Mortality and cardiovascular risk in vasculitis ANCA. Importance of hypertension and renal function. Experience from southern Spain / Mortalidad y riesgo cardiovascular en vasculitis ANCA. Importancia de la hipertension y funcion renal. Experiencia del sur de España

Background: Cardiovascular disease (CVD) is one of the principal causes of death in antineutrophil cytoplasmic antibody-(ANCA)-associated vasculitis (AAV). Objectives: To evaluate the mortality and it's causes and CVD and its vascular risk factors (VRFs) in AAV patients in Andalusia. Methods: A multicenter cohort of 220 AAV patients followed-up from 1979 until June 2020 was studied in Andalussia, south of Spain. The information, including socio-demographic and clinical data was recorded retrospectively through chart review. Data was analysed using Chi2, ANOVA and Cox proportional hazards regresion as uni and multivariate test with a 95% confidence interval (CI). Results: During a mean ± standard deviation follow-up of 96.79 ± 75.83 months, 51 patients died and 30 presented at least one CVE. Independent prognostic factors of mortality were age (HR 1.083, p=0.001) and baseline creatinine (HR 4.41, p=0.01). Independent prognostic factors of CVE were age [hazard ratio (HR) 1.042, p=0.005] and the presence of hypertension (HTN) six months after diagnosis (HR 4.641, p=0.01). HTN, diabetes and renal failure, all of these important VRFs, are more prevalent in AAV patients than it is described in matched general population. Conclusions: Age and baseline renal function, but not CVEs, are predictors of mortality and age and early HTN are independent predictors for having a CVE. CVD screening in AAV patients is demanded.(AU)

Introducción: La enfermedad cardiovascular (ECV) es una de las principales causas de muerte en las vasculitis asociadas a anticuerpos anticitoplasma de neutrófilos (ANCA) (VAA). Objetivos: Evaluar la mortalidad y sus causas, entre ellas la ECV y sus factores de riesgo vascular (FRV) en pacientes con VAA en Andalucía. Métodos: Se estudió una cohorte multicéntrica de 220 pacientes con VAA seguidos desde 1979 hasta junio de 2020 en Andalucía. La información, incluidos los datos sociodemográficos y clínicos, se registró retrospectivamente a través de la revisión de historias clínicas. Los datos se analizaron mediante Chi2, ANOVA y regresión de riesgos proporcionales de Cox de forma uni y multivariante con un intervalo de confianza (IC) del 95%. Resultados: Durante un seguimiento medio y desviación estándar de 96,79 ± 75,83 meses, 51 pacientes fallecieron y 30 presentaron al menos un ECV. Los factores pronósticos independientes de mortalidad fueron la edad (HR 1,083, p=0,001) y la creatinina basal (HR 4,41, p=0,01). Los factores pronósticos independientes de ECV fueron la edad [hazard ratio (HR) 1,042, p=0,005] y la presencia de hipertensión arterial (HTA) seis meses después del diagnóstico (HR 4,641, p=0,01). La prevalencia de HTA, diabetes e insuficiencia renal fue elevada o muy elevada en comparación con la población general emparentada, todos FRCV determinantes para el pronóstico de estos pacientes. Conclusiones: La edad y la función renal basal son predictores de mortalidad y la edad y la HTA de aparición precoz son predictores independientes de tener ECV. Se recomienda el cribado de FRCV en pacientes con vasculitis ANCA.(AU)

Mortality and cardiovascular risk in vasculitis ANCA. Importance of hypertension and renal function. Experience from southern Spain.

BACKGROUND: Cardiovascular disease (CVD) is one of the principal causes of death in antineutrophil cytoplasmic antibody-(ANCA)-associated vasculitis (AAV). OBJECTIVES: To evaluate the mortality and it's causes and CVD and its vascular risk factors (VRFs) in AAV patients in Andalusia. METHODS: A multicenter cohort of 220 AAV patients followed-up from 1979 until June 2020 was studied in Andalussia, south of Spain. The information, including socio-demographic and clinical data was recorded retrospectively through chart review. Data was analysed using Chi2, ANOVA and Cox proportional hazards regresion as uni and multivariate test with a 95% confidence interval (CI). RESULTS: During a mean ± standard deviation follow-up of 96.79 ± 75.83 months, 51 patients died and 30 presented at least one CVE. Independent prognostic factors of mortality were age (HR 1.083, p=0.001) and baseline creatinine (HR 4.41, p=0.01). Independent prognostic factors of CVE were age [hazard ratio (HR) 1.042, p=0.005] and the presence of hypertension (HTN) six months after diagnosis (HR 4.641, p=0.01). HTN, diabetes and renal failure, all of these important VRFs, are more prevalent in AAV patients than it is described in matched general population. CONCLUSIONS: Age and baseline renal function, but not CVEs, are predictors of mortality and age and early HTN are independent predictors for having a CVE. CVD screening in AAV patients is demanded.

