Motor unit number estimation - A Bayesian approach

All muscle contractions are dependent on the functioning of motor units. In diseases such as amyotrophic lateral sclerosis (ALS), progressive loss of motor units leads to gradual paralysis. A major difficulty in the search for a treatment for these diseases has been the lack of a reliable measure of disease progression. One possible measure would be an estimate of the number of surviving motor units. Despite over 30 years of motor unit number estimation (MUNE), all proposed methods have been met with practical and theoretical objections. Our aim is to develop a method of MUNE that overcomes these objections. We record the compound muscle action potential (CMAP) from a selected muscle in response to a graded electrical stimulation applied to the nerve. As the stimulus increases, the threshold of each motor unit is exceeded, and the size of the CMAP increases until a maximum response is obtained. However, the threshold potential required to excite an axon is not a precise value but fluctuates over a small range leading to probabilistic activation of motor units in response to a given stimulus. When the threshold ranges of motor units overlap, there may be alternation where the number of motor units that fire in response to the stimulus is variable. This means that increments in the value of the CMAP correspond to the firing of different combinations of motor units. At a fixed stimulus, variability in the CMAP, measured as variance, can be used to conduct MUNE using the "statistical" or the "Poisson" method. However, this method relies on the assumptions that the numbers of motor units that are firing probabilistically have the Poisson distribution and that all single motor unit action potentials (MUAP) have a fixed and identical size. These assumptions are not necessarily correct. We propose to develop a Bayesian statistical methodology to analyze electrophysiological data to provide an estimate of motor unit numbers. Our method of MUNE incorporates the variability of the threshold, the variability between and within single MUAPs, and baseline variability. Our model not only gives the most probable number of motor units but also provides information about both the population of units and individual units. We use Markov chain Monte Carlo to obtain information about the characteristics of individual motor units and about the population of motor units and the Bayesian information criterion for MUNE. We test our method of MUNE on three subjects. Our method provides a reproducible estimate for a patient with stable but severe ALS. In a serial study, we demonstrate a decline in the number of motor unit numbers with a patient with rapidly advancing disease. Finally, with our last patient, we show that our method has the capacity to estimate a larger number of motor units.

ASPECTOS PSICODINÂMICOS E ADAPTATIVOS DE PAIS E MÃES DE CRIANÇAS DIAGNOSTICADAS COM PARALISIA CEREBRAL

Diante do sofrimento e das questões enfrentadas por pais e mães de crianças diagnosticadas com paralisia cerebral, o presente trabalho estudou aspectos psicodinâmicos e adaptativos. Para tanto, foi realizado estudo clínico, de recorte diagnóstico, em que participaram dois pais e duas mães de crianças com paralisia cerebral que contavam de 1 a 5 anos de idade. Os pais e mães passaram por entrevistas clínicas preventivas orientadas pela EDAO e pela técnica de Desenho-Estória com Tema. A análise do material obtido indicou que todos eles passaram por período de crise após a notícia do diagnóstico da criança e, na atualidade, foram classificados como tendo Adaptação Ineficaz, sendo que os dois pais e uma mãe tinham Adaptação Ineficaz Severa, uma mãe Adaptação Ineficaz Moderada. Com relação aos aspectos intra-píquicos, foram identificadas defesas primitivas nessa lida com o sofrimento ocasionado pela deficiência da criança. Concluiu-se que os recursos internos anteriores ao nascimento da criança e à própria personalidade de cada pai/mãe foi preponderante em sua adaptação e que a paralisia cerebral da criança afetou todos os setores adaptativos dos pais e mães.(AU)

Apoptosis of neurons and oligodendrocytes in the spinal cord of spinal hyperostotic mouse (twy/twy):possible pathomechanism of human cervical compressive myelopathy

Cervical compressive myelopathy is the most serious complication of cervical spondylosis or ossification of the posterior longitudinal ligament (OPLL) and the most frequent cause of spinal cord dysfunction. There is little information on the exact pathophysiological mechanism responsible for the progressive loss of neural tissue in the spinal cord of such patients. In this study, we used the spinal hyperostotic mouse (twy/twy) as a suitable model of human spondylosis, and OPLL to investigate the cellular and molecular changes in the spinal cord. Mutant twy/twy mouse developed ossification of the ligamentum flavum at C2-C3 and exhibited progressive paralysis.

