Experiencias de aprendizaje metacognitivo en estudiantes de enfermería: revisión de alcance / Metacognitive Learning Experiences in Nursing Students: Scoping Review / Experiências de aprendizagem metacognitiva em estudantes de enfermagem: Uma revisão do escopo

Introducción: la metacognición es comprendida como el conocimiento de los propios procesos cognitivos y su autorregulación por parte de los estudiantes. Objetivo: conocer las tendencias de la investigación en torno a la metacognición en los procesos de enseñanza y aprendizaje en programas de Enfermería. Método: se llevó a cabo una revisión de alcance en torno a la metacognición como una estrategia de reflexión de los procesos de enseñanza aprendizaje al interior de los programas de Enfermería. El proceso de búsqueda se realizó en la base de datos Web of Science en el período comprendido entre 2015-2021; la selección de estudios cumplió con los criterios de inclusión. Resultados: el análisis de la información permitió identificar cuatro tendencias: a) la metacognición en procesos de enseñanza y aprendizaje con algunas didácticas grupales, b) la metacognición en procesos de aprendizaje con simulación clínica, c) el aprendizaje reflexivo como estrategia de regulación metacognitiva aplicada a los procesos de evaluación, d) la metacognición en los planes curriculares de los programas académicos en salud. Conclusiones: la revisión evidencia la implementación de metodologías y estrategias metacognitivas en los espacios de formación académica en los programas de Enfermería, que trascienden un modelo educativo tradicional centrado en los contenidos y se dirigen a un modelo centrado en la reflexión consciente y participativa de los estudiantes en el proceso de aprendizaje.

Introduction: Metacognition is understood as the students' knowledge of their own cognitive processes and their self-regulation. Objective: Aknowledge research trends of metacognition in teaching and learning processes in Nursing programs. Method: A scoping review was carried out on metacognition as a strategy for reflection of the teaching-learning processes within Nursing programs. The search process was carried out in the Web of Science database in the period 2015-2021; the selection of studies met the inclusion criteria. Results: The analysis of information allowed the identification of four trends a) metacognition in teaching and learning processes with some group didactics, b) metacognition in learning processes with clinical simulation, c) reflective learning as a metacognitive regulation strategy applied to evaluation processes, d) metacognition in the curricular plans of academic health programs. Conclusions: The review evidences the implementation of metacognitive methodologies and strategies in academic training spaces in Nursing programs, which transcend a traditional educational model focused on contents and moves to a model centered on students' conscious and participative reflection in the learning process.

Introdução: a metacognição é entendida como o conhecimento que os estudantes têm dos seus próprios processos cognitivos e da sua autorregulação. Objetivo: conhecer as tendências da investigação sobre a metacognição nos processos de ensino e aprendizagem nos programas de enfermagem. Método: foi realizada uma revisão de escopo sobre a metacognição como estratégia de reflexão sobre os processos de ensino-aprendizagem nos programas de enfermagem. O processo de busca foi realizado na base de dados Web of Science no período de 2015-2021; Na seleção dos estudos foram atendidos os critérios de inclusão. Resultados: a análise da informação permitiu identificar quatro tendências a) a metacognição nos processos de ensino e aprendizagem com algumas didácticas de grupo, b) a metacognição nos processos de aprendizagem com simulação clínica, c) a aprendizagem reflexiva como estratégia de regulação metacognitiva aplicada aos processos de avaliação, d) a metacognição nos planos curriculares dos programas académicos de saúde.Conclusões: a revisão evidencia a implementação de metodologias e estratégias metacognitivas em espaços de formação académica em programas de Enfermagem, que transcendem de um modelo educativo tradicional centrado nos conteúdos para um modelo centrado na reflexão consciente e participativa dos estudantes no processo de aprendizagem.