Metallodendrimers Unveiled: Investigating the Formation and Features of Double-Decker Silsesquioxane-Based Silylferrocene Dendrimers.

Dendrimers exhibiting reversible redox properties have attracted extensive attention for their potential as electron transfer mediators, catalysts, and molecular sensors. In this study, we introduce intriguing G1 and G2 dendrimers featuring double-decker silsesquioxane cores and silylferrocene moieties. Through a carefully orchestrated sequence of condensation, reduction, and hydrosilylation reactions, these compounds were synthesized and comprehensively characterized spectroscopically and spectrometrically. Our investigation also encompassed the examination of their properties, including thermal stability, solubility in common organic solvents, and electrochemical behavior. We determined that these dendrimers possess the capability to form monolayers on platinum electrodes, which we conclusively demonstrated through the probing of cyclic voltammetry, electrochemical impedance spectroscopy, and scanning electron microscopy imaging. Notably, this study marks the first-ever example of modifying double-decker silsesquioxane cores with ferrocene groups while simultaneously representing one of the scarce instances of dendrimers exhibiting an open double-decker silsesquioxane core.

Self-assembled gold decorated polydopamine nanospheres as electrochemical sensor for simultaneous determination of ascorbic acid, dopamine, uric acid and tryptophan.

Herein, a new sensor based on screen-printed carbon electrodes covalently modified with self-assembled gold-decorated-polydopamine nanospheres (Au-PDNs) is reported. The sensor was applied to the simultaneous determination of the biologically significant molecules ascorbic acid (AA), dopamine (DA), uric acid (UA) and tryptophan (TR). The Au-PDNs were anchored to gold nanoparticles electrodeposited onto the bare electrodes via cysteamine-glutaraldehyde bridges, and were characterized by scanning and transmission electron microscopies. The stepwise fabrication of the electrodes and their electrochemical responses were evaluated by cyclic voltammetry, electrochemical impedance spectroscopy and differential pulse voltammetry. The response of the new device to these analytes is pH-dependent, which allows selecting the best working conditions as a function of the sample characteristics. At pH values of 3.0 and 8.0, it was possible to determine simultaneously AA, UA and TR in presence of DA, and DA, UA and TR in presence of AA respectively, with very wide linear ranges and high sensitivities. The simultaneous determination of AA, DA, UA and TR was possible at pH 6.0 with competitive sensitivities in two consecutive linear ranges, between 10-80 µM and 80-240 µM; 1-160 µM and 160-350 µM; 10-120 µM and 120-350 µM; and 1-160 µM and 160-280 µM, respectively. The obtained limits of detection were 0.2 nM, 0.1 nM, 0.1 nM and 0.1 nM, respectively.

Direct quantification of inorganic iodine in seawater by mixed-mode liquid chromatography-electrospray ionization-mass spectrometry.

Atmospheric iodine plays a relevant role in climate change. Bearing in mind that most of this iodine comes from the oceans, analytical methods capable of determining iodine in a challenging matrix as seawater are necessary. In this work, the first method capable of direct determination of total inorganic iodine in seawater at subnanomolar level based on mixed-mode liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) without any sample treatment is presented. Analytical characteristics of the developed method were studied in terms of linear range, limits of detection and quantification, precision, trueness, matrix effect, and robustness. The detection limit for iodide was as low as 0.16 nM, injecting 5 µL of seawater without any sample treatment and the working linear range of four orders of magnitude was wide enough to cover the broad concentration range observed in seawater samples. Average values for repeatability and intermediate precision were 4.1% and 8.1%, respectively. The suitability of the method was demonstrated through its application to the analysis of several types of samples, including seawater samples taken at different locations along the Spanish Mediterranean coast and some domestic iodized salts. According to the results obtained, the method developed is rapid, easy to apply and to be automated, avoids sample treatment and requires only few microliters of sample. Furthermore, it has a low detection limit and allows the quantification of inorganic iodine over a wide concentration range.

