Browsing by Subject "neuroinflammation"

Mild traumatic brain injury (TBI) is defined as an injury that disrupts the normal functioning of the brain and is the result of external force to the head. It is the most common type of traumatic head injury, and it is common especially in contact sports and within military personnel. Mild TBI typically causes no clear structural changes to the head, but it can induce persistent clinical symptoms, as well as microscopic pathological changes to the brain that may eventually lead to neurodegeneration and increase the risk for several diseases. Mild TBI is a risk factor for several neurodegenerative diseases, including Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, and chronic traumatic encephalopathy. The primary objective of this study was to develop a repetitive mild TBI mouse model for future research purposes in the field of head trauma and neurodegeneration. The injury was induced as a closed head injury with an electromagnetic impactor. Literature and pilot experiments were used to define the parameters of the impactor required to induce a brain injury of desired severity. The characterization criteria of the mild TBI model considered the criteria used to define human mild TBI, as well as long term effects often reported after repetitive mild TBI: neurodegeneration as tau protein related pathology, neuroinflammation, and memory deficits. The secondary objective of this study was to tentatively test a prolyl oligopeptidase (PREP) inhibitor on the behavioral and histological effects of mild TBI. The functioning of the mild TBI model was studied by histopathological and behavioral assessments. After baseline behavioral assessment and repetitive (1 injury every 24 hours altogether 5 times) mild TBI inductions, the mice were monitored for approximately 3 months, during which several rounds of behavioral tests were performed. Barnes maze and novel object recognition tests were used to assess memory functions, and locomotor activity test was used to assess general locomotor activity. After euthanasia, brain histopathology was performed to study the amount of tau protein and the level of neuroinflammation. Due to the low number of animals in the study, the results are directional and need to be confirmed in subsequent studies. The histopathology showed greater amount of neuroinflammation and tau protein in the brains of injured mice, but statistical evaluations could not be made. Memory functions were slightly worse in the injured mice compared to controls, but significance of the results is unclear. Locomotor activity was not influenced by the mild TBIs. PREP inhibition treatment increased the locomotor activity of the mice, but the significance is unclear. The mild TBI model seems promising and the characterization criteria were partially met. The results of the study need to be verified in subsequent studies with a greater amount of animals. The model developed here can be used to study the involvement of head trauma in neurodegeneration, as well as treatment alternatives to changes caused by mild TBIs. As there currently are no curative treatments to neurodegenerative diseases, research regarding neurodegeneration and its risk factors is highly important.

Meningeal lymphatic vessels (mLVs), a recent functionally characterized structure in the meninges, contribute to the clearance of macromolecules, immune cells and metabolic waste from the central nervous system to peripheral lymph nodes. Having been identified as a route of clearance, there is a focus on understanding their role in neurological disease pathology. Here we consider their function in the pathology of traumatic brain injury (TBI) particularly in blood solute clearance, lesion progression and neuroinflammatory response. We use a transgenic model of mLV developmental dysfunction, K14-VEGFR3-Ig, to analyze the progression and severity of a controlled cortical impact (CCI) injury. We show that in mice lacking mLVs there are a higher percentage of microglia cells in an activated state in the hippocampus whereas the progression of hematoma and lesion size does not differ from wild type. Our results suggest that at two months post injury, meningeal lymphatics could be functionally important in modulating microglia activation, which is associated with chronic inflammation.

