The role of WNK in modulation of KCl cotransport activity in red cells from normal individuals and patients with sickle cell anaemia

Abstract: Abnormal activity of red cell KCl cotransport (KCC) is involved in pathogenesis of sickle cell anaemia (SCA). KCC-mediated solute loss causes shrinkage, concentrates HbS, and promotes HbS polymerisation. Red cell KCC also responds to various stimuli including pH, volume, urea, and oxygen tension, and regulation involves protein phosphorylation. The main aim of this study was to investigate the role of the WNK/SPAK/OSR1 pathway in sickle cells. The pan WNK inhibitor WNK463 stimulated KCC with an EC50 of 10.9 ± 1.1 nM and 7.9 ± 1.2 nM in sickle and normal red cells, respectively. SPAK/OSR1 inhibitors had little effect. The action of WNK463 was not additive with other kinase inhibitors (staurosporine and N-ethylmaleimide). Its effects were largely abrogated by pre-treatment with the phosphatase inhibitor calyculin A. WNK463 also reduced the effects of physiological KCC stimuli (pH, volume, urea) and abolished any response of KCC to changes in oxygen tension. Finally, although protein kinases have been implicated in regulation of phosphatidylserine exposure, WNK463 had no effect. Findings indicate a predominant role for WNKs in control of KCC in sickle cells but an apparent absence of downstream involvement of SPAK/OSR1. A more complete understanding of the mechanisms will inform pathogenesis whilst manipulation of WNK activity represents a potential therapeutic approach

Early detection of osteoarthritis in the rat with an antibody specific to type II collagen modified by reactive oxygen species

Background Osteoarthritis (OA) is a disease of the whole joint, with articular cartilage breakdown as a major characteristic. Inflammatory mediators, proteases, and oxidants produced by chondrocytes are known to be responsible for driving cartilage degradation. Nevertheless, the early pathogenic events are still unclear. To investigate this, we employed an antibody that is specific to oxidative post-translationally modified collagen type II (anti-oxPTM-CII) to detect early cartilage pathogenic changes in two rat models of OA. Methods The animals underwent surgery for destabilization of the medial meniscus (DMM) and were sacrificed after 3, 5, 7, 14, and 28 days. Alternatively, anterior cruciate ligament transection with partial meniscectomy (ACLT+pMx) was performed and animals were sacrificed after 1, 3, 5, 7, and 14 days. Joints were stained with toluidine blue and saffron du Gatinais for histological scoring, anti-oxPTM-CII, and anti-collagen type X antibodies (anti-CX). Results We observed positive oxPTM-CII staining as early as 1 or 3 days after ACLT+pMx or DMM surgeries, respectively, before overt cartilage lesions were visible. oxPTM-CII was located mostly in the deep zone of the medial tibial cartilage, in the pericellular and territorial matrix of hypertrophic chondrocytes, and co-localized with CX staining. Staining was weak or absent for the lateral compartment or the contralateral knees except at later time points. Conclusion The results demonstrate that oxidant production and chondrocyte hypertrophy occur very early in the onset of OA, possibly initiating the pathogenic events of OA. We propose to use anti-oxPTM-CII as an early biomarker for OA ahead of radiographic changes

Litter N-content influences soil millipede abundance, species richness and feeding preferences in a semi-evergreen dry forest of Guadeloupe (Lesser Antilles)

Despite the impact of soil millipedes on litter fragmentation in tropical forests, there have been few studies dealing with factors determining their habitat preference in these ecosystems. In a natural secondary dry forest of Guadeloupe on Leptosol, two complementary studies were carried out in order to test the hypothesis that litter N-content strongly influences millipede distribution. Millipede abundance and species richness were described in the field under two tree species, Bursera simaruba and Pisonia subcordata, and were related to the chemical characteristics of their foliage. In addition, a laboratory experiment was done in order to assess millipede feeding preferences regarding the chemical characteristics of leaves from various species. Millipede abundance and species richness were significantly higher under P. subcordata than under B. simaruba, probably due to the higher N content of P. subcordata leaves. Moreover, millipedes fed preferentially on N-rich leaves. The present study confirms that there was a close correlation between the preferred food, its chemical composition and the local distribution of millipede populations

Changes in soil biological quality under legume- and maize-based farming systems in a humid savanna zone of Côte d’Ivoire

Studying the impact of farming systems on soil status is essential in determining the most relevant for a given agroecological zone. A trial was conducted in a West Africa humid savanna, aiming at assessing the short-term effects of farming systems on soil (0-10 cm) organic carbon (SOC) content and some soil microbiological properties. A randomized complete block experimental design with three replications, and the following treatments were used: Mucuna pruriens (Mucuna), Pueraria phaseoloides (Pueraria), Lablab purpureus (Lablab), a combination of these three legumes (Mixed-legumes), maize + urea (Maize-U), maize + triple super phosphate (Maize-Sp), maize + urea + triple super phosphate (Maize-USp), fertilizer-free maize continuous cropping (Maize-Tradi). Results indicated that SOC content was improved over time under legume-based systems. The relative increase was the highest with the legume association and Lablab, where SOC varied from 7.5 to 8.6 g.kg-1 (i.e. 14.7%) and from 7.2 to 8.3 g.kg-1 (i.e. 15.3%) respectively, between the start and the end of the trial. Besides, applying grass and maize residues as mulch on the ground, in association with inorganic fertilizers may be a way of improving SOC content in the short-term. Although legume-based systems exhibited highest values, microbial biomass carbon (MBC) did not show any statistical significant differences between treatments. However, soil C mineralization and soil specific respiration were influenced by the farming systems, with higher mean values under legume-based systems (42 ± 7.6 mg C-CO2.g-1 Corg and 0.4 mg C-CO2.g-1 biomass C, respectively), compared to maize continuous cropping systems (33.1 ± 1.6 mg C-CO2.g-1 Corg and 0.3 mg C-CO2.g-1 biomass C, respectively). Thus, these parameters can be used as sensitive indicators of the early changes in soil organic matter quality. The integration of legumes cover crops in farming systems may contribute to improve soil quality that would lead to sustainable agriculture in Côte d'Ivoire humid savannas