Ortodoncia con sistema de alineación con placas en pacientes con discapacidad / Orthodontics with plate aligning system in disabled patients

En personas con discapacidad se presentan las enfermedades prevalentes de la cavidad bucal con mayor frecuencia. Dentro de éstas, las alteraciones oclusales, como el apiñamiento dentario, son muy frecuentes y se considera responsable de exacerbar la patología gingival, periodontal y la estética, con impacto en la salud bucodental y la calidad de vida de estos pacientes. La técnica basada en el uso de placas alineadoras es sencilla, no invasiva y fundamentalmente preventiva de la enfermedad buco-dental (AU)

Direct, indirect and total effectiveness of bivalent HPV vaccine in women in Galicia, Spain.

Bivalent human papillomavirus (HPV) vaccine was incorporated into the childhood vaccination calendar in Galicia, Spain in 2008. The objectives of this study were to estimate direct, indirect and total effectiveness of HPV vaccine and to identify sexual habits changes in the post-vaccination period in Galicia, Spain.Endocervical scrapings of 745 women attending 7 Health Areas of the Galician Public Health Service were collected in the post-vaccination period, from 2014-2017. Two groups were studied: women born between 1989 and 1993 (n = 397) and women born in 1994 or later (n = 348). Twelve high-risk human papillomavirus (HR-HPV) genotypes were detected by Cobas® 4800 HPV test (Roche Diagnostics, Mannheim, Germany). The Linear Array® HPV Genotyping Test (Roche Diagnostics) was used for HR-HPV genotype detection other than HPV 16/18. Information about sexual habits was collected by a self-filled questionnaire. Post-vaccination data were compared to previously published pre-vaccination data obtained between 2008 and 2010 in Galicia from women of the same age (18-26 years old, n = 523). The Stata 14.2 software was employed for statistical analyses.Data from 392 unvaccinated and 353 vaccinated women were compared. For unvaccinated and vaccinated women, HPV 16/18 prevalence was 9.2% and 0.8%, respectively, and HPV 31/33/45 prevalence was 8.4% and 1.1%, respectively. Direct, indirect and total effectiveness of the HPV vaccine were (%, 95% CI): 94 (72-99), 30 (-11-56) and 95 (79-99), respectively, for HPV 16/18 and 83 (46-94), -10 (-88-33) and 84 (54-94), respectively, for HPV 31/33/45. The number of women with first intercourse before 17 years old and 3 or more sexual partners along life was higher in the post-vaccination period (p < 0.05). A positive impact of bivalent HPV vaccine was observed, both on direct and cross protection. Sexual habits could have changed in the post-vaccination period.

Optimizing Digital Mammographic Image Quality for Full-Field Digital Detectors: Artifacts Encountered during the QC Process.

Early detection of breast cancer through routine mammographic screening has been shown to reduce mortality from breast cancer by up to 30% in multiple studies. However, this reduction of mortality is possible only with careful attention to image quality by the medical physicist, radiologic technologist, and interpreting radiologist. The accepted quality control (QC) processes for analog mammography are well established. However, now that use of digital units is widespread in both the United States and internationally, information regarding the necessary steps and the inherent challenges that might be encountered at each step needs to be elucidated. In this review, the essential steps of the QC process for digital mammography are reviewed, with special attention to the possible problems that can occur during the QC process, many of which can lead to image artifacts. For each of the daily, weekly, monthly, and semiannual QC tests, we review the steps and expected performance and provide examples of some of the common artifacts that may be encountered. Understanding the components of the QC process and recognizing problems that may result in a suboptimal image is critical to ensure optimal image quality in an effort to maximize early detection of breast cancer.

New carbosilane polymers with interacting ferrocenes as support and bioelectrocatalysts of oxidases to develop versatile and specific amperometric biodevices.