Metacognición en el Aprendizaje de la Anatomía / Metacognition in the Learning of Anatomy

RESUMEN: Actualmente la enseñanza y aprendizaje de la Anatomía tiene grandes retos que debe superar: los estudiantes están abrumados por gran cantidad de información; generalmente el aprendizaje está basado en la memoria; la enseñanza esta carente de métodos que permitan hacer consciente el aprendizaje; falta mejorar estrategias evaluativas que demuestren aprendizajes. Por su parte, la metacognición, entendida como el dominio que tiene el sujeto sobre sus propios procesos cognoscitivos busca propiciar en los estudiantes la regulación de sus propios aprendizajes. Este trabajo describe el aporte de la metacognición al aprendizaje de la Anatomía. Esta investigación es de tipo cualitativo. Fue un estudio de caso que analizó cómo un estudiante incorporó la metacognición al aprendizaje de la Anatomía. En el desarrollo del trabajo se indagaron obstáculos de aprendizaje, se diseñaron, aplicaron y analizaron diferentes actividades sobre aprendizaje metacognitivo de la Anatomía. Se incorporó la metacognición al aprendizaje de la Anatomía encontrando que la planeación hace el estudio más organizado, provocando mayor consciencia de cada actividad de aprendizaje y permitiendo obtener mejores resultados. El monitoreo halló que la mayor dificultad en el aprendizaje de la Anatomía se encuentra en la capacidad de orientación espacial. Finalmente la evaluación consideró el progreso en el aprendizaje de la Anatomía justificado en el control del tiempo, en un nuevo lenguaje con términos más adecuados, en la seguridad y confianza frente a las evaluaciones.

SUMMARY: Currently there are important challenges in the teaching and learning of Anatomy. Students are overwhelmed by all of the information and generally, learning is based on memory. Furthermore, teaching instruction lacks methods that encourage cognitive learning. It is necessary to improve evaluation strategies that demonstrate learning. Metacognition, is understood as the command an individual has over their own cognitive processes aims to encourage students to regulate their own learning process. This paper describes the contribution of metacognition to Anatomy learning. This research is qualitative; it was a case study that analyzed how a student incorporated metacognition into Anatomy learning. During the development of this study, learning obstacles were investigated, different activities on metacognitive learning of the Anatomy were designed, applied and analyzed. The research incorporated metacognition into the learning of Anatomy and determined that planning allowed for better study organization, created greater awareness of each learning activity and showed better results. During monitoring, it was found that the greatest difficulty in learning Anatomy, was understanding spatial awareness. Finally the evaluation considered the learning progress of anatomy based on time control, new language, more adequate terms, and greater assurance and confidence during the evaluations.

Bases moleculares de la memoria y su relación con el aprendizaje / Molecular Basis of Memory and its Relation with Learning

El aprendizaje y la memoria son funciones cognitivas estudiadas históricamente desde diferentes campos del saber, de los cuales, podemos mencionar la filosofía, la psicología y las neurociencias. Aportes más recientes provienen de los desarrollos de las ciencias cognitivas y, de manera particular, de la antropología, la lingüística, la semiótica y la inteligencia artificial. En su conjunto, los aportes provenientes de estos diversos campos del conocimiento tienen incidencia importante en la educación. El aprendizaje se refiere al cambio permanente o no del comportamiento, de las ideas, los conceptos, los modelos mentales, los sentimientos, intereses, motivaciones derivados de la experiencia de los sujetos; por su parte, la memoria se refiere al proceso, por el cual ese conocimiento es codificado, almacenado y recordado. El aprendizaje y la memoria son esenciales para el funcionamiento adecuado, adaptación y supervivencia independiente de las diferentes especies animales. En esta revisión, en primer lugar, se presenta una breve reseña de los estudios más sobresalientes sobre aprendizaje y memoria, en términos generales se describen los tipos de aprendizaje y de memoria con relación a la naturaleza de la información y al tiempo de almacenamiento. En segundo lugar, se describen los procesos moleculares que explican el aprendizaje y la memoria implícita y finalmente se presentan algunas reflexiones generales desde el campo conceptual de la educación en relación con los desarrollos presentados en cuanto a la memoria y al aprendizaje...(AU)

Learning and memory are cognitive functions that have been historically studied from different fields of knowledge, namely, philosophy, psychology and neurosciences.Recent contributions come from developments on cognitive sciences and, particularly, anthropology, linguistics, semiotics and artificial intelligence as well. As a whole, contributions from these different fields of knowledge have an important impact on education. Learning refers to a continuous change regarding behavior, ideas, concepts, mental models, feelings, interests, and motivations that are derived from the individuals' experience, while memory encompasses the process by which knowledge is codified, stored and retrieved. Learning and memory are paramount for proper functioning, independent adaptation and survival of different animal species. On one hand, this review shows a brief overview of the most outstanding studies on learning and memory. In general terms, it describes the types of learning and memory in relation to the nature of information and storage time. On the other hand, it presents the molecular processes underlying learning and implicit memory. It finally describes some general reflections from the conceptual perspective on education in relation to the developments of memory and learning...(AU)

Alpha-lipoic acid supplementation protects enzymes from damage by nitrosative and oxidative stress.

