Detección de pacientes de alto riesgo a través del servicio de indicación farmacéutica: INDICA+PRO Implantación (2020-2021) / Detection of high-risk patients through the Pharmaceutical Indication Service: INDICA+PRO Implementation (2020-2021)

JUSTIFICACIÓN: la derivación al médico es una de las posibles actuaciones del farmacéutico a través del servicio de indicación farmacéutica. Dicha derivación permite el cribado de pacientes de alto riesgo desde la farmacia comunitaria (FC). La colaboración multidisciplinar permite establecer criterios de derivación de pacientes consensuados entre profesionales. El estudio INDICA+PRO tiene como objetivo implantar el SIF protocolizado en la FC española a través del uso de protocolos consensuados en una fase previa de codiseño del servicio. OBJETIVOS: describir las características de los pacientes derivados al médico a través del SIF en el estudio INDICA+PRO Implantación. MATERIAL Y MÉTODOS: estudio con diseño híbrido de efectividad-implantación tipo 3 llevado a cabo inicialmente durante 14 meses. La intervención codiseñada constaba de: procedimiento general del SIF establecido por Foro de Atención Farmacéutica en FC, protocolos consensuados entre sociedades médicas (Semergen y SemFyC), asociaciones farmacéuticas (SEFAC y MICOF) y universidad (GIAF-UGR) específicos para síntomas menores (dermatológicos, digestivos, relacionados con el dolor, respiratorios y otros) incluidos en una plataforma digital (SEFAC e_XPERT®) y la formación de los farmacéuticos con el seguimiento de 33 farmacéuticos facilitadores del cambio de práctica. Cada protocolo específico incluía criterios de derivación divididos en: duración del síntoma, edad del paciente, síntomas de alarma, medicamentos, problemas de salud y estado fisiológico del paciente y otros. (AU)

Actuaciones del farmacéutico sobre la demanda de medicamentos a través del servicio de indicación farmacéutica: INDICA+PRO Implantación (2020-2021) / Pharmacist actions on medicines demand through the pharmaceutical indication service: INDICA+PRO Implementation (2020-2021)

JUSTIFICACIÓN: en España, las especialidades farmacéuticas publicitarias sólo están disponibles a través de la farmacia comunitaria (FC). Desde 2019, Foro de Atención Farmacéutica en Farmacia Comunitaria (Foro AF-FC) incluye la demanda de medicamento para un síntoma menor a través del servicio de indicación farmacéutica (SIF) y lo simboliza como “Deme esto para…”. El estudio INDICA+PRO Implantación tiene como objetivo implantar el SIF protocolizado en la FC española incluyendo ambas vías de entrada al servicio (consulta de síntoma menor y demanda de medicamento para el mismo).OBJETIVOS: describir la demanda de medicamentos para un síntoma menor atendidas por el farmacéutico comunitario a través del SIF.MATERIAL Y MÉTODOS: el estudio realizado utiliza un diseño híbrido de efectividad-implantación tipo 3 durante 14 meses inicialmente. La intervención implantada constaba de: procedimiento general del SIF establecido por Foro AF-FC, protocolos consensuados entre sociedades médicas (Semergen y SemFyC), asociaciones farmacéuticas (SEFAC y MICOF) y universidad (GIAF-UGR) específicos para 31 síntomas menores incluidos en una plataforma digital (SEFAC e_XPERT®) y la formación y el seguimiento de los farmacéuticos. Se consideraron los pacientes que acudían a FC demandando un medicamento para un síntoma menor incluido en el estudio (dermatológico, digestivo, relacionado con el dolor, respiratorio u otro). Todos los pacientes recibían seguimiento 10 días tras la consulta en FC.RESULTADOS: 14083 consultas fueron registradas por 687 farmacéuticos pertenecientes a 518 FC. Mayor número de consultas por síntoma menor (84,6 %, n=11911) que por demanda de un medicamento concreto fueron atendidas a través del SIF (15,4 %, n=2172). Entre aquellos pacientes que demandaron un medicamento, el 7,7 % (n=168) fueron derivados al médico. (AU)

Implantación del servicio de cribado de deterioro cognitivo en farmacia comunitaria / Implementation of the cognitive impairment screening service in community pharmacies

