Bioabsorbable magnesium alloys are becoming prominent as temporary functional implants, as they avoid the risks generated by permanent metallic implants such as persistent inflammation and late restenosis. Nevertheless, the overfast corrosion of Mg alloys under physiological conditions hinders their wider application as medical implant materials. Here we investigate a simple one-step process to introduce a cross-linked 3-amino-propyltrimethoxysilane (APTES) silane physical barrier layer on the surface of Mg–Zn–Y–Nd alloys prior to electrostatic spraying with rapamycin-eluting poly(lactic-co-glycolic acid) (PLGA) layer. Surface microstructure was characterized by scanning electron microscope and Fourier transform infrared spectroscopy. Nanoscratch test verified the superior adhesion strength of PLGA coating in the group pretreated with APTES. Electrochemical tests combined with long-term immersion results suggested that the preferable in vitro anticorrosion behavior could be achieved by dense APTES barrier. Cell morphology and proliferation data demonstrated that APTES pretreated group resulted in remarkably preferable compatibility for both human umbilical vein endothelial cells and vascular smooth muscle cells. On the basis of excellent in vitro mechenical property, the animal study on the APTES pretreated Mg–Zn–Y–Nd stent implanted into porcine coronary arteries confirmed benign tissue compatibility as well as re-endothelialization without thrombogenesis or in-stent restenosis at six-month followup.Keywords: biocompatibility; coronary artery stent; in vitro degradation; Mg−Zn−Y−Nd alloy; surface modification;

Along with the role transformation of biomaterials from bioinert substitute to regenerative inducer, the biological effect and mechanism of material-organism interaction become more important. Since most of animal tests and cellular experiments stay on the phenomenon description instead of mechanism interpretation, the development of proteomics technologies provides a golden opportunity to uncover the molecular interaction mechanism between biomaterial-organism on whole scale. This review summarizes current application of proteomics in biological effect and mechanism study of biomaterials, and discusses the development and challenges for future studies.

Bioabsorbable magnesium alloys are widely studied for various implant applications, as they reduce the risks such as severe inflammatory response existing in permanent metallic implants. However, the over-fast corrosion rate of magnesium alloy is usually an obstacle in biomedical applications. Here we report a simple two-step reaction to introduce anticorrosive silane pre-treatment on MgZnYNd alloys before coating with poly (glycolide-co-lactide) (PLGA). The first step is to immerse the NaOH-activated MgZnYNd with bistriethoxysilylethane (BTSE) to form a cross-linked silane coating layer with enhanced corrosion resistance; the second step involves immobilizing amine functional groups for forming hydrogen bond with outer PLGA coating by treating the BTSE-modified MgZnYNd with 3-amino-propyltrimethoxysilane (APTES). We characterized the BTSE-APTES pre-treated PLGA coating on MgZnYNd by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), static contact angle and Acid Orange 7 measurement. Nano-scratch test was to verify that the scratch resistance of the PLGA coating with BTSE-APTES pre-treatment was superior to direct PLGA coating. Standard electrochemical measurements along with the long-term immersion results indicated that the BTSE-APTES pre-treatment rendered better in vitro degradation behavior. Cell adhesion and cell viability tests with both vascular smooth muscle cells (VSMC) and human umbilical vein endothelial cells (EA. hy926) demonstrated that BTSE-APTES pre-treated MgZnYNd substrate had significantly more beneficial effects. The favorable anti-corrosion behavior and biocompatibility of BTSE-APTES pre-treated PLGA coatings on MgZnYNd alloy suggest that the novel two-step silanization procedure may have the great potential to enhance the performance of the magnesium-based biomaterials and provide a valid solution for the conversion modification of cardiovascular implants, taking the magnesium-based bioabsorbable materials closer to clinical application.

