Natural history of circulating miRNAs in Duchenne disease: Association with muscle injury and metabolic parameters.

OBJECTIVES: This study aimed to evaluate whether the expression of circulating dystromiRs and a group of oxidative stress-related (OS-R) miRNAs is associated with muscle injury and circulating metabolic parameters in Duchenne muscular dystrophy (DMD) patients. METHODS: Twenty-four DMD patients were included in this cross-sectional study. Clinical scales to evaluate muscle injury (Vignos, GMFCS, Brooke, and Medical Research Council), enzymatic muscle injury parameters (CPK, ALT, and AST), anthropometry, metabolic indicators, physical activity, serum dystromiRs (miR-1-3p, miR-133a-3p, and miR-206), and OS-R miRNAs (miR-21-5p, miR-31-5p, miR-128-3p, and miR-144-3p) levels were measured in ambulatory and non-ambulatory DMD patients. RESULTS: DystromiRs (except miR-1-3p) and miRNAs OS-R levels were lower (p-value <.05) in the non-ambulatory group than the ambulatory group. The expression of those miRNAs correlated with Vignos scale score (For instance, rho = -0.567, p-value <0.05 for miR-21-5p) and with other scales scores of muscle function and strength. CPK, AST, and ALT concentration correlated with expression of all miRNAs (For instance, rho = 0.741, p-value <.05 between miR-206 level and AST concentration). MiR-21-5p level correlated with glucose concentration (rho = -0.369, p-value = .038), and the miR-1-3p level correlated with insulin concentration (rho = 0.343, p-value = .05). CONCLUSIONS: Non-ambulatory DMD patients have lower circulating dystromiRs and OS-R miRNAs levels than ambulatory DMD patients. The progressive muscle injury is associated with a decrease in the expression of those miRNAs, evidencing DMD progress. These findings add new information about the natural history of DMD.

Genetic analysis of muscular dystrophies: our experience in Mexico.

Muscular dystrophies are a group of well-defined genetic disorders characterized by the variable distribution of muscle wasting and progressive weakness. The diagnosis and treatment of these diseases remain challenging due to genetic heterogeneity and clinical overlapping. Herein, we describe our 10 years' experience with the diagnosis and management of muscular dystrophy patients. In total, 169 patients were screened for pathogenic variants in eleven genes linked to frequent muscular dystrophies using MLPA and NGS sequencing panels. Most frequent muscular dystrophies found in the Mexican population were dystrophinopathies, dysferlinopathies and calpainopathies. Novel variants were found in genes: DMD, CAPN3, DYSF, and FKRP. For Duchenne muscular dystrophy, improvements in early diagnosis and prolonged ambulation were achieved, on the contrary, for limb-girdle muscular dystrophies and congenital muscular dystrophies, uncomplimentary follow-up and lack of detection strategies were observed. For most common muscular dystrophies, improvements in diagnosis and management have been achieved in the last 10 years, due to a collaborative effort done nationwide.

Potential therapeutic impact of omega-3 long chain-polyunsaturated fatty acids on inflammation markers in Duchenne muscular dystrophy: A double-blind, controlled randomized trial.

BACKGROUND & AIMS: Duchenne Muscular Dystrophy (DMD) is the most frequent dystrophy in childhood generated by a deficiency in dystrophin. DMD is a neuromuscular disease and its clinical course comprises chronic inflammation and gradual muscle weakness. Supplementation of omega-3 long chain-Polyunsaturated Fatty Acids (ω-3 long chain-PUFA) reduces inflammatory markers in various disorders. The goal of this research was to analyze the influence of ω-3 long chain-PUFA intake on gene expression and blood inflammatory markers in boys with DMD. METHODS: In a placebo-controlled, double. Blind, randomized trial, boys with DMD (n = 36) consumed 2.9 g/day of ω-3 long chain-PUFA or sunflower oil as control, in capsules, for a period of 6 months. Blood was analyzed at baseline and at months 1, 2, 3, and 6 of supplementation for expression of inflammatory markers in leukocytes and serum. RESULTS: There was high adherence to capsule intake (control: 95.3% ± 7.2%, and ω-3 long chain-PUFA: 97.4% ± 3.7% at month 6). Enrichment of EicosaPentaenoic Acid (EPA) and DocosaHexaenoic Acid (DHA) in erythrocytes increased significantly in patients supplemented with ω-3 long chain-PUFA compared with the placebo group during the 6 months of supplementation. Messenger RNA (mRNA) of the Nuclear Factor kappa beta (NF-κB) and its target genes InterLeukin 1 beta (IL-1ß) and IL-6 was downregulated significantly (p < 0.05) in leukocytes from DMD boys supplemented with ω-3 long chain-PUFA for 6 months, compared to the placebo group. Omega-3 long chain-PUFA intake decreased the serum IL-1ß (-59.5%; p = 0.011) and IL-6 (-54.8%; p = 0.041), and increased the serum IL-10 (99.9%, p < 0.005), in relation to those with placebo treatment. CONCLUSION: Supplementation with ω-3 long chain-PUFA 2.9 g/day is well-tolerated, has a beneficial reductive effect on proinflammatory markers, and increases an anti-inflammatory marker, indicating that ω-3 long chain-PUFA could have a potential therapeutic impact on chronic inflammation in DMD. This research is registered at clinicaltrials.gov (NCT018264229).