Gene therapy strategies for the treatment of spinal cord injury

Spinal cord injury is a complex pathology often resulting in functional impairment and paralysis. Gene therapy has emerged as a possible solution to the problems of limited neural tissue regeneration through the administration of factors promoting axonal growth, while also offering long-term local delivery of therapeutic molecules at the injury site. Of note, gene therapy is our response to the requirements of neural and glial cells following spinal cord injury, providing, in a time-dependent manner, growth substances for axonal regeneration and eliminating axonal growth inhibitors. Herein, we explore different gene therapy strategies, including targeting gene expression to modulate the presence of neurotrophic growth or survival factors and increase neural tissue plasticity. Special attention is given to describing advances in viral and non-viral gene delivery systems, as well as the available routes of gene delivery. Finally, we discuss the future of combinatorial gene therapies and give consideration to the implementation of gene therapy in humans. © 2014 Future Science Ltd.

Características bioativas, funcionais e efeito protetor do resíduo desidratado de camu-camu (Myrciaria dubia H.B.K. (McVaugh)) sobre doenças degenerativas utilizando modelos in vivo C. elegans

Camu-camu (Myrciaria dubia H.B.K. (McVaugh)) is a native Amazon fruit, recognized worldwide as one of the main natural sources of ascorbic acid. Due to its great acidity, this fruit is generally consumed after processing into juice or as ingredient in food preparations. As a co-product of the camu-camu processing, a significant amount of agroindustrial residue is generated. Despite the studies showing the bioactive value and biological potential of the fruit, few studies have approached the possible processing techniques, transformation and preservation of camu-camu fruits and its agroindustrial pomace. Therefore, the present work has the objective of evaluating two different drying processes applied to camu-camu pomace (peel and seeds with residual pulp), freeze drying and hot air drying, in order to obtain a functional fruit product. This thesis was divided into three stages: the first one shows the studies related to the freeze drying and hot air drying, where we demonstrated the impact of the selected drying techniques on the bioactive components of camu-camu, taking the fresh pomace as the control group. Among the investigated conditions, the groups obtained at 50ºC and 4 m/s (SC50) and 80ºC and 6 m/s (SC80) were selected as for further studies, based on their ascorbic acid final content and Folin-Ciocalteau reducing capacity. In addition to SC50 and SC80, the fresh pomace (RF) and freeze dried (RL) samples were also evaluated in these further stages of the research. Overall, the results show higher bioactive concentration in the RF samples, followed by RL, SC50 and SC80. On the second step of the research, the antioxidant, antimicrobial and antienzymatic activities were evaluated and the same tendency was observed. It was also reported, for the first time in the literature, the presence of syringic acid in dried camu-camu pomace. In the third and final stage of the research, it was investigated the effect of dried camu-camu on aging and neuroprotective disorders, using the in vivo model C.elegans. It was observed that camu-camu extracts were able to modulate important signaling genes relevant to thermal and oxidative stresses (p < 0.05). The polar acid, polar basic and polar neutral fractions obtained from the low molecular extracts of SC50 were able to extend the lifespan of wild type N2 C. elegans in 20% and 13% (p < 0.001). Results also showed that the paralysis induced by the β1-42 amyloid was significantly (p < 0.0001) retarded in CL4176 worms. Similarly, the camu-camu extracts attenuated the dopaminergic induction associated to Parkinson’s disease. Finally, a global analysis of the data presented here reveal that the camu-camu pomace, a co-product obtained from the industrial processing of a native Brazilian fruit, is a relevant natural source of health relevant compounds. This thesis, shows for the first time, the multifunctionality of camu-camu pomace, a natural resource still underexploited for scientific, commercial and technological purposes.

