Mouse Model of Respiratory Tract Infection Induced by Waddlia chondrophila

Waddlia chondrophila, an obligate intracellular bacterium belonging to the Chlamydiales order, is considered as an emerging pathogen. Some clinical studies highlighted a possible role of W. chondrophila in bronchiolitis, pneumonia and miscarriage. This pathogenic potential is further supported by the ability of W. chondrophila to infect and replicate within human pneumocytes, macrophages and endometrial cells. Considering that W. chondrophila might be a causative agent of respiratory tract infection, we developed a mouse model of respiratory tract infection to get insight into the pathogenesis of W. chondrophila. Following intranasal inoculation of 2 x 10⁸ W. chondrophila, mice lost up to 40% of their body weight, and succumbed rapidly from infection with a death rate reaching 50% at day 4 post-inoculation. Bacterial loads, estimated by qPCR, increased from day 0 to day 3 post-infection and decreased thereafter in surviving mice. Bacterial growth was confirmed by detecting dividing bacteria using electron microscopy, and living bacteria were isolated from lungs 14 days post-infection. Immunohistochemistry and histopathology of infected lungs revealed the presence of bacteria associated with pneumonia characterized by an important multifocal inflammation. The high inflammatory score in the lungs was associated with the presence of pro-inflammatory cytokines in both serum and lungs at day 3 post-infection. This animal model supports the role of W. chondrophila as an agent of respiratory tract infection, and will help understanding the pathogenesis of this strict intracellular bacterium.     

Inhibition of the Differentiation of Monocyte-Derived Dendritic Cells by Human Gingival Fibroblasts

We investigated whether gingival fibroblasts (GFs) can modulate the differentiation and/or maturation of monocyte-derived dendritic cells (DCs) and analyzed soluble factors that may be involved in this immune modulation. Experiments were performed using human monocytes in co-culture with human GFs in Transwell® chambers or using monocyte cultures treated with conditioned media (CM) from GFs of four donors. The four CM and supernatants from cell culture were assayed by ELISA for cytokines involved in the differentiation of dendritic cells, such as IL-6, VEGF, TGFβ1, IL-13 and IL-10. The maturation of monocyte-derived DCs induced by LPS in presence of CM was also studied. Cell surface phenotype markers were analyzed by flow cytometry. In co-cultures, GFs inhibited the differentiation of monocyte-derived DCs and the strength of this blockade correlated with the GF/monocyte ratio. Conditioned media from GFs showed similar effects, suggesting the involvement of soluble factors produced by GFs. This inhibition was associated with a lower stimulatory activity in MLR of DCs generated with GFs or its CM. Neutralizing antibodies against IL-6 and VEGF significantly (P<0.05) inhibited the inhibitory effect of CM on the differentiation of monocytes-derived DCs and in a dose dependent manner. Our data suggest that IL-6 is the main factor responsible for the inhibition of DCs differentiation mediated by GFs but that VEGF is also involved and constitutes an additional mechanism.     

Behavioural responses of juvenile cuttlefish (Sepia officinalis) to local water movements

Physiological studies have shown that the epidermal head and arm lines in cephalopods are a mechanoreceptive system that is similar to the fish and amphibian lateral lines (Budelmann BU, Bleckmann H. 1988. A lateral line analogue in cephalopods: Water waves generate microphonic potentials in the epidermal head lines of Sepia officinalis and Lolliguncula brevis. J. Comp. Physiol. A 164:1-5.); however, the biological significance of the epidermal lines remains unclear. To test whether cuttlefish show behavioural responses to local water movements, juvenile Sepia officinalis were exposed to local sinusoidal water movements of different frequencies (0.01-1000 Hz) produced by a vibrating sphere. Five behavioural responses were recorded: body pattern changing, moving, burrowing, orienting, and swimming. Cuttlefish responded to a wide range of frequencies (20-600 Hz), but not to all of the frequencies tested within that range. No habituation to repeated stimuli was seen. Results indicate that cuttlefish can detect local water movements (most likely with the epidermal head and arm lines) and are able to integrate that information into behavioural responses.     

A-Kinase Anchoring Protein Lbc Coordinates a p38 Activating Signaling Complex Controlling Compensatory Cardiac Hypertrophy

In response to stress, the heart undergoes a remodeling process associated with cardiac hypertrophy that eventually leads to heart failure. A-kinase anchoring proteins (AKAPs) have been shown to coordinate numerous prohypertrophic signaling pathways in cultured cardiomyocytes. However, it remains to be established whether AKAP-based signaling complexes control cardiac hypertrophy and remodeling in vivo. In the current study, we show that AKAP-Lbc assembles a signaling complex composed of the kinases PKN, MLTK, MKK3, and p38α that mediates the activation of p38 in cardiomyocytes in response to stress signals. To address the role of this complex in cardiac remodeling, we generated transgenic mice displaying cardiomyocyte-specific overexpression of a molecular inhibitor of the interaction between AKAP-Lbc and the p38-activating module. Our results indicate that disruption of the AKAP-Lbc/p38 signaling complex inhibits compensatory cardiomyocyte hypertrophy in response to aortic banding-induced pressure overload and promotes early cardiac dysfunction associated with increased myocardial apoptosis, stress gene activation, and ventricular dilation. Attenuation of hypertrophy results from a reduced protein synthesis capacity, as indicated by decreased phosphorylation of 4E-binding protein 1 and ribosomal protein S6. These results indicate that AKAP-Lbc enhances p38-mediated hypertrophic signaling in the heart in response to abrupt increases in the afterload.     

Micrometric segregation of fluorescent membrane lipids: relevance for endogenous lipids and biogenesis in erythrocytes

Micrometric membrane lipid segregation is controversial. We addressed this issue in attached erythrocytes and found that fluorescent boron dipyrromethene (BODIPY) analogs of glycosphingolipids (GSLs) [glucosylceramide (BODIPY-GlcCer) and monosialotetrahexosylganglioside (GM1BODIPY)], sphingomyelin (BODIPY-SM), and phosphatidylcholine (BODIPY-PC inserted into the plasma membrane spontaneously gathered into distinct submicrometric domains. GM1BODIPY domains colocalized with endogenous GM1 labeled by cholera toxin. All BODIPY-lipid domains disappeared upon erythrocyte stretching, indicating control by membrane tension. Minor cholesterol depletion suppressed BODIPY-SM and BODIPY-PC but preserved BODIPY-GlcCer domains. Each type of domain exchanged constituents but assumed fixed positions, suggesting self-clustering and anchorage to spectrin. Domains showed differential association with 4.1R versus ankyrin complexes upon antibody patching. BODIPY-lipid domains also responded differentially to uncoupling at 4.1R complexes [protein kinase C (PKC) activation] and ankyrin complexes (in spherocytosis, a membrane fragility disease). These data point to micrometric compartmentation of polar BODIPY-lipids modulated by membrane tension, cholesterol, and differential association to the two nonredundant membrane:spectrin anchorage complexes. Micrometric compartmentation might play a role in erythrocyte membrane deformability and fragility.