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Файл:Ординатура / Хирургия / Библиотека им академика М.И. Перельмана / Книга_5326_Библиотеки_им_академика_М_И_Перельмана
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44 Surface Modification of Magnesium and its Alloys for Biomedical Applications
(a)
(c)
(e)
(b)
(d)
(f)
Figure 2.14 L-929 cell morphology after 1 day of incubation on different samples. (Xu, Pan,
et al., 2009). (a) and (b) Naked Mg alloy. (c) and (d) Treated Mg alloy. (e) and (f) Pure Ti.
and uniform. In addition, the outline shape of the magnesium implants was slightly
changed, indicating that the implants were corroded by the body fluid, or the implant
degraded in the body. However, it is hard to distinguish the difference in the degradation between the naked Mg alloy implant and the phosphated Mg alloy implant after
4 weeks implantation because the duration is not long enough to evaluate the in vivo
degradation.
Figure 2.18 shows the optical HE microstructure of the interfaces between the
magnesium implants and new bone 1 to 4 weeks postimplantation. For the naked
Mg alloy, lymphocytic infiltration at the interface was observed 1 week postimplantation, as indicated by ‘‘L’’ in Figure 2.18(a). After 2 weeks implantation, there was a
Phosphate treatment of magnesium alloy implants for biomedical applications 45
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(a) (b)
(c)
(e)
Figure 2.15 L-929 cell morphology after 3 days of incubation on different samples. (Xu, Pan,
et al., 2009). (a) and (b) Naked Mg alloy. (c) and (d) Treated Mg alloy. (e) and (f) Pure Ti.
(d)
(f)
continuous fibroblast band between the implant and the bone, as indicated by ‘‘F’’ in
Figure 2.18(b). However, lymphocyt ic infiltration and plasmablastic infiltration were
not noted. Three weeks later, the fibroblast band became thinner, as shown by ‘‘F’’
in Figure 2.18(c), and a small amount of newly formed osteoid tissue was found, as
indicated by ‘‘N’’ in Figure 2.18(c). At week 4, the surface of the implant was taken
up by newborn bone and bone trabecular, as indicated by ‘‘N’’ in Figure 2.18(d).
Crowded osteoblasts and bone matrix were observed. For the Ca-Pecoated Mg alloy,
connective tissue was seen at the interface 1 week postimplantation, as indicated by
‘‘C’’ in Figure 2.18(e), but lymphocytic infiltration and plasmablastic infiltration
46 Surface Modification of Magnesium and its Alloys for Biomedical Applications
(a)
(b)
(c) (d)
(e)
(f)
Figure 2.16 L-929 cell morphology after 5 days of incubation on different samples (Xu, Pan,
et al., 2009). (a) and (b) Naked Mg alloy. (c) and (d) Treated Mg alloy. (e) and (f) Pure Ti.
were not seen. At week 2, the connective tissue became thinner, and osteoblasts and
bone matrix were noted. Newly formed osteoid tissue with embedded osteocyte
formed in some areas, as indicated by ‘‘N’’ in Figure 2.18(f). Three weeks later, the
connective tissue was replaced by osteoblasts and bone matrix, as indicated by ‘‘N’’
in Figure 2.18(g). Bone trabeculaes connected together, but the alignment of the connected bone trabeculae was disorganized.
Four weeks later, more newborn bone was observed, as indicated by ‘‘N’’ in
Figure 2.18(h). Osteoid tissue and newborn bone trabeculae almost covered the
implant surface completely. Osteoid tissues connected together, and bone trabeculae
Phosphate treatment of magnesium alloy implants for biomedical applications 47
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1800
)
2
1600
1400
1200
1000
800
600
400
200
Cell number per unit area (number / mm
0
Figure 2.17 Growth of L929 cells versus culturing time. *p < 0.05 (Xu, Pan, et al., 2009).
Naked Mg-Mn-Zn
Treated Mg-Mn-Zn
Pure Ti
*
*
*
*
*
*
3 days1 day
Culture time
*
*
5 days
aligned compactly and regularly, in which mature osteocyte was embedded. Ossification was manifest. Table 2.4 summarizes the routine pathological examination results.
