Neurofibromatosis type 2 is an autosomal dominant disease characterized by the formation of schwannomas and other peripheral neuropathies. The nf2 gene encodes the protein Schwannomin, or merlin. Schwannomin (Sch) is a membrane-cytoskeletal linking protein that suppresses cell proliferation at high cell density and modulates cell shape. Sch's tumor suppressive activity is regulated by its localization, conformation, and phosphorylation at serine 518 (S518). Sch's localization is dependent on binding the scaffold protein, paxillin. Phosphorylation of Sch at S518 regulates its conformation and tumor suppressor function. In a negative feedback loop, unphosphorylated Sch restricts cell proliferation downstream of Rac and p21-activated kinase (Pak), whereas Pak-induced phosphorylation inactivates Sch's ability to inhibit Pak and cell proliferation. Little is known about the function of the phosphorylated form of Sch, or the molecular mechanisms leading to its phosphorylation. Here we demonstrate that Sch-S518 phosphorylation is dependent on paxillin-binding and plasma membrane localization in SCs. Phosphorylation of Sch at the plasma membrane is mediated by Cdc42-Pak and results in altered SC morphology and polarity. Moreover, we have identified two extracellular stimuli that trigger Sch-S518 phosphorylation; these are neuregulin (NRG) and laminin, two potent activators of SC proliferation and myelination. NRG promotes Sch-S518 phosphorylation downstream of ErbB2/ErbB3 through PKA, whereas laminin-1 stimulation of β1 integrin promotes Pak- dependent phosphorylation of Sch-S518. Additionally, we find that Sch promotes process formation and elongation in primary and myelinating SCs, independent of Sch S518 phosphorylation. However, Sch phosphorylation was found to influence SC differentiation, as expression of an unphosphorylatable variant, Sch-S518A, facilitated SC myelination, whereas expression of a phospho-mimicking variant, Sch-S518D, reduced the SC's ability to myelinate. Together, these findings have identified receptor-mediated and paxillin-dependent pathways that regulate phosphorylation and inactivation of Sch's tumor suppressor function. Additionally, these results have elucidated novel normal functions for Sch during peripheral nerve development and myelination, and identify novel therapeutic targets for treatment of NF2 and other peripheral neuropathies.

No description available.

No description available.

A estimulação elétrica cerebral é uma alternativa terapêutica para pacientes com dor neuropática refratária à terapia convencional sendo o córtex insular (CI) um alvo promissor para seu tratamento. Este trabalho investigou o efeito da estimulação elétrica do córtex insular (EECI) sobre a sensibilidade dolorosa de ratos submetidos à constrição crônica do nervo isquiático. Ratos foram avaliados nos testes de pressão da pata e por filamentos de von Frey após 15 min de EECI (60 Hz, µs, 1 V). A EECI induziu antinocicepção na pata contralateral ao hemisfério estimulado, sem alterar a resposta da pata ipsilateral. Naloxona e rimonabanto reverteram a antinocicepção induzida pela EECI. Ioimbina e metisergida não interferiram neste efeito, demonstrando participação dos sistemas opióide e canabinóide, mas não adrenérgico e serotonérgico. Este estudo propõe um novo modelo experimental de estimulação cerebral profunda para o tratamento de dor neuropática e caracteriza o CI como um alvo promissor no estudo de novas terapêuticas para controlar dor crônica refratária. / Electrical brain stimulation is a therapeutic alternative for neuropathic pain patients, refractory to conventional treatments and the insular cortex (IC) is a promising target for their treatment. Herein the effects of electrical stimulation of the insular cortex (ICS) on pain sensitivity of rats with chronic constriction injury (CCI) of the sciatic nerve were investigated. Paw pressure test and von Frey filaments evaluated mechanical pain sensitivity at the end of a 15 min session of ICS (60 Hz, 210 µs, 1 V). ICS induced antinociception at the paw contralateral to the stimulated hemisphere, without changing ipsilateral paw threshold. Both naloxone and rimonabanto, but not yohimbine and methysergide reversed ICS-induced antinociception, demonstrating the involvement of opioid and cannabinoid systems, but not adrenergic and serotonergic. This study proposes a new experimental model of deep brain stimulation for the treatment of neuropathic pain and characterizes IC as a promising target to the study of new therapies to control refractory chronic pain.

