Characterisation and evaluation of the regenerative capacity of Stro-4+ enriched bone marrow mesenchymal stromal cells using bovine extracellular matrix hydrogel and a novel biocompatible melt electro-written medical-grade polycaprolactone scaffold.

Many skeletal tissue regenerative strategies centre around the multifunctional properties of bone marrow derived stromal cells (BMSC) or mesenchymal stem/stromal cells (MSC)/bone marrow derived skeletal stem cells (SSC). Specific identification of these particular stem cells has been inconclusive. However, enriching these heterogeneous bone marrow cell populations with characterised skeletal progenitor markers has been a contributing factor in successful skeletal bone regeneration and repair strategies. In the current studies we have isolated, characterised and enriched ovine bone marrow mesenchymal stromal cells (oBMSCs) using a specific antibody, Stro-4, examined their multipotential differentiation capacity and, in translational studies combined Stro-4+ oBMSCs with a bovine extracellular matrix (bECM) hydrogel and a biocompatible melt electro-written medical-grade polycaprolactone scaffold, and tested their bone regenerative capacity in a small in vivo, highly vascularised, chick chorioallantoic membrane (CAM) model and a preclinical, critical-sized ovine segmental tibial defect model. Proliferation rates and CFU-F formation were similar between unselected and Stro-4+ oBMSCs. Col1A1, Col2A1, mSOX-9, PPARG gene expression were upregulated in respective osteogenic, chondrogenic and adipogenic culture conditions compared to basal conditions with no significant difference between Stro-4+ and unselected oBMSCs. In contrast, proteoglycan expression, alkaline phosphatase activity and adipogenesis were significantly upregulated in the Stro-4+ cells. Furthermore, with extended cultures, the oBMSCs had a predisposition to maintain a strong chondrogenic phenotype. In the CAM model Stro-4+ oBMSCs/bECM hydrogel was able to induce bone formation at a femur fracture site compared to bECM hydrogel and control blank defect alone. Translational studies in a critical-sized ovine tibial defect showed autograft samples contained significantly more bone, (4250.63 mm3, SD = 1485.57) than blank (1045.29 mm3, SD = 219.68) ECM-hydrogel (1152.58 mm3, SD = 191.95) and Stro-4+/ECM-hydrogel (1127.95 mm3, SD = 166.44) groups. Stro-4+ oBMSCs demonstrated a potential to aid bone repair in vitro and in a small in vivo bone defect model using select scaffolds. However, critically, translation to a large related preclinical model demonstrated the complexities of bringing small scale reported stem-cell material therapies to a clinically relevant model and thus facilitate progression to the clinic.

Extracellular NK histones promote immune cell anti-tumor activity by inducing cell clusters through binding to CD138 receptor.

BACKGROUND: Natural killer (NK) cells are important anti-tumor cells of our innate immune system. Their anti-cancer activity is mediated through interaction of a wide array of activating and inhibitory receptors with their ligands on tumor cells. After activation, NK cells also secrete a variety of pro-inflammatory molecules that contribute to the final immune response by modulating other innate and adaptive immune cells. In this regard, external proteins from NK cell secretome and the mechanisms by which they mediate these responses are poorly defined. METHODS: TRANS-stable-isotope labeling of amino acids in cell culture (TRANS-SILAC) combined with proteomic was undertaken to identify early materials transferred between cord blood-derived NK cells (CB-NK) and multiple myeloma (MM) cells. Further in vitro and in vivo studies with knock-down of histones and CD138, overexpression of histones and addition of exogenous histones were undertaken to confirm TRANS-SILAC results and to determine functional roles of this material transferred. RESULTS: We describe a novel mechanism by which histones are actively released by NK cells early after contact with MM cells. We show that extracellular histones bind to the heparan sulfate proteoglycan CD138 on the surface of MM cells to promote the creation of immune-tumor cell clusters bringing immune and MM cells into close proximity, and thus facilitating not only NK but also T lymphocyte anti-MM activity. CONCLUSION: This study demonstrates a novel immunoregulatory role of NK cells against MM cells mediated by histones, and an additional role of NK cells modulating T lymphocytes activity that will open up new avenues to design future immunotherapy clinical strategies.

Demethylation of an NF-κB enhancer element orchestrates iNOS induction in osteoarthritis and is associated with altered chondrocyte cell cycle.