Magnesium (Mg)-based biomaterials have shown great potential in clinical applications. However, the cytotoxic effects of excessive Mg2+ and the corrosion products from Mg-based biomaterials, particularly their effects on neurons, have been little studied. Although viability tests are most commonly used, a functional evaluation is critically needed. Here, both methyl thiazolyl tetrazolium (MTT) and lactate dehydrogenase (LDH) assays were used to test the effect of Mg2+ and Mg-extract solution on neuronal viability. Microelectrode arrays (MEAs), which provide long-term, real-time recording of extracellular electrophysiological signals of in vitro neuronal networks, were used to test for toxic effects. The minimum effective concentrations (ECmin) of Mg2+ from the MTT and LDH assays were 3 mmol/L and 100 mmol/L, respectively, while the ECmin obtained from the MEA assay was 0.1 mmol/L. MEA data revealed significant loss of neuronal network activity when the culture was exposed to 25% Mg-extract solution, a concentration that did not affect neuronal viability. For evaluating the biocompatibility of Mg-based biomaterials with neurons, MEA electrophysiological testing is a more precise method than basic cell-viability testing.

•First proteomic analysis of bioeffect mechanism caused by biodegradable Mg•Totally 867 proteins were identified by iTRAQ LC-MS/MS in this work.•65 proteins were focused on because they were regulated within all the culture time.•The 65 proteins were associated with diverse biological processes.•Cell proliferation mechanism in Mg extract was investigated.Magnesium and its alloys gain wide attention as degradable biomaterials. In order to reveal the molecular mechanism of the influence of biodegradable magnesium on cells, proteomics analysis was performed in this work. After mouse fibroblasts (L929) were cultured with or without Mg degradation products (Mg-extract) for 8, 24, and 48 h, changes in protein expression profiles were obtained using isobaric tags for relative and absolute quantitation (iTRAQ) coupled two dimensional liquid chromatography-tandem mass spectrometry (2D LC MS/MS). A total of 867 proteins were identified (relying on at least two peptides). Compared to the control group, 205, 282, and 217 regulated proteins were identified at 8, 24, and 48 h, respectively. 65 common proteins were up or down- regulated within all the three time points, which were involved in various physiological and metabolic activities. Consistent with viability, proliferation, and cell cycle analysis, stimulated energy metabolism as well as protein synthesis pathways were discussed, indicating a possible effect of Mg-extract on L929 proliferation. Furthermore, endocytosis and focal adhesion processes were also discussed. This proteomics study uncovers early cellular mechanisms triggered by Mg degradation products and highlights the cytocompatibility of biodegradable metallic materials for biomedical applications such as stents or orthopaedic implants.

•Mg alloy with MgF2/polydopamine coating demonstrated dramatic corrosion resistance enhancement in vitro.•Vascular endothelial cells grew better than vascular smooth muscle cells on the coating.•The coating method is simple, effective and inexpensive.Mg alloy is of great potential in the application of vascular stent due to its degradation in physical environment and proper mechanical property. However its mechanical integrity does not meet the clinical requirement due to relatively fast degradation. Besides, in order to accelerate the re-endothelialization of Mg-based stents, it needs surface modification to improve the attachment, growth and adhesion of endothelial cells (ECs). To solve the main obstacles, an anti-corrosion and quick endothelialization coating was prepared on novel Mg–Zn–Y‒Nd alloy via a simple two-step immersion method in the present study, first in hydrofluoric acid (HF) then in dopamine tris–Hydrochloric acid (tris–HCl) solution. The coating was uniform and thin, which consisted of two layers—the upper was polydopamine (PDA) layer and the lower was MgF2 layer. The alloy with the coating demonstrated dramatic corrosion resistance enhancement in vitro by immersion test and electrochemical test. Moreover the HF-PDA-treated Mg alloy exhibited great performance of cell adhesion and proliferation. The coating created a favorable environment for ECs to have a competitive advantage over vascular smooth muscle cells (VSMCs), which was preferable for re-endothelialization. The results suggest that HF-PDA-treated Mg–Zn–Y‒Nd alloy has great potential in the application of vascular stent and the surface coating method is of great application value in biodegradable Mg alloy stent due to its simplicity and effectiveness.