BACKGROUND: S-nitrosylation of mitochondrial enzymes involved in energy transfer under nitrosative stress may result in ATP deficiency. We investigated whether α-lipoic acid, a powerful antioxidant, could alleviate nitrosative stress by regulating S-nitrosylation, which could result in retaining the mitochondrial enzyme activity. METHODS: In this study, we have identified the S-nitrosylated forms of subunit 1 of dihydrolipoyllysine succinyltransferase (complex III), and subunit 2 of the α-ketoglutarate dehydrogenase complex by implementing a fluorescence-based differential quantitative proteomics method. RESULTS: We found that the activities of these two mitochondrial enzymes were partially but reversibly inhibited by S-nitrosylation in cultured endothelial cells, and that their activities were partially restored by supplementation of α-lipoic acid. We show that protein S-nitrosylation affects the activity of mitochondrial enzymes that are central to energy supply, and that α-lipoic acid protects mitochondrial enzymes by altering S-nitrosylation levels. CONCLUSIONS: Inhibiting protein S-nitrosylation with α-lipoic acid seems to be a protective mechanism against nitrosative stress. GENERAL SIGNIFICANCE: Identification and characterization of these new protein targets should contribute to expanding the therapeutic power of α-lipoic acid and to a better understanding of the underlying antioxidant mechanisms.

La argumentación metacognitiva en el aula de ciencias / Metacognitive argumentation in the science classroom

Nuestro objetivo central en esta investigación fue describir cualitativamente la categoría argumentación metacognitiva en estudiantes de básica secundaria, con edades comprendidas entre los 14 y los 16 años. Seguimos un enfoque cualitativo en el que diseñamos tres escenarios en perspectiva CTS para la recolección de la información. Los resultados derivados del análisis de la información nos permitieron, a manera de conclusiones, caracterizar tres tendencias en la expresión de la argumentación metacognitiva: a) a partir del sentir-pensar-actuar, b) las centradas en el conocimiento y, c) las centradas en la perspectiva ética de los estudiantes y las estudiantes. Debido al potencial de la categoría aquí investigada en función del logro de aprendizajes en profundidad en ciencias, sería pertinente que futuros estudios profundizaran en la comprensión de las interacciones entre argumentación y metacognición, y enfatizaran en relación a cómo desarrollarla en el aula...

Increased yield of endothelial cells from peripheral blood for cell therapies and tissue engineering.

AIM: Peripheral blood-derived endothelial cells (pBD-ECs) are an attractive tool for cell therapies and tissue engineering, but have been limited by their low isolation yield. We increase pBD-EC yield via administration of the chemokine receptor type 4 antagonist AMD3100, as well as via a diluted whole blood incubation (DWBI). MATERIALS & METHODS: Porcine pBD-ECs were isolated using AMD3100 and DWBI and tested for EC markers, acetylated LDL uptake, growth kinetics, metabolic activity, flow-mediated nitric oxide production and seeded onto titanium tubes implanted into vessels of pigs. RESULTS: DWBI increased the yield of porcine pBD-ECs 6.6-fold, and AMD3100 increased the yield 4.5-fold. AMD3100-mobilized ECs were phenotypically indistinguishable from nonmobilized ECs. In porcine implants, the cells expressed endothelial nitric oxide synthase, reduced thrombin-antithrombin complex systemically and prevented thrombosis. CONCLUSION: Administration of AMD3100 and the DWBI method both increase pBD-EC yield.

Developmental exposure to a commercial PBDE mixture: effects on protein networks in the cerebellum and hippocampus of rats.