OBJECTIVE: To examine the methylation profile of the nuclear factor (NF)-κB enhancer region at -5.8 kb of inducible nitric oxide synthase (iNOS) and the subsequent role in the induction of osteoarthritis (OA) via cell cycle regulation. METHODS: Percentage methylation was determined by pyrosequencing, gene expression by qRT-PCR and cell proliferation was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Transient transfections were induced to determine the effect of the NF-κB enhancer region on cell proliferation and the influence of DNA methylation. RESULTS: In vitro de-methylation with 5-aza-dC showed decreased levels of DNA methylation at CpG sites localised at -5.8 kb, which correlated with higher levels of iNOS expression. In vitro methylation of the NF-κB enhancer region at -5.8 kb increased the percentage of cells at G0/G1 cell cycle phase. Loss of methylation within this region correlated with, enhanced proliferation and increased number of cells at G2/M phase. OA chondrocytes demonstrated up-regulation of the G0/G1 cell cycle progression markers Cyclin D1 and CDK6 in contrast to control cells. We demonstrate the loss of methylation that occurs at specific CpG sites localised at the -5.8 kb NF-κB enhancer region of the iNOS gene in OA chondrocytes permits the binding of this transcription factor activating the expression of iNOS. This results in subsequent altered cell cycle regulation, altered proliferative phenotype and transmission of the pathogenic phenotype to daughter cells. CONCLUSIONS: This study indicates that inhibition of cell cycle progression by iNOS enhancer hypermethylation is capable of reducing pro-inflammatory responses via down-regulation of NF-κB with important therapeutic implications in OA.

Epigenetic regulation of interleukin-8, an inflammatory chemokine, in osteoarthritis.

OBJECTIVE: To determine whether altered IL8 methylation status is associated with increased expression of IL8 in human osteoarthritic (OA) chondrocytes. METHODS: IL8 expression levels and the percentage CpG methylation in human chondrocytes were quantified by qRT-PCR and pyrosequencing in OA patients and in non-OA osteoporotic controls. The effect of CpG methylation on IL8 promoter activity was determined using a CpG-free vector; co-transfections with expression vectors encoding nuclear factor-kappa B (NF-κB), AP-1 and C/EBP were subsequently undertaken to analyse for IL8 promoter activity in response to changes in methylation status. RESULTS: IL8 expression in OA patients was 37-fold higher than in osteoporotic controls. Three CpG sites in the IL8 promoter were significantly demethylated in OA patients. Multiple regression analysis revealed that the degree of methylation of the CpG site located at -116-bp was the strongest predictor of IL8 expression. In vitro DNA methylation was noted to decrease IL8 promoter basal activity. Furthermore, NF-κB, AP-1 and C/EBP strongly enhanced IL8 promoter activity whilst DNA methylation inhibited the effects of these three transcription factors. CONCLUSIONS: The present study demonstrates the key role of DNA methylation status on the expression of IL8 in human chondrocytes. We demonstrate a quantitative relationship between percentage methylation and gene expression within clinical samples. These studies provide direct evidence linking the activation of IL8, DNA demethylation and the induction of the OA process with important therapeutic implications therein for patients with this debilitating disease.

Mente Activa® Improves Impaired Spatial Memory in Aging Rats.

OBJECTIVES: Aging is accompanied by a decline in several aspects of the cognitive function, having negative personal and socioeconomic impacts. Dietary supplements could be beneficial for preventing age-related cognitive decline. In this context, we examined whether the nutritional supplement Mente Activa® has beneficial effects on aging-related cognitive deficits without inducing side effects. METHODS: Mente Activa® was administered to old rats (n= 30 treated rats and n= 30 control rats) during 5 months, and the Morris water maze was used to test the learning capacities of the animals. The first assessment was conducted before the nutritional intervention (age of 18-19 months), to determine the baseline of the performance of animals on this test, and the second assessment was performed at the end of the treatment (23-24 moths). In order to examine possible secondary effects of this nutritional supplement, plasma, heart anatomy and liver parameters were evaluated. RESULTS: Our data indicate that supplemented rats showed less escape latency, distance swum, higher use of spatial search strategies, and crossed the former platform location with higher frequency than control rats. These effects were specific of the treatment, indicating that this nutritional supplement has a beneficial effect on spatial memory. On the other hand, the regular intake of Mente Activa® did not induce any negative effects in plasma parameters and heart size. CONCLUSIONS: Aged rats under a sustained dietary intake of the nutritional supplement Mente Activa® displayed improved learning and memory abilities compared to the non-treated rats. These results suggest the therapeutic potential and safety of use of Mente Activa® for age-related cognitive deficits, particularly, in the onset of the first cognitive dysfunction symptoms.