Compared with the naked Mg alloy, routine pathological examination analysis results such as no lymphocytic infiltration (inflammation) at week 1, thinner connective
tissue and the formation of bone matrix at week 2, more bone matrix and interconnected bone trabecular at week 3, and more newborn bones at week 4, as shown in
Figure 2.18, reveal clearly that the phosphated Mg alloy exhibits better surface
biocompatibility than the naked alloy during the first 4 weeks postoperation.
BMP-2 expression, TGF-b1 expression, and PDGF expression were investigated
at the implant/bone interface to reveal the effect of the phosphating coating on the
bone response at the early stage (Xu,Pan,etal.,2009). As an example, Figure 2.19
shows BMP-2 staining photomicrographs at the inter face s between the implants and
bones after different periods of implantation. Figure 2.20 shows the MOD values in
the BMP-2 expression at the implant/bone interface after different periods of implantation. At week 1, strong positive activities were obs erve d at the interfaces fo r both
the naked alloy implant group and the pho sph ate alloy implant group. At week 2,
positive activities increased and peaked. At weeks 3 and 4, osteoid tissues were
clearly observed at the interface, and the BMP-2 expressions gradually reduced.
At all time intervals, the phos phat ed Mg implant group shows a higher MOD value
in the BMP-2 expression than the naked implant. A significant positive activity was
found at the first 3 weeks as found for the Ca-P coated implant (p < 0.05). Similar
resultshavealsobeenobservedintheTGF-b1expressionandPDGFexpression
(Xu,Pan,etal.,2009).
The BMP plays crucial roles in normal skeletal development as well as bone healing, and is able to activate transcription of genes involved in cellular migration,
48 Surface Modification of Magnesium and its Alloys for Biomedical Applications
Table 2.4 Summary of the routine pathological examination of the
implant/bone interface (Xu, Pan, et al., 2009)
Periods
(weeks) Naked Mg alloy Ca-P coated Mg alloy
1 Lymphocytic infiltration Connective tissue
No lymphocytic infiltration
No plasmablastic
infiltration
2 Continuous fibroblast band
No lymphocytic infiltration
No plasmablastic
infiltration
3 Thinner fibroblast band
Small amount of newly
formed osteoid
4 New bone
Trabecular
Osteoblasts
Thinner connective tissue
Osteoblasts
Newly formed bone matrix
Newly formed osteoid with
osteocyte
No connective tissue
Osteoblasts
Bone matrix
Connecting trabecular
More new bones
New bone trabecular
aligned compactly and
regularly
proliferation, and differentiation. It was reported that endogenous BMP-2 is an indispensable osteogenic stimulus for initiation of fracture healing in mice (Tsujietal.,
2006). In addition, it is believed that TGF-b1 stimulates osteogenesis, angiogenesis,
fibroblast migration, and deposition of matrix (Helm, Dayou b , & Jan e, 20 01 ) and has
osteoinductive properties (Beck et al., 1993; Noda, 1989; Zhang, Ahmad, &
Gronowicz, 2003). PDGF stimulates osteoblas t proliferation, collagen synthesis,
and may play a regulatory role in fracture repair (Helm et al., 2001). The above
immunohistochemical analy sis results strongly demonstrated th at t he phosphatecoated implant provided a high BMP-2expressionduringthefirst 4 weeks postimplantation, and especially statistically significant differences in BMP-2 expression
between the phosphate-coated Mg alloy and the naked Mg alloy after 1 week,
2 weeks, and 3 weeks postoperation. The phosphate-coated implant also exhibited
ahighTGF-b1 expression during the first 4 weeks posti mplantation, and statistically
significant differences in the TGF-b1expressionbetweenthephosphateMgalloy
and the naked Mg alloy after 2 weeks, 3 weeks, and 4 weeks postoperation. Similarly, a s tatistically significant difference was also observed in P DGF expression between the phosphate Mg alloy and the naked Mg alloy after 1 weeks, 2 weeks, and
4 weeks postoperation. All the above in vivo results demonstrated significantly
good osteoconductivity of the phosphate Mg alloy at the early osseous integration
stage.
