Page 28 - IJB-2-1

Preventing bacterial adhesion on scaffolds for bone tissue engineering

cal use with anti-bacterial adhesion properties while bited different scales of porosity, i.e., channels of ca.
preserving its biocompatibility. Nanocrystalline hy- 800 µm and macropores at 0.01–600 µm range, al-
droxyapatite (HA) is a calcium phosphate-based bio- lowing good cellular internalization with adequate cell
ceramic widely used in dental and orthopedic recon- anchorage and cell colonization over the entire surface
structive medicine owing to its biocompatibility, bio- of the scaffolds.
activity and osteoconductivity [47] . Although the inhe-
rent brittleness of HA limits its use in the restoration 2.2 Development of Nanostructured Surfaces
of large bone defects, its applications include dental Albeit a well-established application of nanotechnol-
implants, periodontal treatment, alveolar ridge recon- ogy in electronic and optical engineering, the use of
struction and augmentation, orthopedics, maxillofacial nanostructured materials in medicine and biology is
surgery, and otolaryngology [47,48] . Thus, HA is com- still at its infancy. In this sense, it had demonstrated
mercially available in several physical forms, includ- the major role of surface nanotopography in bacterial
ing powders, particles, granules, dense blocks, self- adhesion and biofilm formation [15-17,50] . Different stu-
setting cements, porous 3D scaffolds, implant coatings dies using modeled nanostructured surfaces have
and composite components. The possibility to provide demonstrated the influence of the nanostructure in the
HA of anti-bacterial adhesion capability would be an inhibition of bacterial adhesion [18,51,52]
.
added value. The research group of Prof. Vallet-Regí Nature constitutes an unexhausted font of inspira-
reported the preparation of stoichiometric HA, Ca 10 tion for scientists and engineers, particularly in bio-
(PO 4) 6(OH) 2, exhibiting zwitterionic surface capable mimetics [13] . Several natural surfaces are able to main-
of inhibiting bacterial adhesion while allowing ost- tain a contaminant-free status despite the innate abun-
eoblast colonization [49] . APTES and CES organosi- dance of contaminants in the surroundings [53–57] . Most
lanes were used to functionalize the surface of HA with of these surfaces owe its non-fouling characteristics to
Θ
⊕

–NH 3 and –COO groups, respectively (Figure 1B). In its superhydrophobic properties, which in turn are
a first approach, the functionalization process was largely due to its nanotopography. Many animals (e.g.,
optimized in HA powders prepared using the con- the wing of cicadae [13] , mosquitos [58] , etc.) and plants
trolled crystallization method. Then, the validity of (e.g., lotus (Nelumbo nucifera [59] )) possess a hierar-
this functionalization method for application in HA chical surface with nanotopologic characteristics that
substrates shaped in several forms was assessed. For significantly increase its hydrophobicity, often to the
this purpose, HA 3D scaffolds were fabricated by RP point of becoming superhydrophobic [60] , and repellent
technique (see Section 3.1 for further description of to microorganism adhesion. Its antibacterial effects are
this technique) and the resulting 3D-HA scaffolds exclusively due to surface nanostructure and not to
were functionalized using APTES and CES. In vitro surface chemical effect. Several surface modification
bacterial adhesion using E. coli under physiological techniques have been widely used in the construction
conditions proved that bacterial adhesion in zwitterio- of artificial antibacterial surfaces based in nanostruc-
nic powder HA and 3D-HA decreased 92% and 99% tured surfaces [22] . These surfaces comprised a range of
respectively with respect to unmodified HA materials polymers, nanotubes and nanoparticle-based surfaces
⊕
Θ
(Figure 2A). The presence of –NH 3 /–COO zwitte- in nanoscale, exhibiting bactericidal or anti-biofouling
rionic pairs onto HA surface accounts for its bacterial effect.
anti-adhesive properties. To evaluate the biocompati- It should be highlighted that the development of
bility of these HA surfaces, in vitro assays were per- surfaces with simultaneous opposite responses toward
formed using HOS cell cultures. Thus, zwitterionic osteoblasts and bacterial proliferation would represent
and pristine HA samples, both as powder and 3D a significant achievement in orthopedic implantolo-
scaffolds were used to carry out the in vitro tests. Os- gy [50] . However, there have been very few studies ana-
teoblastic like-cell spreading was observed in all sam- lyzing surfaces that fulfill both conditions [61,62] . The
ples. High magnification scanning electron microsco- idea of tailoring surfaces with customized and selec-
py (SEM) micrographs showed viable and well-spread tive responses toward specific cell types (eukaryotic
cells, which preserved the typical osteoblast mor- and prokaryotic cells) should be mandatory in the de-
phology (Figure 2B). Regarding cell morphology, sign of biomaterials for TE purposes [63] . In this sense,
there were no differences between zwitterionic and the key role of surface nanotopography in the stimula-
bare HA samples. Moreover, HA-3D scaffolds exhi- tion of osteoblast-like cells while reducing bacterial

24 International Journal of Bioprinting (2016)–Volume 2, Issue 1