Genetic diversity of wild grapevine populations in Spain and their genetic relationships with cultivated grapevines.

The wild grapevine, Vitis vinifera L. ssp. sylvestris (Gmelin) Hegi, considered as the ancestor of the cultivated grapevine, is native from Eurasia. In Spain, natural populations of V. vinifera ssp. sylvestris can still be found along river banks. In this work, we have performed a wide search of wild grapevine populations in Spain and characterized the amount and distribution of their genetic diversity using 25 nuclear SSR loci. We have also analysed the possible coexistence in the natural habitat of wild grapevines with naturalized grapevine cultivars and rootstocks. In this way, phenotypic and genetic analyses identified 19% of the collected samples as derived from cultivated genotypes, being either naturalized cultivars or hybrid genotypes derived from spontaneous crosses between wild and cultivated grapevines. The genetic diversity of wild grapevine populations was similar than that observed in the cultivated group. The molecular analysis showed that cultivated germplasm and wild germplasm are genetically divergent with low level of introgression. Using a model-based approach implemented in the software structure, we identified four genetic groups, with two of them fundamentally represented among cultivated genotypes and two among wild accessions. The analyses of genetic relationships between wild and cultivated grapevines could suggest a genetic contribution of wild accessions from Spain to current Western cultivars.

Vigilancia de infecciones y otros eventos adversos en pacientes en diálisis en el área sur de Gran Canaria / Surveillance for infections and other adverse events in dialysis patients in southern Gran Canaria

Introducción: Las infecciones bacterianas representan un gran desafío en las estrategias de gestión del riesgo, prevención y seguridad del paciente en hemodiálisis de las cuales las infecciones del acceso vascular (AV) representan la primera causa morbi-mortalidad en estos pacientes. Métodos: Estudio prospectivo de incidencia de eventos adversos e infecciones de 7 meses (marzo-septiembre 2008) en las unidades de Hemodiálisis del Área sanitaria Sur de Gran Canaria (Hospital y Centro periférico) utilizando la metodología del Dialysis Surveillance Network del CDC. Resultados: Se vigilaron 1545 pacientes/mes, 60,5% con fístula (FAV), 35,5% con catéter permanente (CP), 3,0% con prótesis y 1,0% con catéter temporal. La incidencia de eventos en ambos centros fue 8,7 casos por 100 pacientes-mes; la tasa de eventos infecciosos fue de 9,1 para FAV y 20,6 para CP en ámbito hospitalario, mientras las tasas de otras infecciones (respiratorias, herida, orina) fueron similares. Se realizó cultivo antes de empezar tratamiento antibiótico en el 91,0% frente a sospecha de bacteriemia y/o infección AV. El 90,0% de tratamientos se ajustaron con antibiograma. Destaca una baja incidencia de bacterias mutirresistentes mientras que las infecciones relacionadas con el AV fueron causadas en proporción similar por bacterias grampositivas y gramnegativas. Conclusiones: El acceso vascular es el principal factor de riesgo para el desarrollo de infecciones. La vigilancia epidemiológica he permitido detectar oportunidades de mejora en ámbitos asistenciales distintos, integrándose como elemento fundamental en el desarrollo de estrategias multidisciplinarias de seguridad del paciente (AU)

Background: Bacterial infections pose a great challenge to risk management activities in the area of chronic haemodialysis, as vascular access related infections are the main cause of mortality among these patients. Methods: Prospective surveillance study lasting 7 months (March-September, 2008) at the two haemodialysis units in a district health area in Gran Canaria, Spain. We have used methodology proposed by CDC´s Dialysis Surveillance Network. Results: 1545 patientsmonth were enrolled, 60,5% having an arterio-venous fistula (AVF), 35,5% permanent catheter (PC), 3,0% graft and 1,0% temporary catheters. Events incidence rate at both centers was 8,6 cases per 100 patients-month, 9,1 rate for FAV and 2,9 rates for CP, So, the greatest incidence of vascular access related infections was for permanent catheter as compared with AFV. Nevertheless the other type of infections (respiratory, urinary tract, skin and chronic ulcers) showed a similar rate. Microbiological cultures before antibiotic treatment were performed in 82,2 %, but increased up to 91,0% when a vascular related infection was suspected. Empiric treatment was adjusted to antibiogram results in 90,0% of occasions. A low incidence of multirresistant microbes was seen. Gram-positive and gram-negative bacteria appeared in a similar proportion. Conclusions: Vascular access is the main risk factor for infectious events. Epidemiological surveillance has allowed us to detect areas of improvement in different settings, appearing as a key element in the risk management and patient safety areas (AU)

Surveillance for infections and other adverse events in dialysis patients in southern Gran Canaria.

