Multivariate Anti-inflammatory Approaches to Rescue Neurogenesis and Cognitive function in Aged Animals

vi CHAPTER 1: INTRODUCTION 1 1.1 Adult neurogenesis 1 1.1.1 Adult neurogenesis and cognitive function 3 1.1.2 Process of adult neurogenesis 8 1.2 Aging and neurogenesis 12 1.2.1 Aging and cognitive function 14 1.2.2 Oxidative stress and process of aging 17 1.2.3 Inflammation and aging 22 CHAPTER 2: MATERIALS AND METHODS 29 2.1 Animals 29 2.2 Surgical Procedures 29 2.2.1 Eyeblink conditioning surgery procedures 29 2.3 Sources of NT-020 30 2.4 Treatments 30 2.4.1 Treatment with rrTNF-α 30 2.4.2 Treatment with anti-TNF-α 31 2.4.3 Oral treatment of NT-020 in rats 31 2.4.4 NT-020 supplemented diet 31 2.5 Behavioral testing 31 2.5.1 Delay eyeblink classical conditioning (EBCC) 31 2.5.2 Morris water maze 32 2.5.3 Radial arm water maze 33 2.6 Immunohistochemistry 34 2.6.1 Tissue processing 34 2.6.2 Immature neuronal marker doublecortin (DCX), mitotic marker Ki67, and MHC class II OX6 immunostaining 34 2.7Stereology 35 2.8 Western blot 36 ii 2.9 Enzyme link immunosorbent assay (ELISA) 37 2.10 Statistical analysis 37 CHAPTER 3: ROLE OF TNF-α INDUCED INFLAMMATION IN DELAY EYEBLINK CONDITIONING IN YOUNG AND AGED RATS 38 3.1 Aging and tumor necrosis factor alpha 38 3.2 Norepinephrine and learning 39 3.3 Specific materials and methods 41 3.3.1 Animals and surgery 41 3.3.2 Treatment with rrTNF-α and anti-rat TNF-α 42 3.3.3 Training of behavior in a delay classical eyeblink conditioning task 43 3.3.4 Design and analysis 43 3.4 Results 44 3.4.1 Young rats 44 3.4.2 Aged rats 47 3.5 Discussion 50 3.5.1 TNF-α in young rats 51 3.5.2 anti-TNF-α in aged rats 52 3.6 References 54 CHAPTER 4: NT-020, A NATURAL THERAPEUTIC APPROACH TO OPTIMIZE SPATIAL MEMORY PERFORMANCE AND INCREASE NEURAL PROGENITOR CELL PROLIFERATION AND DECREASE INFLAMMATION IN THE AGED RAT 59 4.1 NT-020 effects in learning and memory and neurogenesis 59 4.1.2 Process of aging 60 4.2 Experimental Procedures 64 4.2.1 Sources of NT-020 64 4.2.2 Subjects 64 4.2.3 Oral treatment of NT-020 in rats 65 4.2.4 Behavioral Testing 65 4.2.5 Immunohistochemistry 66 4.2.6 Stereology 66 4.2.7 Statistics 67 4.3 Results 67 4.3.1 Cognitive function 67 4.3.2 Neural stem cell proliferation and neurogenesis in subgranular zone (SGZ) of dentate gyrus of the hippocampus is increased in aged rats after one month of NT-020 dietary supplementation 70 4.3.3 NT-020 treatment decreased activated microglia in the dentate gyrus of the hippocampus 73 4.3.4 NT-020 treatment increases proliferation of neural progenitors in the SVZ 75 4.4 Discussion 77 iii 4.5 References 80 CHAPTER 5: NT 020, A NATURAL THERAPEUTIC APPROACH TO AMELIORATE SPATIAL MEMORY PERFORMANCE, AND TO REGULATE DIFFERENT INFLAMMATORY MARKERS 85 5.1 NT-020 effects in spatial learning and memory and inflammatory markers 85 5.1.1 Aging and cognitive function 87 5.1.2 Influence of aging on CD4+T cells 88 5.1.3 Aging, microglia and inflammatory markers 90 5.1.4 Oxidative stress during the process of aging 91 5.2 NT-020 a natural therapeutic formulation 92 5.3 Specific materials and methods 93 5.3.1 Source of NT-020 93 5.3.2 Subjects 93 5.3.3 NT-020 treatment 93 5.3.4 Behavioral Testing 94 5.3.5 Protein expression by Western blot analyses 95 5.3.6 Analysis of TNF-α and IL-1β 96 5.3.7 Statistics 97 5.4 Results 97 5.4.1 Cognitive function 97 5.4.2 CD4+T cells expression in the thymus and hippocampus of young and aged rats after 4 weeks of NIH-31 control diet or NT-020 dietary supplementation 100 5.4.3 Inflammatory markers in the CNS of young and aged rats after 4 weeks of NIH-31 control diet or NT-020 dietary supplementation 102 5.5 Discussion 104 CHAPTER 6: DISCUSSION 106 6.1 Main goal 106 6.2 Cognitive function and neurogenesis 107 6.3 Pro-inflammatory cytokines 113 6.4 Influence of NT-020 diet on adaptive immune system 115 6.5 Predictions 116 6.6 Conclusion 119 REFERENCES 121 ABOUT THE AUTHOR End Page