An Experimental Study On Local Brain Hypothermia Induced By Epidural Cooling For Severe Traumatic Brain Injury In Rats

Research background Severe traumatic brain injury is a frequently-occurring disease that usually causes significant harm to human health and life. The aftermath of such injuries leaves survivors with permanent disabilities resulting in lifelong medical, financial, emotional, familial and social problems. Today, although the advancement of medical sciences is obvious, the rate of disability and fatality after traumatic brain injury has been high for many years, and there are still not an effective treatment. In the last 1990s and the beginning of this century, the lack of progress in achieving brain protection with several pharmacologic agents, while a positive evidence of the therapeutic efficacy of mild to moderate hypothermia from the studies in post-cardiac arrest survivors and neonatal hypoxic encephalopathy, led to renewed interest in therapeutic hypothermia for the severe traumatic brain injuries. There are many ways to induce the therapeutic hypothermia, of which the systemic cooling method is most widely used at present. However, the systemic side effects of whole-body cooling such as chills, electrolyte disturbances, arrhythmia, bleeding tendency, easy to infected and skin lesion, have been reported in some studies. It is important to design and imply techniques of cooling in order to maximize the therapeutic benefit and minimize the complications. At present, the mechanisms that cause the neuroprotection provided by hypothermia still a matter of debate and are probably multiple. Further understanding the mechanisms underlying hypothermia's benefits will lead to more effective treatments to reduce the disability and mortality of severe traumatic brain injuries.Chapter 1. The Efficacy, Security and Complication of the Local Brain Hypothermia Induced by Epidural CoolingObjective To develop an epidural cooling method to induce local brain hypothermia using a self-made cooling coil, and to evaluate the efficacy, security and complications of this epidural local hypothermia technique.Methods SD rats were randomly divided into normal temperature group(Nor), local hypothermia group(LH) and systemic hypothermia group(SH). The local and systemic hypothermia were induced by self-made cooling coil and ice bag, alcohol respectively, the brain temperature were reduced to 31.0-32.0℃and maintained for 10 hours. During cooling, the shivering reaction, ipsilateral and contralateral brain temperature, rectal temperature, respiratory, heart rate and blood pressure of rats were assessed. The blood gas and electrolyte analysis, blood routine and hemorheology of rats were detected after cooling. The neurological evaluation and microscopic examination were carried out at 24 h follow cooling, and the brain water, Na~+, K~+ content and blood-brain barrier permeability were also detected.Results In LH group, local brain hypothermia was produced within minutes from (36.5±0.3)℃to (31.4±0.4)℃in the ipsilateral hemisphere after cooling, while the contralateral brain and rectal temperature, R, HR and MABP were not significantly altered. In SH group, the decrease of bilateral brain and rectal temperature as well as an 57.1% of shivering incidence were observed during cooling. The decrease of HR and platelet count, and the increase of whole blood viscosity under high shear and plasma viscosity were also detected in SH group. There were no significant difference in NSS score, brain water, Na~+, K~+ and Evans Blue content among groups, and no structural alterations were found in brain tissue of both LH and SH group neither.Conclusions This epidural cooling method can achieve the effect of brain hypothermia as well as systemic cooling technique, and dose not cause fluctuations in vital signs and acute neuronal injury. Meanwhile it may developed few complications compared with systemic hypothermia. Chapter 2. Therapeutic Efficacy of Epidural Local Hypothermia on Severe Traumatic Brain Injury and Comparative Study with Systemic Cooling MethodObjective To study the therapeutic efficacy of epidural local hypothermia on mortality, body weight, neurological function, brain edema and histopathological changes on rats after traumatic brain injury, and which were compared with systemic hypothermia at the same time.Methods SD rats were randomly divided into normal control group(Norm), sham operation