BACKGROUND: Bacterial infections pose a major challenge to risk management activities in the area of chronic haemodialysis, as vascular access-related infections are the main cause of mortality among these patients. METHODS: Prospective surveillance study lasting 7 months (March-September, 2008) at two haemodialysis units in a district health area Gran Canaria, Spain. We used the methodology proposed by CDC´s Dialysis Surveillance Network. RESULTS: 1545 patients/month were recorded, 60.5% with an arteriovenous fistula (AVF), 35.5% with a permanent catheter (PC), 3.0% with grafts and 1.0% with temporary catheters. The rate of adverse events was 8.6 cases per 100 patients/month, 9.1 for AVF patients, and 2.9 for PC. Nevertheless, the other types of infections (respiratory, urinary tract, skin and chronic ulcers) showed similar rates. Microbiological cultures were taken in 82.2%, but this rate increased to 91.0% when a vascular access-related infection was suspected. Empirical treatment was adjusted to antibiogram results in 90.0% of occasions. A low incidence of multi-resistant microbes was observed. Gram-positive and gram-negative bacteria appeared in similar proportions. CONCLUSIONS: Vascular access is the main risk factor for infectious events. Epidemiological surveillance has allowed us to detect areas of improvement in different settings, acting as a key element in risk management and patient safety.

The epigenetic effect of glucosamine and a nuclear factor-kappa B (NF-kB) inhibitor on primary human chondrocytes--implications for osteoarthritis.

OBJECTIVE: Idiopathic osteoarthritis is the most common form of osteoarthritis (OA) world-wide and remains the leading cause of disability and the associated socio-economic burden in an increasing aging population. Traditionally, OA has been viewed as a degenerative joint disease characterized by progressive destruction of the articular cartilage and changes in the subchondral bone culminating in joint failure. However, the etiology of OA is multifactorial involving genetic, mechanical and environmental factors. Treatment modalities include analgesia, joint injection with steroids or hyaluronic acid, oral supplements including glucosamine and chondroitin sulfate, as well as physiotherapy. Thus, there is significant interest in the discovery of disease modifying agents. One such agent, glucosamine (GlcN) is commonly prescribed even though the therapeutic efficacy and mechanism of action remain controversial. Inflammatory cytokines, including IL-1ß, and proteinases such as MMP-13 have been implicated in the pathogenesis and progression of OA together with an associated CpG demethylation in their promoters. We have investigated the potential of GlcN to modulate NF-kB activity and cytokine-induced abnormal gene expression in articular chondrocytes and, critically, whether this is associated with an epigenetic process. METHOD: Human chondrocytes were isolated from the articular cartilage of femoral heads, obtained with ethical permission, following fractured neck of femur surgery. Chondrocytes were cultured for 5 weeks in six separate groups; (i) control culture, (ii) cultured with a mixture of 2.5 ng/ml IL-1ß and 2.5 ng/ml oncostatin M (OSM), (iii) cultured with 2mM N-acetyl GlcN (Sigma-Aldrich), (iv) cultured with a mixture of 2.5 ng/ml IL-1ß, 2.5 ng/ml OSM and 2mM GlcN, (v) cultured with 1.0 µM BAY 11-7082 (BAY; NF-kB inhibitor: Calbiochem, Darmstadt, Germany) and, (vi) cultured with a mixture of 2.5 ng/ml IL-1ß, 2.5 ng/ml OSM and 1.0 µM BAY. The levels of IL1B and MMP13 mRNA were examined using qRT-PCR. The percentage DNA methylation in the CpG sites of the IL1ß and MMP13 proximal promoter were quantified by pyrosequencing. RESULT: IL1ß expression was enhanced over 580-fold in articular chondrocytes treated with IL-1ß and OSM. GlcN dramatically ameliorated the cytokine-induced expression by 4-fold. BAY alone increased IL1ß expression by 3-fold. In the presence of BAY, IL-1ß induced IL1B mRNA levels were decreased by 6-fold. The observed average percentage methylation of the -256 CpG site in the IL1ß promoter was 65% in control cultures and decreased to 36% in the presence of IL-1ß/OSM. GlcN and BAY alone had a negligible effect on the methylation status of the IL1B promoter. The cytokine-induced loss of methylation status in the IL1B promoter was ameliorated by both GlcN and BAY to 44% and 53%, respectively. IL-1ß/OSM treatment increased MMP13 mRNA levels independently of either GlcN or BAY and no change in the methylation status of the MMP13 promoter was observed. CONCLUSION: We demonstrate for the first time that GlcN and BAY can prevent cytokine-induced demethylation of a specific CpG site in the IL1ß promoter and this was associated with decreased expression of IL1ß. These studies provide a potential mechanism of action for OA disease modifying agents via NF-kB and, critically, demonstrate the need for further studies to elucidate the role that NF-kB may play in DNA demethylation in human chondrocytes.

