Brain Inflammation and Neurodegeneration After Spinal Cord Injury

Neuronal regulated cell regulated neuronal cell death including apoptotic and non-apoptotic phenotypes, is a key component of the secondary injury leading to delayed tissue damage after traumatic brain injury (TBI). Both caspase-dependent and caspase-independent apoptotic mechanisms have been implicated in post-traumatic neuronal loss, but a complete understanding of these processes in clinically relevant experimental TBI models has yet to be achieved. Recent work in our laboratory has focused on the relative importance of these mechanisms using a mouse controlled cortical impact (CCI) model. Caspase-dependent mechanisms are often initiated in response to release of cytochrome c from the mitochondria into the cytosol where it forms a caspase-activating complex (apoptosome) with apoptotic peptidase activating factor 1 (Apaf-1) leading to sequential activation of caspase-9 and caspase-3 (the main executioner caspase). Caspase-independent mechanisms may be initiated by release of apoptosis inducing factor mitochondria associated 1 (AIFM1 aka AIF) from the mitochondria The AIF-mediated cell death requires its translocation to the nucleus, a step that depends on its interaction with the peptidyl-proplyl isomerase Cyclophilin A (CypA) which transports AIF from the cytosol to the nucleus. Our studies compared the effects of single versus combined inhibition of caspase and AIF pathways in a mouse CCI model, by examining the effects of CypA gene knockout (CypA(-/-)), caspase inhibition with a pan-caspase inhibitor (boc-aspartyl(OMe)-fluoromethylketone, BAF), or combined intervention. TBI caused caspase activation as well as translocation of AIF to the nucleus. Markers of caspase activation including caspase-specific fodrin cleavage fragments and number of FLIVO-positive cells were reduced in BAF-treated CypA(+/+) mice, whereas markers of AIF activation including AIF/H2AX interaction and AIF translocation to the nucleus were attenuated in CypA(-/-) mice. Each single intervention, (CypA(-/-) or BAF-treated CypA(+/+)) reduced the number of apoptotic cells (TUNEL-positive) in the cortex and improved long-term sensorimotor function; CypA(-/-) also attenuated microglial activation after injury. Importantly, BAF-treated CypA(-/-) mice (combined intervention), showed greater effects than either intervention alone on multiple outcomes including: reduction in TUNEL-positive cells, decrease in neuroinflammation, improved motor and cognitive recovery, and attenuation of lesion volume and neuronal loss in the hippocampus. We also used two in vitro neuronal cell death models known to induce AIF-mediated cell death, we also showed that neurons from CypA(-/-) animals were protected and that effects were unrelated to caspase activation. These data indicate that AIF-mediated and caspase-dependent pathways contribute independently and in parallel to secondary injury after TBI and suggest that combined therapeutic strategies directed at multiple PCD pathways may provide superior neuroprotection than those directed at single mechanisms.

Future studies will continue to probe the mechanisms that control AIF-dependent cell death including the role of poly (ADP-ribose) polymerase in the events leading to the activation of the AIF pathway as well as the role of other cell death phenotypes such as various types of regulated necrosis (necroptosis, etc.) in the neuronal loss and neurological deficits following TBI.