JUSTIFICACIÓN: El diagnóstico temprano de deterioro cognitivo permite el control de factores de riesgo modificables para prevenir el desarrollo de demencia en edades más tardías. La farmacia comunitaria es un establecimiento sanitario próximo, accesible y de confianza, en el que el farmacéutico a menudo participa en procesos de prevención de la enfermedad. Esto le convierte en un profesional sanitario adecuado para el cribado de deterioro cognitivo.OBJETIVOS: desarrollar un programa de detección precoz de deterioro cognitivo en pacientes mayores de 50 años con queja subjetiva de memoria desde la farmacia comunitaria. Favorecer la creación de equipos multidisciplinares en la detección precoz de la demencia.MATERIAL Y MÉTODOS: se diseñó un estudio observacional transversal mediante entrevista personal estructurada a pacientes con indicios de pérdida de memoria, el cual fue aprobado por el Comité Ético, con número de registro CEEI21/198. Se invitó a participar de forma voluntaria a farmacias de la provincia. Antes de la puesta en marcha del servicio, los farmacéuticos participantes recibieron una formación específica. En la entrevista personal al paciente, tras firmar el correspondiente consentimiento informado, se realizaron tres cuestionarios validados para evaluar su estado cognitivo: Memory Impairment Screening (MIS), Fluidez Verbal Semántica (FVS) y Short Portable Mental State Questionnaire (SPMSQ) de Pfeiffer. Siempre que fue posible, se realizó un cuarto cuestionario: Test del Informador Breve (TIN Breve). La recogida de datos se llevó a cabo a través de la plataforma informática ATENFARMA y se procedió al tratamiento estadístico de los resultados.RESULTADOS/DISCUSIÓN: durante el periodo de tiempo comprendido entre febrero de 2021 y febrero de 2022, han participado 16 farmacias, en las cuales se cribaron un total de 206 pacientes. De ellos, 58 (28 %) presentaban sospecha de deterioro cognitivo. (AU)

Diseño y desarrollo del servicio de cribado de sufrir fractura osteoporótica del MICOF de Valencia / Design and development of the screening service for osteoporotic fractures at the MICOF in Valencia

JUSTIFICACIÓN: la osteoporosis es una patología asintomática que se desarrolla de manera lenta pero progresiva sin mostrar síntomas ni signos aparentes; una de las consecuencias de esta enfermedad son las fracturas osteoporóticas y dadas sus consecuencias clínicas y morbimortalidad, supone un importante problema de salud pública en los países desarrollados.OBJETIVOS: diseñar y desarrollar un servicio de identificación de personas en riesgo de padecer fracturas osteoporóticas desde la farmacia comunitaria e incorporarlo en el Catálogo de Servicios Profesionales del MICOF.MATERIAL Y MÉTODOS: se ha realizado una revisión bibliográfica de documentos de sociedades científicas con respecto a las herramientas de cribado para evaluar el riesgo de sufrir fracturas osteoporóticas en función de los factores de riesgo individuales. No existe ningún consenso específico aceptado a nivel nacional ni mundial, sin embargo, de entre todas las escalas, la más utilizada en la práctica clínica para evaluar el riesgo de fractura es la escala FRAX®.RESULTADOS: tras la revisión se decide utilizar el algoritmo propuesto por SEFAC en su curso sobre Osteoporosis consistente en identificar y clasificar los factores de riesgo clínicos, y en evaluar el riesgo de padecer fracturas a través de la escala FRAX®. En aquellos pacientes con un riesgo elevado o intermedio se derivará al médico para la valoración de la realización de una densitometría además de una intervención farmacéutica en estilo de vida (nutrición y actividad física) en prevención primaria y secundaria de la osteoporosis. (AU)

Immature granules are not major sites for segregation of constitutively secreted granule content proteins in NIT-1 insulinoma cells.

Immature secretory granules (ISG's) are sites of segregation of proteins destined for secretion by unregulated pathways from those stored in mature secretory granules in endocrine cells. To determine whether significant soluble protein sorting occurs in ISG's, the secretion of soluble versions of the pancreatic protein GP2 (GP2-GPI(-)) and placental alkaline phosphatase (SEAP) was analyzed in NIT-1 cells. By immunofluorescence microscopy, neither protein localized to SG's in transfected cells. Their secretion was secretagogue-independent in pulse-chase radiolabeling experiments even at early times of chase, while a small increase in the secretion of amylase, which is known to enter ISG's, could be detected. Finally, in sucrose gradient fractionation experiments, SEAP was present in light density fractions. We conclude that while some proteins, such as amylase, have a limited intrinsic capacity to enter ISG's, the segregation of proteins secreted via the constitutive pathway from SG content proteins occurs primarily in the trans Golgi network.

Human huntingtin-associated protein (HAP-1) gene: genomic organisation and an intragenic polymorphism.

The huntingtin-associated protein (HAP-1) interacts with the Huntington disease gene product, huntingtin. It is predominantly expressed in the brain and shows an increased affinity for mutant huntingtin. We have sequenced an 18,656bp genomic region encompassing the entire human HAP-1 gene and determined its genomic organisation, with 11 exons spanning 12.1kb. We have also found an intragenic polymorphism within intron 6 of HAP-1. We have recently shown that HAP-1 maps to a region of the genome which has been implicated in a variety of neurological conditions, including progressive supranuclear palsy (PSP), a late-onset atypical parkinsonian disorder. The detailed characterisation of the genomic organisation of HAP-1 and the presence of an intragenic polymorphism will be helpful in evaluating its role in different disorders, using candidate gene approaches.

Deformation simulation algorithms of elastic tissues in "real-time" based in elasticity theory.

In this paper, a new deformable model based in Boundary Elements Method (BEM) is presented. This model is characterised by its robustness and high deformation calculation speed. Experiments developed show that this model could calculate elastic deformations of elastic objects composed by 150 nodes with a 15 Hz. refresh rate (minim refresh rate acceptable in real time interactive systems).

Gene structure and map location of the murine homolog of the Huntington-associated protein, Hap1.

Huntington's Disease (HD) is an inherited progressive neurodegenerative disorder associated with a mutation in a gene expressed in both affected and non-affected tissues. The selective neuropathology in HD is thought to be mediated in part through interactions with other proteins including the Huntington Associated Protein, HAP-1, which is predominantly expressed in the brain. We have mapped its murine homolog, Hap1, to mouse Chr 11 (band D), which shares extensive synteny with human Chr 17 including the region 17q21-q22, where the gene for 'frontotemporal dementia and parkinsonism linked to chromosome 17' has bee mapped. In addition, we have sequenced a 21,984 base pair (bp) genomic clone encompassing the entire Hap1 gene. It is organized as 11 exons and flanked by exons from potentially one or more novel genes. At least three Hap1 transcripts (Hap1-A; Hap1-B; Hap1-C) can be formed by alternative splicing at the 3' end of the gene leading to protein isoforms with novel C-termini.

Atrophin-1, the DRPLA gene product, interacts with two families of WW domain-containing proteins.

Atrophin-1 contains a polyglutamine repeat, expansion of which is responsible for dentatorubral and pallidoluysian atrophy (DRPLA). The normal function of atrophin-1 is unknown. We have identified five atrophin-1 interacting proteins (AIPs) which bind to atrophin-1 in the vicinity of the polyglutamine tract using the yeast two-hybrid system. Four of the interactions were confirmed using in vitro binding assays. All five interactors contained multiple WW domains. Two are novel. The AIPs can be divided into two distinct classes. AIP1 and AIP3/WWP3 are MAGUK-like multidomain proteins containing a number of protein-protein interaction modules, namely a guanylate kinase-like region, two WW domains, and multiple PDZ domains. AIP2/WWP2, AIP4, and AIP5/WWP1 are highly homologous, each having four WW domains and a HECT domain characteristic of ubiquitin ligases. These interactors are similar to recently isolated huntingtin-interacting proteins, suggesting possible commonality of function between two proteins responsible for very similar diseases.

Caveolin transfection results in caveolae formation but not apical sorting of glycosylphosphatidylinositol (GPI)-anchored proteins in epithelial cells.

Most epithelial cells sort glycosylphosphatidylinositol (GPI)-anchored proteins to the apical surface. The "raft" hypothesis, based on data mainly obtained in the prototype cell line MDCK, postulates that apical sorting depends on the incorporation of apical proteins into cholesterol/glycosphingolipid (GSL) rafts, rich in the cholesterol binding protein caveolin/VIP21, in the Golgi apparatus. Fischer rat thyroid (FRT) cells constitute an ideal model to test this hypothesis, since they missort both endogenous and transfected GPI-anchored proteins to the basolateral plasma membrane and fail to incorporate them into cholesterol/glycosphingolipid clusters. Because FRT cells lack caveolin, a major component of the caveolar coat that has been proposed to have a role in apical sorting of GPI-anchored proteins (Zurzolo, C., W. Van't Hoff, G. van Meer, and E. Rodriguez-Boulan. 1994. EMBO [Eur. Mol. Biol. Organ.] J. 13:42-53.), we carried out experiments to determine whether the lack of caveolin accounted for the sorting/clustering defect of GPI-anchored proteins. We report here that FRT cells lack morphological caveolae, but, upon stable transfection of the caveolin1 gene (cav1), form typical flask-shaped caveolae. However, cav1 expression did not redistribute GPI-anchored proteins to the apical surface, nor promote their inclusion into cholesterol/GSL rafts. Our results demonstrate that the absence of caveolin1 and morphologically identifiable caveolae cannot explain the inability of FRT cells to sort GPI-anchored proteins to the apical domain. Thus, FRT cells may lack additional factors required for apical sorting or for the clustering with GSLs of GPI-anchored proteins, or express factors that inhibit these events. Alternatively, cav1 and caveolae may not be directly involved in these processes.

