The major findings of our study are as follows: 1) mice treated with single or multi-dose APC had significantly improved functional recovery reflected in rapid improvement on the rotarod, wire grip task, and balance beam when compared with vehicle-treated groups; 2) mice treated with single or multi-dose APC post-TBI showed significantly reduced lesion volume at 7 days post-injury compared to vehicle-treated groups, with multi-dose APC treatment reducing lesion volume significantly more than single-dose; 3) post-traumatic brain repair was enhanced in the multi-dose APC group, by increased formation of new blood vessels and neurogenesis when compared to the single-dose and vehicle-treated groups.
APC exerts direct neuronal and vascular protective effects in vitro
and in vivo
against divergent inducers of apoptosis and blocks both the intrinsic, mitochondria-mediated and the extrinsic, death receptors-mediated apoptotic pathways (2
). An important point to address is whether neuroprotection and/or brain repair is responsible for the presently observed improvement in functional recovery as seen in the APC treated mice, when compared to vehicle. From prior studies examining the effects of APC and other agents on angiogenesis and neurogenesis following ischemic stroke in rodents, it has been shown that these brain repair processes are not seen until several days after injury (1
). Similarly, angiogenic or neurogenic responses were not found 24 h after injury in brain trauma models in rodents (see for example refs. 7
). Therefore, the early functional improvements seen in the mice after APC treatment is likely attributed to neuroprotection only. To precisely determine the specific role of brain repair, as one of APC's beneficial effects, future studies should examine whether APC given at the time when it no longer reduces lesion volume, but promotes angiogenesis and neurogenesis, for example beginning at 48 hours after injury, may still improve the functional outcome after TBI.
In the current study, with a 1 mm CCI injury, most mice in the study are at, or close to, their pre-injury functional levels at the 7 day endpoint. Therefore, the present model may not be sensitive enough to study the role of brain repair after APC treatment because the functional outcome corrects itself without any treatment within 7 days due to high brain plasticity in young mice. But, a more significant injury model, possibly with a deeper injury, or studies in older mice, which would not allow a rapid spontaneous functional recovery, may offer better opportunities to study the true benefits of neurogenesis and angiogenesis on functional recovery vs. neuroprotective effects of APC.
Another important aspect of APC treatment in TBI remaining to be clarified is the timing of treatment necessary to achieve treatment benefits. First, as seen with tPA treatment of stroke, timing of treatment can be critical, as delayed treatment can actually increase injury severity (30
). From a previous study in an ischemic stroke model, APC given at 6 or 24 hours after ischemia significantly improved functional outcome, reduced cerebral infarction volume, and enhanced repair of damaged brain tissue by promoting cerebral angiogenesis and neurogenesis (22
). In future studies, APC treatment in a TBI model administered at time points further out from injury will help clarify this important issue.
Previous studies have looked at other treatments such as recombinant human erythropoietin, statins, and inducible nitric oxide as possible ways to promote brain repair through angiogenesis and neurogenesis following TBI (7
). While some benefits from these treatments have been shown, presently no definitive treatment has been established. This study demonstrates APC is a neuroprotective agent after TBI and promotes brain repair by stimulating post-TBI angiogenesis and neurogenesis. Until recently, APC has been studied mostly in the treatment of stroke, sepsis, and multiple sclerosis. Our data, however, show that APC may have a promising future in the treatment of TBI, and warrants further investigations to more clearly elucidate the true benefits of APC in the treatment of patients after TBI.
Traumatic brain injury (TBI) is a common cause of death and disability in the United States, often resulting in permanent functional deficits. Numerous pharmacologic agents have been explored as potential therapeutic interventions aimed at ameliorating damage after TBI, but without much success. This study shows a significant decrease in lesion volume, and improvement in functional recovery in mice treated with activated protein C (APC) following TBI. In addition, we found that APC enhances brain repair in mice treated with multiple doses of APC. APC augmented new blood vessel formation, and may play an important role in tissue recovery and functional outcome after head injury. In addition, APC enhanced proliferation and migration of neuroblasts which may also contribute to functional recovery after TBI. Previous studies have looked at other possible treatments to promote brain repair, however, to this point no definitive treatment has been established. This study establishes APC as a protective agent in TBI in addition to its newly reported cerebral repair effects. Our data show that APC has a promising future in the treatment of TBI, and warrants further investigations into the efficacy of APC in the treatment of TBI.