Date of Award

2004

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

Dhandapani Kuppuswamy

Fourth Advisor

Robert P. Thompson

Fifth Advisor

Maria Trojanowska

Abstract

Abnormalities of the tissue kallikrein-kinin system have been identified in the pathogenesis of hypertension and cardiac and renal diseases. The purpose of this study was to investigate the effect of exogenous tissue kallikrein gene delivery on heart and kidney injury in pressure overload and volume overload hypertension using spontaneously hypertensive rats (SHR) and Dahl salt-sensitive (DSS) rats. In order to examine the role of kallikrein in cardiac protection, adenovirus encoding the human tissue kallikrein gene was injected intravenously into SHR. Expression of recombinant human tissue kallikrein was identified in rat serum by ELISA and heart and kidney tissue by RT-PCR/Southern blot one week after gene delivery. Kallikrein gene transfer lowered blood pressure in SHR for the first two weeks, but had no effect from 3-5 weeks. Five weeks post-injection, SHR receiving kallikrein exhibited significantly less left ventricular hypertrophy, as denoted by a considerable decrease in cardiomyocyte size, and collagen accumulation compared to control SHR. The protective morphological alterations by kallikrein were associated with decreased levels of transforming growth factor-β1 (TGF-β1), plasminogen activator inhibitor-1 (PAI-1), and phosphorylated c-jun N -terminal kinase (JNK). In addition, kallikrein gene transfer promoted neovascularization in the hearts of SHR, which normally have diminished capillary density. Furthermore, kallikrein's protective effects on cardiac remodeling and capillary density in SHR were accompanied by increased cardiac nitric oxide (NO) and cGMP production, implicating a role for the NO-cGMP pathway. The effect of kallikrein gene transfer against salt -induced kidney injury was investigated in DSS rats. Four weeks after high salt (4% NaCl) loading, when renal damage was apparent, adenovirus harboring the human tissue kallikrein gene was injected into DSS rats with or without infusion of the B2 receptor antagonist icatibant. Kidneys were examined before and two weeks after gene delivery by histochemical and biochemical analyses. Kallikrein gene delivery attenuated salt-induced renal dysfunction, as evidenced by a decrease in urinary protein and blood urea nitrogen. In addition, kallikrein gene transfer not only attenuated but also reversed inflammatory cell infiltration, myofibroblast accumulation, collagen deposition, and glomerular hypertrophy induced by high salt intake. The morphological changes mediated by kallikrein were associated with reduced intercellular adhesion molecule-1 (ICAM-1) and TGF-β1 expression, and cell cycle inhibitory protein (p21, p27) levels. Moreover, kallikrein restored NO production and suppressed NADH oxidase activity and superoxide formation. Kallikrein's effects were abolished by icatibant, indicating a kinin-B2 receptor-mediated event. These studies provide the novel finding that kallikrein/kinin acts as an anti-inflammatory agent and an anti-oxidant in attenuating and reversing salt-induced inflammation, fibrosis and glomerular hypertrophy. Collectively, these results demonstrate that kallikrein/kinin provides beneficial effects against heart and kidney complications in pressure- and salt-induced hypertension.

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