•Mg2 + and pH have been tested for hemolysis and cytotoxicity of biomedical Mg.•Even 1000 μg/ml Mg2 + cannot cause hemolysis, but hemolysis reaches 53.8% when pH > 11.•Mg2 + > 300 μg/mL induces death of L929 and slight alkaline improves the proliferation.•Pure Mg in normal saline induces high hemolysis, but in PBS causes no hemolysis.•True reason for the hemolysis of Mg is the pH change during degradation.Good hemocompatibility and cell compatibility are essential requirements for coronary stents, especially for biodegradable magnesium alloy stents, which could change the in situ environment after implanted. In this work, the effects of magnesium ion concentration and pH value on the hemolysis and cytotoxicity have been evaluated. Solution with different Mg2 + concentration gradients and pH values of normal saline and cell culture media DMEM adjusted by MgCl2 and NaOH respectively were tested for the hemolysis and cell viability. Results show that even when the concentration of Mg2 + reaches 1000 μg/mL, it has little destructive effect on erythrocyte, and the high pH value over 11 caused by the degradation is the real reason for the high hemolysis ratio. Low concentrations of Mg2 + (< 100 μg/mL) cause no cytotoxicity to L929 cells, of which the cell viability is above 80%, while high concentrations of Mg2 + (> 300 μg/mL) could induce obvious death of the L929 cells. The pH of the extract plays a synergetic effect on cytotoxicity, due to the buffer action of the cell culture medium. To validate this conclusion, commercial pure Mg using normal saline and PBS as extract was tested with the measurement of pH and Mg2 + concentration. Pure Mg leads to a higher hemolysis ratio in normal saline (47.76%) than in buffered solution (4.38%) with different pH values and low concentration of Mg2 +. The Mg extract culture media caused no cytotoxicity, with pH = 8.44 and 47.80 μg/mL Mg2 +. It is suggested that buffered solution and dynamic condition should be adopted in the hemolysis evaluation.

Most related investigations focused on the effects of borate glass on cell proliferation/biocompatibility in vitro or bone repair in vivo; however, very few researches were carried out on other cell behaviors. Three novel borate bioglasses were designed as scaffolds for bone regeneration in this wok. Comparative effects of three bioglasses on the behaviors of osteoblastic MC3T3-E1 cells were evaluated. Excellent cytocompatibility of these novel borate bioglasses were approved in this work. Meanwhile, the promotion on cell proliferation, protein secretion and migration with minor cell apoptosis were also discussed in details, which contributed to the potential clinical application as a new biomaterial for orthopedics.

The mechanical properties, chemical properties and biocompatibility of Mg–3Sn–0.5Mn alloy were tested. A series of in vitro evaluations such as tensile test, static and dynamic immersion test, hemocompatibility test as well as cytotoxicity test were presented, with commercial magnesium alloy WE43 as the control. Mg–3Sn–0.5Mn alloy possesses suitable strength and superior ductility compared with WE43 and AZ31. Static immersion and dynamic degradation tests showed more uniform degradation with a more moderate rate for Mg–3Sn–0.5Mn alloy (0.34 mm/y in static condition and 0.25 mm/y in dynamic condition) compared with WE43 alloy (0.42 mm/y in static condition and 0.33 mm/y in dynamic condition) in Hank's solution. Blood compatibility evaluation suggested that Mg–3Sn–0.5Mn alloy had no destructive effect on erythrocyte and showed excellent anti-thrombogenicity to blood system. Besides, Mg–3Sn–0.5Mn alloy showed no inhibition effect to L929 metabolic activity and mild toxicity to vascular smooth muscle cell (VSMC) in preliminary cell viability assessment. By considering its excellent mechanical strength, corrosion resistance, low ion release rate and good biocompatibility, Mg–3Sn–0.5Mn alloy may be a promising economical candidate as biomedical implant material for load-bearing clinical applications in the future.

Recently more and more researchers query the predictability of cytotoxicity results of biomedical Mg alloys obtained according to ISO 10993 due to significant difference between in vitro and in vivo corrosion. This study aimed to observe the influence of different extraction parameters (time, volume/surface ratio and medium composition) on cytotoxicity results and illustrate whether more predictable results could be obtained by adjusting the extraction parameters. The results showed that longer extraction time and smaller extraction volume/surface ratio improve the sensitivity of screening Mg materials by making inferior Mg materials release relatively more ions to the extract; and more predictable results could not be obtained by the way of simply adding bovine serum albumin (BSA) into the extraction medium to the same level in vivo or simply using fetal bovine serum (FBS) directly as extraction medium, since BSA and FBS accelerated the corrosion of Mg materials during extraction and they affected the cells׳ health states during the test. In order to get more predictable results, in our opinions, it is necessary to establish a database of primary cells׳ hazards (metal ions, pH and H2 gas) tolerance and a set of in vitro corrosion test with high similarity in vivo, which is very difficult to realize now however.