Quantification of Mixed-Linkage ß-Glucan (MLG) in Bacteria.

Prokaryotes are known to produce and secrete a broad range of biopolymers with a high functional and structural heterogeneity, often with critical duties in the bacterial physiology and ecology. Among these, exopolysaccharides (EPS) play relevant roles in the interaction of bacteria with eukaryotic hosts. EPS can help to colonize the host and assist in bacterial survival, making this interaction more robust by facilitating the formation of structured biofilms. In addition, they are often key molecules in the specific recognition mechanisms involved in both beneficial and pathogenic bacteria-host interactions. A novel EPS known as MLG (Mixed-Linkage ß-Glucan) was recently discovered in rhizobia, where it participates in bacterial aggregation and biofilm formation and is required for efficient attachment to the roots of their legume host plants. MLG is the first and, so far, the only reported linear Mixed-Linkage ß-glucan in bacteria, containing a perfect alternation of ß (1 â†’ 3) and ß (1 â†’ 4) bonds. A phylogenetic study of MLG biosynthetic genes suggests that far from being exclusive of rhizobia, different soil and plant-associated bacteria likely produce MLG, adding this novel polymer to the plethora of surface polysaccharides that help bacteria thrive in the changing environment and to establish successful interactions with their hosts.In this work, a quantification method for MLG is proposed. It relays on the hydrolysis of MLG by a specific enzyme (lichenase), and the subsequent quantification of the released disaccharide (laminaribiose) by the phenol-sulfuric acid method. The protocol has been set up and optimized for its use in 96-well plates, which makes it suitable for high-throughput screening (HTS) approaches. This method stands out by its fast processing, technical simplicity, and capability to handle multiple samples and biological replicates at a time.

Store-Operated Calcium Entry Inhibition and Plasma Membrane Calcium Pump Upregulation Contribute to the Maintenance of Resting Cytosolic Calcium Concentration in A1-like Astrocytes.

Highly neurotoxic A1-reactive astrocytes have been associated with several human neurodegenerative diseases. Complement protein C3 expression is strongly upregulated in A1 astrocytes, and this protein has been shown to be a specific biomarker of these astrocytes. Several cytokines released in neurodegenerative diseases have been shown to upregulate the production of amyloid ß protein precursor (APP) and neurotoxic amyloid ß (Aß) peptides in reactive astrocytes. Also, aberrant Ca2+ signals have been proposed as a hallmark of astrocyte functional remodeling in Alzheimer's disease mouse models. In this work, we induced the generation of A1-like reactive astrocytes after the co-treatment of U251 human astroglioma cells with a cocktail of the cytokines TNF-α, IL1-α and C1q. These A1-like astrocytes show increased production of APP and Aß peptides compared to untreated U251 cells. Additionally, A1-like astrocytes show a (75 ± 10)% decrease in the Ca2+ stored in the endoplasmic reticulum (ER), (85 ± 10)% attenuation of Ca2+ entry after complete Ca2+ depletion of the ER, and three-fold upregulation of plasma membrane calcium pump expression, with respect to non-treated Control astrocytes. These altered intracellular Ca2+ dynamics allow A1-like astrocytes to efficiently counterbalance the enhanced release of Ca2+ from the ER, preventing a rise in the resting cytosolic Ca2+ concentration.

Sorcin Activates the Brain PMCA and Blocks the Inhibitory Effects of Molecular Markers of Alzheimer's Disease on the Pump Activity.

Since dysregulation of intracellular calcium (Ca2+) levels is a common occurrence in neurodegenerative diseases, including Alzheimer's disease (AD), the study of proteins that can correct neuronal Ca2+ dysregulation is of great interest. In previous work, we have shown that plasma membrane Ca2+-ATPase (PMCA), a high-affinity Ca2+ pump, is functionally impaired in AD and is inhibited by amyloid-ß peptide (Aß) and tau, two key components of pathological AD hallmarks. On the other hand, sorcin is a Ca2+-binding protein highly expressed in the brain, although its mechanism of action is far from being clear. Sorcin has been shown to interact with the intracellular sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), and other modulators of intracellular Ca2+ signaling, such as the ryanodine receptor or presenilin 2, which is closely associated with AD. The present work focuses on sorcin in search of new regulators of PMCA and antagonists of Aß and tau toxicity. Results show sorcin as an activator of PMCA, which also prevents the inhibitory effects of Aß and tau on the pump, and counteracts the neurotoxicity of Aß and tau by interacting with them.

