Rebecca A. Wachs

Implantable sensor technology: From research to clinical practice

Abstract For decades, implantable sensors have been used in research to provide comprehensive understanding of the biomechanics of the human musculoskeletal system. These complex sensor systems have improved our understanding of the in vivo environment by yielding in vivo measurements of force, torque, pressure, and temperature. Historically, implants have been modified to be used as vehicles for sensors and telemetry systems. Recently, microfabrication and nanofabrication technology have sufficiently evolved that wireless, passive sensor systems can be incorporated into implants or tissue with minimal or no modification to the host implant. At the same time, sensor technology costs per unit have become less expensive, providing opportunities for use in daily clinical practice. Although diagnostic implantable sensors can be used clinically without significant increases in expense or surgical time, to date, orthopaedic smart implants have been used exclusively as research tools. These implantable sensors can facilitate personalized medicine by providing exquisitely accurate in vivo data unique to each patient.

Creation of an injectable in situ gelling native extracellular matrix for nucleus pulposus tissue engineering

BACKGROUND CONTEXT Disc degeneration is the leading cause of low back pain and is often characterized by a loss of disc height, resulting from cleavage of chondroitin sulfate proteoglycans (CSPGs) present in the nucleus pulposus. Intact CSPGs are critical to water retention and maintenance of the nucleus osmotic pressure. Decellularization of healthy nucleus pulposus tissue has the potential to serve as an ideal matrix for tissue engineering of the disc because of the presence of native disc proteins and CSPGs. Injectable in situ gelling matrices are the most viable therapeutic option to prevent damage to the anulus fibrosus and future disc degeneration. PURPOSE The purpose of this research was to create a gentle decellularization method for use on healthy nucleus pulposus tissue explants and to develop an injectable formulation of this matrix to enable therapeutic use without substantial tissue disruption. STUDY DESIGN Porcine nuclei pulposi were isolated, decellularized, and solubilized. Samples were assessed to determine the degree of cell removal, matrix maintenance, gelation ability, cytotoxic residuals, and native cell viability. METHODS Nuclei pulposi were decellularized using serial detergent, buffer, and enzyme treatments. Decellularized nuclei pulposi were solubilized, neutralized, and buffered. The efficacy of decellularization was assessed by quantifying DNA removal and matrix preservation. An elution study was performed to confirm removal of cytotoxic residuals. Gelation kinetics and injectability were quantified. Long-term in vitro experiments were performed with nucleus pulposus cells to ensure cell viability and native matrix production within the injectable decellularized nucleus pulposus matrices. RESULTS This work resulted in the creation of a robust acellular matrix (>96% DNA removal) with highly preserved sulfated glycosaminoglycans (>47%), and collagen content and microstructure similar to native nucleus pulposus, indicating preservation of disc components. Furthermore, it was possible to create an injectable formulation that gelled in situ within 45 minutes and formed fibrillar collagen with similar diameters to native nucleus pulposus. The processing did not result in any remaining cytotoxic residuals. Solubilized decellularized nucleus pulposus samples seeded with nucleus pulposus cells maintained robust viability (>89%) up to 21 days of culture in vitro, with morphology similar to native nucleus pulposus cells, and exhibited significantly enhanced sulfated glycosaminoglycans production over 21 days. CONCLUSIONS A gentle decellularization of porcine nucleus pulposus followed by solubilization enabled the creation of an injectable tissue-specific matrix that is well tolerated in vitro by nucleus pulposus cells. These matrices have the potential to be used as a minimally invasive nucleus pulposus therapeutic to restore disc height.

Decellularized peripheral nerve supports Schwann cell transplants and axon growth following spinal cord injury

Schwann cell (SC) transplantation has been comprehensively studied as a strategy for spinal cord injury (SCI) repair. SCs are neuroprotective and promote axon regeneration and myelination. Nonetheless, substantial SC death occurs post-implantation, which limits therapeutic efficacy. The use of extracellular matrix (ECM)-derived matrices, such as Matrigel, supports transplanted SC survival and axon growth, resulting in improved motor function. Because appropriate matrices are needed for clinical translation, we test here the use of an acellular injectable peripheral nerve (iPN) matrix. Implantation of SCs in iPN into a contusion lesion did not alter immune cell infiltration compared to injury only controls. iPN implants were larger and contained twice as many SC-myelinated axons as Matrigel grafts. SC/iPN animals performed as well as the SC/Matrigel group in the BBB locomotor test, and made fewer errors on the grid walk at 4 weeks, equalizing at 8 weeks. The fact that this clinically relevant iPN matrix is immunologically tolerated and supports SC survival and axon growth within the graft offers a highly translational possibility for improving efficacy of SC treatment after SCI. To our knowledge, it is the first time that an injectable PN matrix is being evaluated to improve the efficacy of SC transplantation in SCI repair.

The Open Source GAITOR Suite for Rodent Gait Analysis

Locomotive changes are often associated with disease or injury, and these changes can be quantified through gait analysis. Gait analysis has been applied to preclinical studies, providing quantitative behavioural assessment with a reasonable clinical analogue. However, available gait analysis technology for small animals is somewhat limited. Furthermore, technological and analytical challenges can limit the effectiveness of preclinical gait analysis. The Gait Analysis Instrumentation and Technology Optimized for Rodents (GAITOR) Suite is designed to increase the accessibility of preclinical gait analysis to researchers, facilitating hardware and software customization for broad applications. Here, the GAITOR Suite’s utility is demonstrated in 4 models: a monoiodoacetate (MIA) injection model of joint pain, a sciatic nerve injury model, an elbow joint contracture model, and a spinal cord injury model. The GAITOR Suite identified unique compensatory gait patterns in each model, demonstrating the software’s utility for detecting gait changes in rodent models of highly disparate injuries and diseases. Robust gait analysis may improve preclinical model selection, disease sequelae assessment, and evaluation of potential therapeutics. Our group has provided the GAITOR Suite as an open resource to the research community at www.GAITOR.org, aiming to promote and improve the implementation of gait analysis in preclinical rodent models.