Background: Chemotherapy Induced Peripheral Neuropathy (CIPN) is a debilitating neuropathy caused by commonly used chemotherapeutics. Clinically, the problem of CIPN is compounded by difficulties with diagnosis and limited treatment options. The pathophysiology of CIPN remains elusive, with current mechanistic postulates focused mainly on the peripheral nervous system. However, animal and human models of non-CIPN neuropathic conditions have shown the brain to be central to the development and maintenance of painful neuropathy. Moreover, evidence suggests that aberrant activity in key regions of the brain and brainstem could denote individual vulnerability for chronic pain states. The impact of the brain on CIPN development is unknown. Assessment of drug efficacy using brain imaging can provide sensitive readouts and is increasingly used in clinical trials. Aims: Firstly, to prospectively explore the structure and function of the brain in cancer patients prior to chemotherapy administration, using functional magnetic resonance imaging (fMRI), in order to determine whether baseline differences exist between patients who progress to CIPN as compared to those who do not. Secondly, to develop a pilot study using fMRI to investigate a topical treatment for CIPN, in order to assess the feasibility of setting up a study with this kind of design. Methods: To address the first aim of this thesis a prospective cohort study (the CIPN fMRI Study) was developed. Cancer patients scheduled to receive neurotoxic chemotherapy treatment including oxaliplatin, carboplatin, carbotaxol, or cisplatin, were recruited from three NHS trusts in Scotland, to undergo a high resolution (3 tesla) functional MRI scan, at a single time point prior to commencement of chemotherapy. During the scan structural, resting state and functional data were collected. Functional data involved the presentation of punctate stimuli (using a 256mN von Frey filament), above the patients’ right medial malleolus. While receiving the punctate stimuli, patients viewed images that had neutral or positive emotional content or a baseline coloured image with no content. Sample size was based on previously successful pain fMRI studies and pragmatic estimates. Acute CIPN was defined clinically by common toxicity criteria as necessitating a chemotherapy dose reduction or cessation. Data were analysed using FMRIB’s Software Library (FSL) version 5, 2015. Standard data pre-processing (brain extraction, registration, B0 unwarping, motion correction, and denosiing with FIX) was carried out. Structural analysis was conducted using FIRST. Resting state analysis utilised FSL’s MELODIC tool, and a non-parametric group comparison was made following a dual regression approach. FEAT was used for both first and second level functional analyses. Group comparisons were made using a mixed effects analysis (z threshold 2·3 and 2, regions considered significant at p < 0·05, cluster corrected). The group was split by sex to explore known sex differences in pain processing. To address the second aim of this thesis, a pilot fMRI randomised controlled trial (MINT3 Study) was designed. Approvals from ethics and research and development were sought and obtained. Data collection forms were developed. An fMRI experiment was proposed and a single pilot scan was conducted and analysed. Results: 30 patients were recruited for the CIPN fMRI study (mean age 60·4 years, 95% Confidence Interval: 57.4-63.4, 17 women). Two patients had lung cancer, nine had gynecological malignancies and 18 had colorectal cancer. 17 patients developed acute CIPN. Structural analysis showed that patients who developed CIPN had a smaller volume of the Nucleus Accumbens (NAc). Resting state analysis did not show clear differences between those who developed CIPN and those who did not. Finally, functional analysis showed that patients who did not develop CIPN had greater activation in the superior frontal gyrus when viewing positive emotional images as compared to those who did progress to CIPN. Region of interest analysis showed that female patients who developed CIPN had greater activity in their mesencephalic pontine reticular formation (MPRF). Male patients who progressed to CIPN had decreased activity in their thalamus. Feasability of the MINT3 study set up and fMRI paradigm was assessed. Interpretation Differences in brain structure and function are evident between patients who developed CIPN and those who did not. Crucially, the regions identified, in particular the NAc, have been postulated to denote a vulnerability for progression to pain states. Although the findings need further confirmation they suggest a paradigm shift in terms of CIPN as a clinical problem. Specifically, it appears that certain individuals can be considered as having increased risk of CIPN development prior to chemotherapy administration. This risk relates to the baseline structure, and function of their brains. Finally, the set up of the MINT3 fMRI study showed that this kind of study design is acceptable in terms of ethical and R&D approvals and a single healthy volunteer pilot.