In this work, the bioelectrocatalytical properties and kinetic characteristics of new oxidase amperometric biosensors based on two different ferrocene functionalized carbosilane polymers, polydiallylmethylsilane (PDAMS) and polymethyldiundecenylsilane (PMDUS) are described. In the development of these biodevices, glucose oxidase has been used as example of oxidase enzyme, and two different immobilization procedures have been studied. The polymer-modified electrodes act as efficient transducers for glucose sensing in anodic and cathodic aerobic conditions and also in anodic anaerobic conditions, and this fact turns them into useful devices for a wide field of applications. PMDUS has shown to be the bioelectrocatalyst with best kinetic and analytical properties in aerobic media while PDAMS was better in anaerobic conditions. The best aerobic biosensor developed displayed a strictly linear range from 0 to 3.0 mM, a detection limit of 7.8 µM and a response time less than 2 s in an ascorbate interference free work potential interval. The apparent Michaelis-Menten constant was calculated to be 1.36 mM according to the Lineweaver-Burk equation.

Anti-apoptotic effect of transforming growth factor-beta1 on human articular chondrocytes: role of protein phosphatase 2A.

OBJECTIVE: To study whether transforming growth factor-beta1 (TGF-beta1) is able to protect human chondrocytes from apoptosis and to analyze the role of phosphatases in the possible anti-apoptotic effect of TGF-beta1. METHODS: Cartilage was obtained from patients with osteoarthritis (OA) who were undergoing joint replacement; normal cartilage was obtained from cadavers who had no history of joint disease. Chondrocytes stimulated with tumor necrosis factor-alpha (TNF-alpha) plus Ro 31-8220 (a specific inhibitor of mitogen-activated kinase phosphatase-1 - MKP-1) were employed as an in vitro model of apoptosis. Apoptosis was assessed by flow cytometry and a cell death immunoassay. Protein phosphatase 2A (PP2A) activity was estimated by measuring the absorbance of a molybdate:malachite green:phosphate reaction complex. MKP-1, bcl-2 and bax expressions were quantified by western blot. RESULTS: In OA cells, TGF-beta1 significantly reduced the percentage of hypo-diploid chondrocytes, as well as the percentage of internucleosomal DNA breakage. However, in normal chondrocytes, TGF-beta1 did not reduce apoptosis, as assessed by both the percentage of hypo-diploid chondrocytes and internucleosomal DNA breakage. MKP-1 expression did not show significant modulation in OA or normal chondrocytes. However, PP2A activity was differentially modulated in normal and OA chondrocytes. In OA chondrocytes, PP2A activity was not altered by TGF-beta1 stimulation; however in normal chondrocytes PP2A activity was significantly activated by TGF-beta1. The preincubation of normal chondrocytes with TGF-beta1 plus the PP2A inhibitor protein, IPP2A, reduced internucleosomal DNA breakage when compared with TGF-beta1 stimulation alone. The bcl-2/bax protein ratio was significantly higher in TGF-beta1 plus IPP2A preincubated normal chondrocytes than in cells stimulated with TGF-beta1 alone. CONCLUSION: By manipulating the degree of PP2A activity, these results show the major role that PP2A plays in the outcome of TGF-beta1 signal transduction. These data suggest that PP2A could be a pivotal regulator of anti-apoptotic TGF-beta1-induced effects.

Differential effects of tumor necrosis factor-alpha and interleukin-1beta on cell death in human articular chondrocytes.