[Care for immigrant patients: facts and professionals' perception in 6 primary health care zones in Navarre]. / Atención al paciente inmigrante: realidad y percepción de los profesionales de 6 zonas de Atención Primaria de Navarra.

BACKGROUND: To describe utilisation of health care services and motives for consultation in Primary Care in the native and the immigrant population, and compare this with the perception of primary care professionals. METHODS: Data was collected on health care activity during the year 2006 for all people registered (N=86,966) in the 6 basic health care zones with the highest proportion of immigrants (14.4%) and on the following variables: country of origin, age, sex, year of inscription in the public health service. The health card and OMI-AP programme databases were used. A qualitative methodology of focus groups and in-depth interviews was employed. RESULTS: Seventy-two point four percent of immigrants requested care from the primary care professionals in 2006, of whom 50% proceeded from Ecuador and 70% were between 25 and 44 years old. Eighty-two percent of the natives made consultations and required more referrals to specialised care than the immigrants of the same age group. The most frequent consultation with natives and with immigrants was "acute respiratory infections" (7 to 23% according to age group). The second most frequent with immigrants was "administrative problems". The consultations with immigrants were not related to preventive aspects such as smoking and there were more consultations (p>0.001) for gynaeco-obstetric episodes (10.7%) and those related to work (19%) or psychosomatic problems (8.5%). The perception of the primary care professionals was that the immigrants carry out more consultations than the natives and generate a certain "disorder" in the clinic. CONCLUSION: Immigrants use healthcare services less than the native population. Nonetheless, this fact is not perceived in this way by the primary care professionals. Fewer preventive activities are carried out with immigrants, who suffer from more labour and psychosomatic problems.

Temporal analysis of natural variation for the rate of leaf production and its relationship with flowering initiation in Arabidopsis thaliana.

Vegetative growth and flowering initiation are two crucial developmental processes in the life cycle of annual plants that are closely associated. The timing of both processes affects several presumed adaptive traits, such as flowering time (FT), total leaf number (TLN), or the rate of leaf production (RLP). However, the interactions among these complex processes and traits, and their mechanistic bases, remain largely unknown. To determine the genetic relationships between them, the natural genetic variation between A. thaliana accessions Fei-0 and Ler has been studied using a new population of 222 LerxFei-0 recombinant inbred lines. Temporal analysis of the parental development under a short day photoperiod distinguishes two vegetative phases differing in their RLP. QTL mapping of RLP in consecutive time intervals of vegetative development indicates that Ler/Fei-0 variation is caused by 10 loci whose small to moderate effects mainly display two different temporal patterns. Further comparative QTL analyses show that most of the genomic regions affecting FT or TLN also alter RLP. In addition, the partially independent genetic bases observed for FT and TLN appear determined by several genomic regions with two different patterns of phenotypic effects: regions with a larger effect on FT than TLN, and vice versa. The distinct temporal and pleiotropic patterns of QTL effects suggest that natural variation for flowering time is caused by different genetic mechanisms involved in vegetative and/or reproductive phase changes, most of them interacting with the control of leaf production rate. Thus, natural selection might contribute to maintain this genetic variation due to its phenotypic effects not only on the timing of flowering initiation but also on the rate of vegetative growth.

Natural variation for seed dormancy in Arabidopsis is regulated by additive genetic and molecular pathways.