Systemic Inflammation in Duchenne Muscular Dystrophy: Association with Muscle Function and Nutritional Status.

Inflammation described in patients with Duchenne muscular dystrophy (DMD) may be related to loss of muscle function or to obesity. It is unknown if circulating proinflammatory cytokines (IL-6, IL-1, and TNF-α) levels are associated with muscle function. The purpose was to evaluate whether an association exists between systemic inflammation with muscle function and nutritional status in DMD patients. In 66 DMD patients without corticosteroid treatment, the following were evaluated in serum: cytokines (IL-1, IL-6, and TNF-α), C-reactive protein (CRP), leptin, adiponectin, and creatine kinase (CK). Muscle function was evaluated using Vignos Scale. Patients with better muscle function had the highest concentration of CK, IL-1, and TNF-α compared with less muscle function. No differences in IL-6 and adiponectin concentration were identified among groups with different levels of muscle function. Also, no differences were observed in the concentration of cytokines among groups with different nutritional status levels (underweight, normal weight, and overweight/obese). However, CRP and leptin were increased in the obese group compared with normal and underweight subjects. Systemic inflammation is increased in patients with better muscle function and decreases in DMD patients with poorer muscle function; nevertheless, systemic inflammation is similar among different levels of nutritional status in DMD patients.

Non-Invasive Biomarkers for Duchenne Muscular Dystrophy and Carrier Detection.

Non-invasive biological indicators of the absence/presence or progress of the disease that could be used to support diagnosis and to evaluate the effectiveness of treatment are of utmost importance in Duchenne Muscular Dystrophy (DMD). This neuromuscular disorder affects male children, causing weakness and disability, whereas female relatives are at risk of being carriers of the disease. A biomarker with both high sensitivity and specificity for accurate prediction is preferred. Until now creatine kinase (CK) levels have been used for DMD diagnosis but these fail to assess disease progression. Herein we examined the potential applicability of serum levels of matrix metalloproteinase 9 and matrix metalloproteinase 2, tissue inhibitor of metalloproteinases 1, myostatin (GDF-8) and follistatin (FSTN) as non-invasive biomarkers to distinguish between DMD steroid naïve patients and healthy controls of similar age and also for carrier detection. Our data suggest that serum levels of MMP-9, GDF-8 and FSTN are useful to discriminate DMD from controls (p < 0.05), to correlate with some neuromuscular assessments for DMD, and also to differentiate between Becker muscular dystrophy (BMD) and Limb-girdle muscular dystrophy (LGMD) patients. In DMD individuals under steroid treatment, GDF-8 levels increased as FSTN levels decreased, resembling the proportions of these proteins in healthy controls and also the baseline ratio of patients without steroids. GDF-8 and FSTN serum levels were also useful for carrier detection (p < 0.05). Longitudinal studies with larger cohorts are necessary to confirm that these molecules correlate with disease progression. The biomarkers presented herein could potentially outperform CK levels for carrier detection and also harbor potential for monitoring disease progression.

Comparison of mutation profiles in the Duchenne muscular dystrophy gene among populations: implications for potential molecular therapies.