Polpa de jambolão (Eugenia jambolana Lam.) fresca e desidratada: características físico-químicas, bioativas e funcionais, efeitos biológicos em Caenorhabditis elegans e uso para produção de frozen yogurt caprino probiótico

This work evaluated the fresh, spray dried (with 10 % of Arabic Gum) and freeze dried jambolan pulp (Eugenia jambolana Lam.) in regard to physicochemical (pH, moisture, water activity, average particle diameter, solubility and color), bioactive [total phenolic content (TPC), monomeric anthocyanin, pronathocyanidin (PA), total elagic acid (TEA), myricetin and cyanidin] and in vitro functionality (antioxidant, antienzymatic and antimicrobial activities]. In addition, the in vivo functionality of jambolan pulp was investigated using the Caenorhabditis elegans model for insulin signaling, longevity and induced neurodegeneration (Alzheimer’s disease and Parkinson’s disease related symptoms). The dried jambolan pulp presented TPC retention (50% to 75%), PA (90% to 98%), TEA (31% to 83%), myricetin (40% to 84%), cyanidin (72% to 84%) and antioxidant activity (15%). The fresh jambolan pulp, the freeze dried pulp and the spray dried jambolan pulp presented high enzymatic inhibitory activity against pancreatic lipase (4,4 to 5,8 mg/mL), alpha-glycosidase (10,3 to 13,8 mg/mL) and alpha-amylase (8,9 to 11,2 mg/mL). They also were active inhibitors against the pathogen S. aureus. The dried jambolan experimental samples were able to increase the expression of several genes linked to the insulin signaling pathways (SIR-2.1, PPTR-1, DAF-16, SOD-3, e CTL) and increased the lifespan in C. elegans (18,07 % - 24,34 %), besides decreasing the amyloid AB1-42 aggregation induced paralysis and MPP+ (1-methyl-4-phenylpyridinium) induced neurodegeneration. Based on that, the jambolan pulp and the spray dried jambolan pulp were further selected for the production of caprine frozen yogurt with the addition of Bifidobacterium animalis subsp. lactis BI-07. The final product were evaluated in regard to their physicochemical (pH, acidity, total solids, protein, total reducing sugars, fat, ashes, overrun, melting test), bioactive (TPC and monomeric anthocyanin, antioxidant activity, probiotic viability and sensory analysis (sensory acceptance). The results showed that samples with probiotic had lowest pH and higher acidity, TPC, anthocyanin and antioxidant activity. It was also observed low overrun (14.2% to 22.6%). vi Samples with probiotic had lower flavor scores. Overall, this research presents the jambolan as a highly functional bioactive-rich fruit with the potential to modulate important biological pathways, extend lifespan and retard the development of neurodegenerative diseases. Jambolan is an underexploited exotic fruit with a high colorant potential and this thesis shows for the first time in the literature important technological, biological and scientific data about this fruit that could be used towards the development of health-oriented food products.