BACKGROUND: Polybrominated diphenyl ethers (PBDEs) are structurally similar to polychlorinated biphenyls (PCBs) and have both central (learning and memory deficits) and peripheral (motor dysfunction) neurotoxic effects at concentrations/doses similar to those of PCBs. The cellular and molecular mechanisms for these neurotoxic effects are not fully understood; however, several studies have shown that PBDEs affect thyroid hormones, cause oxidative stress, and disrupt Ca2+-mediated signal transduction. Changes in these signal transduction pathways can lead to differential gene regulation with subsequent changes in protein expression, which can affect the development and function of the nervous system. OBJECTIVE: In this study, we examined the protein expression profiles in the rat cerebellum and hippocampus following developmental exposure to a commercial PBDE mixture, DE-71. METHODS: Pregnant Long-Evans rats were dosed perinatally with 0 or 30.6 mg/kg/day of DE-71 from gestation day 6 through sampling on postnatal day 14. Proteins from the cerebellum and hippocampus were extracted, expression differences were detected by two-dimensional difference gel electrophoresis, and proteins were identified by tandem mass spectrometry. Protein network interaction analysis was performed using Ingenuity® Pathway Analysis, and the proteins of interest were validated by Western blotting. RESULTS: Four proteins were significantly differentially expressed in the cerebellum following DE-71 exposure, whereas 70 proteins were significantly differentially expressed in the hippocampus. Of these proteins, 4 from the cerebellum and 47 from the hippocampus, identifiable by mass spectrometry, were found to have roles in mitochondrial energy metabolism, oxidative stress, apoptosis, calcium signaling, and growth of the nervous system. CONCLUSIONS: Results suggest that changes in energy metabolism and processes related to neuroplasticity and growth may be involved in the developmental neurotoxicity of PBDEs.

Differentially charged isoforms of apolipoprotein E from human blood are potential biomarkers of Alzheimer's disease.

INTRODUCTION: Alzheimer's disease (AD) is the major cause of dementia among the elderly. Finding blood-based biomarkers for disease diagnosis and prognosis is urgently needed. METHODS: We studied protein distributions in brain tissues, cerebrospinal fluid (CSF), and blood of AD patients by using proteomics and a new proteomic method that we call "2D multiplexed Western blot" (2D mxWd). This method allows us to determine in multiple samples the electrophoretic patterns of protein isoforms with different isoelectric points. RESULTS: Apolipoprotein E (ApoE) displays a unique distribution of electrophoretic isoforms in the presence of AD and also a unique pattern specific to the APOE genotype. CONCLUSIONS: The isoelectric distribution of differentially charged ApoE isoforms was used to determine the presence of AD in a small group of samples. Further studies are needed to validate their use as predictors of disease onset and progression, and as biomarkers for determining the efficacy of therapeutic treatments.

Polymerase I and transcript release factor (PTRF) regulates adipocyte differentiation and determines adipose tissue expandability.

Impaired adipogenesis renders an adipose tissue unable to expand, leading to lipotoxicity and conditions such as diabetes and cardiovascular disease. While factors important for adipogenesis have been studied extensively, those that set the limits of adipose tissue expansion remain undetermined. Feeding a Western-type diet to apolipoprotein E2 knock-in mice, a model of metabolic syndrome, produced 3 groups of equally obese mice: mice with normal glucose tolerance, hyperinsulinemic yet glucose-tolerant mice, and prediabetic mice with impaired glucose tolerance and reduced circulating insulin. Using proteomics, we compared subcutaneous adipose tissues from mice in these groups and found that the expression of PTRF (polymerase I and transcript release factor) associated selectively with their glucose tolerance status. Lentiviral and pharmacologically overexpressed PTRF, whose function is critical for caveola formation, compromised adipocyte differentiation of cultured 3T3-L1cells. In human adipose tissue, PTRF mRNA levels positively correlated with markers of lipolysis and cellular senescence. Furthermore, a negative relationship between telomere length and PTRF mRNA levels was observed in human subcutaneous fat. PTRF is associated with limited adipose tissue expansion underpinning the key role of caveolae in adipocyte regulation. Furthermore, PTRF may be a suitable adipocyte marker for predicting pathological obesity and inform clinical management.

α-Lipoic acid protects mitochondrial enzymes and attenuates lipopolysaccharide-induced hypothermia in mice.