Novel therapeutic approaches are emerging to restore dystrophin function in Duchenne Muscular Dystrophy (DMD), a severe neuromuscular disease characterized by progressive muscle wasting and weakness. Some of the molecular therapies, such as exon skipping, stop codon read-through and internal ribosome entry site-mediated translation rely on the type and location of mutations. Hence, their potential applicability worldwide depends on mutation frequencies within populations. In view of this, we compared the mutation profiles of the populations represented in the DMD Leiden Open-source Variation Database with original data from Mexican patients (n = 162) with clinical diagnosis of the disease. Our data confirm that applicability of exon 51 is high in most populations, but also show that differences in theoretical applicability of exon skipping may exist among populations; Mexico has the highest frequency of potential candidates for the skipping of exons 44 and 46, which is different from other populations (p < 0.001). To our knowledge, this is the first comprehensive comparison of theoretical applicability of exon skipping targets among specific populations.

Detección de la duplicación del gen PMP22 en pacientes con neuropatía periférica: estudio en la población mexicana / Detection of the PMP22 gene duplication in peripheral neuropathy patients: a study in Mexican population

Introducción. La enfermedad de Charcot-Marie-Tooth (CMT) es una neuropatía que afecta los nervios motores y sensitivos, y la CMT1A es el subtipo más frecuente en el mundo. La CMT1A se produce por una duplicación de 1,5 Mb en el locus 17p11.2-p12, donde se localiza el gen PMP22. Para el diagnóstico de CMT1A es importante contar con técnicas moleculares específicas para la determinación de esta mutación. Objetivos. Establecer un método de uso rutinario para detectar la duplicación de PMP22 en la población mexicana y estimar su frecuencia en pacientes con características clínicas para la CMT. Pacientes y métodos. Se analizaron 157 pacientes mexicanos no relacionados entre sí, diagnosticados de CMT por valoración clínica. La determinación de la duplicación de PMP22 se realizó a través de reacción en cadena de la polimerasa en tiempo real mediante el método comparativo 2-ΔΔCT. Resultados. El método 2-ΔΔCT para detectar la duplicación del gen PMP22 mostró ser sensible y fiable. Los resultados fueron consistentes con los obtenidos mediante la técnica de hibridación in situ fluorescente. Se detectó la duplicación de PMP22 en 79 pacientes (50,3%), con un comportamiento similar a lo comunicado en Estados Unidos, Australia, Finlandia, Suecia y España. Sin embargo, se observó que existen diferencias con otras poblaciones. Conclusiones. La técnica de reacción en cadena de la polimerasa cuantitativa se implementó como un diagnóstico molecular de CMT1A eficaz y de bajo coste, por lo que puede utilizarse rutinariamente en México. Esto es esencial para el asesoramiento genético y el tratamiento oportuno de los pacientes con CMT. La frecuencia de la duplicación del gen PMP22 varía entre regiones geográficas, por lo que es importante estimarla en diferentes poblaciones (AU)

Introduction. Charcot-Marie-Tooth disease (CMT) is a neuropathy that affects sensory and motor nerves. The most common CMT subtype is CMT1A due to a PMP22 duplication of a 1.5 Mb fragment on the 17p11.2-p12. The development of a specific molecular technique that detects the PMP22 duplication is necessary for the diagnosis of CMT1A. Aim. To establish a routinary test for detection of the PMP22 gene duplication in Mexican population and to estimate the CMT1A frequency in patients clinically diagnosed as CMT. Patients and methods. A cohort of 157 individuals clinically diagnosed as CMT were analyzed. The detection of the PMP22 gene duplication was performed using the comparative 2-ΔΔCT qPCR method. Results. The comparative 2-ΔΔCT method was sensitive and reliable for the detection of the PMP22 duplication. In order to validate the testing, data was compared with FISH results. Duplication of PMP22 was detected in 79 patients (50.3%). Although CMT1A frequency is different among populations, in Mexican patients it was similar with other populations such as United States, Australia, Finland, Sweden and Spain. Conclusions. The qPCR technique is an accurate and inexpensive method for the diagnosis of CMT1A. This method can be routinely used in México where CMT1A represents ≈ 50% of CMT cases. Molecular diagnosis of CMT1A is essential for the genetic counseling and treatment of patients (AU)

[Detection of the PMP22 gene duplication in peripheral neuropathy patients: a study in Mexican population]. / Detección de la duplicación del gen PMP22 en pacientes con neuropatía periférica: estudio en la población mexicana