Phosphate treatment of magnesium alloy implants for biomedical applications 49
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(a)
(e)
(b) (f)
(c)
(g)
(d)
Figure 2.18 HE microstructure of the bone tissue at the interface between bone tissue and
magnesium implants (Xu, Pan, et al., 2009).
(h)
50 Surface Modification of Magnesium and its Alloys for Biomedical Applications
(a)
(e)
(b) (f)
(c)
(g)
(d)
Figure 2.19 Photomicrographs of BMP-2 expression at interfaces between the implants and
bones after 1, 2, 3, and 4 weeks of implantation. (aed) Naked alloy implants. (eeh) Treated
alloy implants (Xu, Pan, et al., 2009).
(h)
Phosphate treatment of magnesium alloy implants for biomedical applications 51
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0.28
Naked Mg-Mn-Zn
Treated Mg-Mn-Zn
0.24
0.20
0.16
*
0.12
0.08
Mean optical density
0.04
0.00
Figure 2.20 BMP-2 expression at the interface between Mg alloy implant and bone after
different periods of implantation. *p < 0.05 ( Xu, Pan, et al., 2009).
*
1
234
Time (week)
*
For biomaterials application, the surface bioactivity is mainly controlled by the
physical properties and the chemi cal pro pe rtie s of the surface. A porous surface at
the microscale or nanoscale level would contribute greatly to the faster adhesion
and growth of cells, and a porous coating helps in bone cells’ growth and proliferationonthesurfaceoftheimplant, resulting in a significantly stronger bond to the
parent tissue. After the phosphating process, a porous surface structure was successfully prepared on a magnesium sample, as stated in the previous section, which
would definitely contribute to the good surface bioactivi ty. On the other hand, the
surface chemical properties of the biomaterials also play a very important role in
good surface bioactivity. Various calcium phosphate coatings including brushite,
octacalcium phosphate, and HA have been successfully applied to titanium-based
and other alloys (Shirkhanzadeh, 1998; Vijayaraghavan & Bensalem, 1994)
because these compounds contain the same chemical composition or structure as
the mineral composition of natural bone, and the release of Ca
ing hydrolysis can be utilized in the course of forming new bone. Furthermore,
2þ
Ca
is also essential in chemical signaling with the cell. Research has shown
2þ
and HPO
2
4
dur-
that the osteoblast quantit y and activity on the calcium-containing titanium surface
are higher than those on the titanium surface with solely phosphate ions and the
surface of pure Ti. And Ca
2þ
sites on the material surfaces favor protein absorption, such as fibronectin and vitronectin, whic h are important cell attachmentpromoting proteins and have influence over cell attachment and spreading onto
the surface due to positive electricity and chemical and biological function
(Feng, Weng, Yang, Qu, & Zhang, 2004). Mg
2þ
and Zn2þalso play important roles
in controlling the function of osteoblasts and increasing osteoblast adhesion and the
alkaline phosphatase activity of bone cells.
52 Surface Modification of Magnesium and its Alloys for Biomedical Applications
2.9 Future trends
Phosphating treatment as a successful surface biomedical treatment has been used in
titanium implants. Although magnesium and magnesium alloys have not been used
clinically, the phosphating treatment has shown potential application as a surface
modification method for reducing the degradation rate and improving the surface
biocompatibility. However, it has to be pointed out that in the future, the following
areas might be interesting in the development of the phosphating treatment of magnesium implants:
1. Careful selection of the phosphating system, keeping it as simple as possible. Biosafety is
always the first important characteristic for an implant, especially an implant that will
degrade in vivo.
2. Balance between the surface microstructure and the degradation rate. The phosphating treat-
ment should provide magnesium with good protection from fast degradation but also cell
biocompatibility, including cell adhesion and cell proliferation.
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