Physiomimetic biocompatibility evaluation of directly printed degradable porous iron implants using various cell types.

Additively manufactured (AM) degradable porous metallic biomaterials offer unique opportunities for satisfying the design requirements of an ideal bone substitute. Among the currently available biodegradable metals, iron has the highest elastic modulus, meaning that it would benefit the most from porous design. Given the successful preclinical applications of such biomaterials for the treatment of cardiovascular diseases, the moderate compatibility of AM porous iron with osteoblast-like cells, reported in earlier studies, has been surprising. This may be because, as opposed to static in vitro conditions, the biodegradation products of iron in vivo are transported away and excreted. To better mimic the in situ situations of biodegradable biomaterials after implantation, we compared the biodegradation behavior and cytocompatibility of AM porous iron under static conditions to the conditions with dynamic in situ-like fluid flow perfusion in a bioreactor. Furthermore, the compatibility of these scaffolds with four different cell types was evaluated to better understand the implications of these implants for the complex process of natural wound healing. These included endothelial cells, L929 fibroblasts, RAW264.7 macrophage-like cells, and osteoblastic MG-63 cells. The biodegradation rate of the scaffolds was significantly increased in the perfusion bioreactor as compared to static immersion. Under either condition, the compatibility with L929 cells was the best. Moreover, the compatibility with all the cell types was much enhanced under physiomimetic dynamic flow conditions as compared to static biodegradation. Our study highlights the importance of physiomimetic culture conditions and cell type selection when evaluating the cytocompatibility of degradable biomaterials in vitro. STATEMENT OF SIGNIFICANCE: Additively manufactured (AM) degradable porous metals offer unique opportunities for the treatment of large bony defects. Despite the successful preclinical applications of biodegradable iron in the cardiovascular field, the moderate compatibility of AM porous iron with osteoblast-like cells was reported. To better mimic the in vivo condition, we compared the biodegradation behavior and cytocompatibility of AM porous iron under static condition to dynamic perfusion. Furthermore, the compatibility of these scaffolds with various cell types was evaluated to better simulate the process of natural wound healing. Our study suggests that AM porous iron holds great promise for orthopedic applications, while also highlighting the importance of physio-mimetic culture conditions and cell type selection when evaluating the cytocompatibility of degradable biomaterials in vitro.

Additively manufactured biodegradable porous zinc.

Additively manufacturing (AM) opens up the possibility for biodegradable metals to possess uniquely combined characteristics that are desired for bone substitution, including bone-mimicking mechanical properties, topologically ordered porous structure, pore interconnectivity and biodegradability. Zinc is considered to be one of the promising biomaterials with respect to biodegradation rate and biocompatibility. However, no information regarding the biodegradability and biocompatibility of topologically ordered AM porous zinc is yet available. Here, we applied powder bed fusion to fabricate porous zinc with a topologically ordered diamond structure. An integrative study was conducted on the static and dynamic biodegradation behavior (in vitro, up to 4 weeks), evolution of mechanical properties with increasing immersion time, electrochemical performance, and biocompatibility of the AM porous zinc. The specimens lost 7.8% of their weight after 4 weeks of dynamic immersion in a revised simulated body fluid. The mechanisms of biodegradation were site-dependent and differed from the top of the specimens to the bottom. During the whole in vitro immersion time of 4 weeks, the elastic modulus values of the AM porous zinc (E = 700-1000 MPa) even increased and remained within the scope of those of cancellous bone. Indirect cytotoxicity revealed good cellular activity up to 72 h according to ISO 10,993-5 and -12. Live-dead staining confirmed good viability of MG-63 cells cultured on the surface of the AM porous zinc. These important findings could open up unprecedented opportunities for the development of multifunctional bone substituting materials that will enable reconstruction and regeneration of critical-size load-bearing bone defects. STATEMENT OF SIGNIFICANCE: No information regarding the biodegradability and biocompatibility of topologically ordered AM porous zinc is available. We applied selective laser melting to fabricate topologically ordered porous zinc and conducted a comprehensive study on the biodegradation behavior, electrochemical performance, time-dependent mechanical properties, and biocompatibility of the scaffolds. The specimens lost 7.8% of their weight after4 weeks dynamic biodegradation while their mechanical properties surprisingly increased after 4 weeks. Indirect cytotoxicity revealed good cellular activity up to 72 h. Intimate contact between MG-63 cells and the scaffolds was also observed. These important findings could open up unprecedented opportunities for the development of multifunctional bone substituting materials that mimic bone properties and enable full regeneration of critical-size load-bearing bony defects.