Huntingtin-associated protein 1 (HAP1) binds to a Trio-like polypeptide, with a rac1 guanine nucleotide exchange factor domain.

Huntington's disease (HD) occurs when the widely expressed protein huntingtin contains an expanded glutamine repeat. The selective degeneration and neuronal morphologic abnormalities of HD may involve interactions with proteins that bind to huntingtin, such as HAP1. The biological significance of this interaction is unclear because neither HAP1 nor huntingtin have significant homology to known proteins. Therefore, we sought to identify HAP1-binding proteins. Using the yeast two-hybrid system, we isolated a rat cDNA encoding part of a protein that interacts with HAP1, and we confirmed the specificity of this interaction using an in vitro protein-binding assay. We called the protein Duo because it is closely related to the human protein Trio but is shorter. Northern blot analysis indicates brain-specific expression of Duo. Human Duo contains a guanine nucleotide exchange factor (GEF) domain that is likely to be rac1-specific, a pleckstrin homology (PH) domain and spectrin-like repeat units. These data support the hypothesis that huntingtin is involved in vesicle trafficking and cytoskeletal functions, and raise the possibility of a role for huntingtin in the regulation of a ras-related signaling pathway.

Huntington's disease and dentatorubral-pallidoluysian atrophy: proteins, pathogenesis and pathology.

Each of the glutamine repeat neurodegenerative diseases has a particular pattern of pathology largely restricted to the CNS. However, there is considerable overlap among the regions affected, suggesting that the diseases share pathogenic mechanisms, presumably involving the glutamine repeats. We focus on Huntington's disease (HD) and Dentatorubral-pallidoluysian atrophy (DRPLA) as models for this family of diseases, since they have striking similarities and also notable differences in their clinical features and pathology. We review the pattern of pathology in adult and juvenile onset cases. Despite selective pathology, the disease genes and their protein products (huntingtin and atrophin-1) are widely expressed. This presents a central problem for all the glutamine repeat diseases-how do widely expressed gene products give rise to restricted pathology? The pathogenic effects are believed to occur via a "gain of function" mechanism at the protein level. Mechanisms of cell death may include excitotoxicity, metabolic toxicity, apoptosis, and free radical stress. Emerging data indicate that huntingtin and atrophin-1 may have distinct protein interactions. The specific interaction partners may help explain the selective pathology of these diseases.

Huntingtin-associated protein 1 (HAP1) interacts with the p150Glued subunit of dynactin.

Huntington's disease (HD) is an inherited neurodegenerative disease caused by expansion of a polyglutamine repeat in the HD protein huntingtin. Huntingtin's localization within the cell includes an association with cytoskeletal elements and vesicles. We previously identified a protein (HAP1) which binds to huntingtin in a glutamine repeat length-dependent manner. We now report that HAP1 interacts with cytoskeletal proteins, namely the p150 Glued subunit of dynactin and the pericentriolar protein PCM-1. Structural predictions indicate that both HAP1 and the interacting proteins have a high probability of forming coiled coils. We examined the interaction of HAP1 with p150 Glued . Binding of HAP1 to p150 Glued (amino acids 879-1150) was confirmed in vitro by binding of p150 Glued to a HAP1-GST fusion protein immobilized on glutathione-Sepharose beads. Also, HAP1 co-immunoprecipitated with p150 Glued from brain extracts, indicating that the interaction occurs in vivo . Like HAP1, p150 Glued is highly expressed in neurons in brain and both proteins are enriched in a nerve terminal vesicle-rich fraction. Double label immunofluorescence experiments in NGF-treated PC12 cells using confocal microscopy revealed that HAP1 and p150 Glued partially co-localize. These results suggest that HAP1 might function as an adaptor protein using coiled coils to mediate interactions among cytoskeletal, vesicular and motor proteins. Thus, HAP1 and huntingtin may play a role in vesicle trafficking within the cell and disruption of this function could contribute to the neuronal dysfunction and death seen in HD.

Secretory granule content proteins and the luminal domains of granule membrane proteins aggregate in vitro at mildly acidic pH.

