No description available.

No description available.

Light and electron microscopical techniques were employed to make qualitative and quantitative observations on peripheral nerve morphology. Sural nerve biopsies from 4 groups of diabetic patients with different syndromes of polyneuropathy and 6 non-diabetic controls were evaluated. Observations indicated that unequal rates of successful fibre regeneration may underlie an apparent difference in myelinated fibre (MF) loss between painful and painless diaebetic polyneuropathy. MF and unmyelinated fibre (UF) degeneration/regeneration per se are unlikely to be the cause of neuropathic pain in diabetic polyneuropathy since evidence of each was present in patients with remission from painful symptoms and in patients with painless neuropathy. Axonal atrophy may have a role in neuropathic pain generation. Abnormal endoneurial capillary morphology was manifest in all the diabetic patients. Alterations in capillary basement membrane thickness were interdependent with the type and extent of neuropathological change and the clinical expression of the neuropathy. Two patients presented with intractible vomiting due to diabetic gastropathy. The findings of a study of their anterior and posterior abdominal vagus nerves, obtained as biopsy specimens at vagotomy and gastroenterostomy to prevent stomal ulceration, were compared with those of 2 diabetic and 2 non-diabetic patients undergoing vagotomy for duodenal ulceration. Autovagotomy, and its causative role in gastropathy, was not supported since there was persistence of intact and regenerating fibres and evidence of similar MF and UF changes in both groups of diabetic patients. Male Sprague-Dawley rats were employed to make a comparison of the effects of conventional insulin therapy (CIT) and continuous subcutaneous insulin infusion therapy (CSII) in streptozotocin-induced diabetes. CSII was superior to CIT in diurnal and circadian blood glucose homeostasis and in ameliorating a diabetes-related deficit in body weight and skeletal length. Although CSII performed better, neither form of therapy was completely successful in reversing the effects of untreated-diabetes on parameters indexing MF morphology in the tibial nerve. Potentially serious degenerative changes in MFs following the initiation of either form of insulin therapy were found to correlate with the number of detected hypoglycaemic episodes. Normal maturation of MFs in the sural nerve involved the formation of a proximo-distal taper for fibre and axon size. MFs and axons in diabetic rats did not assume a size taper. The effect of diabetes was manifest proximally in the nerve with no evidence of selective involvement of axon or Schwann cell. Male albino Lewis rats were made diabetic with streptozotocin and used to study the effect of early (within 3 weeks= EIT) and delayed (after 6 months= DIT) pancreatic islet transplantation on MF morphology. Optimal diurnal and circadian blood glucose control was achieved for 6 months following EIT and DIT. A body weight deficit in diabetic rats was ameliorated by EIT but not DIT. EIT completely prevented the occurrence of abnormal fibre and axon morphology in the tibial nerves of diabetic rats. DIT completely reversed a deficit in fibre size but not axon size. If these observations were applicable to man the best therapeutic option would be for candidates to receive islet transplantation soon after the onset of diabetes. The occurrence of neuropathy might then be prevented.

abstract: The use of a non-invasive form of energy to modulate neural structures has gained wide spread attention because of its ability to remotely control neural excitation. This study investigates the ability of focused high frequency ultrasound to modulate the excitability the peripheral nerve of an amphibian. A 5MHz ultrasound transducer is used for the study with the pulse characteristics of 57msec long train burst and duty cycle of 8% followed by an interrogative electrical stimulus varying from 30μsecs to 2msecs in pulse duration. The nerve excitability is determined by the compound action potential (CAP) amplitude evoked by a constant electrical stimulus. We observe that ultrasound's immediate effect on axons is to reduce the electrically evoked CAP amplitude and thereby suppressive in effect. However, a subsequent time delayed increased excitability was observed as reflected in the CAP amplitude of the nerve several tens of milliseconds later. This subsequent change from ultrasound induced nerve inhibition to increased excitability as a function of delay from ultrasound pulse application is unexpected and not predicted by typical nerve ion channel kinetic models. The recruitment curve of the sciatic nerve modified by ultrasound suggests the possibility of a fiber specific response where the ultrasound inhibits the faster fibers more than the slower ones. Also, changes in the shape of the CAP waveform when the nerve is under the inhibitive effect of ultrasound was observed. It is postulated that these effects can be a result of activation of stretch activation channels, mechanical sensitivity of the nerve to acoustic radiation pressure and modulation of ion channels by ultrasound. The neuromodulatory capabilities of ultrasound in tandem with electrical stimulation has a significant potential for development of neural interfaces to peripheral nerve. / Dissertation/Thesis / Masters Thesis Bioengineering 2016

