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Objective: To determine the Dynamin-related protein 1 (DRP1) regulation of mitochondrial fission in chondrocytes under pathological conditions, an area which is underexplored in osteoarthritis pathogenesis.Design: DRP1 protein expression was determined by immunohistochemistry (IHC) or immunofluorescence (IF) staining of cartilage sections. IL-1β-induced DRP1 mRNA expression in chondrocytes was quantified by qPCR and protein expression by immunoblotting. Mitochondrial fragmentation in chondrocytes was visualized by MitoTracker staining or IF staining of mitochondrial marker proteins or by transient expression of mitoDsRed. Mitochondrial reactive oxygen species (ROS) levels were determined by MitoSOX staining. Apoptosis was determined by lactate dehydrogenase (LDH) release assay, Caspase 3/7 activity assay, propidium iodide (PI), and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining and IF staining of cleaved caspase 3. Cytochrome c release was determined by confocal microscopy. Surgical destabilization of the medial meniscus (DMM) was used to induce osteoarthritis (OA) in mice.Results: Expression of DRP1 and mitochondrial damage was high in human OA cartilage and in the joints of mice subjected to DMM surgery which also showed increased chondrocytes apoptosis. IL-1β-induced mitochondrial network fragmentation and chondrocyte apoptosis via modulation of DRP1 expression and activity and induce apoptosis via Bax-mediated release of Cytochrome c. Pharmacological inhibition of DRP1 activity by Mdivi-1 blocked IL-1β-induced mitochondrial damage and apoptosis in chondrocytes. Additionally, IL-1β-induced activation of extracellular signal-regulated kinase 1/2 (ERK1/2) is crucial for DRP1 activation and induction of mitochondrial network fragmentation in chondrocytes as these were blocked by inhibiting ERK1/2 activation.Conclusions: These findings demonstrate that ERK1/2 is a critical player in DRP1-mediated induction of mitochondrial fission and apoptosis in IL-1β-stimulated chondrocytes. [ABSTRACT FROM AUTHOR]

Objective: Lysosomes are the major catabolic organelle of the cell and regulate the macromolecular and organelle turnover and programmed cell death. Here, we investigated the lysosome dysfunction in cartilage and its role in chondrocytes apoptosis and the associated mechanism.Design: Lysosomal acidification in Osteoarthritis (OA) and aged cartilage was determined by LysoSensor staining. Lysosomal function in chondrocytes was blocked by siRNA mediated depletion of Lysosomal Associated Membrane Protein 2 (LAMP2) or with lysosome inhibitors. Chondrocyte apoptosis was determined by LDH release, Caspase-3/7 activation, TUNEL and PI uptake assays. Loss of mitochondrial membrane potential (MMP/ΔΨM) and mitochondrial superoxide level was determined by JC-1 and MitoSOX staining, respectively. Colocalization of mitochondria with BCL2 associated X (BAX) and Cytochrome c was determined by immunostaining. Destabilization of medial meniscus (DMM) was performed to induce OA in mice.Results: Lysosomal acidification was found to be significantly decreased in aged mouse and human and mouse OA cartilage which also showed increased chondrocyte apoptosis. Inhibition of lysosomal function resulted in increased oxidative stress, accumulation of dysfunctional mitochondria and apoptosis in chondrocytes in monolayer and in cartilage explant cultures. Depletion of LAMP2 expression or treatment of chondrocytes with lysosomal function inhibitors increased the expression and mitochondrial translocation of BAX leading to Cytochrome c release. Lysosomal dysfunction-induced apoptosis in chondrocytes was not blocked by antioxidants MitoTempo or Diphenyleneiodonium (DPI) but was abrogated by inhibiting BAX.Conclusion: Lysosomal dysfunction induce apoptosis in chondrocytes through BAX-mediated mitochondrial damage and release of Cytochrome c. Our data points to lysosomal function restoration and/or BAX inhibition in chondrocytes as a therapeutic approach for OA. [ABSTRACT FROM AUTHOR]

Structure-based constitutive models might help in exploring mechanisms by which arterial wall histology is linked to wall mechanics. This study aims to validate a recently proposed structure-based constitutive model. Specifically, the model’s ability to predict mechanical biaxial response of porcine aortic tissue with predefined collagen structure was tested. Histological slices from porcine thoracic aorta wall ( n = 9) were automatically processed to quantify the collagen fiber organization, and mechanical testing identified the non-linear properties of the wall samples ( n = 18) over a wide range of biaxial stretches. Histological and mechanical experimental data were used to identify the model parameters of a recently proposed multi-scale constitutive description for arterial layers. The model predictive capability was tested with respect to interpolation and extrapolation. Collagen in the media was predominantly aligned in circumferential direction (planar von Mises distribution with concentration parameter b M = 1.03 ± 0.23 ), and its coherence decreased gradually from the luminal to the abluminal tissue layers (inner media, b = 1.54 ± 0.40 ; outer media, b = 0.72 ± 0.20 ). In contrast, the collagen in the adventitia was aligned almost isotropically ( b A = 0.27 ± 0.11 ), and no features, such as families of coherent fibers, were identified. The applied constitutive model captured the aorta biaxial properties accurately (coefficient of determination R 2 = 0.95 ± 0.03 ) over the entire range of biaxial deformations and with physically meaningful model parameters. Good predictive properties, well outside the parameter identification space, were observed ( R 2 = 0.92 ± 0.04 ). Multi-scale constitutive models equipped with realistic micro-histological data can predict macroscopic non-linear aorta wall properties. Collagen largely defines already low strain properties of media, which explains the origin of wall anisotropy seen at this strain level. The structure and mechanical properties of adventitia are well designed to protect the media from axial and circumferential overloads. [ABSTRACT FROM AUTHOR]