INTRODUCTION: Charcot-Marie-Tooth disease (CMT) is a neuropathy that affects sensory and motor nerves. The most common CMT subtype is CMT1A due to a PMP22 duplication of a 1.5 Mb fragment on the 17p11.2-p12. The development of a specific molecular technique that detects the PMP22 duplication is necessary for the diagnosis of CMT1A. AIM: To establish a routinary test for detection of the PMP22 gene duplication in Mexican population and to estimate the CMT1A frequency in patients clinically diagnosed as CMT. PATIENTS AND METHODS: A cohort of 157 individuals clinically diagnosed as CMT were analyzed. The detection of the PMP22 gene duplication was performed using the comparative 2-ΔΔCT qPCR method. RESULTS: The comparative 2-ΔΔCT method was sensitive and reliable for the detection of the PMP22 duplication. In order to validate the testing, data was compared with FISH results. Duplication of PMP22 was detected in 79 patients (50.3%). Although CMT1A frequency is different among populations, in Mexican patients it was similar with other populations such as United States, Australia, Finland, Sweden and Spain. CONCLUSIONS: The qPCR technique is an accurate and inexpensive method for the diagnosis of CMT1A. This method can be routinely used in Mexico where CMT1A represents ≍ 50% of CMT cases. Molecular diagnosis of CMT1A is essential for the genetic counseling and treatment of patients.

TITLE: Deteccion de la duplicacion del gen PMP22 en pacientes con neuropatia periferica: estudio en la poblacion mexicana.Introduccion. La enfermedad de Charcot-Marie-Tooth (CMT) es una neuropatia que afecta los nervios motores y sensitivos, y la CMT1A es el subtipo mas frecuente en el mundo. La CMT1A se produce por una duplicacion de 1,5 Mb en el locus 17p11.2-p12, donde se localiza el gen PMP22. Para el diagnostico de CMT1A es importante contar con tecnicas moleculares especificas para la determinacion de esta mutacion. Objetivos. Establecer un metodo de uso rutinario para detectar la duplicacion de PMP22 en la poblacion mexicana y estimar su frecuencia en pacientes con caracteristicas clinicas para la CMT. Pacientes y metodos. Se analizaron 157 pacientes mexicanos no relacionados entre si, diagnosticados de CMT por valoracion clinica. La determinacion de la duplicacion de PMP22 se realizo a traves de reaccion en cadena de la polimerasa en tiempo real mediante el metodo comparativo 2­ΔΔCT. Resultados. El metodo 2­ΔΔCT para detectar la duplicacion del gen PMP22 mostro ser sensible y fiable. Los resultados fueron consistentes con los obtenidos mediante la tecnica de hibridacion in situ fluorescente. Se detecto la duplicacion de PMP22 en 79 pacientes (50,3%), con un comportamiento similar a lo comunicado en Estados Unidos, Australia, Finlandia, Suecia y España. Sin embargo, se observo que existen diferencias con otras poblaciones. Conclusiones. La tecnica de reaccion en cadena de la polimerasa cuantitativa se implemento como un diagnostico molecular de CMT1A eficaz y de bajo coste, por lo que puede utilizarse rutinariamente en Mexico. Esto es esencial para el asesoramiento genetico y el tratamiento oportuno de los pacientes con CMT. La frecuencia de la duplicacion del gen PMP22 varia entre regiones geograficas, por lo que es importante estimarla en diferentes poblaciones.

Estudios de imagen y electromiografía en las miopatías inflamatorias / Imaging techniques and electromyography in inflammatory myopathies

Las diferentes técnicas de imagen son empleadas en el diagnóstico, evaluación y seguimiento de las miopatías inflamatorias (MI); de ellas, las que mayor información proporcionan son el ultrasonido y las imágenes por resonancia magnética. El electrodiagnóstico en las MI se basa principalmente en la electromiografía (EMG), que presenta diferentes patrones sobre la base del tiempo de evolución de la enfermedad. En la fase aguda se documentan potenciales polifásicos con duración y amplitud disminuidas, mientras que en la fase crónica se encuentran patrones mixtos neuropáticos y miopáticos. Posterior al tratamiento con esteroides tiende a normalizarse la EMG (AU)

Different imaging techniques are used for the diagnosis, evaluation and follow-up of inflammatory myopathies; of these techniques, the ones that provide the largest amount of information are ultrasonography and magnetic resonance imaging. Electrodiagnosis in inflammatory myopathies is based mainly on electromyography (EMG), which shows different patterns according to the different disease presentations. In the acute phase, polyphasic potentials with diminished amplitude and duration are seen, whereas in its chronic phases, mixed neuropathic and myopathic patterns are found. After corticosteroid treatment, EMG tends to return to normal (AU)

[Imaging techniques and electromyography in inflammatory myopathies]. / Estudios de imagen y electromiografía en las miopatías inflamatorias.