Background Chemotherapy-induced peripheral neuropathy (CIPN) is a common dose-limiting toxicity with significant sequelae impacting prognosis and quality of life. The natural history and pathophysiological mechanisms of CIPN are unclear. Equally, the lack of systematic approach to diagnosis and assessments contribute to difficulty identifying at risk patients with implications on symptom burden. Effective management of CIPN is also difficult due to limited treatment options. To try and address this challenging clinical problem, this thesis aimed to adopt a translational approach to: 1) characterisation and early identification of the development of CIPN in cancer patients receiving neurotoxic chemotherapy and 2) explore topical treatment options in patients with chronic peripheral neuropathic pain. Methodology In the CIPN study, a mixed cohort of colorectal, gynaecological and lung cancer patients receiving neurotoxic chemotherapy (platinum agents and taxanes) were assessed prospectively, at baseline (prior to initiating chemotherapy), during cycles (every 3 weeks) and post-treatment (every 3 months) for up to 12 months (cumulative 261 assessments). Comprehensive longitudinal clinical characterisation consisted of the integration of quantitative sensory testing (QST), objective measure of function (grooved pegboard test), patient-reported outcomes and in vivo confocal microscopy to provide insight into the clinical course and potential psychophysical biomarkers of CIPN during and after chemotherapy. In the pilot intervention study, patients with chronic, complex cancer treatment related peripheral neuropathic pain received a single application of high concentration 8% capsaicin patch. Assessments conducted at baseline, 4 weeks and 12 weeks included patient-reported outcomes and QST with an exploratory application of in vivo confocal microscopy in a case. Results In the CIPN study, 33 patients when compared to 33 age and gender matched healthy controls displayed thermal hyperalgesia, sensorimotor impairment and increased resting heart rate prior to initiating neurotoxic chemotherapy. Characterisation of somato-sensory profile demonstrated dysfunction of the various types of primary afferent nerves (Aβ, Aδ and C). Assessing the change over time from baseline to during cycles and post-treatment follow up, revealed signs and symptoms as early as cycle 2 with an increase in the later cycles and 3 months post-treatment follow up. A greater burden was observed at 12 months in comparison to baseline. Significant changes were observed in QST parameters indicating both small and large fibre deficits. Interesting associations were observed for example with tactile deficits in the upper and lower limb and patient-reported outcomes. The repeated measures model provided an opportunity to distil the relationship between subjective and objective measures of CIPN. The subclinical findings at baseline however did not translate to obvious predictors of CIPN development. The exploratory use of in vivo confocal microscopy (45 healthy controls, 9 patients) demonstrated correlation with current assessment tools (QST). Analysis from the pilot intervention study of 20 patients revealed clinically significant improvement in pain in a subset at 4 and 12 weeks post-treatment. Conclusion Overall the combination of subjective and objective measures utilised in the prospective characterisation of this mixed cohort of cancer patients provided a useful paradigm for qualifying and quantifying the trajectory of CIPN related peripheral nerve damage and symptom burden with additional contribution from the novel in vivo confocal microscopy work. In capturing the varied spectrum of phenotypes, this approach may provide insight into the complexities of the underlying neurobiological mechanisms. The baseline subclinical sensory, motor and autonomic nerve dysfunction implicate a cancer-mediated process possibly contributing to CIPN. Although the preliminary investigation of baseline predictors of CIPN was inconclusive, thermal pain threshold warrant further investigation. These findings highlight the need to further address prediction and risk stratification in larger studies. The exploratory intervention study suggests that patients with chronic neuropathic pain may receive some benefit in pain severity, function and mood with effect continuing at 12 weeks post-treatment. This research warrants further investigation in larger cohorts.

