Sheep Gait Biomechanics and the Assessment of Musculoskeletal Conditions: A Systematic Review

Acknowledgments: This research work was supported by the Portuguese Foundation for Science and Technology (FCT) and Centro2020 through Project References UID/Multi/04044/2013; PAMI – ROTEIRO/0328/2013 (Nº 022158) and BioMaTE project - A novel bio-manufacturing system to produce bioactive scaffolds for tissue engineering reference PTDC/EMS-SIS/7032/2014.In recent years, sheep have been reported as the ideal animal model to study osteoporosis, hence it is important to identify instruments, tools and ideal parameters needed to assess the effects of different treatments. In previous studies conducted in other animal models with osteoporosis, the most common parameters used for evaluation concerned primarily bone properties, such as the dual X-ray absorptiometry. However, biomechanical gait analysis as an integrative functional parameter and a non-invasive method, will be an important tool in research and clinical applications. This research review was performed using the PubMed database and included studies related to sheep with outcome measures concerning functional performance assessed during gait in vivo; and excluded studies related with cardiovascular disease and sperm properties, which include other animal species, with outcomes not related with functional locomotor evaluation. Only studies related with bone properties were analyzed. The most frequent and relevant included parameters were the following: mean peak vertical ground force reaction, gait cycle and stance/swing phase duration, percentage of stance/swing phase in a gait cycle, stride length and the stifle joint angles during a gait cycle. Gait biomechanical parameters have been established for the assessment of some clinical orthopedic condition using sheep models but not currently for osteoporosis.info:eu-repo/semantics/publishedVersio

An Investigation into Noxious Mechanosensation, and the Role of Peripheral Neuron Subpopulations in Pain

This thesis uses transgenic mice to explore the role of candidate and known mechanotransducers in acute mechanical pain. It also utilises transgenics to ablate whole populations of sensory neurons in mice to establish what role these also have in pain, both under normal and inflammatory conditions. The water and ion channel Aquaporin 1 (Aqp1) is preferentially expressed in the small diameter neurons of the peripheral nervous system (PNS). These are responsible for nociception, and Aqp1 has previously been implicated in pain sensation. Its role in acute mechanical pain has not fully been explored. By using global Aqp1 knockout (Aqp1KO) mice and mechano-clamp electrophysiology I am the first to demonstrate that Aqp1 contributes to the mechanically activated (MA) currents associated with pain sensing in nociceptors. However, it does not produce MA currents when expressed in naïve cells. Aqp1 is necessary for normal mechanical pain in vivo as Aqp1KO animals have an increased mechanical pain threshold. Thus, it is unlikely that Aqp1 is a pore-forming component of a noxious mechanotransducer but may form part of a membrane complex essential to mechanical pain sensation. Piezo2 is a known mammalian mechanotransducer and is responsible for light touch sensation and proprioception. It’s contribution to mechanical pain under pathological conditions is established but it’s role in acute mechanical pain remains controversial. I generate mice with a nociceptor-specific Piezo2 deletion and again use a combination of electrophysiological and behavioural assays to demonstrate that Piezo2 is not required for acute noxious mechanosensation. Thus, my data confirms that the mechanotransducer responsible for mechanical pain remains ambiguous. Finally I study the role of the cutaneous population of Parvalbumin-positive (PV+) sensory neurons in pain. This population is required for innocuous mechanical sensation including vibration sensing. By genetically ablating PV+ neurons to generate ‘PVDTA’ mice, I provide evidence that these neurons are necessary for negatively regulating the thermal, mechanical, and inflammatory pain response, as behaving animals are hypersensitive to these insults. I am the first to propose that cutaneous PV+ neurons are responsible for closing the so-called ‘pain gate’ in the dorsal horn of the spinal cord. Further evidence for this comes from an in vivo electrophysiological study of dorsal horn neurons in PVDTA mice, which exhibit increased excitability as a consequence of noxious stimulation. In vivo DRG neuron imaging in animals expressing a reporter protein in PV+ sensory neurons show that these neurons are capable of responding to noxious stimuli, thus solidifying this hypothesis

Role of Bone Marrow Mesenchymal Stem Cells in Motor recovery of spinal cord injury in rats