Timing of germination is presumably under strong natural selection as it determines the environmental conditions in which a plant germinates and initiates its postembryonic life cycle. To investigate how seed dormancy is controlled, quantitative trait loci (QTL) analyses has been performed in six Arabidopsis thaliana recombinant inbred line populations by analyzing them simultaneously using a mixed model QTL approach. The recombinant inbred line populations were derived from crosses between the reference accession Landsberg erecta (Ler) and accessions from different world regions. In total, 11 delay of germination (DOG) QTL have been identified, and nine of them have been confirmed by near isogenic lines (NILs). The absence of strong epistatic interactions between the different DOG loci suggests that they affect dormancy mainly by distinct genetic pathways. This was confirmed by analyzing the transcriptome of freshly harvested dry seeds of five different DOG NILs. All five DOG NILs showed discernible and different expression patterns compared with the expression of their genetic background Ler. The genes identified in the different DOG NILs represent largely different gene ontology profiles. It is proposed that natural variation for seed dormancy in Arabidopsis is mainly controlled by different additive genetic and molecular pathways rather than epistatic interactions, indicating the involvement of several independent pathways.

Molecular markers for establishing distinctness in vegetatively propagated crops: a case study in grapevine.

Distinctness, uniformity and stability (DUS) testing of varieties is usually required to apply for Plant Breeders' Rights. This exam is currently carried out using morphological traits, where the establishment of distinctness through a minimum distance is the key issue. In this study, the possibility of using microsatellite markers for establishing the minimum distance in a vegetatively propagated crop (grapevine) has been evaluated. A collection of 991 accessions have been studied with nine microsatellite markers and pair-wise compared, and the highest intra-variety distance and the lowest inter-variety distance determined. The collection included 489 different genotypes, and synonyms and sports. Average values for number of alleles per locus (19), Polymorphic Information Content (0.764) and heterozygosities observed (0.773) and expected (0.785) indicated the high level of polymorphism existing in grapevine. The maximum intra-variety variability found was one allele between two accessions of the same variety, of a total of 3,171 pair-wise comparisons. The minimum inter-variety variability found was two alleles between two pairs of varieties, of a total of 119,316 pair-wise comparisons. In base to these results, the minimum distance required to set distinctness in grapevine with the nine microsatellite markers used could be established in two alleles. General rules for the use of the system as a support for establishing distinctness in vegetatively propagated crops are discussed.

Effect of nitric oxide on mitochondrial respiratory activity of human articular chondrocytes.

OBJECTIVE: To investigate the effect of nitric oxide (NO) on mitochondrial activity and its relation with the apoptosis of human articular chondrocytes. MATERIALS AND METHODS: Mitochondrial function was evaluated by analysing respiratory chain enzyme complexes, citrate synthase (CS) activities, and mitochondrial membrane potential (Delta psi m). The activities of the mitochondrial respiratory chain (MRC) complexes (complex I: NADH CoQ(1) reductase, complex II: succinate dehydrogenase, complex III: ubiquinol cytochrome c reductase, complex IV: cytochrome c oxidase) and CS were measured in human articular chondrocytes isolated from normal cartilage. The Delta psi m was measured by 5,5',6,6'-tetracholoro-1,1',3,3'-tetraethylbenzimidazole carbocyanide iodide (JC-1) using flow cytometry. Apoptosis was analysed by flow cytometry. The mRNA expression of caspases was analysed by ribonuclease protection analysis and the detection of protein synthesis by western blotting. Sodium nitroprusside (SNP) was used as an NO compound donor. RESULTS: SNP at concentrations higher than 0.5 mmol/l for 24 hours induced cellular changes characteristic of apoptosis. SNP elicited mRNA expression of caspase-3 and caspase-7 and down regulated bcl-2 synthesis in a dose and time dependent manner. Furthermore, 0.5 mM SNP induced depolarisation of the mitochondrial membrane at 5, 12, and 24 hours. Analysis of the MRC showed that at 5 hours, 0.5 mM SNP reduced the activity of complex IV by 33%. The individual inhibition of mitochondrial complex IV with azide modified the Delta psi m and induced apoptosis. CONCLUSIONS: This study suggests that the effect of NO on chondrocyte survival is mediated by its effect on complex IV of the MRC.

The biological action of hyaluronan on human osteoartritic articular chondrocytes: the importance of molecular weight.

