A Computational Approach for the Design of Epidural Electrical Spinal Cord Stimulation Strategies to Enable Locomotion after Spinal Cord Injury

Spinal cord injury (SCI) is a major cause of paralysis with currently no effective treatment. Epidural electrical stimulation (EES) of the lumbar spinal cord has been shown to restore locomotion in animal models of SCI, but has not yet reached the same level of efficacy in humans. The mechanisms through which EES promotes locomotion, and the causes underlying these inter-species differences remain largely unknown, although essential to fully exploit the therapeutic potential of this neuromodulation strategy. Here, we addressed these questions using a deductive approach based on computer simulations and hypothesis-driven experiments, and proposed complementary strategies to enhance the current efficacy of EES-based therapies. In the first part of this thesis, we studied the mechanisms through which EES enables locomotion in rat models of SCI. Performing simulations and behavioral experiments, we provided evidence that EES modulates proprioceptive afferents activity, without interfering with the ongoing sensory signals. We showed that this synergistic interaction allows muscle spindle feedback circuits to steer the unspecific excitation delivered by EES to functionally relevant pathways, thus allowing the formation of locomotor patterns. By leveraging this understanding, we developed a stimulation strategy that allowed adjusting lesion-specific gait deficits, hence increasing the therapeutic efficacy of EES. In the second part of this thesis, we evaluated the influence of trunk posture on proprioceptive feedback circuits during locomotion, and thus on the effect of EES, in rat models of SCI. By combining modeling and experiments, we showed that trunk orientation regulates leg proprioceptive signals, as well as the motor patterns produced during EES-induced stepping. We exploited these results to develop a control policy that by automatically regulating trunk orientation significantly enhanced locomotor performance. In the last part of this thesis, we investigated the causes underlying species-specific effects of EES. Hypothesis-driven simulations suggested that in humans continuous EES blocks the proprioceptive signals traveling along the recruited fibers. We corroborated this prediction by performing experiments in rats and people with SCI. In particular, we showed that EES disrupts the conscious perception of leg movements, as well as the afferent modulation of sensorimotor circuits in humans, but not in rats. We provide evidence that in humans, due to this phenomenon, continuous EES can only facilitate locomotion to a limited extent. This was insufficient to provide clinically relevant improvements in the tested participants. Finally, we proposed two sensory-compliant stimulation strategies that might overcome these limitations, and thus augment the therapeutic efficacy of EES. In this thesis we elucidated key mechanisms through which EES promotes locomotion, we exposed critical limitations of continuous EES strategies when applied to humans, and we introduced complementary strategies to maximize the efficacy of EES therapies. These findings have far-reaching implications in the development of future strategies and technologies supporting the recovery of locomotion in people with SCI using EES

Evaluating Nutraceuticals for Selective Toxicity Toward Leukemia Stem Cells

Targeting leukemia stem cells (LSCs) is critical to improving the poor outcome of acute myeloid leukemia (AML) patients. Nutraceuticals (i.e., food derived bioactive compounds) provide a wealthy resource for novel anti-cancer, and specifically anti-AML drug discovery. With the advent of novel LSC cell lines, preliminary screening of these compounds against LSC-like cells can be achieved rapidly. To identify potential novel anti-LSC therapeutics, we created and screened a unique library consisting of 288 nutraceuticals in an MTS assay against TEX leukemia cells, a surrogate LSC line and K562, a control cell line which does not possess LSC activity. Here, we identified diosmetin, a flavonoid found in citrus fruits and various green plants, as a novel anti- LSC agent (EC50: 6.0 ± 1.7μM). To confirm its activity, diosmetin (10μM) reduced clonogenic growth of primary AML cells (n = 4) with no effect on normal CD34 positive bone marrow derived stem cells (n = 3) observed in colony forming cell assays. A dose-response and time course analysis performed via the Annexin/PI assay and flow cytometry revealed that diosmetin induced apoptosis, as evidenced by the accumulation of ANN+/PI- cells. Apoptosis was further confirmed by a subG1 peak after performing cell cycle analysis. Utilizing the Database for Annotation, Visualization and Integrated Discovery (DAVID) tool, we determined that the estrogen receptor (ER) was a potential molecular target for diosmetin’s anti-leukemia activity. To assess the role of estrogen receptors, we measured ERα and ERβ protein levels in diosmetin sensitive and insensitive cell lines. Interestingly, diosmetin sensitive cell lines display significantly elevated ERβ protein levels compared to diosmetin insensitive cells. However, this pattern was not observed for ERα. Similar results were observed through quantitative PCR measures, as TEX cells displayed levels of ESR2 (ERβ) mRNA, with no observed levels of ESR1 (ERα) mRNA levels. The opposite results were observed in K562 cells. Through ER reporter assays, it was demonstrated that diosmetin acts as a partial agonist in ERβ reporter cells, increasing luciferase activity with increasing doses of diosmetin in ERβ reporter cells. Moreover, we find that caspase 8 but not caspase 9 is elevated following diosmetin treatment, consistent with the extrinsic pathway of apoptosis and our observed increased in TNF-α, similar to previous reports highlighting the link between ERβ agonists and cancer cell death. In summary, these studies highlight that estrogen receptors, specifically ERβ, is a novel LSC therapeutic target, and the potential role of nutraceuticals as promising compounds for future drug discovery endeavours