Development of an apoptosis-assisted decellularization method for maximal preservation of nerve tissue structure

Preservation of tissue structure is often a primary goal when optimizing tissue and organ decellularization methods. Many current protocols nonetheless rely on detergents that aid extraction of cellular components but also damage tissue architecture. It may be more beneficial to leverage an innate cellular process such as apoptosis and promote cell removal without the use of damaging reagents. During apoptosis, a cell detaches from the extracellular matrix, degrades its internal components, and fragments its contents for easier clearance. We have developed a method that leverages this process to achieve tissue decellularization using only mild wash buffers. We have demonstrated that treating peripheral nerve tissue with camptothecin induced both an early marker of apoptosis, cleaved caspase-3 expression, as well as a late stage marker, TUNEL+ DNA fragmentation. Clearance of the cellular components was then achieved in an apoptosis-dependent manner using a gentle wash in hypertonic phosphate buffered saline followed by DNase treatment. This wash paradigm did not significantly affect collagen or glycosaminoglycan content, but it was sufficient to remove any trace of the cytotoxic compound based on conditioned media experiments. The resulting acellular tissue graft was immunogenically tolerated in vivo and exhibited an intact basal lamina microarchitecture mimicking that of native, unprocessed nerve. Hence, ex vivo induction of apoptosis is a promising method to decellularize tissue without the use of harsh reagents while better preserving the benefits of native tissue such as tissue-specific composition and microarchitecture. STATEMENT OF SIGNIFICANCE Tissue decellularization has expanded the ability to generate non-immunogenic organ replacements for a broad range of health applications. Current technologies typically rely on the use of harsh agents for clearing cellular debris, altering the tissue structure and potentially diminishing the pro-regenerative effects. We have developed a method for effectively, yet gently, removing cellular components from peripheral nerve tissue while preserving the native tissue architecture. The novelty of this process is in the induction of programmed cell death - or apoptosis - via a general cytotoxin, thereby enabling antigen clearance using only hypertonic wash buffers. The resulting acellular nerve scaffolds are nearly identical to unprocessed tissue on a microscopic level and elicit low immune responses comparable to an isograft negative control in a model of subcutaneous implantation.

Implantation methodology development for tissue-engineered-electronic-neural-interface (TEENI) devices

Regenerative peripheral-nerve interfaces are a novel method for integrating with the peripheral nervous system. These devices have the potential to isolate and transduce both afferent (sensory) and efferent (motor) neural signals to produce fine control of advanced prosthetics. We have developed a novel regenerative device comprised of microfabricated polyimide electrode threads supported by a hydrogel scaffold containing methacrylated hyaluronic acid, collagen I, and laminin to enable intimate contact with regenerating axons. While this advanced device holds theoretical promise for establishing a stable chronic neural interface, it also requires a novel surgical approach in comparison to current existing methods of peripheral neural interface technologies. Here we describe the development of the surgical methodology required for successful chronic implantation of the TEENI device in the rat sciatic nerve.

Histological evaluation of chronically implanted tissue-engineered-electronic-neural-interface (TEENI) devices

Neural-interface devices have the potential to isolate and transduce both afferent (sensory) and efferent (motor) neural signals of the peripheral nerve to and from electrical signals in instrumentation for stimulation and recording to produce fine control of advanced prosthetics. In order to potentiate the full spectrum of possible applications, the persistent foreign-body response needs to be addressed. Here we describe the cellular and extracellular components of chronically implanted polyimide threads suspended within a tricomponent hydrogel. The results of these experiments will contribute to design modifications for future fabrications of tissue-engineered-electronic-nerve-interface (TEENI) devices.

Design, fabrication, and characterization of a scalable tissue-engineered-electronic-nerve-interface (TEENI) device

In this study, we describe a novel peripheral-nerve interface which makes use of highly flexible multi-electrode arrays that are integrated into hydrogel-based scaffolds to form a hybrid tissue-engineered electronic construct. This tissue-engineered electronic nerve interface (TEENI) is designed to be scalable to high channel counts using multiple polyimide-based “threads” that are evenly distributed through a volume of the nerve equal to its diameter times the distance between one or more nodes of Ranvier. Such scalability could greatly increase the precision and resolution of motor-control and sensory-feedback signals exchanged between amputees and advanced upper-limb prosthetic devices.

Design, fabrication and characterization of miniature passive wireless force sensors

The majority of the population will have low back pain in their lifetime. Low back pain correlates to degenerative pain disease (DDD) which has been linked to exposure to mechanical loads. Certain occupations involving heavy lifting also correlate with DDD. Analytical models of the spine provide estimates of loading in the spine. However, data is lacking explicitly measuring mechanical forces in the spine. This data is necessary to truly characterize the translation from overload to onset of DDD. There is a need for an implantable device that is better able to characterize the in vivo mechanical environment present in the spine.

Behavior compensation reduces loads in the cervical spine measured with a force-sensing implant

Low back pain is growing health concern affecting 31 million Americans at any given time. Many of the risk factors for low back pain are mechanical in nature. Characterizing the axial loads, muscle recruitment, and behavior compensation during everyday motion an important step in minimizng the mechanical risk factors. The purpose of this study was to use an interbody implant to analyze the intradiscal forces in an in vivo model during everyday activities and to monitor the effect of behavior compensation on the resulting loads.