Hypothermia is a key symptom of sepsis, but the mechanism(s) leading to hypothermia during sepsis is largely unknown and thus no effective therapy is available for hypothermia. Therefore, it is important to investigate the mechanism and develop effective therapeutic methods. Lipopolysaccharide (LPS)-induced hypothermia accompanied by excess nitric oxide (NO) production leads to a reduction in energy production in wild-type mice. However, mice lacking inducible nitric oxide synthase did not suffer from LPS-induced hypothermia, suggesting that hypothermia is associated with excess NO production during sepsis. This observation is supported by the treatment of wild-type mice with α-lipoic acid (LA) in that it effectively attenuates LPS-induced hypothermia with decreased NO production. We also found that LA partially restored ATP production, and activities of the mitochondrial enzymes involved in energy metabolism, which were inhibited during sepsis. These data suggest that hypothermia is related to mitochondrial dysfunction, which is probably compromised by excess NO production and that LA administration attenuates hypothermia mainly by protecting mitochondrial enzymes from NO damage.

PPARα activation induces N(ε)-Lys-acetylation of rat liver peroxisomal multifunctional enzyme type 1.

Peroxisomes are ubiquitous subcellular organelles that participate in metabolic and disease processes, with few of its proteins undergoing posttranslational modifications. As the role of lysine-acetylation has expanded into the cellular intermediary metabolism, we used a combination of differential centrifugation, organelle isolation by linear density gradient centrifugation, western blot analysis, and peptide fingerprinting and amino acid sequencing by mass spectrometry to investigate protein acetylation in control and ciprofibrate-treated rat liver peroxisomes. Organelle protein samples isolated by density gradient centrifugation from PPARα-agonist treated rat liver screened with an anti-N(ε)-acetyl lysine antibody revealed a single protein band of 75 kDa. Immunoprecipitation with this antibody resulted in the precipitation of a protein from the protein pool of ciprofibrate-induced peroxisomes, but not from the protein pool of non-induced peroxisomes. Peptide mass fingerprinting analysis identified the protein as the peroxisomal multifunctional enzyme type 1. In addition, mass spectrometry-based amino acid sequencing resulted in the identification of unique peptides containing 4 acetylated-Lys residues (K¹55, K¹7³, K¹9°, and K58³). This is the first report that demonstrates posttranslational acetylation of a peroxisomal enzyme in PPARα-dependent proliferation of peroxisomes in rat liver.

Diggin' on u(biquitin): a novel method for the identification of physiological E3 ubiquitin ligase substrates.

The ubiquitin-proteasome system (UPS) plays a central role in maintaining protein homeostasis, emphasized by a myriad of diseases that are associated with altered UPS function such as cancer, muscle-wasting, and neurodegeneration. Protein ubiquitination plays a central role in both the promotion of proteasomal degradation as well as cellular signaling through regulation of the stability of transcription factors and other signaling molecules. Substrate-specificity is a critical regulatory step of ubiquitination and is mediated by ubiquitin ligases. Recent studies implicate ubiquitin ligases in multiple models of cardiac diseases such as cardiac hypertrophy, atrophy, and ischemia/reperfusion injury, both in a cardioprotective and maladaptive role. Therefore, identifying physiological substrates of cardiac ubiquitin ligases provides both mechanistic insights into heart disease as well as possible therapeutic targets. Current methods identifying substrates for ubiquitin ligases rely heavily upon non-physiologic in vitro methods, impeding the unbiased discovery of physiological substrates in relevant model systems. Here we describe a novel method for identifying ubiquitin ligase substrates utilizing tandem ubiquitin binding entities technology, two-dimensional differential in gel electrophoresis, and mass spectrometry, validated by the identification of both known and novel physiological substrates of the ubiquitin ligase MuRF1 in primary cardiomyocytes. This method can be applied to any ubiquitin ligase, both in normal and disease model systems, in order to identify relevant physiological substrates under various biological conditions, opening the door to a clearer mechanistic understanding of ubiquitin ligase function and broadening their potential as therapeutic targets.

Cardiac mitochondrial proteomic expression in inbred rat strains divergent in survival time after hemorrhage.