Síndrome de Guillain-Barré: epidemiologia, prognóstico e fatores de risco

Introduction. Guillain-Barré syndrome (GBS) is an immune-mediated polyneuropathy and the principal cause of acute neuromuscular paralysis. The most prominent GBS subtypes are: acute inflammatory demyelinating polyneuropathy (AIDP), acute motor axonal neuropathy (AMAN), acute motor-sensory axonal neuropathy (AMSAN) and Fisher syndrome (FS). Differences in geographical distribution of variants have been reported. In Brazil, there are few studies describing the characteristics of GBS, but none on the frequency of GBS variants and their clinical manifestations. Infection-induced aberrant immune response resulting from molecular mimicry and formation of cross-reacting antibodies, contribute to complement activation. Functional biallelic polymorphism in immunoglobulin receptors that influence the affinity of IgG subclasses and the type of immune response have been described, suggesting genetic susceptibility to developing disease. It remains unclear whether individuals carrying different FCGR alleles have differential risk for GBS and⁄or disease severity. The goals of this study were: (1) To characterize GBS and describe the clinical findings in a cohort of patients with GBS from the state of Rio Grande do Norte, Brazil; (2) to determine whether polymorphism in FCGR were associated with development of GBS, and (3) to tease out whether the global gene expression studies could be a tool to identify pathways and transcriptional networks which could be regulated and decrease the time of disease. Methods. Clinical and laboratory data for 149 cases of GBS diagnosed from 1994 to 2013 were analyzed. Genomic DNA and total RNA were extracted from whole blood. Antigangliosides antibodies were determined in the sera. In addition, we also assessed whether FCGR polymorphism are present in GBS (n=141) and blood donors (n=364), and global gene expressions were determined for 12 participants with GBS. Blood samples were collected at the diagnosis and post-recovery. Results. AIDP was the most frequent variant (81.8%) of GBS, followed by AMAN (14.7%) and AMSAN (3.3%). The incidence of GBS was 0.3 ⁄ 100,000 people for the state of Rio Grande do Norte and cases occurred at a younger age. GBS was preceded by infections, with the axonal variant associated with episodes of diarrhea (P = 0.025). Proximal weakness was more frequent in AIDP, and distal weakness predominant in the axonal variant. Compared to 42.4% of cases with AIDP (P<0.0001), 84.6% of cases with the axonal variant had nadir in <10 days. Individuals with the axonal variant took longer to recover deambulation (P<0.0001). The mortality of GBS was 5.3%. A worse outcome was related to an axonal variant (OR17.063; P=0.03) and time required to improve one point in the Hughes functional scale (OR 1.028; P=0.03). The FCGR genotypes and allele frequencies did not differ significantly between the patients with GBS and the controls (FCGR2A p=0.367 and FCGR3A p=0.2430). Global gene expression using RNAseq showed variation in transcript coding for protein isoforms during acute phase of disease. Conclusions. The annual incidence of GBS was 0.3 per 100,00 and there was no seasonal pattern. A predominance of the AIDP variant was seen, and the incidence of the disease decreased with age. The distribution of weakness is a function of the clinical variants, and individuals with the axonal variant had a poorer prognosis. Early diagnosis and variant identification leads to proper intervention decreasing in long-term morbidity. FCGR polymorphisms do not seem to influence susceptibility to GBS in this population. This study found deregulated genes and signs of transcriptional network alterations during the acute and recovery phases in GBS. Identification of pathways altered during disease might be target for immune regulation and with potential to ameliorate symptoms.

Parasite-host interaction of the protoplast isolates of Entomophthora egressa with the eastern hemlock looper and the eastern spruce budworm

Hemocytes of the insects Lambdina fiscellaria fiscellaria and Choristoneura fumiferana did not adhere to the protoplasts of ~he fungus EntomoEhthora egressa. Hemocyte reaction for both insect species to test-particles was not suppressed by the protoplasts. The spherule cells of _-L. fiscellaria fiscellaria adhered to the spherical hyphal bodies and hyphae of ~· ~gressa. The granular cells of -c. fumiferana adhered to the hyphae of ~· egress~. Protoplasts exposed to papain were attacked by the granular ·cells of -c. fumiferana. Spent growth medium of both protoplast isolates produced paralysis when injected into -c. fumiferana larvae. Evidence suggests that heat-stable proteins may be involved. Protoplast isolates showed differences in the growth rates and regeneration sequences using coagulated egg yolk medium, a highly modified version of Grace's insect tissue . culture medium (MGM) and modifications of MGM and in the presence of C0₂. The isolates also differed in the changes that they induced in MGM composition during protoplast growth and in the rates of glucose utilization and protein secretion. The serum of c. fumiferana larvae contained protein(s) which we believe adhere to the cell membranes of the protoplasts of E. egressa. Evidence is presented for hemocyteplasn~ interaction in the presence of protoplasts. Components in the larval serum were found to influence protoplast growth patterns. The possibility of antiprotoplast serum activity is presented. Melanin, toxic levels of ninhydrinpositive compounds and antiprotoplast proteins may have been involved in this activity. The granular cells of -L. fiscellaria fiscellaria and Q• fumiferana adhered to the hyphae of ,Rhizopus ~i$rican~. Spores of Absidia repens and the bacteria Escherichia coli and Bacillus cereus adhered to the granular cells of both species of· insects. The granular cells and plasmatocytes of -c. fumiferana were capable of phagocytosing -B. cereus. Adhesion of .A... . repens spores to c. fumiferana granular cells ~ . - was stimulated by N-acetylglucosamine and glucosamine, moderately reduced by D-fucose, D-arabinose, D-mannose, D-galatose and sucrose and mildly reduced by D-glucose, D-fructose and trehalose. There was no evidence of humoral opsonins in larval hemolymph favoring test-particle-hemocyte interaction. Granular cells of c. fumiferana exposed to papain had reduced affinities for A. repens spores.