OBJECTIVES: The intra-articular injection of hyaluronan (HA) was originally used in the treatment of osteoarthritis (OA) to increase the viscosity of synovial fluid. However, some findings suggest that the activity of HA cannot be solely explained by its biomechanical properties. The aim of this study was to analyze the in vitro biological effects of HA on human OA chondrocytes and the impact of its molecular weight (MW) on those effects. METHODS: Cells were isolated from cartilage obtained during joint replacement surgery in OA patients. The chondrocytes were cultured for 24 hours to detect prostaglandin E2 (PGE2) and for 48 hours to measure nitric oxide (NO), after which they were pre-incubated with HA and stimulated with interleukin-1 (IL-1) at 5 ng/ml. Two commercial HA preparations with different MWs were used: Hyalgan (500-730 kDa, HA, Bioibérica S.A.) and Synvisc (hylan of 6,000 kDa, Biomatrix Inc). NO was detected by the Greiss reaction and PGE2 was quantified by a commercial EIA in the supernatant. Apoptosis was induced by an NO donor (sodium nitroprusside, SNP) and the effect of HA on apoptosis was quantified by flow cytometry. RESULTS: Neither HA preparation studied had any effect on the basal production of NO or PGE2. However, the 500-730 kDa HA at 200 microg/ml reduced the synthesis of both IL-1-induced NO and PGE2 by 70% and 45% respectively. Furthermore both HA preparations at 200 microg/ml decreased the apoptosis induced by SNP, 500-730 kDa to 40% and 6,000 kDa to 36%. CONCLUSION: HA may induce biological effects in addition to acting as a viscoelastic substance. This study suggests that HA preparations are different due to differences in biological activity resulting from MW.

Plan de cuidados al paciente geriátrico con incontinencia urinaria de urgencia / Care's plan for the geriatric patient with urgency's urinary incontinence

La incontinencia urinaria de urgencia afecta a una población muy amplia, y especialmente a la población anciana, teniendo gran repercusión en los ámbitos físico, psicológico y, social de las personas y afectando significativamente en su calidad de vida. El proceso fisiológico de envejecer produce, entre otras modificaciones, una pérdida de elasticidad del músculo detrusor, siendo ésta la causa principal que desencadena la incontinencia urinaria de urgencia en la población anciana. Desde el ámbito enfermero es importante conocer la fisiopatología de la incontinencia urinaria de urgencia y sus formas de tratamiento, para poder ayudar al paciente geriátrico a adaptarse a las nuevas demandas de autocuidado que aparecen como consecuencia de este problema de salud. En este artículo se propone un plan de cuidados dirigido al anciano con incontinencia urinaria de urgencia, que incluye una valoración focalizada y las intervenciones en los diagnósticos enfermeros que se asocian con mayor frecuencia a esta situación (AU)

[Community collection of syringes by parenteral drug users complementary to syringe exchange programs]. / RECOJE: recogida comunitaria de jeringas por usuarios de drogas por vía parenteral como complemento a los programas de intercambio.

OBJECTIVES: To contribute to harm reduction in injecting drug users (IDUs) by the implementation of a programme to withdraw abandoned syringes from public places, as well as to collaborate on the social reinsertion of IDUs. METHODS: Descriptive study of the Syringe Withdrawal Community (RECOJE in Spanish), carried out from 1997 to 1999 by IDUs, the We Are Helpful Association (ASUT in Spanish) and the Social and Health Prevention and Attention Service (SAPS in Spanish) of the Red Cross, in Ciutat Vella (Barcelona, Spain). RESULTS: 4,332 syringes were withdrawn, 849 in 1997, 1,324 in 1998 and 2,159 in 1999. The mean number of syringes withdrawn each time was 57 33. 45 different IDUs worked in RECOJE. 74.2% of the abandoned syringes were found in parks, gardens, car parks and open spaces. CONCLUSIONS: RECOJE can be a valid programme for harm reduction complementary to the exchange syringe programmes. It could improve the self-esteem, external image and organization of IDUs, contributing to their social reinsertion.

[Analysis of arterial pressure variability among individuals with white coat hypertension and essential arterial hypertension using blood pressure ambulatory monitoring]. / Análisis de la variabilidad de la presión arterial entre individuos con hipertensión de bata blanca e hipertensión arterial esencial mediante monitorización ambulatoria de la presión arterial.