Diez años del Registro Uruguayo de Peritonitis en Diálisis Peritoneal / Ten years of the Uruguayan Registry of Peritonitis in Peritoneal Dialysis

La peritonitis es una complicación grave de la diálisis peritoneal (DP), por lo que interesa conocer la incidencia y sensibilidad antibiótica de los gérmenes causantes. En Uruguay, desde el 1° de enero de 2004, se realiza un registro nacional de las peritonitis en DP, gérmenes, sensibilidad y evolución. Método: se analizaron los registros desde el 1° de enero de 2004 al 31 de diciembre de 2013. El registro fue aprobado por comités de ética institucionales. Resultados: en el período se registraron 850 peritonitis, con una incidencia que descendió de 0,49/paciente-año (2004-2005) a 0,37/paciente-año (2013). La incidencia de Staphylococcus aureus y Staphylococcus coagulasa negativo (SCoN) fue menor en 2009-2013 vs 2004-2005 (0,2 vs 0,12 peritonitis/paciente-año, test Poisson p<0,05). En 2009-2013: 14/54 S. aureus y 26/71 SCoN fueron meticilinorresistente, similar al período previo. El 98% de los gérmenes gramnegativos fueron sensibles a amikacina. En 145/467 (31%) episodios no se identificó germen. Se logró cura primaria en 71% de las peritonitis por grampositivos y en 45% por gramnegativos (chi2 p<0,05). En 2013 se observó mayor incidencia de peritonitis en los centros en los que no se controló el estado de portador nasal. Comentarios y conclusiones: se justifica implementar el control de portador de Staphylococcus aureus. La incidencia de peritonitis por S. aureus y SCoN meticilinorresistentes, la incidencia sostenida de gérmenes gramnegativos (con peor evolución), y el elevado porcentaje de cultivos sin desarrollo justifica mantener el protocolo antibiótico empírico inicial con vancomicina y amikacina. El descenso de la incidencia de S. aureus + SCoN podría ser atribuido a una mejor educación de los pacientes en DP.

Abstract Peritonitis is a severe complication of peritoneal dialysis (PD), so it is important to learn about the incidence and antibiotic sensitivity of the germs that cause it. In Uruguay, since January 1, 2004, a national record is kept for peritonitis in PD, germs, sensitivity and evolution. Method: the records from January 1, 2004 through December 31, 2013 were analyzed. The registry was approved by institutional ethical committes. Results: during the above mentioned period, 850 cases of peritonitis were recorded, and incidence dropped from 0.49/patient-year (2004-2005) to 0.37/patient-year (2013). Incidence of Staphylococcus aureus and coagulase-negative staphylococci (SCoN) was lower in 2009-2013 vs 2004-2005 (0.2 vs 0.12 peritonitis/patient-year, test Poisson p<0.05). In 2009-2013: 14/54 S. aureus and 26/71 SCoN were methicillin-resistant, similar to the previous period. 98% of Gram-negative were sensitive to amikacin. No germ was identified in 145/467 (31%) of episodes. Primary cure was achieved in 71% of peritonitis for Gram-positive and 45% for Gram-negative bacteria (chi2 p<0.05). In 2013 a greater incidence of peritonitis was observed in those centers where the nasal carriage was not controlled. Comments and conclusions: controlling Staphylococcus aureus nasal carriages is worth doing. The incidence of peritonitis by methicillin-resistant S. aureus y SCoN, the sustained incidence of Gram-negative germs (with a worse evolution), and the high percentage of cultures with no development justify keeping the initial empirical antibiotic protocol with vancomycin and amikacin. Reduction in the incidence of S. aureus + SCoN could be explained by a greater education in PD patients.

