Time course study of long-term biocompatibility and foreign body reaction to intraneural polyimide-based implants

The foreign body reaction (FBR) against an implanted device is characterized by the formation of a fibrotic tissue around the implant. In the case of interfaces for peripheral nerves, used to stimulate specific group of axons and to record different nerve signals, the FBR induces a matrix deposition around the implant creating a physical separation between nerve fibers and the interface that may reduce its functionality over time. In order to understand how the FBR to intraneural interfaces evolves, polyimide non-functional devices were implanted in rat peripheral nerve. Functional tests (electrophysiological, pain and locomotion) and histological evaluation demonstrated that implanted devices did not cause any alteration in nerve function, in myelinated axons or in nerve architecture. The inflammatory response due to the surgical implantation decreased after 2 weeks. In contrast, inflammation was higher and more prolonged in the device implanted nerves with a peak after 2 weeks. With regard to tissue deposition, a tissue capsule appeared soon around the devices, acquiring maximal thickness at 2 weeks and being remodeled subsequently. Immunohistochemical analysis revealed two different cell types implicated in the FBR in the nerve: macrophages as the first cells in contact with the interface and fibroblasts that appear later at the edge of the capsule. Our results describe how the FBR against a polyimide implant in the peripheral nerve occurs and which are the main cellular players. Increasing knowledge of these responses will help to improve strategies to decrease the FBR against intraneural implants and to extend their usability

Dexamethasone Reduces the Foreign Body Reaction to Intraneural Electrode Implants in the Peripheral Nerve of the Rat

Intraneural electrodes must be in intimate contact with nerve fibers to have a proper function, but this interface is compromised due to the foreign body reaction (FBR). The FBR is characterized by a first inflammatory phase followed by a second anti-inflammatory and fibrotic phase, which results in the formation of a tissue capsule around the implant, causing physical separation between the active sites of the electrode and the nerve fibers. We have tested systemically several anti-inflammatory drugs such as dexamethasone (subcutaneous), ibuprofen and maraviroc (oral) to reduce macrophage activation, as well as clodronate liposomes (intraperitoneal) to reduce monocyte/macrophage infiltration, and sildenafil (oral) as an antifibrotic drug to reduce collagen deposition in an FBR model with longitudinal Parylene C intraneural implants in the rat sciatic nerve. Treatment with dexamethasone, ibuprofen, or clodronate significantly reduced the inflammatory reaction in the nerve in comparison to the saline group after 2 weeks of the implant, whereas sildenafil and maraviroc had no effect on infiltration of macrophages in the nerve. However, only dexamethasone was able to significantly reduce the matrix deposition around the implant. Similar positive results were obtained with dexamethasone in the case of polyimide-based intraneural implants, another polymer substrate for the electrode. These results indicate that inflammation triggers the FBR in peripheral nerves, and that anti-inflammatory treatment with dexamethasone may have beneficial effects on lengthening intraneural interface functionality

Material Biocompatibility and Applications in Metabolic Monitoring

The present dissertation focuses and expands on the optimization and application of poly(lactic-co-glycolic acid) (PLGA)/polyvinyl alcohol (PVA) composite coatings and initiates metabolic studies in small animals to identify novel biomarkers to be utilized in exhaustion prediction. The composites are used to coat implantable biosensors and improve their biocompatibility. The objectives of the work are: i) investigate species differences related to the foreign body reaction (FBR) between small and large animals; ii) develop composite coatings to prevent the FBR in large animals; iii) develop composites loaded with combinations of dexamethasone, vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF) to promote angiogenesis around implanted biosensors; and iv) apply multi-analyte monitoring to exhaustion prediction. The onset and severity of fibrosis was identified as a key difference between minipigs and rats, with minipigs demonstrating earlier onset and more severe chronic inflammation. In order to counter this, dexamethasone release must be continuous, with no lag phase. The effective dexamethasone dosing regime was 100 μg during the first day and 10 μg/day thereafter. A novel method for the preparation of microspheres containing insoluble drugs was developed to achieve homogeneous drug distribution, high loading and low burst release. Dexamethasone microspheres prepared by this method were utilized in microsphere/hydrogel composite coatings which successfully prevented FBR in a large animal model for a period of one month. Combinations of dexamethasone, VEGF and PDGF were investigated for the first time to prevent FBR and promote angiogenesis. It was determined that VEGF has to be delivered at higher doses than PDGF and an increase in dexamethasone must be accompanied by proportional increase in growth factors. An array of biomarkers that can be used in exhaustion prediction was successfully identified, with prediction times ranging from 10 to 20 minutes in normal as well as type 1 diabetic rats. It was discovered that multi-analyte biomarkers based on glucose and lactate are far more responsive in the subcutaneous tissue (an implantation-friendly compartment) than in the blood. In conclusion, the outcomes of this work contribute to the advancement biomaterials, as well as applications in exercise physiology

Haemocompatiblity improvement of metallic surfaces by covalent immobilization of heparin-liposomes

Stainless steel surfaces were processed by means of plasma enhanced chemical vapor deposition (PE-CVD) fed with acrylic acid vapors in order to functionalize them with carboxyl groups, which were subsequently activated for covalent immobilization of heparin-loaded (HEP) NH2 group-functionalized (Fun) nanoliposomes (NLs). Empty Fun or HEP non-functionalized (control) NLs were used as controls. NLs were characterized for mean diameter, surface charge and heparin encapsulation/release. Different lipid compositions were used for NL construction; PC/Chol (2:1 mol/mol) or PC/Chol (4:1 mol/mol) (fluid type vesicles) [ which allow gradual release of heparin] and DSPC/Chol (2:1 mol/mol) (rigid type vesicles). Surface haemocompatibility was tested by measuring blood clotting time. Platelet adhesion on surfaces was evaluated morphologically by SEM and CLSM. The haemocompatibility of plasma-processed surfaces was improved (compared to untreated surfaces); Fun-HEP NL-coated surfaces demonstrated highest coagulation times. For short surface/blood incubation periods, surfaces coated with Fun-HEP NLs consisting of PC/Chol (2:1) had higher coagulation times (compared to DSPC/Chol NLs) due to faster release of heparin. Heparin release rate from the various NL types and surface platelet adhesion results were in agreement with the corresponding blood coagulation times. Concluding, covalent immobilization of drug entrapping NLs on plasma processed surfaces is a potential method for preparation of controlled-rate drug-eluting metallic stents or devices

Covalent immobilization of liposomes on plasma functionalized metallic surfaces

A method was developed to functionalize biomedical metals with liposomes. The novelty of the method includes the plasma-functionalization of the metal surface with proper chemical groups to be used as anchor sites for the covalent immobilization of the liposomes. Stainless steel (SS-316) disks were processed in radiofrequency glow discharges fed with vapors of acrylic acid to coat them with thin adherent films characterized by surface carboxylic groups, where liposomes were covalently bound through the formation of amide bonds. For this, liposomes decorated with polyethylene glycol molecules bearing terminal amine-groups were prepared. After ensuring that the liposomes remain intact, under the conditions applying for immobilization; different attachment conditions were evaluated (incubation time, concentration of liposome dispersion) for optimization of the technique. Immobilization of calcein-entrapping liposomes was evaluated by monitoring the percent of calcein attached on the surfaces. Best results were obtained when liposome dispersions with 5 mg/ml (liposomal lipid) concentration were incubated on each disk for 24 h at 37 degrees C. The method is proposed for developing drug-eluting biomedical materials or devices by using liposomes that have appropriate membrane compositions and are loaded with drugs or other bioactive agents