Vol.30 (2007) No.1
Developmental Activity of the Renin-
Angiotensin System during the “Critical
Period” Modulates Later L-NAME–Induced
Hypertension and Renal Injury
Kimiko ISHIGURO1), Hiroyuki SASAMURA1), Yusuke SAKAMAKI1),
Hiroshi ITOH1), and Takao SARUTA1)
The incidence of hypertension and hypertensive renal disease is increasing worldwide, and new strategies
to prevent these diseases need to be investigated. The aims of this study were 1) to examine if transient
exposure to an angiotensin receptor blocker (ARB) during an early period in hypertension development con-
fers protection against subsequent worsening of hypertension and renal injury induced by the NO synthase
inhibitor NG-nitro-L-arginine methyl ester (L-NAME), and 2) conversely, to examine the effects of transient
exposure to angiotensin II (Ang II) during the same period. First, spontaneously hypertensive rats (SHR)
were treated transiently from age 3 to 10 weeks with an ARB (candesartan cilexetil), a calcium channel
antagonist or a vasodilator, then taken off treatment for 2 months. Administration of L-NAME at age 18
weeks caused severe hypertension and renal injury. However, the rats that had been exposed to the ARB
not only had a lower blood pressure, but also failed to show signs of renal injury or increase of oxidative
stress. Furthermore, the elevation of components of the renin-angiotensin-aldosterone system was also
suppressed in these rats. In the second study, Wistar-Kyoto rats (WKY) and SHR were exposed to Ang II
from age 4 to 8 weeks. The follow-up showed that the blood pressures in the WKY remained elevated com-
pared to controls, while the SHR had heightened increases in blood pressure, renal renin mRNA, and urinary
8-hyroxydeoxyguanosine after L-NAME administration. Together, these experiments demonstrate that tran-
sient treatment of rats during an early phase in the development of hypertension with an ARB suppresses
the renin-angiotensin-aldosterone system and confers long-term protection against subsequent L-NAME–
induced renal injury and increases in renal oxidative stress. Conversely, developmental exposure to Ang II
during this “critical” period had the opposite effect, predisposing rats to higher blood pressure, renal injury,
and oxidative stress after L-NAME administration. (Hypertens Res 2007; 30: 63–75)
Key Words: angiotensin, hypertension, renal injury, renin-angiotensin-aldosterone system, oxidative stress
The incidence of hypertension was estimated as approxi-
mately 26.4% in 2004 and continues to increase worldwide,
with a projected value of 29.2% by 2025 (1). Renal injury is
an important complication of hypertension, and hypertensive
nephrosclerosis is the leading cause of end-stage renal disease
in many countries. It is therefore an important research goal to
develop new methods to prevent the incidence of hyperten-
sion and hypertensive renal injury.
It has been shown that the renin-angiotensin system plays a
From the 1)Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan.
Address for Reprints: Hiroyuki Sasamura, M.D., Ph.D., Department of Internal Medicine, School of Medicine, Keio University, 35 Shinanomachi, Shin-
juku-ku, Tokyo 160–8582, Japan. E-mail: email@example.com
Received May 1, 2006; Accepted in revised form September 4, 2006.
Hypertens Res Vol. 30, No. 1 (2007)
central role in the pathogenesis of both hypertension and
hypertensive renal injury. Recent studies have also high-
lighted a role for oxidative stress in the pathophysiology of
hypertension and hypertensive end-organ damage (2, 3).
Therefore, the development of interventions to prevent exces-
sive activation of the renin-angiotensin system, and to perma-
nently attenuate the development of oxidative stress, would
be important steps toward the ultimate goal of preventing
hypertension and hypertensive renal disease.
Previously, we reported that treatment of stroke-prone
spontaneously hypertensive rats (SHRSP) or Dahl salt-sensi-
tive (Dahl-S) rats during a critical period in the development
of hypertension (from age 3 to 10 weeks) with an angiotensin
inhibitor resulted in an attenuation of hypertension and renal
damage (4, 5). Both of these rat models can be categorized as
genetic (i.e., hereditary) models of hypertensive renal disease.