OBJECTIVE: The death of chondrocytes by apoptosis is characteristic of degenerative joint diseases, such as osteoarthritis (OA). Tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) have been shown to play an important role in the development of OA. In this study we analyzed the effects of TNF-alpha and IL-1beta on cell death in normal human chondrocytes. METHODS: Normal human chondrocytes were isolated from knee cartilage obtained at autopsy from 30 adult cadaveric donors. The cells were stimulated with TNF-alpha (10 ng/ml) or IL-1beta (5 ng/ml) in the presence or absence of Ro 31-8220 (Ro: a structurally related analog of bisindolylmaleimide that inhibits mitogen-activated protein kinase phosphatase 1 [MKP-1]) (Ro; 10 microM), an MKP-1 inhibitor, which induces apoptosis in chondrocytes. Apoptosis was evaluated by flow cytometry (propidium iodide) and nuclear morphology was evaluated with 4',6'-dianidino-2-phenylindole dihydrochloride. The expressions of caspase-8, -7 and -3 and Bcl-2 were analyzed by Western blot and the activation of caspase-3 and -8 was measured by flow cytometry. Prostaglandin E2 (PGE2) was evaluated by enzyme-linked immunosorbent assay. RESULTS: At 24 h the percentage of apoptotic (hypodiploid) nuclei induced by TNF-alpha+Ro was higher than the level induced by Ro alone. The combination of IL-1beta (5 ng/ml) with Ro did not show a synergistic effect. A morphological analysis demonstrated that treatment with TNF-alpha+Ro resulted in a large number of cells with condensed nuclei and DNA fragmentation. Western blot studies indicated that IL-1beta+Ro did not induce the time-dependent activation of caspase-8, -7 and -3 as seen with TNF-alpha+Ro. As quantified by flow cytometry, TNF-alpha+Ro induced a higher level of caspase-3 and -8 activation than that seen with IL-1beta+Ro. Pre-incubation for 2h with caspase inhibitors for caspase-3, -7, -8 and pan-caspase significantly decreased the hypodiploid DNA peak induced by treatment with TNF-alpha+Ro at 24 h. Indomethacin increased the cell death induced by IL-1beta+Ro; however, apoptosis induced by TNF-alpha+Ro was not modified by indomethacin. CONCLUSIONS: These results confirm that TNF-alpha and IL-1beta regulate apoptosis differently in this human chondrocyte model and that the differing effects of these cytokines are PGE2-independent. Indomethacin potentiates the effect of IL-1 on cell death and this may explain the reported effect of indomethacin on the progression of joint destruction.

Salidas terapéuticas: espacio lúdico para la integración / Therapeutic outings: a playing space for integration

Las actividades de ocio y tiempo libre deben ser una parte importante de cualquier programa integral de rehabilitación en personas con enfermedad mental grave (AU)

A Program for preparation for leisure time it seems to us a esencial element of a Rehabilitation Unit for patients with an important mental illness (AU)

Mitochondrial activity is modulated by TNFalpha and IL-1beta in normal human chondrocyte cells.

OBJECTIVE: Pro-inflammatory cytokines play an important role in osteoarthritis (OA). In osteoarthritic cartilage, chondrocytes exhibit an alteration in mitochondrial activity. This study analyzes the effect of tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta) on the mitochondrial activity of normal human chondrocytes. MATERIALS AND METHODS: Mitochondrial function was evaluated by analyzing the activities of respiratory chain enzyme complexes and citrate synthase, as well as by mitochondrial membrane potential (Deltapsim) and adenosine triphosphate (ATP) synthesis. Bcl-2 family mRNA expression and protein synthesis were analyzed by RNase protection assay (RPA) and Western-blot, respectively. Cell viability was analyzed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) and apoptosis by 4', 6-diamidino-2-phenylindole dihydrochloride (DAPI) stain. Glycosaminoglycans were quantified in supernatant by a dimethyl-methylene blue binding assay. RESULTS: Compared to basal cells, stimulation with TNFalpha (10 ng/ml) and IL-1beta (5 ng/ml) for 48 h significantly decreased the activity of complex I (TNFalpha=35% and IL-1beta=35%) and the production of ATP (TNFalpha=18% and IL-1beta=19%). Both TNFalpha and IL-1beta caused a definitive time-dependent decrease in the red/green fluorescence ratio in chondrocytes, indicating depolarization of the mitochondria. Both cytokines induced mRNA expression and protein synthesis of the Bcl-2 family. Rotenone, an inhibitor of complex I, caused a significant reduction of the red/green ratio, but it did not reduce the viability of the chondrocytes. Rotenone also increased Bcl-2 mRNA expression and protein synthesis. Finally, rotenone as well as TNFalpha and IL-1beta, reduced the content of proteoglycans in the extracellular matrix of normal cartilage. CONCLUSION: These results show that both TNFalpha and IL-1beta regulate mitochondrial function in human articular chondrocytes. Furthermore, the inhibition of complex I by both cytokines could play a key role in cartilage degradation induced by TNFalpha and IL-1beta. These data could be important for understanding of the OA pathogenesis.