group(Sham), severe traumatic brain injury group(sTBI), local hypothermia therapy group(LHT) and systemic hypothermia therapy (SHT). Feeney's weight-drop (25 g, 60 cm) model was used to induce severe traumatic brain injury in rats. The epidural local hypothermia or systemic cooling were conducted 30min post injury and last for 10 hours. The body weight measurement, NSS score, beam balance test, beam walking test and Morris water maze were used in different time point post injury, as well as the brain water, Na~+, K~+ content and HE stain were detected to evaluate the therapeutic efficacy of hypothermia.Results The mortality of rats in sTBI, LHT and SHT group were 32.32%,15.19% and 19.28% respectively. The body weight loss of rats were less obvious in LHT and SHT group than sTBI group. The neurological function observed on rats in LHT and SHT group were better than sTBI group detected by NSS score, beam balance test, beam walking test and Morris water maze. At 12h,1d and 3d post injury, an increase of brain water, Na~+ content while a decrease of brain K~+ content were detected in rats of sTBI group, indicating a brain edema. Both LHT and SHT group developed less extent of brain edema in the same day. Rats developed a severe histopathological damage after severe traumatic brain injury, and both of local hypothermia therapy and systemic hypothermia therapy could reduce the extent of pathological damage of brain tissue in rats. In addition, no significant differences were found in mortality, body weight loss, neurological dysfunction and brain edema between LHT and SHT group.Conclusions The same as systemic hypothermia, the epidural cooling technique could slow down the weight loss, improve neurological deficits, reduce brain edema and ameliorate the histopathological damage of brain tissue after head injury. Both of local hypothermia and systemic hypothermia provide the same definite therapeutic efficacy on severe traumatic brain injury. Chapter 3. Effect of Epidural Local Hypothermia on Expression of PKC-delta, Synaptophysin and GAP-43 after Severe TraumaticBrain Injury in RatsObjective To study the expression of PKC-delta, after severe traumatic brain injury in rats for an investigation on the mechanisms of neuroprotection by hypothermia.Methods SD rats were randomly divided into normal control group(Norm), sham operation group(Sham), severe traumatic brain injury group(sTBI) and local hypothermia therapy group(LHT). Feeney's weight-drop model was used. The epidural local cooling were conducted 30 min post injury and last for 10 hours. The ultrastructure changes of brain tissues were observed under transmission electron microscope. The expressions of PKC-δwere detected by western-blotting. TUNEL staining were conducted at 12 h, 1 d and 3 d post injury. The expressions of synaptophysin and GAP-43 were detected at 7 d, 14 d and 28 d post injury by western-blotting and immunohistochemisty.Results Rats in sTBI group developed a serious ultrastructure damage, and the structures of nucleus, mitochondria and blood-brain barrier were improved in LHT group. The amount of constitutively activated C-terminal catalytic fragment(CF-PKC-δ) increased, as well as the translocation to the membrane of PKC-5, both of which were hallmarks of PKC-8 activation, were observed after severe traumatic brain injury. These events were blocked by hypothermia, implicating the inhibition of PKC-δactivation by hypothermia. Meanwhile, an increase of the number of apoptotic cells in brain tissue were observed by TUNEL staining in the acute stage(3 days post injury) of severe traumatic brain injury, and could be reduced by hypothermia. In addition, a reduction of synaptophysin and GAP-43 expressions due to severe traumatic brain injury were investigated in the late stage, and hypothermia could raise the expressions of synaptophysin and GAP-43, which were coincident with the results of Morris Water Maze in rats.Conclusions Hypothermia could reduce the ultrastructure damage caused by severe traumatic brain injury. Hypothermia protects against the traumatic damage in part dues to the suppressing PKC-δactivation by blockage of CF-PKC-δproducing and PKC-δtranslocation to the membrane. Meanwhile, hypothermia could inhibit the apoptosis after injury. In addition, hypothermia might promote nerve regeneration and remodeling in late stage of TBI by the way of increased expressions of synaptophysin and GAP-43 in brain tissue.