Unlike the central nervous system, axonal regeneration does occur in the peripheral nervous system, however, despite this, functional recovery from nerve transection injury remains dismal. This has been attributed to factors intrinsic to the motor or sensory cell body and to elements in the local site of injury including nerve gaps, scar and a limited time frame in which supportive growth factors and extracellular matrix molecules are expressed. The aim of this thesis is to review the mechanisms behind axonal damage and regeneration in the peripheral nervous system and investigate surgical, pharmacological and biological approaches to overcome limitations in regeneration and functional recovery. By taking a broad approach to the topic, I hoped to gain a greater understanding of the inhibitory and regenerative processes at play and provide a contribution to the understanding in the field of peripheral nerve surgery. / Experimental Surgery

No description available.

Peripheral nerve injuries can cause serious health problems and result in lifelong disabilities. Although researchers have been studying peripheral nerve injuries, patients may not regain complete function of their muscles even after surgeries to repair their nerves are performed. However, animal studies have shown that after peripheral nerve cut and repair (muscle self-reinnervation), stretch-reflex in the affected muscles does not recover, which may affect the muscle electromyographic (EMG) activity of all muscle synergists, as well as joint kinematics. The aim of this study is to determine the effects of the self-reinnervation of the lateral gastrocnemius (LG) and soleus (SO) muscles in the hind limb of felines on the mean EMG activity of the intact synergist medial gastrocnemius (MG), as well as the moments at the knee and ankle joints during different walking conditions: level (0%), downslope (-50%), and upslope (+50%). The EMG activity and joint kinematics were recorded on the three different walking conditions before and 12 weeks after the self-reinnervation of LG and SO when these muscles recovered their activity. The self-reinnervation of the two muscles caused the MG EMG activity to increase for all walking conditions. However, the changes in the knee and ankle moments differed depending on the three different walking conditions. It was concluded that the changes in EMG and joint moments after self-reinnervation could be caused by the absence of stretch-reflex in the affected muscles and/or changes in physiological properties of muscles.

The present study has developed poly ε-caprolactone (PCL)/ poly lactic acid (PLA) films with specific internal structure suitable to prepare nerve guide conduit for peripheral nerve repair. The film preparation method has been carried out using an environmental chamber to prepare the solvent cast films with the specific surface structure. Different cellular behaviour of neuronal cell cultures was seen on the pitted films with different pits configurations (size and distribution). The consistent surface morphology provided a reliable surface structure for further in vitro and in vivo studies. The effect of a medical grade sterilisation process using gamma radiation at eight doses (0-45kGy) on PCL/PLA films was explored. It has been shown that material properties, including mechanical strength, were significantly affected, while cellular behaviour and responses (NG108-15) were improved. Grooved films with three groove shapes (Sloped, Square, and V shape) were prepared using patterned silicon substrates, photolithography and wet/dry etching. The groove patterns were successfully transferred and good mechanical strength was observed for grooved PCL/ PLA. Oriented growth of NG108-15 cells was observed on the patterned films with an improved alignment and organisation on SL and V shape grooved films. UV-ozone treatment was used to increase hydrophilicity of PCL/PLA films to improve Schwann cells behaviour. No negative effect was observed on cell growth and proliferation on the treated films however the mechanical properties were reduced. Schwann cells expressed typical long spindle-shape morphology with cell-to-cell interaction in longitudinal direction on the treated grooved films. Consistent to in vitro experiment with NG108-15, Schwann cells alignment was also improved on SL and V shape grooves. A three-week in vivo study was carried out to test grooved and non-grooved conduits in a rat sciatic nerve model. The grooved conduits showed better regeneration, with SL-grooved film showing a significant improvement of nerve regeneration. A separate in vivo study evaluated the effect of wall-thickness on nerve regeneration. However, it was shown that the wall thickness had no positive effect, and the conduit with improved mechanical strength adversely affected the nerve regeneration. In conclusion, a nerve guide conduit was developed with the optimised surface structure to support nerve regeneration. The promising in vitro and in vivo studies together with the suitable biomechanical properties and specific surface structure and morphology indicate that the grooved PCL/PLA conduit is a viable treatment for peripheral nerve repair.