Different imaging techniques are used for the diagnosis, evaluation and follow-up of inflammatory myopathies; of these techniques, the ones that provide the largest amount of information are ultrasonography and magnetic resonance imaging. Electrodiagnosis in inflammatory myopathies is based mainly on electromyography (EMG), which shows different patterns according to the different disease presentations. In the acute phase, polyphasic potentials with diminished amplitude and duration are seen, whereas in its chronic phases, mixed neuropathic and myopathic patterns are found. After corticosteroid treatment, EMG tends to return to normal.

Estrategias para el diagnóstico clinico y molecular de Charcot-Marie-Tooth 1A: estudio en pacientes mexicanos / Strategies for clinical and molecular diagnosis of Charcot-Marie-Tooth 1A among Mexican patients

Antecedentes: La neuropatía periférica de Charcot-Marie-Tooth (CMT) es la enfermedad hereditaria más común del sistema nervioso periférico humano. El subtipo más frecuente, CMT1A, es asociado a una duplicación de un fragmento de ~1.5 Mb en 17p11.2-p12, que incluye al gen PMP22. Objetivo: Describir diferentes estrategias para el diagnóstico clínico y molecular de CMT1A en pacientes del Instituto Nacional de Rehabilitación. Material y métodos: A 17 pacientes estudiados clínica y electrofisiológicamente que reunieron los criterios para CMT1, se les realizó el estudio molecular mediante electroforesis capilar para detectar la duplicación del gen PMP22. Resultados: Los estudios clínico, bioquímico y electrofisiológico ofrecieron los criterios para establecer el diagnóstico de CMT1. Con la electroforesis capilar se detectó la duplicación del gen PMP22 en siete pacientes que fueron diagnosticados clínica y electrofisiológicamente como CMT1, pudiendo llegar al diagnóstico de CMT1A. Todas las duplicaciones identificadas fueron corroboradas mediante hibridación in situ fluorescente. Conclusión: Los resultados nos permiten asegurar que la electroforesis capilar es un método fácil y confiable para detectar la duplicación del gen PMP22. Además, el aplicar diferentes estrategias tanto clínicas, electrofisiológicas y moleculares en este tipo de pacientes, nos permitieron establecer el diagnóstico correcto y ofrecer asesoramiento genético adecuado.

BACKGROUND: Charcot-Marie-Tooth (CMT) is the most common inherited disorder of the human peripheral nerve. The mos tfrequent subtype, CMT1A, is associated with duplication of approximately 1.5 Mb fragment in 17p11-p12, that includes the PMP22 gene. OBJECTIVE: The aim of this study was to describe different strategies used for clinical and molecular CNT1A diagnoses among patients attending the National Rehabilitation Institute of Mexico (INR). MATERIAL AND METHODS: 17 patients had clinical and electrophysiological features compatible with CMT1. A molecular study using capillary electrophoresis (CE) was performed and a PMP22 gene duplication was detected RESULTS: Clinical, biochemical and electrophysiological studies constituted the inclusion criteria to establish a CMT1 diagnosis. With CE the duplication of the PMP22 gene was observable and we established a possible CMT1A diagnosis in seven patients. All duplications detected by capillary electrophoresis were corroborated using FISH. CONCLUSION: CE is a feasible and reliable method to detect PMP22 gene duplication. Using different clinical, electrophysiological and molecular strategies in this patient population allowed us to establish an accurate diagnosis and offer suitable genetic counseling.

[Strategies for clinical and molecular diagnosis of Charcot-Marie-Tooth 1A among Mexican patients]. / Estrategias para el diagnóstico clinico y molecular de Charcot-Marie-Tooth 1A. Estudio en pacientes mexicanos.