Resumo A peritonite é uma complicação grave da diálise peritoneal (DP), sendo, portanto, importante conhecer a incidência e a sensibilidade antibiótica dos gérmens causadores. No Uruguai, desde 1 de janeiro de 2004, realiza-se um registro nacional das peritonites em DP, com dados sobre gérmens, sensibilidade e evolução. Método: foram analisados os registros do período 1 de janeiro de 2004 - 31 de dezembro de 2013. O registro foi aprovado pelos comitês de ética das instituições envolvidas. Resultados: foram registradas 850 peritonites no período estudado; a incidência diminuiu de 0,49/paciente-ano no período 2004-2005 a 0,37/paciente-ano em 2013. A incidência de Staphylococcus aureus e Staphylococcus coagulase negativo (SCoN) foi menor no período 2009-2013 comparada com 2004-2005 (0,2 vs 0,12 peritonite/paciente-ano, teste de Poisson p<0,05). No período 2009-2013: 14/54 S. aureus e 26/71 SCoN foram resistentes à meticilina, similar ao período prévio. 98% dos gérmens gramnegativos eram sensíveis a amicacina. Não se pode identificar o gérmen em 145/467 (31%) episódios. Em 71% das peritonites por grampositivos e em 5% por gramnegativos (chi2 p<0,05) foi possível obter cura primária. Em 2013 foi observada uma maior incidência de peritonite nos centros em que não se realizava controle de portador nasal. Comentários e conclusões: justifica-se a realização de controle de portador de Staphylococcus aureus. A incidência de peritonite por S. aureus e SCoN resistentes à meticilina, a incidência constante de gérmens gramnegativos (com pior evolução), e a alta porcentagem de cultivos sem crescimento justificam manter o protocolo antibiótico empírico inicial com vancomicina e amicacina. A redução da incidência de S. aureus + SCoN poderia ser atribuída a melhor educação dos pacientes em DP.

Deterioro funcional asociado a la hospitalización en pacientes mayores de 65 años / Hospitalisation as a risk for functional decline in older adults

El deterioro funcional es un riesgo asociado a la hospitalización en las personas mayores, con elevada prevalencia (35-70%) y graves consecuencias. OBJETIVO: Determinar la incidencia del deterioro funcional relacionado con la hospitalización (DFH) en personas mayores ingresadas en el Área Médica del Complejo Hospitalario Universitario de Albacete. MÉTODO: Se ha realizado un estudio de cohortes, cuya variable principal ha sido el DFH (pérdida de capacidad para realizar actividades cotidianas durante el ingreso y tras el alta); se han incluido otras variables (demográficas, relacionadas con el ingreso, patologías asociadas, estado cognitivo). Los datos se recogieron por entrevista presencial al paciente y cuidadores, llamada telefónica tras el alta y revisión de historia clínica. RESULTADOS: Se incluyeron 104 pacientes, de los que un 51,9% eran mujeres; la edad media fue 79,97 años (dt = 7,89), IC 95% [78,43; 81,5] y la estancia media de 10,11 días (dt = 7,65) IC 95% [8,62; 11,6]. El primer día del ingreso 43 (41,4%) pacientes presentaban estado mental intacto. Se produjo DFH en las primeras 24 horas en 60 (57,7%) pacientes; tras el alta hospitalaria, el deterioro existía en un 32,6% de los 92 pacientes que pudieron ser evaluados. En pacientes que antes del ingreso eran independientes para las actividades cotidianas, un 19% quedó con dependencia importante. El DFH se relacionó de forma estadísticamente significativa con los antecedentes de caídas y la edad. CONCLUSIONES: El DFH se produce en un porcentaje elevado de los mayores de 65 años; entre los previamente independientes, casi la quinta parte queda en situación de dependencia