Made available in DSpace on 2014-06-11T19:22:13Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-06-22Bitstream added on 2014-06-13T19:07:27Z : No. of bitstreams: 1 franco_rg_me_botfm.pdf: 2218416 bytes, checksum: e0192057697a7c16907ca66d4f8f696a (MD5) / A neurorrafia término-lateral é técnica consagrada em casos onde não há coto proximal após lesão de nervo periférico. O coto distal pode ser suturado na lateral de qualquer nervo ou receber enxerto de nervo unindo a sua face lateral à face lateral de outro nervo, atuando como uma ponte. O número de enxertos em ponte poderia aumentar o resultado da reinervação muscular? Para responder esta pergunta realizou-se este trabalho. Foram operados 100 ratos Wistar divididos em cinco grupos experimentais com 20 animais cada. O GN (grupo controle de normalidade) não sofreu nenhum procedimento. No GD (grupo controle de desnervação) o nervo fibular foi seccionado e ambos os cotos foram suturados em músculos adjacentes e distantes entre si. O G1 recebeu um enxerto (ponte de nervo sural) tendo uma de suas extremidades suturadas na face lateral do nervo tibial (NT) e a outra na face lateral do segmento distal do nervo fibular (NF). No G2 e no G3 foram realizados os mesmos procedimentos que no G1. O G3 com 3 e o G2 com 2 enxertos. Em todos os casos foram utilizadas neurorrafias término-laterais (NTL) tipo “embracing” sem retirada de janela de epineuro. Os animais foram sacrificados após 120 dias. Foram realizados testes de marcha (“Walking-tracks”) e testes eletrofisiológicos além de aferição da massa dos animais e dos músculos tibiais craniais direitos (MTCD). Segmentos de nervo de interesse também foram coletados. Foram realizadas análises histomorfométricas das fibras musculares e nervosas. Após análise global dos atributos do MTCD como massa, IMM (índice de massa muscular), área, menor e maior diâmetro das fibras musculares, número de fibras por campo, análise funcional (IFC –índice funcional do ciático), análise eletrofisiológica e análise morfométrica dos segmentos nervosos mais importantes, concluímos que não houve benefício com o aumento do número de pontes / The end-to-side neurorrhaphy technique is enshrined in cases where there is no proximal stump after peripheral nerve injury. The distal end can be sutured to the side of any nerve. We also can use a nerve graft joining the lateral face of the distal end to the lateral face of another nerve, serving as a bridge. The number of bridges would increase the outcome of muscle reinnervation? To answer this question took place this work. Were operated on 100 rats divided into five groups with 20 animals each. GN (normal control group) received no further procedure. For GD (control group denervation) the peroneal nerve was transected and both stumps were sutured to adjacent muscles and far between. G1 received one graft (sural nerve bridge) having one end stitched on the side of the tibial nerve (NT) and the other on the side of the distal segment of the peroneal nerve (NF). In G2 and G3 were performed the same procedures as in G1. G3 with three and G2 with two grafts. In all cases were used end-to-side neurorraphies (NTL) performed as embracing without removing epineural window. The animals were sacrificed after 120 days. We performed tests of gait (Walking-tracks) and electrophysiological tests in addition to measuring the mass of animals and also the mass of cranial tibial muscles (MTCD). Nerve segments of interest were also collected. We performed histomorphometric analysis of muscle fibers and nerves. After comprehensive analysis of attributes such as mass MTCD, IMM (body mass index), area, smaller and larger diameter of muscle fibers, number of fibers per field, functional analysis (IFC, sciatic functional index), electrophysiological and morphometric analysis of the most important nerve segments, we conclude that there was no benefit to increasing the number of bridges