Epileptic patients experience spontaneous recurrent seizures and interictal epileptiform discharges that lead to brain injuries, triggering neuroinflammation and waste product accumulation. Due to the detrimental effect of waste products on brain homeostasis, their removal from the central nervous system is (CNS) is crucial. Meningeal lymphatic vessels (mLVs) located in dura matter contribute to CNS clearance by the drainage of metabolites, waste products, and immune cells from subarachnoid space into cervical lymph nodes. Therefore, because of its role in brain homeostasis, the study of mLVs in different neurological conditions and diseases, including TLE, has gotten increased attention in the last decade. In this study, we sought to understand mLVs role in neuroinflammation and changes in rapid eye movement (REM) sleep stage during epilepsy. For this purpose, we induced mLVs ablation followed by kainic acid (KA) epilepsy model in mice. Shortly, animals were inoculated with AAV-VEGFR3-1-4 to induce mLVs ablation and subsequently challenged with KA to induce status epilepticus. Simultaneously, a control group of animals were injected with a sham AAV and later injection of KA. Afterward, spontaneous EEG activity was registered continuously, and data analysed to compare durations of REM sleep. Also, immunohistochemistry of brain samples was performed to investigate neuroinflammatory changes between experimental groups. Ex-vivo analyses of Iba1 and GFAP expression in brain tissue did not show statistically significant changes in neuroinflammation between experimental groups. However, we observed a trend towards lower expression of inflammatory markers in mLVs ablated animals. The analysis of REM sleep duration shows a progressive reduction of this sleep stage in both groups during the first recording period with a subsequent stabilization during the second one. Our data also indicate that mLVs ablated animals present prolonged REM sleep duration compared to the control group. Although this data contradicts our initial hypothesis it is consistent with the well-established negative correlation between neuroinflammation and REM sleep duration. Future studies should consider a deeper analysis of the glial cell profile for a better understanding of the effect of mLVs dysfunction on epileptic pathology. Moreover, the impact of mLVs ablation on REM sleep duration should be characterized in healthy animals.

The APOE4 gene variant of apolipoprotein E (apoE) has been identified as a predisposing factor for late-onset Alzheimer’s disease (AD). ApoE is known to interact with one of the classically characterized hallmarks of AD, the amyloid-beta (Aβ) protein. Aβ activates the classical complement pathway by binding to C1q that may sustain inflammation, hamper Aβ clearance and therefore promote accumulation of Aβ deposits in the brain and affect the integrity of the blood-brain barrier. Depending on context, binding of apoE to C1q can either activate or inhibit complement activation. ApoE can also interact with the complement regulator factor H (FH) which colocalizes with C1q on Aβ plaques. The interactions between these proteins and their role in the AD pathogenesis has not been fully explored. This thesis aimed at unraveling the complex formation between Aβ1-42, apoE, FH, and C1q, and their role in complement activation to provide novel insights on the mechanisms contributing to AD pathophysiology. By conducting enzyme-linked immunosorbent assays and Western Blotting of native PAGE, I was able to show that FH bound to apoE and forms a complex with Aβ1-42/apoE in an isoform- specific manner (apoE2 > apoE3 > apoE4) whereas C1q bound all Aβ1-42/apoE complexes with same affinity. FH did not bind Aβ1-42 alone, thus the results indicated that binding of FH in Aβ1-42/apoE complex took place via apoE2 and apoE3. In the absence of FH and C1q, immobilized Aβ1-42/apoE4 complexes led to higher serum complement activation levels when compared to Aβ1-42/apoE2 and Aβ1-42/apoE3 complexes or Aβ1-42 alone. C1q activated serum complement especially in the presence of Aβ1-42/apoE3 and Aβ1-42/apoE4. Further functional analysis with flow cytometry revealed that in fluid phase the presence of FH could restrict Aβ1-42-induced complement activation, the subsequent formation of C5a and activation of C5a receptor (C5aR) expressed on Human embryonic kidney (HEK) 293 cells. The presence of all apoE isoforms on Aβ1-42/C1q complexes led to reduced C5aR activation. Surprisingly, however, the reduction was the largest with apoE4, and the presence of C1q on Aβ1-42 alone could also decrease C5aR activation. The results of this thesis elucidated the effects of Aβ1-42, apoE, C1q and FH in complement activation and proposed a mechanism by which FH regulates complement activation and cell-mediated inflammation through isoform-specific binding to apoE associated with Aβ1-42. Further work is required to analyze the functional effects of these complexes, whether Aβ1-42, apoE, FH, and C1q can all bind simultaneously to form a large complex and whether this complex affects C5aR activation. This study provides important insights on how APOE4 may predispose to the neurotoxicity and neuroinflammatory changes in AD pathophysiology early in the disease process. Yet, the findings are only preliminary and therefore further studies are required to validate these results and to demonstrate whether promoting FH binding to apoE/Aβ complex could provide a novel target for AD therapy.