Functional decline: (FD) is a risk associated with hospital admission in older people, due to its high prevalence (35-70%) and its serious consequences. AIM: To determine the incidence of FD in the elderly after hospital admission at the Geriatric and Internal Medicine wards of a tertiary teaching hospital (Albacete, Spain). METHOD: A cohort study has been designed, whose primary focus was FD, defined as the loss of independence to perform the activities of daily living between preadmission status and discharge. Demographic characteristics, comorbidity, length of hospital stay and cognitive status have been analysed. Data collection was performed by interviews with patients and caregivers during hospitalization and after discharge (by phone), as well as by revision of clinical records. RESULTS: 104 patients were evaluated, of which 51.9% were female; the average age was 79.97 years (dt = 7.89) IC 95% [78.43, 81.5] and the average length of stay was 10.11 days (dt = 7.65) IC 95% [8.62,11.6]. The proportion of patients who showed a normal cognitive status on the first in-hospital day was 41.4% (43 patients). FD was present in 60 (57.7%) patients in the first day of hospitalisation; when discharged, 32.6% of 92 patients who could be evaluated showed FD.19% of patients who were previously independent in activities of daily living developed a serious dependence after discharge. FD was associated statistically with age and a history of previous falls. CONCLUSIONS: FD takes place in a high percentage of the elderly PATIENTS: Among the previously independent patients, 19% remains in a situation of dependence after discharge

[Hospitalisation as a risk for functional decline in older adults]. / Deterioro funcional asociado a la hospitalización en pacientes mayores de 65 años.

UNLABELLED: FUNCTIONAL DECLINE: (FD) is a risk associated with hospital admission in older people, due to its high prevalence (35-70%) and its serious consequences. AIM: To determine the incidence of FD in the elderly after hospital admission at the Geriatric and Internal Medicine wards of a tertiary teaching hospital (Albacete, Spain). METHOD: A cohort study has been designed, whose primary focus was FD, defined as the loss of independence to perform the activities of daily living between preadmission status and discharge. Demographic characteristics, comorbidity, length of hospital stay and cognitive status have been analysed. Data collection was performed by interviews with patients and caregivers during hospitalization and after discharge (by phone), as well as by revision of clinical records. RESULTS: 104 patients were evaluated, of which 51.9% were female; the average age was 79.97 years (dt=7.89) IC 95% [78.43, 81.5] and the average length of stay was 10.11 days (dt=7.65) IC 95% [8.62,11.6]. The proportion of patients who showed a normal cognitive status on the first in-hospital day was 41.4% (43 patients). FD was present in 60 (57.7%) patients in the first day of hospitalisation; when discharged, 32.6% of 92 patients who could be evaluated showed FD. 19% of patients who were previously independent in activities of daily living developed a serious dependence after discharge. FD was associated statistically with age and a history of previous falls. CONCLUSIONS: FD takes place in a high percentage of the elderly patients. Among the previously independent patients, 19% remains in a situation of dependence after discharge.

Using a multidisciplinary training program to reduce peritonitis in peritoneal dialysis patients.

OBJECTIVES: The present study evaluated the tool used to assess patients' skills and the impact on peritonitis rates of a new multidisciplinary peritoneal dialysis (PD) education program (PDEP). METHODS: After the University Hospital Ethics Committee approved the study, the educational and clinical records of PD patients were retrospectively analyzed in two phases. In phase I, an Objective Structured Assessment (OSA) was used during August 2008 to evaluate the practical skills of 25 patients with adequate Kt/V and no mental disabilities who had been on PD for more than 1 month. Test results were correlated with the prior year's peritonitis rate. In phase II, the new PDEP, consisting of individual lessons, a retraining schedule, and group meetings, was introduced starting 1 September 2008. Age, sex, years of education, time on PD, number of training sessions, and peritonitis episodes were recorded. Statistical analyses used t-tests, chi-square tests, and Poisson distributions; a p value of less than 0.05 was considered significant. RESULTS: In phase I, 25 patients [16 men, 9 women; mean age: 54 ± 15 years (range: 22 - 84 years); mean time on PD: 35 ± 30 months (range: 1 - 107 months)] were studied. The OSA results correlated with peritonitis rates: patients who passed the test had experienced significantly lower peritonitis rates during the prior year (p < 0.05). In phase II, after the new PDEP was introduced, overall peritonitis rates significantly declined (to 0.28 episodes/patient-year from 0.55 episodes/patient-year, p < 0.05); the Staphylococcus peritonitis rate also declined (to 0.09 episodes/patient-year from 0.24 episodes/patient-year, p < 0.05). CONCLUSIONS: The OSA is a reliable tool for assessing patients' skills, and it correlates with peritonitis rates. The multidisciplinary PDEP significantly improved outcomes by further lowering peritonitis rates.