A hanseníase é doença crônica causada pelo Mycobaterium leprae e possui como característica a alteração de sensibilidade cutânea, causando deformidades de mãos e pés. O tratamento compreende o uso de medicamentos (PQT) e o Ministério da Saúde (MS) preconiza o uso de palmilhas tipo Plataforma para Tarso (PT) como tratamento complementar. Há na literatura muitos artigos sobre o uso de palmilhas para melhora do equilíbrio e redistribuição da pressão plantar em pacientes neuropáticos, mas não foram encontrados estudos que incluíssem as palmilhas PT. Neste estudo comparou-se o equilíbrio do paciente hanseniano com valores de normalidade pré-definidos e também comparou-se prospectivamente a influência da palmilha PT no paciente com hanseníase, por meio de testes de equilíbrio estático e dinâmico realizados com o aparelho Balance Master (NeuroCom Int. Inc.) antes e após 3 meses de uso. Foram selecionados 40 pacientes em tratamento no HCFMRP, dos quais 19 mantiveram o seguimento à pesquisa (68,4% do sexo masculino, com média etária de 51,95), e todos com mesma classificação operacional Multibacilar. Os testes delinearam o comportamento destes pacientes com relação aos controles involuntário de equilíbrio (sistemas sensoriais-teste modifCTSIB) e voluntário de excursão do Centro de Gravidade Corporal (COG; teste LOS), bem como a estabilidade da marcha (teste WA). Os resultados obtidos com o modifCTSIB mostraram que os pacientes com hanseníase, para o controle do equilíbrio (de acordo com a relação entre testes com olhos abertos e fechados, e para superfície estável e instável; valor p<0,01 para todas as correlações aplicadas), são mais dependentes do sistema visual que do somatossensorial, em relação à normalidade. As palmilhas PT não influenciaram na modulação dos sistemas sensoriais (p>0,05). No entanto o uso das palmilhas interferiu negativamente no teste LOS, para as variáveis distância final percorrida e máxima excursão do COG somente no ponto para frente e para esquerda (p<0,01), e favoreceu o controle direcional do COG no mesmo ponto (p=0,02). Na análise dos prontuários foram identificados déficits sensitivos com maior frequência nos antepés esquerdos. O teste WA revelou que os pacientes apresentaram a marcha mais lenta em relação ao padrão de normalidade, e aumentaram a velocidade de marcha (p=0,04) e o comprimento do passo (p=0,04) após o uso das palmilhas. Concluiu-se que as palmilhas tipo PT favoreceram a estabilidade para a marcha e para o controle voluntário do equilíbrio. E ainda foi observado que os pacientes tornaram-se dependentes das palmilhas, fator importante a ser considerado pelo serviço de Saúde após a alta medicamentosa / Leprosy is a chronic disease caused by the Mycobacterium leprae with sensitivity impairment as a characteristic that could lead to deformities of hands and feet. The Ministry of Health of Brazil (MS) recommends the use of platform for Tarsus (PT) insoles as a complementary treatment to the Multi-drug Therapy (MDT). Many articles in literature discussed the use of insoles for improving balance and redistribution of plantar pressure in neuropathic patients, but there are no studies that included PT insoles. This study compared the balance control values for the leprosy patient with normal standards and also prospectively compared the influence of PT insole in patients with leprosy, by means of static and dynamic tests performed with the Balance Master (NeuroCom Int. Inc.) device, before and after 3 months. 40 patients following treatment in HCFMRP were selected, of which 19 completed the follow-up to the survey (68.4% male, mean age 51.95), all classified operationally as Multibacillary. The tests outlined the behavior of these patients with regard to involuntary balance control (modifCTSIB sensory test) and voluntary excursion of the Body Center of Gravity (COG; LOS test) as well as the gait stability (WA test). The results obtained with the modifCTSIB showed that patients with leprosy are more dependent on the visual system than the somatosensation for balance controlwhen compared to normal values (according to the relationship between tests with open and closed eyes, and stable and unstable surface; p <0.01 for all applied correlations). The insoles PT did not influence the modulation of the sensory systems (p> 0.05). However the use of insoles interfered negatively in the LOS test for the variables end point and maximum excursion of the COG only on the forward to left position (p <0.01), and favored the directional control of COG at the same position (p= 0.02). The analysis of the patients file revealed that sensitivity deficits were identified more frequently in the left forefeet, which might explain the observed differences for the excursion of the COG only in this region. The WA test showed that patients have slower walking patters compared to normal values but had their walking speed (p = 0.04) and the length of their step increased (p = 0.04) after the use of the insoles. It was concluded that the PT insoles favored gait stability and voluntary control of body balance. It was also observed that patients become dependent on the insoles, an important factor to be considered by the Health service after the PQT discharge