BACKGROUND: Charcot-Marie-Tooth (CMT) is the most common inherited disorder of the human peripheral nerve. The mos tfrequent subtype, CMT1A, is associated with duplication of approximately 1.5 Mb fragment in 17p11-p12, that includes the PMP22 gene. OBJECTIVE: The aim of this study was to describe different strategies used for clinical and molecular CNT1A diagnoses among patients attending the National Rehabilitation Institute of Mexico (INR). MATERIAL AND METHODS: 17 patients had clinical and electrophysiological features compatible with CMT1. A molecular study using capillary electrophoresis (CE) was performed and a PMP22 gene duplication was detected RESULTS: Clinical, biochemical and electrophysiological studies constituted the inclusion criteria to establish a CMT1 diagnosis. With CE the duplication of the PMP22 gene was observable and we established a possible CMT1A diagnosis in seven patients. All duplications detected by capillary electrophoresis were corroborated using FISH. CONCLUSION: CE is a feasible and reliable method to detect PMP22 gene duplication. Using different clinical, electrophysiological and molecular strategies in this patient population allowed us to establish an accurate diagnosis and offer suitable genetic counseling.

[Peripheral hereditary neuropathies: Charcot-Marie-Tooth types 1 and 2]. / Neuropatías periféricas hereditarias: Charcot-Marie-Tooth tipos 1 y 2.

OBJECTIVE: We report 14 cases of patients with Charcot-Marie-Tooth types 1 (CMT) and type 2 (CMT2). The objective of this study was to determine the diagnostic value of structural studies of peripheral nerve and striated muscle biopsies and to discuss correlations with clinical symptoms and electrophysiologic findings. MATERIAL AND METHODS: Nerve and muscle biopsies were obtained and processed according to standards methods for transmission electron microscopy. Diagnosis of CMT was based on clinical electrophysiologic findings. RESULTS: Our results demonstrated that myelinated fibers showed moderate-to- severe demyelinization and remyelinization, folding complex, tomacula formations, and presence of moderate vacuoles into myelin. The axonal cytoskeleton shows an important decrease of neurofilaments and microtubules in myelinated and unmyelinated nerves. Striated muscle showed different degenerative changes in Z disc. One important finding was presence of crystals inclusions in mitochondrion. In 11 cases, it was possible to find relevant changes in mitochondria-like hyperplasia. In this study, we had four cases with > 20 years of evolution; these cases showed relevant degenerative changes in nerve and muscle. Nerves presented great demyelination with damage in cytoskeleton of the axon, while fibers of muscle had necrosis and hyalinization. CONCLUSIONS: According to our results, it is possible to distinguish morphologically CMT subtypes, due to the fact that in CMT2 we found a 50% decrease of filaments and microtubules that were reduced in myelinic axons; this finding does not occur in CMT1.

Estandarización del método y condiciones de registro de potenciales evocados somatosensoriales en cirugías correctivas de deformidades de la columna vertebral: estudio neurofisiológico en 82 casos quirúrgicos / Standardization of method and record conditions of somatosensorial evoked potentials in corrective surgery for deformities of the spine: neurophysiological study in 82 operated cases

Se han reportado alteraciones neurológicas posteriores a intervenciones quirúrgicas en corrección de deformidades de columna vertebral, debido al riesgo de lesión que se presenta, se ha utilizado la maniobra del "despertar" para valorar la integridad neurológica, siendo ésta insuficiente, por lo que se han establecido técnicas neurofisiológicas como potenciales evocados somatosensoriales (PES) valorando dicha integridad, este método sin ser invasivo, valora la vía somatosensorial evitando el riesgo de lesión medular. Sin embargo se han observado también en ocasiones falsas positivas y negativas con el mismo. Se estudiaron 82 pacientes, promedio de edad 15.4, a los que se les realizó SEP registrando en corteza y cuello y estimulación en tibial bilateral, no observando diferencia significativa en los registros antes y después de la cirugía p > 0.05. En 9 pacientes hubo falsas positivas y en 7 pacientes los potenciales fueron anormales, presentándose manifestaciones neurológicas posquirúrgicas desde parestesias hasta lesión medular. Debido al riesgo de lesión neurológica en dichas intervenciones se debe monitorizar no sólo con SEP la vía sensitiva, sino también la vía motora con algún otro método, para disminuir la incidencia de falsas positivas y negativas, y dar información más confiable al cirujano, para beneficio del paciente.