Graduation Year

2017

Document Type

Dissertation

Degree

Ph.D.

Degree Name

Doctor of Philosophy (Ph.D.)

Degree Granting Department

Medicine

Major Professor

Paula C. Bickford, Ph.D.

Committee Member

Brian Giunta, M.D., Ph.D.

Committee Member

Stanley Stevens, Ph.D.

Committee Member

Daniel Lee, Ph.D.

Committee Member

Marcia Gordon, Ph.D.

Committee Member

Kevin Nash, Ph.D.

Keywords

Aging, Proteomics, Neuroinflammation, Microglia

Abstract

The last century has seen a steady increase in the extension of the average lifespan. This has concomitantly produced higher incidences of age-related chronic degenerative diseases like Alzheimer’s and Parkinson’s diseases. Age is the single greatest risk factor for the development of not just these degenerative conditions but cancer as well. The aged niche undergoes a number of maladaptive changes that allow underlying conditions to present and progress. Exactly which changes, contribute to the progression of which disease is currently an area of intense study. However, these answers often present therapeutic targets for disease prevention. Age is characterized by a progressive loss of tissue function that eventually leads to the death of the organism. At the cellular level, aged tissues are characterized by a loss of resident stem cell populations, senescence, and low-grade inflammation. While aging is heterogeneous in terms of its ultimate effect on tissue function the underlying changes have a degree of overlap. Cells often experience increased oxidative stress and a diminished activity in pathways like NRF2 whose role it is to provide resistance to such stress. Aged cells also have some change in their overall chromatin and nucleosome structure contributing to observable changes in gene expression and regulation. When these disruptions occur in tissues that can affect the larger organism such as the hypothalamus they affect the organism as a whole and contribute to syndromes seen in age such as insulin resistance. The immune system, in particular, is sensitive to both the cell-autonomous and systematic changes that occur with age. Immune and endocrine signaling pathways have a considerable amount of overlap, and mounting evidence points to the role of inflammation in the metabolic syndromes that occur with age. Immune dysfunction in the CNS has a special significance because of the dual roles of microglia. These cells exist not just to protect against foreign invasion but play vital roles in the maintenance of brain architecture and in processes central to cognition like long-term potentiation and the differentiation of stem cells in the hippocampus. The aged microglial phenotype contributes to the decline that occurs normally with age but can also be central to the progression of underlying pathologies including several degenerative diseases. Therapies targeting the maintenance of microglial function with age hold the potential to delay disease onset and possibly preserve cognitive function further into life.

Top-down research approaches are well suited for the study of interactions between complex systems. The rapid improvement of mass spectrometry over the past decade has allowed researchers to examine more complex samples with fewer preparation steps and improved accuracy. This approach has thus far worked very well in the study of aging with the number of “Omics” techniques in aging models increasing rapidly. We use both label-free mass spectrometry and the more traditional real-time PCR to analyze signaling pathways and systems in both tissue homogenates and isolated cells from aged animals. By analyzing inflammatory and neurogenic pathways in animals treated with polyphenolic compounds we were able to postulate that the improved behavioral effect of these compounds is likely related to the decrease of pro-inflammatory cytokines and a restoration of WNT signaling. Proteomic analysis of aged microglia revealed widespread changes in chromatin structure and cellular machinery responsible for the regulation of transcription. In addition, we uncovered a shift in the underlying metabolic state of aged microglia and identified several pathways upstream of these changes. These upstream pathways included mTOR, a well-studied nutrient sensing pathway that plays a role in regulating microglial phenotype. Modulation of identified pathways through the use of both genetic (siRNA) and pharmacological (allosteric inhibitor) was able to recapitulate the aged phenotype in normal cells, confirming the role of these pathways in pathological changes.

Download

Share

COinS