Mejoría de la calidad de vida y efectos adversos en pacientes con enfermedad arterial obstructiva crónica tratados con alprostadil-alfa-ciclodextrina por vía venosa periférica / Improvement in quality of life and adverse effects in patients with chronic arterial occlusive disease treated with alprostadil alpha-cyclodextrin through the peripheral venous route

Objetivos. Evaluar las complicaciones locales y sistémicas que se presentan en pacientes tratados con alprostadil-alfa-ciclodextrina (AAC) por vía venosa periférica, así como los cambios que se producen en la calidad de vida de estos pacientes. Método. Estudio descriptivo observacional prospectivo. Se incluyeron todos los pacientes ingresados en la Unidad de Angiología y Cirugía Vascular del Complejo Hospitalario Universitario de Albacete que cumplían los criterios de indicación terapéutica para la administración de AAC entre septiembre de 2001 y abril de 2002. Cada paciente recibió el tratamiento según un protocolo previamente consensuado en la Unidad. Se recogieron variables sociodemográficas, enfermedades asociadas, tipo y nivel de la enfermedad arterial, complicaciones locales y sistémicas en la administración de cada uno de los ciclos, calidad de vida al inicio y final del tratamiento. Resultados. Se estudiaron 44 pacientes. El 65,5% presentaba enfermedad arterial en su estadio más avanzado. Las complicaciones sistémicas más frecuentes fueron las reacciones alérgicas y las náuseas; el tratamiento tuvo que ser suspendido en el 11,4% de los casos. Como complicaciones locales aparecieron dolor y eritema en la vía venosa (media de 1,29 por cada 10 dosis de tratamiento); se cambió la vía 0,97 veces de media cada 10 dosis, el 38,6% de las veces debido a flebitis. Los resultados del cuestionario EUROQOL muestran mejoría en movilidad, capacidad para el cuidado personal, capacidad para realizar actividades cotidianas, dolor y ansiedad/depresión. Conclusiones. El tratamiento con AAC según el protocolo establecido ofrece seguridad a los pacientes y produce una notable mejora en la calidad de vida percibida

Objectives. To evaluate local and systemic complications in patients treated with alprostadil alpha-cyclodextrin (AAC) through the peripheral venous route, as well as changes in the quality of life of these patients. Method. A descriptive, observational, prospective study was performed. All patients admitted to the Angiology and Vascular Surgery Unit of the University Hospital of Albacete who fulfilled the criteria for therapeutic indication of AAC administration between September 2001 and April 2002 were included. Each patient received the treatment according to a protocol previously established through consensus in the unit. Sociodemographic variables, associated disease, type and degree of arterial disease, local and systemic complications with administration of each of the cycles, and quality of life at the beginning and end of the treatment were studied. Results. Forty-four patients were studied. A total of 65.5% had arterial disease in the most advanced stage. The most frequent systemic complications were allergic reactions and nausea; treatment had to be discontinued in 11.4% of patients. Local complications consisted of pain and erythema in the area of the venous line (mean 1.29 every 10 treatment doses); there was a mean of 0.97 line changes every 10 doses and 38.6% of the changes was due to phlebitis. The results of the EUROQOL questionnaire showed improvement in: mobility, ability to perform personal care, ability to perform daily activities, pain, and anxiety/depression. Conclusions. Treatment with AAC according to the established protocol is safe and improves perceived quality of life