Desenvolvimento de um exoesqueleto para movimentação/reabilita-ção de paraplégicos

People bedridden by paralysis/motor disability are subject to several problems due to lack of movement. Then, it is necessary to use equipments that enable the people bedridden stand up and walk, so as to reduce the problems due to lack of movement and the time of rehabilitation, impacting directly on its quality of life. The aim of this work is the development of an exoskeleton to make the movement of people with paralysis / motor impairment in the lower limbs, without help of third parties, to be activated by the user. To provide support, stability and security to the deficient, it was decided to use a structure formed by four legs, being each leg consisting of a parallel chain. The gait was obtained by combining the movement of two mechanisms: crank/rocker, responsible for oscillatory motion of the leg, and cam/follower, responsible for the foot motion on the desired trajectory. To achieve the aim of this work was conducted a study about the types of exoskeletons for locomotion/rehabilitation of people with lower limb paralysis and presented a study on the movement of the lower limb joints. It also presented a mathematical model to obtain the desired path to the foot of the exoskeleton, the static model and the design of the structure elements. In the sequence is presented the simulation of movements of a people during the human gait, experimental tests and the comparison with the human gait developed by a people without disabilities.

Setting the Rules of the Game: Mega-Regionals and the Role of the WTO

Faced with a WTO in a state of paralysis, large developed trading nations have shifted their attentions to other fora to pursue their trade policy objectives. In particular, preferential trade agreements (PTAs) are now being used to promote the regulatory disciplines that were previously rejected by developing countries at the multilateral level. These so-called ‘deep’ or ‘21st century’ PTAs address a variety of issues, from technical norms, procurement, investment protection and intellectual property rights to social and environmental protection. Moreover, recently, developed countries have sought to negotiate PTAs which are large in scale, both in terms of economic size and geographical reach, including the so-called ‘mega-regional’ PTAs, such as the EU-US Transatlantic Trade and Investment Partnership, the EU-Japan PTA, the Transpacific Partnership, and the China-backed Regional Comprehensive Economic Partnership. These mega-regional PTAs are distinctive not just in terms of their sheer size and the breadth and depth of issues addressed, but also because some of their proponents readily admit that one of the central aims pursued by such agreements is to design global rules on new trade issues. In other words, these agreements are being conceived as alternatives to multilateral rule making at the WTO level. The proliferation of 21st century trade deals raises important questions concerning the continued relevance of the WTO as a global rule-making venue, and the impact that the regulatory disciplines promoted in such agreements will have on both developing and developed countries. This paper discusses the emerging features of an international trading system that is increasingly populated by large-scale PTAs and discusses some of the points of tension that arise from such practice. Firstly, it examines instances of horizontal tension resulting from the proliferation of PTAs, particularly the extent to which such PTAs represent a threat or multilateral trade governance. Secondly, it looks at an example of vertical tension by examining the manner in which the imposition of regulatory disciplines through trade agreements can undermine the ability of countries, especially developing countries, to pursue legitimate public interest objectives. Finally, the paper considers a number of steps that could be considered to address some of the adverse effects associated with the fragmentation of the international trading system, including the option of embracing variable geometry within the WTO framework and the need to develop mechanisms that provide flexibility for developing countries in the implementation of regulatory disciplines.

Étude de la toxicité causée par le gène C9orf72 dans la Sclérose Latérale Amyotrophique

