR analysis: (e-h) mRNA expression level of ALP, OCN, RUNX-2 and COL-I, respectively.
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