Additively manufactured functionally graded biodegradable porous zinc.

Topological design provides additively manufactured (AM) biodegradable porous metallic biomaterials with a unique opportunity to adjust their biodegradation behavior and mechanical properties, thereby satisfying the requirements for ideal bone substitutes. However, no information is available yet concerning the effect of topological design on the performance of AM porous zinc (Zn) that outperforms Mg and Fe in biodegradation behavior. Here, we studied one functionally graded and two uniform AM porous Zn designs with diamond unit cell. Cylindrical specimens were fabricated from pure Zn powder by using a powder bed fusion technique, followed by a comprehensive study on their static and dynamic biodegradation behaviors, mechanical properties, permeability, and biocompatibility. Topological design, indeed, affected the biodegradation behavior of the specimens, as evidenced by 150% variations in biodegradation rate between the three different designs. After in vitro dynamic immersion for 28 days, the AM porous Zn had weight losses of 7-12%, relying on the topological design. The degradation rates satisfied the desired biodegradation time of 1-2 years for bone substitution. The mechanical properties of the biodegraded specimens of all the groups maintained within the range of those of cancellous bone. As opposed to the trends observed for other biodegradable porous metals, after 28 days of in vitro biodegradation, the yield strengths of the specimens of all the groups (σy = 7-14 MPa) increased consistently, as compared to those of the as-built specimens (σy = 4-11 MPa). Moreover, AM porous Zn showed excellent biocompatibility, given that the cellular activities in none of the groups differed from the Ti controls for up to 72 h. Using topological design of AM porous Zn for controlling its mechanical properties and degradation behavior is thus clearly promising, thereby rendering flexibility to the material to meet a variety of clinical requirements.

Additively manufactured functionally graded biodegradable porous iron.

Additively manufactured (AM) functionally graded porous metallic biomaterials offer unique opportunities to satisfy the contradictory design requirements of an ideal bone substitute. However, no functionally graded porous structures have ever been 3D-printed from biodegradable metals, even though biodegradability is crucial both for full tissue regeneration and for the prevention of implant-associated infections in the long term. Here, we present the first ever report on AM functionally graded biodegradable porous metallic biomaterials. We made use of a diamond unit cell for the topological design of four different types of porous structures including two functionally graded structures and two reference uniform structures. Specimens were then fabricated from pure iron powder using selective laser melting (SLM), followed by experimental and computational analyses of their permeability, dynamic biodegradation behavior, mechanical properties, and cytocompatibility. It was found that the topological design with functional gradients controlled the fluid flow, mass transport properties and biodegradation behavior of the AM porous iron specimens, as up to 4-fold variations in permeability and up to 3-fold variations in biodegradation rate were observed for the different experimental groups. After 4 weeks of in vitro biodegradation, the AM porous scaffolds lost 5-16% of their weight. This falls into the desired range of biodegradation rates for bone substitution and confirms our hypothesis that topological design could indeed accelerate the biodegradation of otherwise slowly degrading metals, like iron. Even after 4 weeks of biodegradation, the mechanical properties of the specimens (i.e., E = 0.5-2.1 GPa, σy = 8-48 MPa) remained within the range of the values reported for trabecular bone. Design-dependent cell viability did not differ from gold standard controls for up to 48 h. This study clearly shows the great potential of AM functionally graded porous iron as a bone substituting material. Moreover, we demonstrate that complex topological design permits the control of mechanical properties, degradation behavior of AM porous metallic biomaterials. STATEMENT OF SIGNIFICANCE: No functionally graded porous structures have ever been 3D-printed from biodegradable metals, even though biodegradability is crucial both for full tissue regeneration and for the prevention of implant-associated infections in the long term. Here, we present the first report on 3D-printed functionally graded biodegradable porous metallic biomaterials. Our results suggest that topological design in general, and functional gradients in particular can be used as an important tool for adjusting the biodegradation behavior of AM porous metallic biomaterials. The biodegradation rate and mass transport properties of AM porous iron can be increased while maintaining the bone-mimicking mechanical properties of these biomaterials. The observations reported here underline the importance of proper topological design in the development of AM porous biodegradable metals.

Additively manufactured biodegradable porous magnesium.

