Synovial Fluid of Patient With Rheumatoid Arthritis Enhanced Osmotic Sensitivity Through the Cytotoxic Edema Module in Synoviocytes

Rheumatoid arthritis (RA) is an autoimmune disease that causes inflammation of the synovial membrane ultimately leading to permanent damage in the affected joints. For this study, synovial fluids from 16 patients diagnosed with either RA or osteoarthritis (OA) were used to examine volume regulation and cooperative water channels, both of which are involved in the cytotoxic edema identified in RA-fibroblast-like synoviocytes (FLS). The osmolarity and inflammatory cytokine interleukin (IL)-6 of synovial fluids from RA patients were mildly enhanced compared to that from OA patients. RA-FLS demonstrated the enhanced property of regulatory volume increase in response to IL-6 and synovial fluids from RA patients. Although there was no difference in the protein expression of the volume-associated protein sodium–potassium–chloride cotransporter1 (NKCC1), its activity was increased by treatment with IL-6. Membrane localization of NKCC1 was also increased by IL-6 treatment. Additionally, both the protein and membrane expressions of aquaporin-1 were increased in RA-FLS by IL-6 stimulation. The IL-6-mediated enhanced osmotic sensitivity of RA-FLS likely involves NKCC1 and aquaporin-1, which mainly constitute the volume-associated ion transporter and water channel elements. These results suggest that RA-FLS provide enhanced electrolytes and concomitant water movement through NKCC1 and aquaporin-1, thereby inducing cellular swelling ultimately resulting in cytotoxic edema. Attenuation of cytotoxic edema and verification of its related mechanism will provide novel therapeutic approaches to RA treatment within the scope of cytotoxic edema.

Characterization of high fludioxonil resistance in Botrytis cinerea isolates from calibrachoa flowers

The fungicide fludioxonil is one of the most effective single-site fungicides available for managing flower blight caused by Botrytis cinerea on fruit and ornamental crops. Though low and moderate levels of resistance to fludioxonil have been reported in the pathogen across the United States and Europe, high resistance has only been reported from greenhouses in China. In this study, two B. cinerea isolates with high resistance (EC50 >100 µg/mL) to fludioxonil were detected on ornamental calibrachoa flowers grown in a greenhouse. These isolates exhibited stable resistance for over 20 generations, produced symptoms on calibrachoa flowers sprayed with label rates of fludioxonil, and displayed in vitro fitness penalties with decreased mycelial growth (p<0.0001) and sporulation (p<0.0001) compared to sensitive isolates. Highly resistant isolates were identified as MDR1h, containing the ΔLV497 deletion in mrr1. However, resistance levels and in vitro fitness parameter characteristics were not consistent with this phenotype. One isolate contained the mutation L267V between HAMP domains 1 and 2 of the Bos-1 gene, and both isolates exhibited high osmotic sensitivity and reduced glycerol accumulation in the presence of fludioxonil, indicating that high resistance of these isolates may be associated with the HOG1 MAPK pathway.

Osmotic Stress Affects Nuclear Morphology and Genome Architecture

The spatial organization of the genome influences its function [1]. Therefore, physical signals that deform the nucleus and the genome within may directly affect gene transcription and translation. In articular chondrocytes, nuclear deformation in response to osmotic stress is not sensitive to actin organization [2]. However, articular chondrocytes differ from most mammalian cells in that they remain round with cortically organized actin in monolayer culture. Adherent cells such as adipose stem cells (ASCs) spread in monolayer culture, forming a more typical, highly bundled actin cytoskeleton. These actin bundles exert tensile stress on the nucleus so we hypothesized that the osmotic sensitivity of the cell nucleus would be modulated by actin organization in ASCs. The osmotic sensitivity of the nucleus was quantified by measuring changes in the size and shape of the nucleus and the spatial arrangement of the chromatin within using 3D confocal microscopy.

Nonlinear Osmotic Properties of the Cell Nucleus

Osmotic stress affects biological function in articular chondrocytes and plays an important role in mechanotransduction in articular cartilage. One potential pathway for osmotic sensitivity in chondrocytes is direct deformation of the nucleus by osmotic stress [1,2]. However, the mechanism of this phenomenon is currently unclear. The nucleus is not contained within a semi-permeable lipid bilayer as are most osmotically sensitive organelles. It is conceivable that fixed charges in chromatin might attract dissolved ions and render it osmotically sensitive. However, this model cannot account for the abolition of the osmotic sensitivity of the nucleus by permeabilization of the cell membrane [3]. The goal of this study was to characterize the osmotic sensitivity of the chondrocyte nucleus and determine the underlying mechanism.