Mitochondrial proteomic characterization of human normal articular chondrocytes.

OBJECTIVE: Mitochondrial dysfunctions have been associated with apoptosis, aging and osteoarthritis (OA). Chondrocyte mitochondrial proteins are attractive targets for the study of the metabolism of cartilage degradation. The copurification of "contaminating" proteins has been the major problem in all phases of mitochondrial proteome research. Therefore, we set up a procedure for the proteomic analysis of human chondrocyte mitochondrial proteins. METHOD: Four types of protein extracts were obtained from primary cultured chondrocytes isolated from healthy donors: (1) initial total chondrocyte extract (CE), (2) cytosol-enriched supernatant fraction (CY), (3) crude mitochondria fraction (CM), and (4) pure mitochondria fraction (PM). Mitochondria were purified by density gradient ultracentrifugation. Mitochondrial proteins were separated by means of two-dimensional gel electrophoresis (2-DE) and silver stained. Protein spots were then identified by mass spectrometry using MALDI-TOF/TOF technology. RESULTS: The best 2-DE reference map of mitochondrial proteome was constructed employing PM fraction. Thirty-nine percent of the identified proteins were functionally distributed in the mitochondria, 14% in the endoplasmic reticulum and 36% in the cytoplasm. Examining their biological function, 22% are involved in protein targeting, 12% in signaling, 12% in glycolysis, 10% in RNA, DNA or protein synthesis, 10% in oxidative phosphorylation and 4% in redox. The analysis of mitochondrial Mn-superoxide dismutase (SODM) revealed an age-dependent decrease of this protein. CONCLUSION: PM fraction allowed the obtention of a high quality proteomic map for the study of mitochondrial proteins in human articular chondrocytes. This proteomic approach may be also efficient to analyze both quantitative and qualitative modulations of the mitochondrial proteome in human chondrocytes during aging and pathological conditions such as OA.

Cytokines, tumor necrosis factor-alpha and interleukin-1beta, differentially regulate apoptosis in osteoarthritis cultured human chondrocytes.

OBJECTIVE: This study addresses the effects of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) on cell death in human chondrocytes. METHODS: Osteoarthritis (OA) human chondrocytes stimulated with Actinomycin-D (ActD) were used as a cellular apoptotic model. Caspase family mRNA expression and protein synthesis were analyzed by the ribonuclease protection assay and Western-blot, respectively. Cell viability and apoptosis were evaluated using the 3-[4,5-dimethylthiazol-2yl] 2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry, respectively. Prostaglandin E2 (PGE2) and nitric oxide (NO) were evaluated by enzyme-linked immunosorbent assay (ELISA) and the Griess method, respectively. RESULTS: TNF-alpha and IL-1beta differentially affected the pattern of caspase mRNA expression by human chondrocytes. TNF-alpha induced a gradual increase in caspase-1 and -8 mRNA levels that was not seen with IL-1beta. The time sequence of caspase-3 and -7 inductions by TNF-alpha differs from that induced by IL-1beta. Cell viability was not modified by TNF-alpha or IL-1beta in cultured chondrocytes. Then, we employed ActD as a model to facilitate cell death. Treatment with TNF-alpha and ActD (TNF-alpha/ActD) increased cell death induced by ActD (23%). Treatment with IL-1beta and ActD (IL-1beta/ActD) did not modulate ActD-induced cell death. Similarly, IL-1beta/ActD did not induce an increase in the activation of caspase-3 and -7 and poly (ADP-ribose) polymerase (PARP) cleavage observed by the incubation with TNF-alpha/ActD. These different effects were not due to bcl-2 or mcl-1 levels. Inhibition of PGE2 synthesis by indomethacin increased the cell death induced by IL-1beta/Act-D (59%). An inhibitor of caspase-8 significantly reduced only the TNF-alpha/ActD-induced cell death (58%). CONCLUSION: TNF-alpha and IL-1beta differentially regulate the apoptotic pathway in human chondrocytes. This difference is dependent on PGE2 and caspase-8 levels.

Electrocatalytical properties of polymethylferrocenyl dendrimers and their applications in biosensing.

