Yvonne Moussy

Use of hydrogel coating to improve the performance of implanted glucose sensors.

In order to protect implanted glucose sensors from biofouling, novel hydrogels (146-217% water by mass) were developed based on a copolymer of hydroxyethyl methacrylate (HEMA) and 2,3-dihydroxypropyl methacrylate (DHPMA). The porosity and mechanical properties of the hydrogels were improved using N-vinyl-2-pyrrolidinone (VP) and ethyleneglycol dimethacrylate (EGDMA). The results of SEM and DSC FT-IT analyses showed that the hydrogel (VP30) produced from a monomeric mixture of 34.5% HEMA, 34.5% DHPMA, 30% VP and 1% EDGMA (mol%) had an excellent pore structure, high water content at swelling equilibrium (W eq=166% by mass) and acceptable mechanical properties. Two kinds of VP30-coated sensors, Pt/GOx/VP30 and Pt/GOx/epoxy-polyurethane (EPU)/VP30 sensors were examined in glucose solutions during a period of 4 weeks. The Pt/GOx/VP30 sensors produced large response currents but the response linearity was poor. Therefore, further studies were focused on the Pt/GOx/EPU/VP30 sensors. With a diffusion-limiting epoxy-polyurethane membrane, the linearity was improved (2-30 mM) and the response time was within 5 min. Eight Pt/GOx/EPU/VP30 sensors were subcutaneously implanted in rats and tested once per week over 4 weeks. All of the implanted sensors kept functioning for at least 21 days and 3 out of 8 sensors still functioned at day 28. Histology revealed that the fibrous capsules surrounding hydrogel-coated sensors were thinner than those surrounding Pt/GOx/EPU sensors after 28 days of implantation.

Coil-type implantable glucose biosensor with excess enzyme loading.

As part of our overall long-term objective of designing a glucose sensor for long-term subcutaneous implantation, a coil-type implantable glucose sensor loaded with excess glucose oxidase (GOD) inside the coils of a 0.125mm diameter coiled platinum-iridium wire has been developed. The excess GOD was immobilized in a glutaraldehyde/bovine serum albumin (BSA) gel reinforced with cotton and located inside the coils chamber of the sensor. The excess GOD increased the lifetime of the sensor. Based on this coil-type design, various coil-type glucose sensors with cellulose acetate (CA), poly(vinyl chloride)(PVC), polyurethane (PU), poly(bisphenol A carbonate) (PC) and Nafion outer membranes were investigated and compared. Comparatively, Nafion based biosensors provided the best long-term response stability. However, Nafion can still not meet the lifetime requirement of the coil-type sensor with high enzyme loading because the observed function failure of these sensors was indeed caused by outer membrane damage rather than loss of enzyme activity. Additional experiments also revealed that hydrogen peroxide accumulation occurred in the GOD impregnated cotton when the sensors were not polarized which could cause a small false positive measurement. However, this artifact can be easily avoided by using an appropriate measurement technique.

A dexamethasone-loaded PLGA microspheres/collagen scaffold composite for implantable glucose sensors

We have developed a new dexamethasone (Dex)-loaded poly(lactic-co-glycolic acid) microspheres/porous collagen scaffold composite for implantable glucose sensors. The scaffolds were fabricated around the sensing element of the sensors and crosslinked using nordihydroguaiaretic acid (NDGA). The microspheres containing Dex were incorporated into the NDGA-crosslinked collagen scaffold by dipping in microsphere suspension in either water or Pluronic. The loading efficiencies of Dex in the microspheres and the scaffold were determined using high performance liquid chromatography. The microspheres/scaffold composite fabricated using microspheres in the hydrogel solution had a better loading efficiency than when microspheres were in water suspension. The composite fabricated using the hydrogel also showed a slower and more sustained drug release than the standard microspheres in vitro during a 4 week study and did not significantly affect the function of the sensors in vitro. The sensors with the composite that were still functional retained above 50% of their original sensitivity at 2 weeks. Histology showed that the inflammatory response to the Dex-loaded composite was much lower than for the control scaffold at 2 and 4 weeks after implantation. The Dex-loaded composite system might be useful to reduce inflammation to implanted glucose sensors and therefore extend their function and lifetime.