We have previously identified inbred rat strains differing in survival time to a severe controlled hemorrhage (StaH). In efforts to identify cellular mechanisms and ultimately genes that are important contributors to enhanced STaH, we conducted a study to characterize potential differences in cardiac mitochondrial proteins in these rats. Inbred rats from three strains [Brown Norway/Medical College of Wisconsin (BN); Dark Agouti (DA), and Fawn Hooded Hypertensive (FHH)] with different StaH (DA = FHH > BN) were assigned to one of three treatment groups (n = 4/strain): nonoperated controls, surgically catheterized rats, or rats surgically catheterized and hemorrhaged 24 h postsurgery. Rats were euthanized 30 min after handling or 30 min after initiation of a 26 min hemorrhage. After euthanasia, hearts were removed and mitochondria isolated. Differential protein expression was determined using 2D DIGE-based Quantitative Intact Proteomics and proteins identified by MALDI/TOF mass spectrometry. Hundreds of proteins (791) differed among inbred rat strains (P ≤ 0.038), and of these 81 were identified. Thirty-eight were unique proteins and 43 were apparent isoforms. For DA rats (longest STaH), 36 proteins increased and 30 decreased compared with BN (shortest STaH). These 81 proteins were associated with lipid (e.g., acyl CoA dehydrogenase) and carbohydrate (e.g., fumarase) metabolism, oxidative phosphorylation (e.g., ubiquinol-cytochrome C reductase), ATP synthesis (F1 ATPase), and H2S synthesis (3-mercaptopyruvate sulfurtransferase). Although we cannot make associations between these identified mitochondrial proteins and StaH, our data do provide evidence for future candidate proteins with which to consider such associations.

Diabetes mellitus in patients with Alzheimer's disease: clinical description and correlation with the APOE genotype in a sample population from the province of Antioquia, Colombia.

INTRODUCTION: Alzheimer's disease is a multifactorial disease affecting approximately twenty million people worldwide. Numerous variables are associated with increased risk of developing this severe neurological disorder. Among the risk factors, diabetes mellitus, and the ε4 isoform of the APOE gene have been amply demonstrated as increasing the risk of developing this disease. OBJECTIVE: To determine if a correlation exists between APOE genotype, diabetes mellitus and Alzheimer's disease. MATERIALS AND METHODS: Clinical studies were carried out by surveying the clinical histories in a group of patients in the province of Antioquia, Colombia. Forty-three Alzheimer's patients were compared with 43 control subjects, paired by age and gender. Commercially available methods were used to determine whether the patients had diabetes, and restriction enzyme-based genotyping was used to determine the APOE genotypes. RESULTS: The most common non-neurological comorbidities were: arterial hypertension, acute myocardial infarction, chronic obstructive pulmonary disease and hypothyroidism. From the many variables investigated, two were conclusive: (1) the presence of Alzheimer's disease was higher in patients with diabetes mellitus, and (2) no correlation between late-onset sporadic Alzheimer's disease and APOE was found in the target population. CONCLUSIONS: To detect any association with the APOE genotype, a study involving much a larger population samples must be undertaken.

CLN5 and CLN8 protein association with ceramide synthase: biochemical and proteomic approaches.

Four patients with juvenile neuronal ceroid lipofuscinoses, a childhood neurodegenerative disorder that was previously described as CLN9 variant, are reclassified as CLN5 disease. CLN5-deficient (CLN5(-/-) ) fibroblasts demonstrate adhesion defects, increased growth, apoptosis, and decreased levels of ceramide, sphingomyelin, and glycosphingolipids. The CLN8 protein (CLN8p) corrects growth and apoptosis in CLN5(-/-) cells. Related proteins containing a Lag1 motif (CerS1/2/4/5/6) partially corrected these deficits, with CerS1, which is primarily expressed in brain, providing the best complementation, suggesting CLN5p activates CerS1 and may co-immunoprecipitate with it. CLN8p complements CLN5-deficient cells, consolidating the interrelationship of CLN5p/CLN8p, whose potential roles are explored as activators of (dihydro)ceramide synthases. Homozygosity mapping using microarray technology led to identification of CLN5 as the culprit gene in previously classified CLN9-defective cases. Similar to CLN5(-/-) cells, ceramide synthase activity, C16/C18:0/C24:0/C24:1 ceramide species, measured by MS is decreased in CLN8(-/-) cells. Comparison of normal versus CLN5(-/-) cell CerS1-bound proteins by immunoprecipitation, differential gel electrophoresis, and MS revealed absence of γ-actin in CLN5(-/-) cells. The γ-actin gene sequence is normal in CLN5(-/-) derived DNA. The γ-actin-bound proteins, vimentin and histones H2Afz/H3F3A/Hist1H4, were absent from the γ-actin protein complex in CLN5(-/-) cells. The function of CLN5p may require vimentin and the histone proteins to bind γ-actin. Defective binding could explain the CLN5(-/-) cellular phenotype. We explore the role of the CLN5/CLN8 proteins in ceramide species specific sphingolipid de novo synthesis, and suggest that CLN5/CLN8 proteins are more closely related than previously believed.

