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Up-regulation of proBDNF in ischemic brain and the detrimental role of proBDNF on cellular survival has already been established. We propose that the up-regulated proBDNF may trigger the harmful events and evoke a secondary ischemic damage after ischemia. This study aimed to establish the neuroprotective effects of anti-proBDNF antibody in a rat photothrombotic ischemic model. Photothrombotic ischemic model was performed on Sprague Dawley rats and anti-proBDNF antibodies were administered intraperitoneally to the ischemic rats at a dose of 5 mg/kg after 6 hours (6 h) and on 3 days (3d) after ischemia. Behavioural tests were performed for sensorimotor functional analyses. Animals were euthanized at 7d for histochemical and biochemical studies. We observed higher proBDNF expression around the ischemic infarct. Higher level of apoptosis and inflammation was evident at 7d after ischemia on brain sections. Interestingly, the anti-proBDNF treatment instigated significant reduction of the infarction size as detected by Haematoxylin and Eosin (H&E) staining. Similar reduction of apoptotic signaling proteins in western blot and immunostaining after anti-proBDNF treatment was found. Up-regulation of synaptic protein expression was also observed after this treatment. Significant sensorimotor functional improvements were also noticed at 7d after anti-proBDNF treatment. We conclude that anti-proBDNF treatment is anti-apoptotic and anti-inflammatory, and plays advantageous role in promoting cellular growth and improving sensorimotor function after ischemic insult. Taken together, our study suggests that this anti-proBDNF treatment can be considered as a therapeutic approach for ischemic recovery. Refereed/Peer-reviewed

Stroke is accompanied by severe inflammation in the brain. The role of mature brain-derived neurotrophic factor (mBDNF) in ischemic stroke has received intensive attention, but the function of its precursor proBDNF is less understood. Recent studies showed that mBDNF and proBDNF in the ischemic brain are upregulated, but the significance of mBDNF and proBDNF in the lymphocytes in ischemic stroke is not known. Here, we propose that the expression levels of mBDNF and proBDNF in lymphocytes correlate with those in the brain after ischemic stroke and therefore can be surrogate markers for the ischemic brain. Using a photothrombotic model in rats and ELISA assay technique, we found that proBDNF and mBDNF in peripheral lymphocytes were upregulated but produced differential time courses after ischemia. The levels of mBDNF and proBDNF in lymphocytes at early stages of stroke (1 day), showed a strong positive correlation with those in the brain. The levels of p75, sortilin, were also increased in a time-dependent manner after ischemic stroke; however, the levels of p-TrkB in the ischemic brain at 6 h, 1 and 3 days were significantly reduced in the brain. The present study suggests that the levels of proBDNF and mBDNF in the blood lymphocytes in acute ischemic stroke reflect those in the brain at early stages. Refereed/Peer-reviewed

Brain derived neurotrophic factor (BDNF) is a member of the neurotrophin family, best characterized for its survival and differentiative effects in the central nervous system. Pro-BDNF, known as the precursor of BDNF, is believed to have opposite functions to mature BDNF (mBDNF). The opposing effects of Pro-BDNF and mBDNF have led researchers to propose a ‘yin’ (Pro-BDNF) and ‘yang’ (mBDNF) model of which, the specific mechanism of its opposing functions is unclear and requires further investigation. In order to elucidate pro-BDNF's explicit role, we established a pro-BDNF knockout (KO) mouse model. This BDNF pro-domain KO mouse model showed significant weight loss, impaired righting reflex, abnormal motor behaviours and short lifespan (less than 22 days), mimicking a Huntington's disease (HD)-like phenotype. ELISA results showed BDNF pro-domain KO not only blocked pro-BDNF, but also significantly affected the level of mBDNF. Abnormal morphologic changes were found in the dentate gyrus (DG) of the hippocampus in pro-BDNF KO mice, and western blot confirmed significant cell apoptosis in pro-BDNF KO mice brains. Furthermore, the expression of glutamic acid decarboxylase 65/67 (GAD65/67) was significantly reduced in pro-BDNF KO mice, indicating impaired inhibitory neurotransmission. Heterozygous (Het) mice showed impaired learning and memory capability and depressive-like behaviours, compared with wild type (WT) mice. Overall, these results support that pro-domain of BDNF is an indispensable part of the BDNF gene; without the proper formation of pro-BDNF, mBDNF cannot be produced successfully and function correctly on its own. Our study also supports the BDNF hypothesis in the pathogenesis of HD Refereed/Peer-reviewed

Treatment with mature brain-derived neurotrophic factor (mBDNF) promotes functional recovery after ischemia in animal trials but the possible role of its precursor protein proBDNF and its receptors or the factors responsible for the conversion of proBDNF to mBDNF in ischemic stroke are not known. The main aim of this study was to characterize the time-dependent expression of genes and/or proteins related to BDNF processing and signaling after ischemia as well as the sensorimotor behavioral dysfunction in a photothrombotic ischemic model in rats. Characterization of different genes and proteins related to BDNF processing and signaling was performed using qPCR, immunoblotting and enzyme-linked immunosorbent assays. We showed in this study that some sensory and motor functional deficiencies appeared in the ischemic group at day 1 and persisted until day 14. Most changes in gene expression of BDNF and its processing enzymes occurred within the first 24 h in the ipsilateral cortex, but not in the contralateral cortex. At the protein level, proBDNF expression was increased at 6 h, mBDNF expression was increased between 15 h and 1 day while p75 receptor protein expression was increased between 6 h and 3 days in the ipsilateral cortex, but not in the contralateral cortex. Therefore, cerebral ischemia in rats led to the up-regulation of genes and/or proteins of BDNF, proBDNF and their processing enzymes and receptors in a time-dependent manner. We propose that the balance between BDNF and proBDNF and their associated proteins may play an important role in the pathogenesis and recovery from ischemia. Refereed/Peer-reviewed

Thesis (PhD)--University of South Australia, 2018. Includes bibliographical references (pages 157-201) This dissertation have tested the hypothesis that the precursor of brain-derived neurotrophic factor (proBDNF) and its related signaling p75 receptor play a detrimental role in the ischemic injury of the brain. The remarkable up-regulation of proBDNF and p75 proteins was found in neurons of ischemic brain. Two interventions were used to obstruct the function of proBDNF and p75 by peripheral administration of anti-proBDNF antibodies and recombinant fusion protein of p75 extracellular domain-immunoglobulin fragment C in ischemic rats. Both treatments significantly reduced infarcted area, apoptosis and inflammation of ischemic brains, and improved the sensorimotor functions. Thus, this thesis proposes a new therapeutic target for ischemic stroke.