INTRODUCTION: Spinal cord injury is a major catastrophy with no solutions for cure, and realistic expectations and possibilities for spinal cord repair are being acknowledged recently. The incidence of spinal cord injuries is approximately 30 per million. The incidence of spinal cord injuries is highest in the age group of 16 to 30 years. More than 80% of spinal cord injury occurs in males. Unfortunately most of the victims are young, in the prime of life, left with permanent paralysis of limbs, bladder, bowel and sexual functions. This results in far reaching social, vocational and psychological consequences. Prevention of complications, stabilisation of spine and active rehabilitation and long term follow up constitute the mainstay of the management of persons with spinal cord injury. Early rehabilitation in an organised multidisciplinary SCI team has been shown to be beneficial with lower mortality, decreased pressure sores, and slightly greater chance of neurological recovery and shorter lengths of stay in the hospital. Surgical stabilisation helps in early mobilisation, but there are no significant differences in neurological level changes between early surgery, late surgery and no surgery. There was no evidence that routine early surgical intervention or decompression improved neurological outcomes. The national spinal cord injury study NASCIS trials I, II, and III on early steroids to limit cellular damage from secondary injury processes suggested this to be part of the early management of SCI. The conclusions of this study have been challenged due to a variety of reasons. AIM OF THE STUDY: To study the effect of Bone marrow Mesenchymal Stem Cell transplantation for motor recovery in rat models of spinal cord injury. OBJECTIVES: 1. Bone marrow mesenchymal stem cell (MSC)culture and characterization. 2. Transdifferentiation of MSC to neuronal cells. 3. MSC transplantation to injured spinal cord segment of rat models. 4. Outcome assessment of transplantation by BBB scale, and EMG. MATERIALS & METHODS: All experiments were conducted with the approval from Institutional Review Board and the Animal Ethics Committee. Materials required: Cell culture facilities: Phosphate buffered saline (PBS), Gibco,Catalog no: 10010. RosetteSep antibody cocktail, Stem Cell Technologies Inc, catalog. No: 15168. Ficoll-Paque, GE Healthcare Bio-Sciences, catalog .no: 17-1440-03. T75 culture flask, Greiner. Alpha - Minimum essential medium, Gibco, Fetal bovine serum, Gibco, catalog.no: 10082. L-Glutamine, Gibco, Catalog.no: 25030. 0.25%Trypsin-EDTA, Gibco, Catalog.No: 25200. Dulbaco modified eagle medium, Gibco,catalog.No: 11995. DiMethylSulphoxide, DMSO Sigma, Catalog no: C6295 Butylated hydroxyanisole (BHA), ACROS. Potassium chloride (Kcl) , Qualigens. Valproic acid, sigma, catalog. No:P4543. Forskolin, Sigma. Hydrocortisone, Sigma, catalog.No: H6909. Insulin, Sigma, catalog.No: I9278. bFGF, Invitrogen, catalog.no: 13256-029. 30 Hanks Balanced salt solution (HBSS), Gibco, catalog no: 14175. 100U/mL penicillin, 100μg/ml streptomycin and 25ng/ml of amphotericin-B. Centrifuge, REMI. Laminar flow hood, Kartos International. CO2 incubator, Thermo Electron Corporation. Inverted phase-contrast microscope. Animal experiments: Albio Wistar rats, Animal house with air-conditioned facilities, Operation theatre for anaesthesia and surgery. Ketamine. Xylazine. Operating microscope. Surgical instruments: Bone rongeur. Surgical scissors. Equipment for drop weight device and transducers to record forces. Transcranial cortical stimulator. EMG recorder. Gait analysis set up for rats and video recorder for BBB score. 3 D Injection device for cell transplantation. CONCLUSIONS: Spinal cord injury results in permanent paralysis, loss of bowel and bladder control and other autonomic functions. Despite extensive research, there is no treatment available for this condition. This study demonstrated the role of bone marrow mesenchymal stem cells in spinal cord injury in rat models. 1. Bone MarrowMesenchymal Stem Cells can be cultured from the bone marrow sample and can be characterised to be used for transplantation. 2. Bone marrow Mesenchymal Stem Cells can be trans differentiated into neural cells in vitro using appropriate inducing media. However this observation needs further confirmation. 3. Drop weight model of spinal cord injury gives a consistent comparable contusion model for research studies. 4. Transcranial stimulation can be used to elicit motor evoked potential to demonstrate functional continuity of corticospinal tract in rat models of spinal cord injury. 5. Transplantation of bone marrow stromal stem cells can improve the motor out come in rat models of spinal cord injury as measured by BBB motor score and electrophysiological studies

DESIGN OF A GAIT ACQUISITION AND ANALYSIS SYSTEM FOR ASSESSING THE RECOVERY OF MICE POST-SPINAL CORD INJURY

Current methods of determining spinal cord recovery in mice, post-directed injury, are qualitative measures. This is due to the small size and quickness of mice. This thesis presents a design for a gait acquisition and analysis system able to capture the footfalls of a mouse, extract position and timing data, and report quantitative gait metrics to the operator. These metrics can then be used to evaluate the recovery of the mouse. This work presents the design evolution of the system, from initial sensor design concepts through prototyping and testing to the final implementation. The system utilizes a machine vision camera, a well-designed walkway enclosure, and image processing techniques to capture and analyze paw strikes. Quantitative results gained from live animal experiments are presented, and it is shown how the measurements can be used to determine healthy, injured, and recovered gait

