Graduation Year
2020
Document Type
Dissertation
Degree
Ph.D.
Degree Name
Doctor of Philosophy (Ph.D.)
Degree Granting Department
Biology (Cell Biology, Microbiology, Molecular Biology)
Major Professor
Brant Burkhardt, Ph.D., M.Sc
Committee Member
Cecilia Nunes, Ph.D.
Committee Member
Sandy Westerheide, Ph.D.
Committee Member
Margaret Park, Ph.D.
Keywords
Autophagy, IGFBP2, NFAT, TCM
Abstract
Type 1 diabetes (T1D) is an autoimmune disease resulting in the loss of pancreatic β cells and subsequent loss of insulin production. Exogenous insulin coupled with continuous glucose monitoring is the only treatment for T1D. Therefore, novel interventional therapies are needed that can inhibit the autoimmune destruction and increase β cell proliferation and function. Based on initial in vitro studies, traditional Chinese medicine (TCM) extract, Cornus officinalis, may be able to serve in this function. CO has been used for 2 millennia for its glucose regulating potential and has shown efficacy in animal models but rarely studied in the context of T1D. Ethnopharmacology can simultaneously target multiple mechanisms using a single herb due to numerous bioactive ingredients, an advantage compared to conventional medicine. It is hypothesized that CO treatment may provide a β cell restorative and protective therapy for T1D by inhibiting the immune response and enhancing existing β cells. We analyzed our CO via HPLC/MS and discovered over 300 compounds including known bioactive ingredients, loganin, morroniside, ellagic acid and cornuside. Our preliminary evidence demonstrated a dose- and time- dependent increase in β cell proliferation and protection from cytokine-induced death upon CO treatment. CO also induced proliferation and formed pseudoislets during a long-term treatment of 1.1B4 cells. We evaluated the metabolic effect of CO via the Agilent Seahorse XF Analyzer and at 2h displayed an increase in maximal respiratory capacity. However, the molecular mechanism in which CO is inducing this phenotype has yet to be elucidated. Therefore, we employed transcriptomics by RNA-seq and discovered a 2-fold increase in calcium-dependent transcription factor, NFATc. NFAT is a transcription factor for β cell proliferation, function and insulin secretion. From this, we utilized a global and phosphoproteome mass spectrometry (MS) approach. Our results from the global analysis showed a 7-20-fold increase in β cell functional regulator, IGFBP2. IGFBP2 has shown T2D protective effects and regulation of glucose metabolism. Ingenuity pathway analysis (IPA) predicted an increase in Nrf2-mediated oxidative stress pathway at 12 and 24h. Furthermore, Nrf2 is indirectly regulated by P62, which was significantly hyperphosphorylated from our MS phosphoproteome results. Nrf2 transcriptionally regulates antioxidant enzymes upon oxidative stress caused by proinflammatory cytokines in T1D while P62 binds to LC3 to create the autophagasome for degradation of toxic material via autophagy. Autophagy promotes β cell survival that would otherwise lead to cell death. Altogether, CO induces β cell proliferation, function and immune protection. Future studies will examine the in vivo effect of CO.
Scholar Commons Citation
Tawfik, Arielle E., "Biological and Proteomic Characterization of Cornus officinalis on Human 1.1B4 Pancreatic β Cells: Exploring Use for T1D Interventional Application" (2020). USF Tampa Graduate Theses and Dissertations.
https://digitalcommons.usf.edu/etd/9040
