Characterization of traumatic brain injury mechanisms and development studied via immunohistochemistry and magnetic resonance spectroscopy

Traumatic brain injury remains a serious issue in the United States despite a large research effort over the years. Uncertainty regarding injury mechanisms and cascades drives the current study, which seeks to address many unknowns. The first phase of this study focuses on determining a direct relationship between metabolites via magnetic resonance spectroscopy (MRS) and axonal damage assessed by immunohistochemistry in a Gottingen minipig in vivo model. Two injury devices were designed and fabricated. One imparts rotational acceleration in the median plane while the other imparts purely translation acceleration. The animals (n=15) undergo baseline 7T MR scans prior to injury, immediately post-injury, and twenty-four hours post injury. MRS is performed within ~200 mm voxel placed in the genu of the corpus callosum. Relevant metabolites include glutamate, N-acetylaspartate, myoInositol, creatine, and lactate. No clear trends were found for any of the metabolites for either time point for these few tests. Further testing is being conducted to see the meaning of the metabolite differences in terms of underlying damage characterized by immunohistochemistry. This work will be adapted to characterize longitudinal TBI development. High-speed biplane x-ray studies looking at head kinematics, brain response, intracranial pressure, and brain injury will support the development of an FE model of the minipig head. Once validated, the model can be used to scale graded injury metrics for prediction of brain injury in the human. Ultimately, a new head injury criterion will be developed for better prediction of the potential for traumatic brain injury. INTRODUCTION raumatic brain injury (TBI) continues to be a leading cause of death and disability in the United States, accounting for 30.5% of all injury-related deaths ("Traumatic Brain Injury in the United States" 2002-2006). Despite the large body of research dedicated to head injury, there is still a great deal of uncertainty regarding the mechanisms and pathogenesis involved. Some of the classic experimental brain injury models are fluid percussion (Lindgren and Rinder 1966), inertial rotation (Unterharnscheidt and Higgins 1969), controlled cortical impact (Lighthall, 1988), and weight drop (Marmarou et al., 1994). These models have limitations such as some of the animals used, especially rodents, do not have brain morphology similar to humans, and some models do not use physiologically relevant injury methods. T This study seeks to answer many of the unknowns surrounding TBI. Specifically, a mild traumatic brain injury threshold needs to be established that relates input kinematics to underlying damage and metabolite concentration changes. A purely linear impact is being used to avoid brain stem injury by limiting neck flexion. After short term injury development is characterized, longitudinal development will be observed up to two weeks after injury. Sibling studies will be used to relate input kinematics, neuronal damage, and metabolite change in one animal to the relative brain/skull kinematics in the other. High speed bi-plane x-ray with neutral density targets in the brain will be used to track relative motion. Ultimately, the current study will produce an FE model of the minipig head that can be used in conjunction with a model of the human head, such as SIMon, to scale graded injury metrics relating linear and angular head accelerations to levels of neuronal damage to be expected in the human. A new injury criterion will be developed, or perhaps an existing modified, to relate impact conditions to the level and risk of injury. In order to address criticisms of previous experimental head injury models, a Gottingen minipig is being used. As shown in Figure 1, the brain of the Gottingen minipig is similar to a human’s in terms of exiting angle of the brain stem into the spinal cord and brain morphology. It has a pronounced falx cerebri and distinct hemispheres, which do not exist in rodent models, and even other minipig models. The brain also has well developed convolutions. Further, the skull of this minipig is not as thick as other swine models. Figure 1: Gottingen minipig brain after perfusing with 4% paraformaldehyde (top). Falx cerebri (bottom). The first phase of this study focuses on determining a direct relationship between short term neuronal damage as assessed using magnetic resonance spectroscopy (MRS) and immunohistochemistry in the Gottingen minipig model. METHODS Input acceleration thresholds: Thresholds were estimated for the Gottingen minipig based on the literature. For the angular acceleration threshold, Ommaya 1984 (Ommaya, 1984) was referenced because proposed tolerances for rotation in the sagittal plane for sub-human primates were developed and scaled to the human. Brain density and material properties were assumed to be equal. Both brain radii and brain mass scaling were done in order to create a range for the minipig. The calculated thresholds for AIS 3 are shown in Table 1. For linear acceleration, the current Head Injury Criteria (HIC) threshold for the National Highway Traffic Safety Administrations’ New Car Assessment Program for a 50 percentile male anthropomorphic test device was taken as a starting point. A value of 700 for 15 ms was used because it corresponds to a 5% risk of AIS 4. As mild traumatic brain injury is of interest, this value was used as an upper boundary for linear acceleration. This value was scaled based on a method proposed by Mertz and Irwin, 2003 that uses length scaling to calculate HIC for different age and gender groups. Brain radius was used to determine the length scaling factor. The linear acceleration threshold corresponding to scaled HIC is shown in Table 1. Table 1. Acceleration and speed thresholds for the minipig based on scaling from the literature. AIS 3 was used for angular thresholds an AIS 4 was used for linear threshold. Since an initial objective is to conduct tests near threshold (below AIS 3), values less than a quarter of those shown in Table 1 were selected as starting-level inputs. Angular Linear Speed >30 rad/s ------Acceleration 24-31 krad/s 300G or 2943 m/s Animal model: Gottingen minipigs were divided into three main groups: control, rotational injury, and linear injury (n=20 total). There are one hour and twenty-four hour survival animals so that injury progression can be observed. In some cases, injury might be seen at the 24 hour interval, but not within the first hour. The overall division of the animals will be decided once injury results are observed. The division of animals tested as of November 2011 is shown in Table 2. Table 2. Minipig division to date. # of Minipigs 24 hr rotational injury 1