Abstract: Woody plants growing in seasonal climates normally form one growth ring each year. However, under severe stress conditions they may not complete annual wood production all the way down to the root collar resulting in continuously missing outer rings at lower stem sections (CMORs). Here we test whether CMORs occur at different distributional margins of woody plant species, where stress levels are expected to be high. We tested 13 species (202 individuals) of trees and shrubs growing at elevational and latitudinal shrub- and tree-lines, under conditions that are normally associated with a reduction of radial growth such as drought, cold or nutrient deficiency. Samples were collected from Alaska, north western Russia, Central Europe, Scandinavia and south eastern Spain. Annual radial growth was measured along several disks or cores of each individual applying the serial sectioning technique. Individuals of nine species showed CMORs. The proportion of individuals with CMORs within a sampled species and site ranged from 0% to 80%. The number of CMORs within an individual increased with decreasing sampling height on the stem. Significant correlations existed between the amount of CMORs and (i) cambial age, (ii) stem length, (iii) stem proportion/length below peat surface, and (iv) herbivore-caused defoliation intensity depending on the species and sampling location. Our results suggest that CMORs can be associated with a cambium that may be inactive, yet functional, for up to 18 years. We conclude that CMORs can occur in a wide variety of species, growth forms and biomes, suggesting a general strategy of woody plant growth. To avoid miss-dating in dendrochronological studies, we further recommend the use of the serial sectioning technique for woody plants growing in unfavorable environments, above all, when sample size is limited and when no established chronology for cross-dating is available. [Copyright &y& Elsevier]

A multiplicative and a semi-mechanistic, BWB-type [Ball, J. T. , Woodrow, I. E. , Berry, J. A. , 1987. A model predicting stomatal conductance and its contribution to the control of photosynthesis under different environmental conditions. In: Biggens, J. (Ed. ), Progress in Photosynthesis Research, vol. IV. Martinus Nijhoff, Dordrecht, pp. 221–224. ] algorithm for calculating stomatal conductance (g s) at the leaf level have been parameterised for two crop and two tree species to test their use in regional scale ozone deposition modelling. The algorithms were tested against measured, site-specific data for durum wheat, grapevine, beech and birch of different European provenances. A direct comparison of both algorithms showed a similar performance in predicting hourly means and daily time-courses of g s, whereas the multiplicative algorithm outperformed the BWB-type algorithm in modelling seasonal time-courses due to the inclusion of a phenology function. The re-parameterisation of the algorithms for local conditions in order to validate ozone deposition modelling on a European scale reveals the higher input requirements of the BWB-type algorithm as compared to the multiplicative algorithm because of the need of the former to model net photosynthesis (A n). [Copyright &y& Elsevier]

Published ozone exposure–response relationships from experimental studies with young trees performed at different sites across Europe were re-analysed in order to test the performance of ozone exposure indices based on AOTX (Accumulated exposure Over a Threshold of X nmol mol−1) and AFstY (Accumulated Stomatal Flux above a threshold of Y nmol m−2 s−1). AFst1. 6 was superior, as compared to AOT40, for explaining biomass reductions, when ozone sensitive species with differing leaf morphology were included in the analysis, while this was not the case for less sensitive species. A re-analysis of data with young black cherry trees, subject to different irrigation regimes, indicated that leaf visible injuries were more strongly related to the estimated stomatal ozone uptake, as compared to the ozone concentration in the air. Experimental data with different clones of silver birch indicated that leaf thickness was also an important factor influencing the development of ozone induced leaf visible injury. [Copyright &y& Elsevier]

Abstract: Seasonal trends in leaf gas exchange and ozone-induced visible foliar injury were investigated for three ozone sensitive woody plant species. Seedlings of Populus nigra L., Viburnum lantana L., and Fraxinus excelsior L. were grown in charcoal-filtered chambers, non-filtered chambers and open plots. Injury assessments and leaf gas exchange measurements were conducted from June to October during 2002. All species developed typical ozone-induced foliar injury. For plants exposed to non-filtered air as compared to the charcoal-filtered air, mean net photosynthesis was reduced by 25%, 21%, and 18% and mean stomatal conductance was reduced by 25%, 16%, and 8% for P. nigra, V. lantana, and F. excelsior, respectively. The timing and severity of the reductions in leaf gas exchange were species specific and corresponded to the onset of visible foliar injury. [Copyright &y& Elsevier]