Blood Pressure Regulation by Vasoactive Peptide Genes Transgenic

Blood Pressure Regulation by Vasoactive Peptide Genes: Transgenic And Knockout Animal Models Dowhan Ahn Department of Physiology Kosin University College of Medicine

Vasoactive Peptides BP • • Angiotensin Endothelin Vasopressin Urotensin II BP • Natriuretic Peptide • Bradykinin • Adrenomedullin

DNA microinjection

Gene targeting

BP measurements • Catherization : accurate, anesthized • Tail-cuff : less accurate, awake • Radiotelemetry : most accurate, awake

The vasoactive pepide systems 1. 2. 3. 4. RAS KKS NPS ES

The Renin-Angiotensin System Angiotensinogen Renin (*kiren) Angiotensin I ACE (*pril) Angiotensin II AT II receptors (*sartan)

Actions of AT II • TPR BP • Na retention • Hypertrophy and remodeling of blood vessels & heart

Transgenic mice with the rat renin and angiotensinogen genes fused with mouse MT-I promotor (Ohkubo H et al: PNAS, 1990)

Angiotensinogen-deficient mice: hypotension (Tanimoto K: JBC, 1994)

Rescue of renovascular abnormalities and hypotension of angiotensinogendeficeint mice (Ishida J: BBRC, 1998) By mating MTAgt mice with Agt-KO mice Control Knockout Rescue

Transgenic rats (TGR 27) with mouse Ren-2 genes : fulminant hypertension (Mullins JJ et al: Nature, 1990; Lee MA et al: AJP, 1996) • Agt level • ACE • AT I & II ( ) • Renin (plasma and kidney) & Prorenin

Renin (Ren-1 C)-deficient mice: hypotension (Yanai K: JBC, 1999)

ACE knockout mice (Cole JM et al: AJP, 2003)

Mapping of AT receptor type 1 (Zhuo J et al: Kidney Int, 1998)

AT II receptor type 1 A-deficient mice (Ito M et al: PNAS, 1995; Sugaya T: JBC, 1995) * Hyperreninemia

AT II receptor 1 a/1 b double knockout mice (Oliverio MI et al: PNAS, 1998) WT KO WT double KO

AT II receptor 2 -null mice: hypertension (Ichiki T et al: Nature, 1995) (Gross V et al: J Hypertens, 2000) Similar AT II concentrations Up-regulation of the AT 1 receptor Alterations in central nervous system functions

The Kallikrein-Kinin System LMW Kininogen tissue kallikrein Bradykinin B 2 receptor (icatibant) kininase I (or ACE) d. Arg-Bradykinin B 1 receptors

• HMW kininogen and plasma kallikrein - liver>> platelet, PMN - inflammation and blood coagulation • LMW kininogen and tissue kallikrein - kidney, endothelial cells - control of organ perfusion and BP; stimulation of endothelial mediators

Kinin transgenics and KO • Kininogen overexpression ? • Kininogen knockout ? (human-1 gene; rat-3 genes; mouse-2 genes) • Kininogen-deficient rats (Norway- Katholiek strain; do not export synthesized kininogen) – hypertensive in response to Na. Cl loading only

Transgenic mice with human tissue kallikrein genes (Chao J &Chao L: Hypertension, 1996)

Mapping of B 2 receptor (Zhuo J et al: Kidney Int, 1998)

B 2 receptor transgenic mice: life-long hypotension (Wang D et al: AJP, 1999) Control Blood pressure, mm. Hg 98. 6 ± 0. 5 Transgenic 85. 4 ± 0. 2

Hypertension in B 2 receptor knockouts - controversial (Cervenka l et al: Hypertension, 1999; Milia Af et al: Hypertension, 2001)

The Natriuretic Peptide System pro-Natriuretic Peptide NEP (ACE inhibitors) Natriuretic Peptide NPR-A (agonists: *ritide) NPR-B NPR-C

NPs • Endogenous antagonists of the RAS • Kindey IMCD- urodilatin • Reduction in cardiac preload BP • Any increase in the NP (therefore, the lowering effect of BP) can not overcome an activated RAS

ANP transgenic mice (Steinhelper ME et al: Hypertension, 1990) (Veress AT et al: AJP, 1995) • w/o natriuresis and diuresis

ANP knockout mice: salt-sensitive hypertension (Melo LG: AJP, 1998) hyperreninemia

Mapping of NPR (Kim SW: Kidney Int, 2001)

NPR-A knockouts: hypertension (Oliver PM et al: PNAS, 1997)

NPR-C knockouts: skeletal deformities (Matsukawa: PNAS, 1999) +/+ -/ - • ANP half-life, min 1. 44 ± 0. 1 2. 40 ± 0. 1 • Plasma ANP 114. 5 ± 5. 9 88. 5 ± 9. 0 • Plasma BNP 21. 6 ± 3. 8 22. 1 ± 1. 9 • BP, mm. Hg 118. 7 ± 1. 9 110. 4 ± 2. 3

The Endothelin System pre-pro. Endothelin endopeptidase pro. Endothelin (Big Endothelin) ECE Endothelin (sarafotoxin 6 b) ETRA or ETRB (*sentan)

Vasoconstrictive effect of ET-1 (Pallone TL et al: AJP, 2003) • Low dose (GFR, RPF unchanged)- BP & natriuresis & diuresis • High dose (GFR, RPF decreased) - BP & antinatriuresis & antidiuresis

ET-1 and ET-2 transgenic mice (Hocher B et al: JCI, 1997) (Hocher B et al: Hypertension, 1996) Plasma ET level /

Endothelium-restricted overexpression of human ET-1 (Amiri F et al: Circulation, 2004) No significant increase in BP WT TG

Mapping of ETRB (Zhuo J et al: Kidney Int, 1998)

Global KO of the ETS

Rescue of congenital ETRB-deficient rats: salt-sensitive hypertension (Gariepy CE et al: JCI, 1998)

Conditional gene targeting

Collecting duct–specific knockout of ET-1: hypertension and sodium retention (Ahn D et al: JCI, 2004)

Conclusion & Comment • The vasoactive peptide systems play an important role in BP regulation and salt homeostasis. – interplay ? • Other physiologic factors such as NO and prostanoids also interact with the VPS. –interplay ?