OBJECTIVES: To compare patients with "white coat" hypertension (WCH) and essential hypertension for variability in blood pressure (BP) taken in the clinic. DESIGN: Crossover study. SETTING: La Orden Health Centre, Huelva. PATIENTS: 126 people with light-to-moderate hypertension, de novo or not, being monitored but not treated by drugs. Two groups were formed: WCH defined by mean daily systolic and distolic pressures below 135 and 85 mmHg, and hypertension when the BP was over 140 and/or 90 mmHg. MEASUREMENTS AND MAIN RESULTS: BP was taken in the clinic in two periods: by day (7 a.m. to midnight) and at night (00.01 to 06.59), with readings every 15 and 30 minutes, respectively. Variability was compared by analysing: a) variability indices (VI), and b) coefficients of variation (CV) in BP. A regression test and multiple correlation was used. Both groups (hypertension and WCH) contained 63 people (average age: 53.4 +/- 10 and 50 +/- 10). No significant differences between the two groups in the VI were observed, except greater variability in the VI of the nocturnal systolic pressure of the hypertension group. The CV was higher in the WCH group for all pressures, with differences for 24-hour and day-time systolic pressure. The VI of systolic pressure correlated significantly with its overall clinical systolic pressure in the 24-hour period, with the day and night-time readings and with age in the day-time period. CONCLUSIONS: Variability in the blood pressure of people with essential hypertension and WCH does not differ. Variability in systolic pressure shows positive correlation with clinical systolic pressure and, to a lesser extent, with age.

Prolactin levels in patients with systemic lupus erythematosus: a case controlled study.

Recent accumulated evidence suggests that prolactin (PRL) is an important immunomodulator and might have a role in the pathogenesis of systemic lupus erythematosus (SLE). Our aim was to assess the frequency of hyperprolactinemia in women with SLE and to evaluate its correlation with disease activity. PRL plasma levels were measured in 36 women with SLE and 20 age-matched healthy controls. We excluded patients with renal and/or hepatic failure, pregnant patients and patients taking drugs which could increase PRL levels. Disease activity was assessed using the SLE disease activity index (SLEDAI). Patients with a score > 10 were considered active. In patients and controls, PRL levels were determined by radioimmunoassay (RIA) during the first part of the menstrual cycle (between the 5th and 8th day) (normal value < 20 ng/ml). Ten of 36 (27.7%) SLE patients had high PRL levels (> 20 ng/ml). The mean PRL level was higher in SLE than in the control group (17.1+/-12.9 s.d. vs 9.9+/-3.5, P < 0.01). Patients with active disease had a trend to higher mean PRL levels than inactive patients although this difference was not statistically significant (21.1+/-4.8 vs 14.8+/-6.9, P = 0.09). No correlation was found between PRL levels and SLEDAI score. Furthermore, no significant correlation was found between PRL levels and any clinical or serological finding.

[Therapeutic compliance in dyslipidemias. A trial of the efficacy of health education]. / Cumplimiento terapéutico de las dislipemias. Ensayo sobre la eficacia de la educación sanitaria.

OBJECTIVE: To analyse the efficacy of health education (HE) through group session with postal back-up in furthering compliance with therapy for Lipaemia. DESIGN: Controlled clinical trial, with random distribution. SETTING: Primary care. PATIENTS: 110 patients with Hypercholesterolaemia, with new diagnoses or not in treatment, in which medical treatment with statins was indicated as a start or change in medication. INTERVENTIONS: They were distributed in two groups at random, with observation four months after being included in the study and appointments after one, two and four months. 1. Control group (CG): 55 patients who received HE from their family doctor. 2. Intervention group (IG): 55 patients whose HE was monitored: a) a group HE session and b) back-up by letter sent to their homes. MEASUREMENTS AND RESULTS: Patients whose consumption was between 80 and 110% of the amount prescribed were defined as compliant. The pill count was recorded. The percentages of compliant patients and mean compliance (chi squared, Student's t) were analysed. 108 individuals, 41 men and 67 women, completed the trial. There was no difference between the two groups as to age, sex, evolution time, number of diseases and dosage of medicines consumed. 71.3% were compliant (CI, 62.8-79.8%), CG = 61.8% and IG 81.1% (p < 0.05). The mean percentage of compliance was 86.1 +/- 14.3 overall, with CG = 83.8 +/- 14 and IG 88.5 +/- 14 (p = NS). CONCLUSIONS: The HE intervention with a group session and postal back-up is an effective way of improving therapeutic compliance in cases of hypercholesterolaemia.