Date of Award

2018

Embargo Period

8-1-2024

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biochemistry and Molecular Biology

College

College of Graduate Studies

First Advisor

Julie Chao

Second Advisor

Lee Chao

Third Advisor

Robin Muise-Helmericks

Fourth Advisor

Yi-Te Hsu

Fifth Advisor

Hongkuan Fan

Abstract

Kallistatin, a plasma protein, consists of two essential structural elements, an active site and a heparin-binding domain. Kallistatin exerts pleiotropic effects on angiogenesis, inflammation, fibrosis and tumor growth. This study aims to decipher the role and mechanism of kallistatin in vascular injury, senescence and aging. First, we determined the effect of kallistatin on endothelial-mesenchymal transition (EndMT), a process mediating vascular injury, organ fibrosis and cancer progression. Recombinant human kallistatin via its heparin-binding site blocked TGF-β-induced EndMT, associated with downregulated microRNA (miR)-21-Akt signaling and oxidative stress in human endothelial cells. Kallistatin’s active site is essential for stimulating antioxidant genes, endothelial nitric oxide synthase (eNOS) and sirtuin 1 (SIRT1) by interacting with a tyrosine kinase. These findings indicate that kallistatin suppresses EndMT by inhibiting miR-21-Akt signaling and oxidative stress. Secondly, we determined the role and mechanism of kallistatin in vascular senescence and aging. Kallistatin antagonized TNF- α-induced senescence and oxidative stress, and inhibited miR-34a, a senescence inducer, leading to elevated SIRT1/eNOS synthesis in human endothelial progenitor cells. Kallistatin administration in streptozotocin (STZ)-induced diabetic mice attenuated aortic senescence associated with reduced miR-34a and elevated SIRT1/eNOS levels. Consistently, kallistatin delayed stress-induced organismal aging in Caenorhabditis elegans by inhibiting miR-34 and elevating the longevity gene, sir-2.1 (SIRT1 homolog) synthesis. Therefore, kallistatin reduces vascular senescence and aging by inhibiting miR-34a-SIRT1/eNOS pathway. Thirdly, we determined the role of endogenous kallistatin in endothelial senescence, oxidative stress and inflammation by generating two strains of kallistatin knockout mice, endothelial cell-specific and general kallistatin knockout mice. Kallistatin via inducing an endo-protective miRNA Let-7g antagonized miR-34a-SIRT1-eNOS pathway and inhibited senescence, oxidative stress and inflammation in human endothelial cells. Conversely, kallistatin deficiency in mouse lung endothelial cells aggravated H2O2-induced senescence, oxidative stress and inflammation associated with downregulated Let-7g and antioxidant genes and upregulated miR-34a synthesis, indicating that kallistatin protects against endothelial senescence by modulating Let-7g mediated miR-34a-SIRT1-eNOS pathway. Moreover, systemic depletion of kallistatin exacerbated aortic oxidative stress and renal fibrosis in STZ-induced diabetic mice. These studies indicate that kallistatin plays a novel role in protection against vascular injury, senescence and aging by regulating Let-7g, miR-34a and miR-21 synthesis, and antioxidant gene expression.

Rights

All rights reserved. Copyright is held by the author.

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