Showing papers by "Montserrat Camps published in 2010"

Regulation of the severity of neuroinflammation and demyelination by TLR-ASK1-p38 pathway

Abstract: Apoptosis signal-regulating kinase 1 (ASK1) is an evolutionarily conserved mitogen-activated protein kinase (MAPK) kinase kinase which plays important roles in stress and immune responses. Here, we show that ASK1 deficiency attenuates neuroinflammation in experimental autoimmune encephalomyelitis (EAE), without affecting the proliferation capability of T cells. Moreover, we found that EAE upregulates expression of Toll-like receptors (TLRs) in activated astrocytes and microglia, and that TLRs can synergize with ASK1-p38 MAPK signalling in the release of key chemokines from astrocytes. Consequently, oral treatment with a specific small molecular weight inhibitor of ASK1 suppressed EAE-induced autoimmune inflammation in both spinal cords and optic nerves. These results suggest that the TLR-ASK1-p38 pathway in glial cells may serve as a valid therapeutic target for autoimmune demyelinating disorders including multiple sclerosis.

Isoform-selective phosphoinositide 3-kinase inhibitors induce apoptosis in chronic lymphocytic leukaemia cells.

Abstract: Phosphoinositide 3-kinase (PI3K) has been reported to be constitutively active in chronic lymphocytic leukaemia (CLL), and to contribute to increased cell survival and resistance to apoptosis (Barragan et al, 2002; Pleyer et al, 2009). PI3Ks generate phosphoinositide lipids in response to extracellular stimuli, regulating survival, proliferation, differentiation and migration (Manning & Cantley, 2007). PI3K class I are heterodimers that consist of a catalytic subunit and a regulatory subunit, and are further divided into class IA (p110a, b and d), activated downstream tyrosine kinase receptors and class IB (p110c), activated downstream G protein-coupled receptors (Engelman et al, 2006). p110a and b are widely distributed whereas p110d and c are mainly, but not exclusively, expressed in leucocytes. LY294002 and wortmannin have been largely used as PI3K inhibitors. However, neither of them can discriminate between the different isoforms of PI3K. Recently, isoform-selective PI3K inhibitors have been described (Jackson et al, 2005; Marone et al, 2008; Draghetti et al, 2009). Here, we have examined the effect of isoform-selective PI3K inhibitors (Table I) in the viability of CLL cells and lymphocytes from healthy donors. Chronic lymphocytic leukaemia was diagnosed according to standard clinical and laboratory criteria. Blood samples were obtained from the Hospital de Bellvitge, L’Hospitalet de Llobregat, Spain. Mononuclear cells from peripheral blood samples were isolated and cultured as previously described (Barragan et al, 2002). First, the ability of isoform-selective PI3K inhibitors to induce apoptosis in CLL cells was examined. Apoptosis was determined by annexin V-fluorescein isothiocyanate (Bender MedSystems, Vienna, Austria) and propidium iodide (PI; Bender MedSystems) double staining, as previously described (Barragan et al, 2002), and cell viability was measured as the percentage of annexin V and PI double-negative cells. Samples were acquired and data was analysed by using the FACSCalibur and CellQuest software (Becton Dickinson, Mountain View, CA, USA). Thus, CLL cells from 5 different patients were incubated with a range of concentrations (5–50 lmol/l) of isoform-selective PI3K inhibitors for 24 and 48 h and the cell viability was measured. PI3K inhibitors induced apoptosis in a doseand time-dependent manner (Fig 1A, n = 5), except MSC1822169B (p110c inhibitor), which did not induce apoptosis in CLL cells. The inhibitors that showed a stronger effect on cell viability were MSC1902994A (p110a inhibitor), MSC1829899A (p110d inhibitor) and MSC1831419A (p110b and d inhibitor). LY294002 was used as a control for broad PI3K inhibition. Incubation of CLL cells with 20 lmol/l LY294002 for 24 and 48 h reduced cell viability to 58 ± 6% and 50 ± 10%, respectively (n = 5). The isoform-selective PI3K inhibitors were then tested in samples from 22 CLL patients. Surprisingly, CLL cells from five of the 22 patients showed resistance to PI3K inhibitor-induced apoptosis when incubated with 10 lmol/l isoform-selective PI3K inhibitors. The samples were grouped into two classes, designated as sensitive and resistant, according to whether the apoptosis induced by all the isoform-selective PI3K inhibitors at 48 h was higher or lower than 20% when compared to the