Proteomic analysis of mice expressing human ApoE demonstrates no differences in global protein solubility between APOE 3 and APOE 4 young mice.

Apolipoprotein E (ApoE) is a major lipid carrier protein. In humans, ApoE is expressed in three polymorphic isoforms, which are encoded by three different alleles APOE2, APOE3, and APOE4. In the brains of Alzheimer's disease (AD) patients, each one of these three allelic isoforms is found in several "isoelectric" protein isoforms (qPI), i.e. protein isoforms resulting from PTMs altering the net charge (q) of the polypeptide. AD is a complex disease in which multiple causes and several risk factors affect the onset and disease outcome. A major risk factor for AD is ApoE4; therefore, it is important to characterize the different ApoE qPIs. We have implemented a detergent-based method for isolation and quantitation of protein isoforms, and we found differences in the solubility of protein isoforms depending on the type of solvent used. In this manuscript, we describe these methods and applied them to young human-ApoE targeted replacement mice. Our results indicate that there are no significant differences in the hippocampus proteome of these mice as a function of the APOE genotype.

Diabetes mellitus in patients withAlzheimer´s disease: clinical descriptionand correlation with the APOEgenotype in a sample population from the province of Antioquia, Colombia / Diabetes mellitus en pacientes con enfermedad de Alzheimer: descripción clínica y correlación con el genotipo APOE en una muestra de población del departamento de Antioquia, Colombia

Introduction. Alzheimer´s disease is a multifactorial disease affecting approximately twenty million people worldwide. Numerous variables are associated with increased risk of developing this severe neurological disorder. Among the risk factors, diabetes mellitus, and the ε4 isoform of the APOE gene have been amply demonstrated as increasing the risk of developing this disease. Objective. To determine if a correlation exists between APOE genotype, diabetes mellitus and Alzheimer´s disease. Materials and methods. Clinical studies were carried out by surveying the clinical histories in a group of patients in the province of Antioquia, Colombia. Forty-three Alzheimer´s patients were compared with 43 control subjects, paired by age and gender. Commercially available methods were used to determine whether the patients had diabetes, and restriction enzyme-based genotyping was used to determine the APOE genotypes. Results. The most common non-neurological comorbidities were: arterial hypertension, acute myocardial infarction, chronic obstructive pulmonary disease and hypothyroidism. From the many variables investigated, two were conclusive: (1) the presence of Alzheimer´s disease was higher in patients with diabetes mellitus, and (2) no correlation between late-onset sporadic Alzheimer´s disease and APOE was found in the target population. Conclusions. To detect any association with the APOE genotype, a study involving much a larger population samples must be undertaken.