A major unresolved issue in the field of secretory granule biogenesis is the extent to which the aggregation of granule content proteins is responsible for the sorting of regulated from constitutively secreted proteins. The aggregation process is postulated to take place in the trans-Golgi network and immature secretory granules as the proteins encounter mildly acidic pH and high calcium concentrations. We have developed in vitro assays that reconstitute the precipitation out of solution of secretory granule content proteins of anterior pituitary gland and adrenal medulla. In the assays, all of the major granule content polypeptides form a precipitate as the pH is titrated below 6.5, and this precipitate can be recovered in the pellet fraction after centrifugation. Addition of calcium is required for the aggregation of chromaffin granule content. In contrast to the proteins secreted by the regulated pathway, the constitutively secreted proteins IgG, albumin, and angiotensinogen, when added to the assays, remain predominantly in the supernatant. Among the individual proteins tested, prolactin is found to aggregate homophilically under these conditions and can drive the co-aggregation of other proteins, such as the chromogranins. Soluble forms of granule membrane proteins, including dopamine beta-hydroxylase and peptidyl glycine alpha-amidating enzyme also co-aggregated with granule content proteins. The results are consistent with the idea that spontaneous aggregation of proteins occurring under ionic conditions similar to those at the sites of granule formation is a property restricted to those proteins packaged in secretory granules. In addition, the association of luminal domains of membrane proteins with content proteins in vitro raises the possibility that analogous interactions between membrane-bound and content proteins also occur during granule formation in intact cells.

Apical plasma membrane proteins are not obligatorily stored in secretory granules in exocrine cells.

Exocrine cells are epithelial cells in which secretory granules undergo fusion with the apical plasma membrane upon secretagogue stimulation. Several apical plasma membrane proteins have been found in secretory granules in cells from pancreas and salivary glands raising the possibility that incorporation into secretory granules followed by exocytosis of the granules accounts for their insertion into the apical plasma membrane. To test this hypothesis, we have expressed the influenza hemagglutinin (HA) in pancreatic AR42J cells, which make zymogen-like granules upon incubation with dexamethasone. The influenza virus HA is known to be specifically targeted to the apical plasma membrane of epithelial cells that lack a regulated pathway and is also known to be excluded from secretory granules in virally-infected pituitary AtT20 cells. Localization of the protein by immunofluorescence microscopy revealed that it accumulated at the plasma membrane of the transfected AR42J cells. HA was not observed in the amylase-rich secretory granules. By immunolabeling of ultrathin cryosections of the transfected cells, HA was also found exclusively on the cell surface, with label over secretory granules not exceeding that seen in control, untransfected cells. In addition, in cell fractionation experiments performed on radiolabeled AR42J cell transformants, HA was not detectable in the secretory granule fractions. These results indicate that HA is not efficiently stored in mature secretory granules and is likely to reach the cell surface via constitutive transport pathways.

Exocrine granule specific packaging signals are present in the polypeptide moiety of the pancreatic granule membrane protein GP2 and in amylase: implications for protein targeting to secretory granules.

The mechanisms for segregation of secretory and membrane proteins incorporated into storage granules from those transported constitutively have been thought to be conserved in diverse cell types, including exocrine and endocrine cells. However, GP2, the major protein of pancreatic zymogen granule membranes, in its native glycosyl phosphatidylinositol (GPI)-linked form, is incorporated into secretory granules when expressed in exocrine pancreatic AR42J cells, but not in the endocrine cells such as pituitary AtT20. To determine whether the protein moiety of GP2 contains the cell-type specific information for packaging into granules, a secretory form of GP2 (GP2-GPI-), with the GPI attachment site deleted, was generated and introduced into AR42J and AtT20 cells. Like native GP2, GP2-GPI- localized to the zymogen-like granules of AR42J cells and underwent regulated secretion. In AtT20 cells expressing GP2-GPI-, however, the protein was secreted by the constitutive pathway. Thus, a granule packaging signal is present in the luminal portion of GP2 that is functional only in the exocrine cells. However, this cell-type dependent sorting process is not limited to GP2 or membrane proteins. Amylase, a major content protein of pancreatic acinar and serous salivary gland granules, was also secreted exclusively by the constitutive pathway when expressed in AtT20 cells. The cell-type specific targeting of GP2 to granules correlated with its behavior in an in vitro aggregation assay where it co-aggregated more effectively with content proteins from pancreatic zymogen granules than with those from pituitary granules.(ABSTRACT TRUNCATED AT 250 WORDS)