The mitofusin 1 and 2 (MFN and MFN2) proteins reside in the outer mitochondrial membrane and have been shown to regulate mitochondrial network architecture by mediating tethering and fusion of mitochondria. Mitochondria normally form a tubular and branched reticular network dynamically regulated by a balance of fusion and fission events. Absence of either Mfn1 or Mfn2 results in a fragmented mitochondrial network. Züchner et al. previously described mutations in the gene mitofusin 2 (MFN2) as the cause of the major autosomal-dominant, axonal form of Charcot-Marie-Tooth neuropathy (CMT2A). CMT type 2 (CMT2) is characterized by chronic axonal degeneration of peripheral nerves leading to the loss of functional nerve fibers. Mutations in MFN2 are the most common cause of CMT2, and in Chapter 2 we report the results from a genetic screen of MFN2 in a CMT2 patient cohort. The original finding that mutations in MFN2 cause CMT2A led to investigations focused on deficiencies of mitochondrial fusion and transport, specifically in the context of long axonal processes affected in CMT. While some experimental work supports disrupted mitochondrial transport in the etiology of CMT2A, other studies on CMT2A patient fibroblasts and cell models suggest abnormal mitochondrial fusion and dynamics do not underlie the etiology of this. In the first half of Chapter 3, we present some of our initial investigations prior to de Brito and Scorrano’s report published in 2008 regarding a novel role for Mfn2 in tethering the endoplasmic reticulum (ER) to mitochondria. In Mfn2 null mouse embryonic fibroblasts (MEFs) regions of contact between mitochondria and the endoplasmic reticulum (ER) are significantly reduced. These regions of contact are thought to form specialized subdomains of the ER, called mitochondrial associated membranes (MAM). Besides observing a fragmented ER network in Mfn2 knockout (KO) mouse embryonic (MEF) cells, de Brito and Scorrano presented several lines of evidence which suggest that the underlying pathogenic mechanism in CMT2A stems from disrupted ER-mitochondria. As this observation had not been replicated in the literature, we describe our attempts to replicate these finding in the last half of Chapter 3. The MAM represents a sub-domain of the ER in close association with the mitochondrial outer membrane. The movement of phosphatidylserine (PS) from the MAM domains of the ER to mitochondria and its subsequent decarboxylation to phosphatidylethanolamine (PE) by the enzyme PS decarboxylase (Pisd) has been well characterized and is known to depend on the existence of an outer mitochondrial membrane protein. As PE has curvature inducing and fusogenic biophysical characteristics, a deficiency in PE would be an attractive mechanism contributing to the morphological and fusion defects observed in Mfn2 null cell models. We hypothesized that loss of Mfn2 would lead to specific decreases in mitochondrial and cellular levels of PE. Chapter 4 describes experiments designed to test this hypothesis. We observed significantly lower levels of PE in Mfn2 null cells, yet observe similar changes in Mfn1 null cells. Likewise, other lipid species such as ether linked PE (ePE) are decreased. To investigate how CMT2A mutations in MFN2 influence cellular phospholipid profiles, we then profiled cellular phospholipids of CMT2A patients and control lymphoblasts. We hypothesized that mutations in MFN2 would result in decreased levels of PE. In Chapter 5, we report the results of a phospholipid screen which reveal changes in ePE in CMT2A patient lymphoblasts, without the drastic decreases in PE previously observed in Mfn2 null lines. In conclusion, our data indicates an important role for both mitofusins in the mitochondrial synthesis of PE. In the context of CMT2A mutations, ePE levels are specifically reduced. Future studies may reveal how deficiencies in ePE might have important functional consequences in the pathogenesis of CMT2A.