Introducción. La enfermedad de Alzheimer es compleja y afecta, aproximadamente, a 20 millones de personas en todo el mundo. Muchas variables parecen aumentar el riesgo de desarrollar esta alteración neurológica. Entre los factores de riesgo, se ha demostrado ampliamente que la diabetes mellitus y la isoforma ε4 del gen APOE tienen incidencia positiva en el desarrollo de la enfermedad. Se reporta un estudio en el cual se investigó la posible correlación entre APOE, diabetes mellitus y la enfermedad de Alzheimer, en un grupo específico de pacientes del departamento de Antioquia, Colombia. Objetivo. Determinar si existe una correlación entre APOE, diabetes mellitus y la enfermedad de Alzheimer, en un grupo de pacientes de Antioquia, Colombia. Materiales y métodos. Se buscaron y analizaron las historias clínicas de los pacientes con diagnóstico de enfermedad de Alzheimer. Se seleccionaron aquellos que cumplían los criterios de inclusión. Se utilizaron métodos comercialmente disponibles para confirmar la presencia de diabetes mellitus. La genotipificación de APOE se hizo con un método basado en la PCR y la digestión con enzimas de restricción, en muestras de todos los participantes en el estudio. Resultados. En este estudio se analizan 43 casos de enfermedad de Alzheimer y 43 individuos sanos controles, pareados por edad y sexo. Las enfermedades concomitantes no neurológicas más comunes fueron: hipertensión arterial, infarto agudo del miocardio, enfermedad pulmonar obstructiva crónica e hipotiroidismo. Conclusiones. De las diferentes variables investigadas, dos arrojaron resultados concluyentes: i) la presencia de la enfermedad de Alzheimer es más frecuente en pacientes con diabetes mellitus, y 2) no se encontró correlación entre la enfermedad de Alzheimer de inicio tardío esporádico y el genotipo de APOE. Es importante indicar que debe llevarse a cabo un estudio con un tamaño de población mayor, para determinar cualquier posible correlación o inferencia con el genotipo de APOE.

Acute phase response, inflammation and metabolic syndrome biomarkers of Libby asbestos exposure.

Identification of biomarkers assists in the diagnosis of disease and the assessment of health risks from environmental exposures. We hypothesized that rats exposed to Libby amphibole (LA) would present with a unique serum proteomic profile which could help elucidate epidemiologically-relevant biomarkers. In four experiments spanning varied protocols and temporality, healthy (Wistar Kyoto, WKY; and F344) and cardiovascular compromised (CVD) rat models (spontaneously hypertensive, SH; and SH heart failure, SHHF) were intratracheally instilled with saline (control) or LA. Serum biomarkers of cancer, inflammation, metabolic syndrome (MetS), and the acute phase response (APR) were analyzed. All rat strains exhibited acute increases in α-2-macroglobulin, and α1-acid glycoprotein. Among markers of inflammation, lipocalin-2 was induced in WKY, SH and SHHF and osteopontin only in WKY after LA exposure. While rat strain- and age-related changes were apparent in MetS biomarkers, no LA effects were evident. The cancer marker mesothelin was increased only slightly at 1 month in WKY in one of the studies. Quantitative Intact Proteomic profiling of WKY serum at 1 day or 4 weeks after 4 weekly LA instillations indicated no oxidative protein modifications, however APR proteins were significantly increased. Those included serine protease inhibitor, apolipoprotein E, α-2-HS-glycoprotein, t-kininogen 1 and 2, ceruloplasmin, vitamin D binding protein, serum amyloid P, and more 1 day after last LA exposure. All changes were reversible after a short recovery regardless of the acute or long-term exposures. Thus, LA exposure induces an APR and systemic inflammatory biomarkers that could have implications in systemic and pulmonary disease in individuals exposed to LA.

Analysis of proteins using DIGE and MALDI mass spectrometry.

Difference gel electrophoresis (DIGE) is a common technique for characterizing differential protein expression in quantitative proteomics. Usually a combination of enzymatic digestion and peptide analysis by mass spectrometry is used to identify differentially expressed proteins following separation and statistical analysis by DIGE. In this chapter, methods for gel spot picking, enzymatic digestion, and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) for protein identification of DIGE-analyzed proteins are discussed. Two examples are given: first, a specific protein is used to test the sensitivity of the 2D DIGE/MALDI MS combination for protein quantification and identification, and second, several proteins with and without the labels typically used in DIGE are identified to demonstrate that these labels do not alter MS-based protein identification. Technical variations of protein gel spot preparation, in-gel digestion, and mass spectral protein identification are discussed.

Analysis of protein posttranslational modifications using DIGE-based proteomics.

Difference gel electrophoresis (DIGE) is most often used to assess relative changes in the expression levels of individual proteins in multiple complex samples, and this information is valuable in making inferences about relative protein activity. However, a protein's activity is not solely dependent upon its expression level. A change in activity may also be influenced by myriad posttranslational modifications (PTMs), including palmitoylation, ubiquitination, oxidation, and phosphorylation. In this chapter, we describe the use of DIGE to determine specific PTMs by introducing specific labels or changes in pI and/or molecular weight.