While cancer chemotherapy continues to significantly contribute to the number of cancer survivors, exposure to these drugs can often result in chemotherapy-induced peripheral neuropathy (CIPN), a consequence of peripheral nerve fiber dysfunction or degeneration. CIPN is characterized by sensory symptoms in the hands and feet, such as numbness, burning, and allodynia, resulting in an overall decrease in quality of life. Paclitaxel (Taxol), a microtubule poison that is commonly used to treat breast, lung, and ovarian cancers, has been found to cause CIPN in 59-78% of cancer patients. There is currently no effective preventative or therapeutic treatment for this side effect, which can be a dose-limiting factor for chemotherapy or delay treatment. Our collaborators in the laboratory of Dr. M. Imad Damaj have shown that nicotine, a nicotinic acetylcholine receptor (nAChR) agonist, and R-47, an α7 nAChR silent agonist, can prevent and reverse paclitaxel-induced peripheral neuropathy in mice. With regard to cancer, this work demonstrates that nicotine and R-47 do not enhance A549 and H460 human non-small cell lung cancer cell viability, colony formation, or proliferation alone, and they do not attenuate paclitaxel-induced growth arrest, apoptosis, or DNA fragmentation. Most importantly, nicotine and R-47 do not increase the growth of A549 tumors or interfere with the antitumor activity of paclitaxel in tumor-bearing mice. These data suggest that targeting nAChRs may be a safe and efficacious approach for the prevention and treatment of CIPN in cancer patients.

Objectives: The incidence of type 2 diabetes mellitus (DM2) is on the rise worldwide. The primary objective was to determine the prevalence of nutrient inadequacy and excessiveness in persons with DM2 with and without diabetic peripheral neuropathy (DPN). Study Design: A validated semi-quantitative food frequency questionnaire was used to determine the prevalence of inadequacy of nutrients with an estimated average requirement; the mean intake of nutrients with an adequate intake; and the proportion of persons not meeting the recommendations for the acceptable macronutrient distribution range (AMDR). Results: Differences were observed in the prevalence of inadequacy of vitamin A and the proportion of persons not meeting the AMDR for total fat, linoleic acid and carbohydrate. Conclusion: The aforementioned nutrients may have a significant role in the progression/development of DPN and should be studied in further detail. We recommend a balanced diet and use of a multi-vitamin for persons with DM2.

Objectives: The incidence of type 2 diabetes mellitus (DM2) is on the rise worldwide. The primary objective was to determine the prevalence of nutrient inadequacy and excessiveness in persons with DM2 with and without diabetic peripheral neuropathy (DPN). Study Design: A validated semi-quantitative food frequency questionnaire was used to determine the prevalence of inadequacy of nutrients with an estimated average requirement; the mean intake of nutrients with an adequate intake; and the proportion of persons not meeting the recommendations for the acceptable macronutrient distribution range (AMDR). Results: Differences were observed in the prevalence of inadequacy of vitamin A and the proportion of persons not meeting the AMDR for total fat, linoleic acid and carbohydrate. Conclusion: The aforementioned nutrients may have a significant role in the progression/development of DPN and should be studied in further detail. We recommend a balanced diet and use of a multi-vitamin for persons with DM2.