A novel porous collagen scaffold around an implantable biosensor for improving biocompatibility. II. Long-term in vitro/in vivo sensitivity characteristics of sensors with NDGA- or GA-crosslinked collagen scaffolds

We have developed a new 3D porous and biostable collagen scaffold for implantable glucose sensors. The scaffolds were fabricated around the sensors and crosslinked using nordihydroguaiaretic acid (NDGA) or glutaraldehyde (GA) to enhance physical and biological stability. The effect of the scaffolds on sensor function and biocompatibility was examined during long-term (>or=28 days) in vitro and in vivo experiments and compared with control bare sensors. To evaluate the effect of the sensor length on micromotion and sensor function, we also fabricated short and long sensors. 3D porous scaffold application around glucose sensors did not significantly affect the long-term in vitro sensitivity of the sensors. The scaffolds, crosslinked by either NDGA or GA, remained stable around the sensors during the 4 week in vitro study. In the long-term in vivo study, the sensitivity of the short sensors was higher than the sensitivity of long sensors presumably because of less micromotion in the subcutis of the rats. The sensors with NDGA-crosslinked scaffolds had a higher sensitivity than the sensors with GA-crosslinked scaffolds. Histological examination showed that NDGA-crosslinked scaffolds retained their physical structure with reduced inflammation when compared with the GA-crosslinked scaffolds. Therefore, the application of NDGA-crosslinked collagen scaffolds might be a good method for enhancing the function and lifetime of implantable biosensors by minimizing the in vivo foreign body response.

Distribution of [3H]Dexamethasone in Rat Subcutaneous Tissue after Delivery from Osmotic Pumps

Inflammation surrounding implantable glucose sensors may be controlled through local release of dexamethasone at the site of implantation. In the present study, we evaluated the distribution of dexamethasone in rat subcutaneous tissue during the first 2.5 days after local release. Osmotic pumps containing [3H]dexamethasone were implanted into the subcutaneous tissue of rats. Digital autoradiography was used to measure the distribution of the [3H]dexamethasone within the subcutaneous tissue at 6, 24, and 60 h after implantation. Measured concentration profiles, near the catheter tip through which the agent was released, were compared to mathematical models of drug diffusion and elimination. The results demonstrate that the majority of the [3H]dexamethasone delivered into the subcutaneous tissue was found within a 3 mm region surrounding the catheter tip. There was good agreement between the experimental data and the mathematical model. The diffusion coefficient for dexamethasone in subcutaneous tissue was found to be D = 4.11 ± 1.77 × 10−10 m2/s, and the elimination rate constant was found to be k = 3.65 ± 2.24 × 10−5 s−1. The diffusion coefficient and elimination rate constants for dexamethasone in subcutaneous tissue have not been previously reported. The use of a mathematical model may be useful in predicting the effectiveness of local delivery of dexamethasone around implantable glucose sensors.

Transport characteristics of a novel local drug delivery system using nordihydroguaiaretic acid (NDGA)-polymerized collagen fibers

The use of nordihydroguaiaretic acid (NDGA)‐polymerized collagen fibers as a novel local drug delivery system is introduced. The drug loading of these biocompatible fibers is illustrated with the anti‐inflammatory agents dexamethasone and dexamethasone 21‐phosphate. Capillary zone electrophoresis was used to measure the amount of drug released from the fibers into phosphate buffered saline with time. From these measurements and the use of a mathematical model, we were able to determine the diffusion coefficients for dexamethasone (D = 1.86 × 10‐14 m2/s) and dexamethasone 21‐phosphate (D = 2.36 × 10‐13 m2/s) in the NDGA collagen fibers. These values have not been previously reported. These fibers can be used to load other agents as well. The diffusion coefficient of any agent loaded in these fibers can be determined using the techniques and mathematical method described. The rate of drug release from the fibers can be controlled using a PLGA coating. The overall importance of this paper is the potential broad application of this novel drug delivery system for the treatment of various human diseases.

Diffusion of [3H]dexamethasone in rat subcutaneous slices after injection measured by digital autoradiography.

A relatively simple method for the determination of the diffusion coefficient of a substance that has been injected into tissue is described. We illustrate this method using [3H]dexamethasone injected into the subcutaneous tissue of rats. Digital autoradiography was used to measure the distribution of the [3H]dexamethasone within the subcutaneous tissue at 2.5 and 20 min after injection. Measured concentration profiles of the injection were compared to a mathematical model of drug diffusion from an injection. There was good agreement between the experimental data and the mathematical model. The diffusion coefficient found using this simple injection method was (4.01 ± 2.01) × 10−10 m2/s. This D value was very close to the value of D = (4.11 ± 1.77) × 10−10 m2/s found previously using different mathematical and experimental techniques with osmotic pumps implanted for 6, 24, and 60 h in rats (1). The simple method given here for the determination of the diffusion coefficient is general enough to be applied to other substances and tissues as well.