The electrochemical characterization of polymethylferrocenyl dendrimers deposited onto a platinum electrode and their applications as hydrogen peroxide and glucose sensor are described. The redox dendrimers consist of flexible poly(propileneimine) dendrimer cores functionalised with octamethylferrocenyl units. Amperometric biosensors for glucose were prepared by immobilization of glucose oxidase onto these modified electrodes. The influence of the dendrimer generation and the thickness of the dendrimer layer, the effect of the substrate concentration, and the interferences and reproducibility on the response of the sensors were investigated.

Phosphatase-1 and -2A inhibition modulates apoptosis in human osteoarthritis chondrocytes independently of nitric oxide production.

OBJECTIVE: To characterise the role of phosphatase-1 and -2A (PP1/2A) in the modulation of apoptosis in human osteoarthritis (OA) chondrocytes. METHODS: Human OA chondrocytes were isolated from cartilage obtained from the femoral heads of patients undergoing joint replacement surgery. Cell viability was evaluated by MTT assay. Apoptosis was quantified by ELISA, which measures DNA fragmentation. Nitric oxide (NO) production was evaluated by the Greiss method, and inducible nitric oxide synthase (iNOS) protein synthesis was studied by western blotting. RESULTS: Inhibition of PP1/2A by the specific inhibitor okadaic acid (OKA) dose and time dependently caused a reduction of cell viability (OKA at 50 nmol/l: a reduction to 60% and 43% at 48 and 72 hours, respectively). Genomic DNA from chondrocytes treated with OKA at 50 and 100 nmol/l for 48 hours displayed increased internucleosomal DNA fragmentation by 11 and 13 fields, respectively. Light microscopy and DAPI studies showed that OKA induced DNA condensation and fragmentation, typical of death by apoptosis. The caspase inhibitors Z-VAD-FMK and Z-DEVD-FMK increased cell viability, reduced by OKA at 50 nmol/l to 87% and 73%, respectively. OKA did not increase iNOS protein synthesis or NO production. CONCLUSION: PP1/2A modulate apoptosis in human OA chondrocytes; this is independent of NO production but dependent on caspases.

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.

Papel de la mitocondria en la artrosis / The role of mitochondria in osteoarthritis

No disponible

EP2/EP4 signalling inhibits monocyte chemoattractant protein-1 production induced by interleukin 1beta in synovial fibroblasts.

BACKGROUND: Besides its proinflammatory properties, prostaglandin E(2) (PGE(2)) acts as a regulator of the expression of inducible genes. Inhibition of PGE(2) synthesis might thus result in a paradoxical deleterious effect on inflammation. OBJECTIVE: To examine the effect of PGE(2) on monocyte chemoattractant protein-1 (MCP-1) expression in cultured synovial fibroblasts (SF) stimulated with interleukin (IL)1beta. METHODS: MCP-1 expression was assessed in SF stimulated with IL1beta in the presence of PGE(2) or different NSAIDs by RT-PCR or northern blot and immunocytochemistry. Expression of cyclo-oxygenase (COX) isoforms was studied by western blot techniques. The role of PGE(2) receptors (EP) in PGE(2) action was assessed employing EP receptor subtype-specific agonists. RESULTS: PGE(2) significantly inhibited IL1beta induced MCP-1 expression in SF in a dose dependent manner. IL1beta increased COX-2 and did not alter COX-1 synthesis in SF. 11-Deoxy-PGE(1), an EP(2)/EP(4) agonist, reproduced PGE(2) action on MCP-1 expression. Butaprost, a selective EP(2) agonist, was less potent than PGE(2). Sulprostone, an EP(1)/EP(3) agonist, had no effect on IL1beta induced MCP-1 expression. Inhibition of endogenous PGE(2) synthesis by NSAIDs further enhanced MCP-1 mRNA expression in IL1beta stimulated SF, an effect prevented by addition of exogenous PGE(2). CONCLUSION: Activation of EP(2)/EP(4) receptors down regulates the expression of MCP-1 in IL1beta stimulated SF, while PGE(2) pharmacological inhibition cuts off this signalling pathway and results in a superinduction of MCP-1 expression. The data suggest that NSAIDs may intercept a natural regulatory circuit controlling the magnitude of inflammation, which questions their continuous administration in inflammatory joint diseases.