Electrical spinal cord stimulation must preserve proprioception to enable locomotion in humans with spinal cord injury

Epidural electrical stimulation (EES) of the spinal cord restores locomotion in animal models of spinal cord injury but is less effective in humans. Here we hypothesized that this interspecies discrepancy is due to interference between EES and proprioceptive information in humans. Computational simulations and preclinical and clinical experiments reveal that EES blocks a significant amount of proprioceptive input in humans, but not in rats. This transient deafferentation prevents modulation of reciprocal inhibitory networks involved in locomotion and reduces or abolishes the conscious perception of leg position. Consequently, continuous EES can only facilitate locomotion within a narrow range of stimulation parameters and is unable to provide meaningful locomotor improvements in humans without rehabilitation. Simulations showed that burst stimulation and spatiotemporal stimulation profiles mitigate the cancellation of proprioceptive information, enabling robust control over motor neuron activity. This demonstrates the importance of stimulation protocols that preserve proprioceptive information to facilitate walking with EES

A biomimetic electrical stimulation strategy to induce asynchronous stochastic neural activity

Objective.Electrical stimulation is an effective method for artificially modulating the activity of the nervous system. However, current stimulation paradigms fail to reproduce the stochastic and asynchronous properties of natural neural activity. Here, we introduce a novel biomimetic stimulation (BioS) strategy that overcomes these limitations.Approach.We hypothesized that high-frequency amplitude-modulated bursts of stimulation could induce asynchronous neural firings by distributing recruitment over the duration of a burst, without sacrificing the ability to precisely control neural activity. We tested this hypothesis using computer simulations and ex vivo experiments.Main results.We found that BioS bursts induce asynchronous, stochastic, yet controllable, neural activity. We established that varying the amplitude, duration, and repetition frequency of a BioS burst enables graded modulation of the number of recruited fibers, their firing rate, and the synchronicity of their responses.Significance.These results demonstrate an unprecedented level of control over artificially induced neural activity, enabling the design of next-generation BioS paradigms with potentially profound consequences for the field of neurostimulation

Mechanisms Underlying the Neuromodulation of Spinal Circuits for Correcting Gait and Balance Deficits after Spinal Cord Injury

Epidural electrical stimulation of lumbar segments facilitates standing and walking in animal models and humans with spinal cord injury. However, the mechanisms through which this neuromodulation therapy engages spinal circuits remain enigmatic. Using computer simulations and behavioral experiments, we provide evidence that epidural electrical stimulation interacts with muscle spindle feedback circuits to modulate muscle activity during locomotion. Hypothesis-driven strategies emerging from simulations steered the design of stimulation protocols that adjust bilateral hindlimb kinematics throughout gait execution. These stimulation strategies corrected subject-specific gait and balance deficits in rats with incomplete and complete spinal cord injury. The conservation of muscle spindle feedback circuits across mammals suggests that the same mechanisms may facilitate motor control in humans. These results provide a conceptual framework to improve stimulation protocols for clinical applications. Computer simulations validated experimentally revealed that epidural electrical stimulation of lumbar segments facilitates motor control through the modulation of muscle spindle feedback circuits. Simulations established a clinical framework to design stimulation protocols correcting gait symmetry and balance deficits after injury

Shared human–robot proportional control of a dexterous myoelectric prosthesis

Myoelectric prostheses allow users to recover lost functionality by controlling a robotic device with their remaining muscle activity. Such commercial devices can give users a high level of autonomy, but still do not approach the dexterity of the intact human hand. Here we present a method to control a robotic hand, shared between user intention and robotic automation. The algorithm allows user-controlled movements when high dexterity is desired, but also assisted grasping when robustness is paramount. This combination of features is currently lacking in commercial prostheses and can greatly improve prosthesis usability. First, we design and test a myoelectric proportional controller that can predict multiple joint angles simultaneously and with high accuracy. We then implement online control with both able-bodied and amputee subjects. Finally, we present a shared control scheme in which robotic automation aids in object grasping by maximizing the contact area between the hand and the object, greatly increasing grasp success and object hold times in both a virtual and a physical environment. Our results present a viable method of prosthesis control implemented in real time, for reliable articulation of multiple simultaneous degrees of freedom