Transcutaneous Implantation Methods for Improving the Long-Term Performance of Glucose Sensors in Rats

Translation of sensor design and function in animal models to human use is an ongoing challenge due to tissue anatomical and physiological differences between species, even at presumably analogous implant locations. Nevertheless, preclinical testing of sensors for long-term glucose monitoring in animals is required for evaluating sensor function in order to improve sensor design. Long-term glucose sensor testing in common laboratory animals (e.g., mice and rats) is especially difficult due to their small size, as well as limited site availability for sensor placement without disturbance or removal by the subject. However, improvements in sensor design and implantation methods to improve sensor survival in these animals could accelerate our understanding of the role of tissue reactions to sensor components, as well as allow reliable testing of biomaterials and various drug or growth factor delivery systems to potentially minimize or modulate tissue reactions. In this study, methods to secure a wire-type subcutaneous sensor in rats for a long period of time (ges28 d), utilizing new implantation techniques and devices were evaluated. Anchoring devices were incorporated into the sensor design and appropriate implantation methods were used to: (1) minimize potential membrane damage caused by animal motion; (2) prevent removal of the entire sensor or sensor wires by the animal; and (3) allow exterior access to wires for periodic sensor performance testing. The anchoring devices for securing sensors to the skin internally, which were sequentially investigated and improved (Protocol A to C), included a modified 22 gauge intravenous winged catheter (Protocol A), Silastic tubing (Protocol B) or silk suture loops held in place by Silastic tubing (Protocol C). The results show that after four weeks implantation, 60% (n = 10), 70% (n = 10), and 92% (n = 12) of the implanted devices survived (Protocols A, B, and C, respectively). Functional testing showed that 30% (n = 10), 40% (n = 10), and 58% (n = 12) of the sensors still worked well four weeks after implantation (Protocols A, B, and C, respectively). No infections were visibly evident at the sites of sensor implantation at any time during the testing period for all protocols. Protocol C shows promise as a viable method for future sensor studies because of the anchoring devices small size and because it was nearly impossible for rats to remove or damage the sensors.

Transport Characteristics of a Novel Local Drug Delivery System Using Nordihydroguaiaretic Acid (NDGA)-Polymerized Collagen Fibers

The use of nordihydroguaiaretic acid (NDGA)-polymerized collagen fibers as a novel local drug delivery system is introduced. The drug loading of these biocompatible fibers is illustrated with the anti-inflammatory agents dexamethasone and dexamethasone 21-phosphate. Capillary zone electrophoresis was used to measure the amount of drug released from the fibers into phosphate buffered saline with time. From these measurements and the use of a mathematical model, we were able to determine the diffusion coefficients for dexamethasone (D = 1.86 x 10(-14) m2/s) and dexamethasone 21-phosphate (D = 2.36 x 10(-13) m2/s) in the NDGA collagen fibers. These values have not been previously reported. These fibers can be used to load other agents as well. The diffusion coefficient of any agent loaded in these fibers can be determined using the techniques and mathematical method described. The rate of drug release from the fibers can be controlled using a PLGA coating. The overall importance of this paper is the potential broad application of this novel drug delivery system for the treatment of various human diseases.

Two Methods of Determining [3H]Dexamethasone Distribution in Rat Subcutaneous Tissue

Several recent reports suggest that controlled local release of dexamethasone may be useful for preventing inflammation around an implantable glucose sensor [1,2]. This decrease in inflammation is expected to increase glucose sensor function and lifetime. Local delivery of dexamethasone would permit high interstitial drug concentrations at the site of glucose sensor implantation without producing high systemic drug levels. Although dexamethasone is a commonly used anti-inflammatory agent, its local concentration, diffusion coefficient and rate of elimination have not been reported following subcutaneous release. The ability of dexamethasone to penetrate subcutaneous tissue can be measured and quantified by comparison to mathematical models [3]. This method allows a reliable estimate of the drug concentration in the tissue near the implanted glucose sensor.Copyright