La Sclérose Latérale Amyotrophique (SLA) est une maladie neurodégénérative qui affecte les neurones moteurs. 10% des cas sont des cas familiaux et l’étude de ces familles a mené à la découverte de plusieurs gènes pouvant causer la SLA, incluant SOD1, TARDBP et FUS. L’expansion de la répétition GGGGCC dans le gène C9orf72 est, à ce jour, la cause la plus connue de SLA. L’impact de cette expansion est encore méconnu et il reste à déterminer si la toxicité est causée par un gain de fonction, une perte de fonction ou les deux. Plusieurs gènes impliqués dans la SLA sont conservés entre le nématode Caenorhabditis elegans et l’humain. C. elegans est un vers transparent fréquemment utilisé pour des études anatomiques, comportementales et génétiques. Il possède une lignée cellulaire invariable qui inclue 302 neurones. Aussi, les mécanismes de réponse au stress ainsi que les mécanismes de vieillissement sont très bien conservés entre ce nématode et l’humain. Donc, notre groupe, et plusieurs autres, ont utilisé C. elegans pour étudier plusieurs aspects de la SLA. Pour mieux comprendre la toxicité causée par l’expansion GGGGCC de C9orf72, nous avons développé deux modèles de vers pour étudier l’impact d’une perte de fonction ainsi que d’un gain de toxicité de l’ARN. Pour voir les conséquences d’une perte de fonction, nous avons étudié l’orthologue de C9orf72 dans C. elegans, alfa-1 (ALS/FTD associated gene homolog). Les vers mutants alfa-1(ok3062) développent des problèmes moteurs causant une paralysie et une dégénérescence spécifique des neurones moteurs GABAergiques. De plus, les mutants sont sensibles au stress osmotique qui provoque une dégénérescence. D’autre part, l’expression de la séquence d’ARN contenant une répétition pathogénique GGGGCC cause aussi des problèmes moteurs et de la dégénérescence affectant les neurones moteurs. Nos résultats suggèrent donc qu’un gain de toxicité de l’ARN ainsi qu’une perte de fonction de C9orf72 sont donc toxiques pour les neurones. Puisque le mouvement du vers peut être rapidement évalué en cultivant les vers dans un milieu liquide, nous avons développé un criblage de molécules pouvant affecter le mouvement des vers mutants alfa-1 en culture liquide. Plus de 4 000 composés ont été évalués et 80 ameliore la mobilité des vers alfa-1. Onze molécules ont aussi été testées dans les vers exprimant l’expansion GGGGCC et huit diminuent aussi le phénotype moteur de ces vers. Finalement, des huit molécules qui diminent la toxicité causée par la perte de fonction de C9orf72 et la toxicité de l’ARN, deux restaurent aussi l’expression anormale de plusieurs transcrits d’ARN observée dans des cellules dérivées de patient C9orf72. Avec ce projet, nous voulons identifier des molécules pouvant affecter tous les modes de toxicité de C9orf72 et possiblement ouvrir de nouvelles avenues thérapeutiques

Clinical and functional outcomes of acute lower extremity compartment syndrome at a Major Trauma Hospital

Background: Acute lower extremity compartment syndrome (CS) is a condition that untreated causes irreversible nerve and muscle ischemia. Treatment by decompression fasciotomy without delay prevents permanent disability. The use of intracompartmental pressure (iCP) measurement in uncertain situations aids in diagnosis of severe leg pain. As an infrequent complication of lower extremity trauma, consequences of CS include chronic pain, nerve injury, and contractures. The purpose of this study was to observe the clinical and functional outcomes for patients with lower extremity CS after fasciotomy. Methods: Retrospective chart analysis for patients with a discharge diagnosis of CS was performed. Physical demographics, employment status, activity at time of injury, injury severity score, fracture types, pain scores, hours to fasciotomy, iCP, serum creatine kinase levels, wound treatment regimen, length of hospital stay, and discharge facility were collected. Lower extremity neurologic examination, pain scores, orthopedic complications, and employment status at 30 days and 12 months after discharge were noted. Results: One hundred twenty‑four patients were enrolled in this study. One hundred and eight patients were assessed at 12 months. Eighty‑one percent were male. Motorized vehicles caused 51% of injuries in males. Forty‑one percent of injuries were tibia fractures. Acute kidney injury occurred in 2.4%. Mean peak serum creatine kinase levels were 58,600 units/ml. Gauze dressing was used in 78.9% of nonfracture patients and negative pressure wound vacuum therapy in 78.2% of fracture patients. About 21.6% of patients with CS had prior surgery. Nearly 12.9% of patients required leg amputation. Around 81.8% of amputees were male. Sixty‑seven percent of amputees had associated vascular injuries. Foot numbness occurred in 20.5% of patients and drop foot palsy in 18.2%. Osteomyelitis developed in 10.2% of patients and fracture nonunion in 6.8%. About 14.7% of patients underwent further orthopedic surgery. At long‑term follow‑up, 10.2% of patients reported moderate lower extremity pain and 69.2% had returned to work. Conclusion: Escalation in leg pain and changes in sensation are the cardinal signs for CS rather than reliance on assessing for firm compartments and pressures. The severity of nerve injury worsens with the delay in performing fasciotomy. Standardized diagnostic protocols and wound treatment strategies will result in improved outcomes from this complication.

