calcein-am and Brain-Injuries

The biochemical mechanisms of explosive blast-induced traumatic brain injury and the subsequent long-term neurobehavioral abnormalities are still not completely understood. We studied the biochemical mechanism of blast traumatic brain injury using our recently reported in-vitro model system with a shock tube. Primary blast exposure of in-vitro models leads to neurobiological changes in an overpressure dose-dependent and time-dependent manner. Lactate dehydrogenase was released significantly into the extracellular medium without cell death after blast exposure, indicating compromised cell membrane integrity. We further explored the integrity of cell membrane after blast exposure by fluorescent dye uptake/release techniques in SH-SY5Y human neuroblastoma cells. Our data indicate that blast exposure leads to an overpressure-dependent transient increase in the release of preloaded calcein AM into the culture medium with proportional intracellular decrease. Uptake of an extracellular nucleic acid-binding dye TO-PRO-3 iodide was also increased significantly after blast exposure, indicating that the increased molecular transport is bidirectional and nuclear membrane integrity is also affected by blast exposure. These results suggest that blast exposure perturbs the integrity of the neuronal cell membrane, leading to increased bidirectional transport of molecules--a potential mechanism that can lead to traumatic brain injury.

Topics: Blast Injuries; Brain Injuries; Carbocyanines; Cell Line; Cell Membrane Permeability; Cell Survival; Fluoresceins; Humans; Models, Biological; Neurons

When the brain suffers injury, microglia migrate to the damaged sites and become activated. These activated microglia are not detected several days later and the mechanisms underlying their disappearance are not well characterized. In this study, we demonstrate that interleukin (IL)-13, an anti-inflammatory cytokine, selectively induces cell death of activated microglia in vitro. Cell death was detected 4 days after the coaddition of IL-13 with any one of the microglial activators, lipopolysaccharide (LPS), ganglioside, or thrombin. This cell death occurred in a time-dependent manner. LPS, ganglioside, thrombin, or IL-13 alone did not induce cell death. Among anti-inflammatory cytokines, IL-4 mimicked the effect of IL-13, while TGF-beta did not. Cells treated with IL-13 plus LPS, or IL-13 plus ganglioside, showed the characteristics of apoptosis when analyzed by electron microscopy and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining. Electron micrographs also showed microglia engulfing neighboring dead cells. We propose that IL-13 and IL-4 induce death of activated microglia, and that this process is important for prevention of chronic inflammation that can cause tissue damage.

Topics: Animals; Animals, Newborn; Brain Injuries; Cell Death; Cell Size; Cells, Cultured; Encephalitis; Ethidium; Fluoresceins; Fluorescent Dyes; Gangliosides; Gliosis; In Situ Nick-End Labeling; Intercalating Agents; Interleukin-13; Interleukin-4; Lipopolysaccharides; Microglia; Microscopy, Electron; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta