The co-culture of bovine brain capillary endothelial cells and rat primary glial cells was established as an in vitro blood-brain barrier model to investigate the mechanisms by which the Gram-positive bacterial cell wall components lipoteichoic acid and muramyl dipeptide induced injury of blood-brain barrier structure and function. We found that highly purified lipoteichoic acid disrupted blood-brain barrier integrity in a concentration- and time-dependent manner indirectly, through glia activation. Low trans-endothelial electrical resistance and high permeability to fluorescein isothiocyanate-inulin observed in the presence of lipoteichoic acid-activated glial cells were potentiated by muramyl dipeptide and could be reversed only when glial cells were activated by lipoteichoic acid at 10 microg/ml but not with a higher lipoteichoic acid concentration (30 microg/ml). Immunocytochemistry analysis revealed no evident changes in the distribution of the cytoskeleton protein F-actin and tight junction proteins occludin and claudin after lipoteichoic acid treatment. However, the tight junction associated protein AHNAK clearly revealed the morphological alteration of the endothelial cells induced by lipoteichoic acid. Lipoteichoic acid-activated glial cells produced nitric oxide and pro-inflammatory cytokines (tumor necrosis factor-alpha and interleukin-1beta) that contributed to lipoteichoic acid-induced blood-brain barrier disruption, since the direct treatment of the endothelial monolayer with tumor necrosis factor-alpha or interleukin-1beta increased blood-brain barrier permeability, whereas the pre-treatment of lipoteichoic acid-activated glial cells with antibodies against these two cytokines blocked lipoteichoic acid effects. Additionally, nitric oxide was also involved in blood-brain barrier damage, since the nitric oxide donor itself (diethylenetriamine-nitric oxide adduct) increased blood-brain barrier permeability and inducible nitric oxide synthase inhibitor (1400W) partially reversed lipoteichoic acid-induced trans-endothelial electrical resistance decrease.

Rat soleus muscle was submitted to 15 days of spinal cord ventral root section (deefferentation) in order to specifically disturb motor innervation. The electrophoretic analysis of the deefferented soleus muscle showed a reduction in myosin heavy chain (MHC) 1 (53% versus 92%) and new expressions in MHC 2D/X (12%) and MHC 2B (22%) relative concentrations, while MHC 2A composition did not evolve significantly. Single skinned fibers from control and deefferented soleus were identified according to their MHC composition and their functional properties (calcium/strontium activations and maximal shortening velocities) were established. Control soleus contained 72% of slow (S) fibers expressing MHC 1 and slow functional properties, and 28% of hybrid fast (HF) fibers, coexpressing MHC 1 with MHC 2A predominantly, and presenting fast functional properties. In deefferented soleus, four fiber types were discriminated: S and HF, and new fast types expressing MHC 2A, and/or MHC 2D/X plus MHC 2B. Moreover, deefferented soleus fibers were very much atrophied and presented alterations in Ca activation properties, the effects being more marked on the fast fibers. All the data support the use, on the rat, of deefferented muscle as a model for motoneuron disease.