Tooth Mobility, Periodontal Ligament Space, and the Alveolus During Periodontal Health, Disease, and Disuse

Thesis (Ph.D.)--University of Washington, 2016-07

Surgical treatment of papillary thyroid carcinoma without lymph nodal involvement

Background. In the treatment of differentiated thyroid cancer (DTC), in absence of enlarged lymph nodes, the role of routine central lymph node dissection (RCLD) remains controversial. The aim of this study is to analyze data resulting from total thyroidectomy (TT) not combined with RCLD in the treatment of DTC. Methods. We retrospectively evaluated the clinical records of 80 patients treated between January 1996 and December 2003 with TT without RCLND, in absence of suspected enlarged lymph nodes at preoperative ultrasonography and intraoperatively during neck exploration. In this series, 75 patients (93.7%) underwent radioiodine (RAI) ablation, followed by Thyroid Stimulating Hormone (TSH) suppression therapy. In case of locoregional lymph nodal recurrence, a central (VI) and ipsilateral (III-IV) selective lymph node dissection was performed. Results. Incidence of permanent hypoparathyroidism (iPTH < 10 pg/ml) and unilateral temporary vocal fold paralysis were respectively 2.55% and 2.55%. Locoregional recurrence, with positive cervical lymph nodes, after a 10.3 ± 4.7 years mean follow-up was observed in 3 patients (3.75%). They were submitted to a central (VI) and ipsilateral (III-IV) selective neck dissection without significant complications. Conclusions. In our series, TT not combined with RCLD was associated to a low locoregional recurrence rate, even if the lack of a control group treated with RCLD does not allow any generalized assumption. RCLD may be indicated in high risk patients, in whom lymph nodal recurrence is more frequent. More prospective randomized studies are needed to better define the role of RCLD and postoperative radioiodine ablation.

Identification and Characterization of New Small Molecule Inhibitors of Picornavirus Replication

The Picornaviridae family consists of positive-strand RNA viruses that are the causative agents of a variety of diseases in humans and animals. Few drugs targeting picornaviruses are available, making the discovery of new antivirals a high priority. Here, we identified and characterized three compounds from a library of kinase inhibitors that block replication of poliovirus, coxsackievirus B3, and encephalomyocarditis virus. The antiviral effect of these compounds is not likely related to their known cellular targets because other inhibitors targeting the same pathways did not inhibit viral replication. Using an in vitro translation-replication system, we showed that these drugs inhibit different stages of the poliovirus life cycle. A4(1) inhibited the formation of a functional replication complex, while E5(1) and E7(2) affected replication after the replication complex had formed. A4(1) demonstrated partial protection from paralysis in a murine model of poliomyelitis. Poliovirus resistant to E7(2) had a single mutation in the 3A protein. This mutation was previously found to confer resistance to enviroxime-like compounds, which target either PI4KIIIβ (major enviroxime-like compounds) or OSBP (minor enviroxime-like compounds), cellular factors involved in lipid metabolism and shown to be important for replication of diverse positive-strand RNA viruses. We classified E7(2) as a minor enviroxime-like compound, because the localization of OSBP changed in the presence of this inhibitor. Interestingly, both E7(2) and major enviroxime-like compound GW5074 interfered with the viral polyprotein processing. Multiple attempts to isolate resistant mutants in the presence of A4(1) or E5(1) were unsuccessful, showing that effective broad-spectrum antivirals could be developed on the basis of these compounds. Studies with these compounds shed light on pathways shared by diverse picornaviruses that could be potential targets for the development of broad-spectrum antiviral drugs.