Design, implementation and validation of an exoskeletal robot for locomotion studies in rodents

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Mechanical Engineering, 2012.Cataloged from PDF version of thesis.Includes bibliographical references (p. 214-226).Growing interest in robotic treatment of patients with neurological injury motivates the development of therapeutic robots for basic research into recovery. Though humans are the ultimate beneficiaries, basic research frequently involves rodent models of neurological injury, which motivates robotic devices that can interact with rats or mice. Currently, available apparatus for locomotion studies of rodents is built upon treadmills, which simplify the design and implementation but also restrict the scope of possible experiments. This is largely due to the treadmill's single-dimensional movement and the lack of accommodation for natural or voluntary movement of the animal. In order to open up new possibilities for locomotion studies in rodents, this work introduces newly developed apparatus for locomotion research in rodents. The key concept is to allow maximal freedom of voluntary movement of the animal while providing forceful interaction when necessary. Advantages and challenges of the proposed machine over other existing designs are discussed. Design and implementation issues are presented and discussed, emphasizing their impact on free, voluntary, movement of the animal. A live-animal experiment was conducted to verify the design principles. Unconstrained natural movement of the animal was compared with movement with the overground robot attached. The compact, overground design and backdrivable implementation of this robot allow novel experiments that involve open-space, free (or interactive) locomotion of the animal.by Yun Seong Song.Ph.D

Afferent information modulates spinal network activity in vitro and in preclinical animal models

Primary afferents are responsible for the transmission of peripheral sensory information to the spinal cord. Spinal circuits involved in sensory processing and in motor activity are directly modulated by incoming input conveyed by afferent fibres. Current neurorehabilitation exploits primary afferent information to induce plastic changes within lesioned spinal circuitries. Plasticity and neuromodulation promoted by activity-based interventions are suggested to support both the functional recovery of locomotion and pain relief in subjects with sensorimotor disorders. The present study was aimed at assessing spinal modifications mediated by afferent information. At the beginning of my PhD project, I adopted a simplified in vitro model of isolated spinal cord from the newborn rat. In this preparation, dorsal root (DR) fibres were repetitively activated by delivering trains of electrical stimuli. Responses of dorsal sensory-related and ventral motor-related circuits were assessed by extracellular recordings. I demonstrated that electrostimulation protocols able to activate the spinal CPG for locomotion, induced primary afferent hyperexcitability, as well. Thus, evidence of incoming signals in modulating spinal circuits was provided. Furthermore, a robust sensorimotor interplay was reported to take place within the spinal cord. I further investigated hyperexcitability conditions in a new in vivo model of peripheral neuropathic pain. Adult rats underwent a surgical procedure where the common peroneal nerve was crushed using a calibrated nerve clamp (modified spared nerve injury, mSNI). Thus, primary afferents of the common peroneal nerve were activated through the application of a noxious compression, which presumably elicited ectopic activity constitutively generated in the periphery. One week after surgery, animals were classified into two groups, with (mSNI+) and without (mSNI-) tactile hypersensitivity, based on behavioral tests assessing paw withdrawal threshold. Interestingly, the efficiency of the mSNI in inducing tactile hypersensitivity was halved with respect to the classical SNI model. Moreover, mSNI animals with tactile hypersensitivity (mSNI+) showed an extensive neuroinflammation within the dorsal horn, with activated microglia and astrocytes being significantly increased with respect to mSNI animals without tactile hypersensitivity (mSNI-) and to sham-operated animals. Lastly, RGS4 (regulator of G protein signaling 4) was reported to be enhanced in lumbar dorsal root ganglia (DRGs) and dorsal horn ipsilaterally to the lesion in mSNI+ animals. Thus, a new molecular marker was demonstrated to be involved in tactile hypersensitivity in our preclinical model of mSNI. Lastly, we developed a novel in vitro model of newborn rat, where hindlimbs were functionally connected to a partially dissected spinal cord and passively-driven by a robotic device (Bipedal Induced Kinetic Exercise, BIKE). I aimed at studying whether spinal activity was influenced by afferent signals evoked during passive cycling. I first demonstrated that BIKE could actually evoke an afferent feedback from the periphery. Then, I determined that spinal circuitries were differentially affected by training sessions of different duration. On one side, a short exercise session could not directly activate the locomotor CPG, but was able to transiently facilitate an electrically-induced locomotor-like activity. Moreover, no changes in reflex or spontaneous activity of dorsal and ventral networks were promoted by a short training. On the other side, a long BIKE session caused a loss in facilitation of spinal locomotor networks and a depression in the area of motor reflexes. Furthermore, activity in dorsal circuits was long-term enhanced, with a significant increase in both electrically-evoked and spontaneous antidromic discharges. Thus, the persistence of training-mediated effects was different, with spinal locomotor circuits being only transiently modulated, whereas dorsal activity being strongly and stably enhanced. Motoneurons were also affected by a prolonged training, showing a reduction in membrane resistance and an increase in the frequency of post-synaptic currents (PSCs), with both fast- and slow-decaying synaptic inputs being augmented. Changes in synaptic transmission onto the motoneuron were suggested to be responsible for network effects mediated by passive training. In conclusion, I demonstrated that afferent information might induce changes within the spinal cord, involving both neuronal and glial cells. In particular, spinal networks are affected by incoming peripheral signals, which mediate synaptic, cellular and molecular modifications. Moreover, a strong interplay between dorsal and ventral spinal circuits was also reported. A full comprehension of basic mechanisms underlying sensory-mediated spinal plasticity and bidirectional interactions between functionally different spinal networks might lead to the development of neurorehabilitation strategies which simultaneously promote locomotor recovery and pain relief