The aim of the present study was to examine whether tran-
sient exposure to an angiotensin receptor blocker (ARB) can
confer long-term protection against the later development of
acquired hypertension and renal damage induced by the
administration of a nitric oxide (NO) synthase inhibitor, NG-
nitro-L-arginine methyl ester (L-NAME), and to examine the
Fig. 1. Effects of transient exposure to the ARB candesartan cilexetil on blood pressure in L-NAME–treated SHR. Control:
untreated; Veh: rats treated with vehicle (tap water); Cand trans: rats treated with candesartan cilexetil from age 3 to 10 weeks;
Cand sust: rats treated with candesartan cilexetil from age 3 to age 21 weeks; Nica trans: rats treated with nicardipine from age
3 to age 10 weeks; Hydra trans: rats treated with hydralazine from age 3 to age 10 weeks; L-NAME: treated with L-NAME from
age 18 to age 21 weeks. Blood pressures in the candesartan-treated groups were significantly reduced compared to the other
four groups at all time points after week 12 (p<0.01, symbols omitted for clarity).
Table 1. Systolic Blood Pressure (SBP) and Heart Rate (HR) in the Different Groups in Experiment 1
SBP (mmHg) 3 w
HR (beats/min) 3 w
SBP (mmHg) 10 w
HR (beats/min) 10 w
SBP (mmHg) 18 w
HR (beats/min) 18 w
SBP (mmHg) 20 w
HR (beats/min) 20 w
Veh: rats treated with vehicle (tap water); Cand trans: rats treated with candesartan cilexetil from age 3 to 10 weeks; Cand sust: rats
treated with candesartan cilexetil from age 3 to age 21 weeks; Nica trans: rats treated with nicardipine from age 3 to age 10 weeks;
Hydra trans: rats treated with hydralazine from age 3 to age 10 weeks. *p<0.05, **p<0.01 vs. L-NAME (+) Veh. w, weeks.
Hypertens Res Vol. 30, No. 1 (2007)
hypertension. Further studies are required to confirm the
importance of this putative “reno-vascular amplifier” mecha-
nism in the development of hypertension in different models
An interesting finding of this study was that the rats in both
the candesartan (transient) and (sustained) groups showed a
clear resistance to the L-NAME–induced increases in oxida-
tive stress. Increases in oxidative stress have been considered
to cause end-organ damage which can occur not only in
hypertension, but also in other related diseases, including dia-
betes, hyperlipidemia, and obesity (18–20). At present, it is
unclear whether the changes in oxidative stress levels are a
cause or a consequence of the hypertension and renal damage,
and further studies are required to examine this issue in
It should be noted that the rats in the candesartan (sus-
tained) group had a tendency toward reduced body weights
compared to the animals in the other groups. Because of these
effects on the body weight, the abilities of candesartan to
attenuate heart weights and media thickness were statistically
significant in the absence of body weight correction, but lost
statistical significance when the values were corrected for
body weight. Recently, Kouyama et al. reported that AT1a
knock-out mice exhibited attenuation of diet-induced weight
gain. The mechanism appears to involve a modulation of adi-
pocytokine production by the AT1a receptor, resulting in
reduced adiposity through increased energy expenditure,
without a change in food intake (21). However, it is unlikely
that the observed decreases in the heart weights and aortic
media thickness in the present study were caused by reduced
food intake and malnutrition, because the changes in body
weights could be explained by reduced adipose tissue.
The results of this study may have important clinical impli-
cations. Although the pathophysiology of hypertension in the
SHR may not be identical to that in human essential hyperten-
sion, our study supports the view that inhibition of the renin-
angiotensin system before the appearance of hypertension
(i.e., at the pre-hypertension stage) could have long-term
effects on the later development of hypertension and hyper-
tensive end-organ damage. This would provide a rationale for
the treatment of hypertension at an earlier stage in order to
attenuate the severity of hypertension and hypertensive renal
injury in susceptible individuals. In this connection, a recent
clinical study (the TROPHY study) has suggested that treat-
ment of prehypertensive patients is feasible and may reduce
the risk of subsequent development of hypertension (15).
Focusing on interventions before the appearance of hyperten-
sion or renal damage may be important for the prevention of
hypertension and renal injury at a later time point.
In summary, the results of this study suggest that transient
treatment with an ARB during a “critical period” in hyperten-
sion development causes suppression of the renin-angio-
tensin-aldosterone system, and provides long-lasting
protection against the later development of acquired hyper-
tension, renal injury, and oxidative stress in the L-NAME
model. Exposure to Ang II has the opposite effect, enhancing
the activity of the renin-angiotensin-aldosterone system, and
predisposing rats to increased blood pressure, renal injury,
and oxidative stress. These results support the hypothesis that
the activity of the renin-angiotensin during this stage in
hypertension development may play an important role in
determining the susceptibility of individuals to hypertension,
renal injury, and oxidative stress in later life.
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