An ideal bone substituting material should be bone-mimicking in terms of mechanical properties, present a precisely controlled and fully interconnected porous structure, and degrade in the human body to allow for full regeneration of large bony defects. However, simultaneously satisfying all these three requirements has so far been highly challenging. Here we present topologically ordered porous magnesium (WE43) scaffolds based on the diamond unit cell that were fabricated by selective laser melting (SLM) and satisfy all the requirements. We studied the in vitro biodegradation behavior (up to 4 weeks), mechanical properties and biocompatibility of the developed scaffolds. The mechanical properties of the AM porous WE43 (E = 700-800 MPa) scaffolds were found to fall into the range of the values reported for trabecular bone even after 4 weeks of biodegradation. Scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), electrochemical tests and µCT revealed a unique biodegradation mechanism that started with uniform corrosion, followed by localized corrosion, particularly in the center of the scaffolds. Biocompatibility tests performed up to 72 h showed level 0 cytotoxicity (according to ISO 10993-5 and -12), except for one time point (i.e., 24 h). Intimate contact between cells (MG-63) and the scaffolds was also observed in SEM images. The study shows for the first time that AM of porous Mg may provide distinct possibilities to adjust biodegradation profile through topological design and open up unprecedented opportunities to develop multifunctional bone substituting materials that mimic bone properties and enable full regeneration of critical-size load-bearing bony defects. STATEMENT OF SIGNIFICANCE: The ideal biomaterials for bone tissue regeneration should be bone-mimicking in terms of mechanical properties, present a fully interconnected porous structure, and exhibit a specific biodegradation behavior to enable full regeneration of bony defects. Recent advances in additive manufacturing have resulted in biomaterials that satisfy the first two requirements but simultaneously satisfying the third requirement has proven challenging so far. Here we present additively manufactured porous magnesium structures that have the potential to satisfy all above-mentioned requirements. Even after 4 weeks of biodegradation, the mechanical properties of the porous structures were found to be within those reported for native bone. Moreover, our comprehensive electrochemical, mechanical, topological, and biological study revealed a unique biodegradation behavior and the limited cytotoxicity of the developed biomaterials.

Additively manufactured biodegradable porous iron.

Additively manufactured (AM) topologically ordered porous metallic biomaterials with the proper biodegradation profile offer a unique combination of properties ideal for bone regeneration. These include a fully interconnected porous structure, bone-mimicking mechanical properties, and the possibility of fully regenerating bony defects. Most of such biomaterials are, however, based on magnesium and, thus, degrade too fast. Here, we present the first report on topologically ordered porous iron made by Direct Metal Printing (DMP). The topological design was based on a repetitive diamond unit cell. We conducted a comprehensive study on the in vitro biodegradation behavior (up to 28 days), electrochemical performance, time-dependent mechanical properties, and biocompatibility of the scaffolds. The mechanical properties of AM porous iron (E = 1600-1800 MPa) were still within the range of the values reported for trabecular bone after 28 days of biodegradation. Electrochemical tests showed up to ≈12 times higher rates of biodegradation for AM porous iron as compared to that of cold-rolled (CR) iron, while only 3.1% of weight loss was measured after 4 weeks of immersion tests. The biodegradation mechanisms were found to be topology-dependent and different between the periphery and central parts of the scaffolds. While direct contact between MG-63 cells and scaffolds revealed substantial and almost instant cytotoxicity in static cell culture, as compared to Ti-6Al-4V, the cytocompatibility according to ISO 10993 was reasonable in in vitro assays for up to 72 h. This study shows how DMP could be used to increase the surface area and decrease the grain sizes of topologically ordered porous metallic biomaterials made from metals that are usually considered to degrade too slowly (e.g., iron), opening up many new opportunities for the development of biodegradable metallic biomaterials. STATEMENT OF SIGNIFICANCE: Biodegradation in general and proper biodegradation profile in particular are perhaps the most important requirements that additively manufactured (AM) topologically ordered porous metallic biomaterials should offer in order to become the ideal biomaterial for bone regeneration. Currently, most biodegradable metallic biomaterials are based on magnesium, which degrade fast with gas generation. Here, we present the first report on topologically ordered porous iron made by Direct Metal Printing (DMP). We also conducted a comprehensive study on the biodegradation behavior, electrochemical performance, biocompatibility, and the time evolution of the mechanical properties of the implants. We show that these implants possess bone-mimicking mechanical properties, accelerated degradation rate, and reasonable cytocompatibility, opening up many new opportunities for the development of iron-based biodegradable materials.