Recent and advanced animal models used in the Screening of analgesics and anti-inflammatory activity

Non-Steroidal anti-inflammatory drugs (NSAIDs) are consisting of three major anti-pyretic, anti-inflammatory and anti-analgesics properties. They have reduced the sensation of pain, body temperature, and inflammation. It is also used for the treatment of the long-term health problems like arthritis (rheumatoid arthritis, osteoarthritis, and lupus). NSAIDs highly protect the lining of the stomach and intestines from the damaging effects of acid promote blood clotting by activating blood platelets, and promote normal function of the kidney. Incompatible with the action of NSAIDs many different types of drugs and plant use for the treatment of the analgesic, inflammation and pyretic activity. Diclofenac inhibit the cyclooxygenase (COX-2) enzyme with the greater potency that it (COX-1). NSAIDs are generally used in the management of pain because of the integrated role of the COX pathway that is recognition of pyretic, inflammation and analgesic. Introduction to painful procedures and/or stressors during the early neonatal period can reprogram the underlying neurocircuitry involved in nociception and neuropathic pain perception. The reprogramming of these systems can result in an enduring elevation in sympathy towards mechanical and thermal stimuli. During adolescence, hind paw mechanical removal thresholds were evaluated using an electronic von Frey Anesthesiometer. Animals challenged neonatally with LPS (nLPS) had increased pain sensitivity on this measure which was related with decreased Oprm1 expression in the prefrontal cortex (PFC) and periaqueductal gray (PAG) of both male and female rats. There was no effect of inflammatory treatment on either anxiety or depressive-like behavior suggesting that affective functioning did not account for differences in mechanical pain sensitivity

Development Of Carbon Based Neural Interface For Neural Stimulation/recording And Neurotransmitter Detection

Electrical stimulation and recording of neural cells have been widely used in basic neuroscience studies, neural prostheses, and clinical therapies. Stable neural interfaces that effectively communicate with the nervous system via electrodes are of great significance. Recently, flexible neural interfaces that combine carbon nanotubes (CNTs) and soft polymer substrates have generated tremendous interests. CNT based microelectrode arrays (MEAs) have shown enhanced electrochemical properties compared to commonly used electrode materials such as tungsten, platinum or titanium nitride. On the other hand, the soft polymer substrate can overcome the mechanical mismatch between the traditional rigid electrodes (or silicon shank) and the soft tissues for chronic use. However, most fabrication techniques suffer from low CNT yield, bad adhesion, and limited controllability. In addition, the electrodes were covered by randomly distributed CNTs in most cases. In this study, a novel fabrication method combining XeF2 etching and parylene deposition was presented to integrate the high quality vertical CNTs grown at high temperature with the heat sensitive parylene substrate in a highly controllable manner. Lower stimulation threshold voltage and higher signal to noise ratio have been demonstrated using vertical CNTs bundles compared to a Pt electrode and other randomly distributed CNT films. Adhesion has also been greatly improved. The work has also been extended to develop cuff shaped electrode for peripheral nerve stimulation. Fast scan cyclic voltammetry is an electrochemical detection technique suitable for in-vivo neurotransmitter detection because of the miniaturization, fast time response, good sensitivity and selectivity. Traditional single carbon fiber microelectrode has been limited to single detection for in-vivo application. Alternatively, pyrolyzed photoresist film (PPF) is a good candidate for this application as they are readily compatible with the microfabrication process for precise fabrication of microelectrode arrays. By the oxygen plasma treatment of photoresist prior to pyrolysis, we obtained carbon fiber arrays. Good sensitivity in dopamine detection by this carbon fiber arrays and improved adhesion have been demonstrated