Extract
Note: Please read the complete
full text with Figures and Tables at
and an arterial catheter (PE30) was inserted into it to measure
arterial blood pressure with a pressure transducer (Transpac,
North Chicago, IL, USA). Left ventricle catheterization was
performed to monitor systemic hemodynamic function
parameters including the heart rate (HR), the left ventricular
systolic pressure (LVSP), the left ventricular end-diastolic
pressure (LVEDP), and the maximal velocity of pressure
increase (±dp/dtmax) with another pressure transducer (P23XL,
Statham; Nihon Kohden, Tokyo, Japan) connected to the
Polygraph System RM-6000 (Nihon Kohden) and the MacLab data acquisition program (MacLab/8S; Analog
Digital Instruments, Castle Hill, NSW, Australia). The left kidney
was then exposed through a midline incision. A perivascular
transonic ultrasonic transmit-time flow probe (1RB;
Transonic Systems, Ithaca, NY, USA) was mounted on the left
renal artery for measurement of renal blood flow, with the
signals transmitted to a transmit-time flow meter (T206;
Transonic Systems). A blunt superficial laser-Doppler probe
(MLD-1; Nankai University, Tianjin, China) was placed on
the kidney surface, and mounted on micromanipulators
(Narishige Scientific Instrument Laboratory, Tokyo, Japan)
so that movement artifacts were avoided. The probe was
connected to a laser-Doppler flowmeter (LDM; Nankai
University) to measure renal cortical microperfusion.
Details of the validation of the transit time laser-Doppler method
are given elsewhere[18,19].
After surgery, the animals were allowed to recover for 30
min. Then, animals were injected intravenously with vehicle
control (saline), and MLB at doses of 10, 30, and 60 mg/kg
consecutively, with 45 min between injections.
Mean values for each determination were analyzed over
a 0.5-min to 1-min period. Renal vascular resistance was
calculated from the mean arterial pressure and the
corresponding renal blood flow.
Statistical analysis Data were given as mean±SD, from 8
animals in each group. The statistical significance of
differences in the hemodynamic parameters was assessed using
one-way analysis of variance (ANOVA). Student¡¯s
t-test was used for comparison of the parameters with their baseline
values. Statistical significance was set at
P<0.05.
Results
Effects on renal microcirculation and hemodynamics
There was no difference between groups with respect to the
baseline values of renal hemodynamic and microcirculation
parameters (Figure 1). Neither vehicle nor MLB
administration had any significant effect on renal blood flow or renal
vascular resistance. In contrast, MLB administration
increased renal cortical microperfusion significantly, whereas
the vehicle had no effect (Figure 1, Tables 1, 2). This
microcirculation improving effect reached its peak 15 min after
injection and returned to baseline after 45 min. Fifteen
minutes after injection with 10, 30, or 60 mg/kg MLB, renal
cortical microperfusion increased by 38.7%±27.3%, 51.4%±22.2%,
and 62.4%±20.2%, respectively (changes relative to baseline;
P<0.01), whereas renal blood flow (2.3%±19.9%, 1.6%±
20.4%, 3.7%±9.7%, respectively) and renal vascular
resistance (-8.4%±13.8%, -1.6%±12.4%, -1.4%±9.1%, respectively)
did not change significantly.
Effects on systemic hemodynamics There was no
difference between groups with respect to the baseline values of
systemic hemodynamic parameters (Figure 2). Vehicle
administration had no effect on these parameters. Although
MLB administration had some effect on these parameters at
some time points, this effect was neither time-dependent nor
dose-dependent. Fifteen minutes after injection with MLB
60 mg/kg, when the effect of MLB on renal cortical
micro-perfusion had reached its peak, mean arterial pressure
(1.9%±10.2% vs baseline), heart rate (0.2%±5.4%
vs baseline), LVSP (0.4%±7.8%
vs baseline), LVEDP (7.8%±33.4%
vs baseline) and
+dp/dtmax (3.0%±6.8%
vs baseline) had not changed in a statistically significant way, whereas
-dp/dtmax increased slightly (5.7%±6.2%;
P<0.05 vs baseline).
Discussion
The kidneys play a central role in the regulation of the
body¡¯s salt and water balance. A highly regulated
microcirculatory and interstitial environment is essential for
optimum function of the kidneys. Although the renal function
improving property of MLB has been studied extensively,
there has been no report concerning its effect on renal
microcirculation. We demonstrated here, to our knowledge
for the first time, that MLB could ameliorate renal
microcirculation while causing no other significant changes to
hemo-dynamics.
In our study, the effect of intravenously administered
MLB on renal cortical microperfusion was dose-dependent,
and reverted back to the baseline level 45 min after MLB
administration. Li et al reported the pharmacokinetic
parameters of MLB after iv administration in 6 beagle dogs, and
showed that MLB was distributed and eliminated
quickly[21]. The mean
T1/2b values for MLB at doses of 3 mg/kg, 6 mg/kg,
and 12 mg/kg were 43±9 min, 42±7 min, and 42±10 min,
respectively. Therefore, the time-response curve of MLB
was correlated with its serum concentration-time profiles.
This effect of MLB on renal circulation is consistent with
previous studies. However, some differences exist. Yokozawa
et al reported that MLB increased renal blood
flow[20], which we did not find in our study. The different animal models
and techniques we used may account for this difference.
Yokozawa et al used rats with renal failure, which had
significantly lower renal blood flow than normal rats, whereas
we used normal Sprague-Dawley rats with normal renal blood
flow. The techniques we used to measure renal blood flow
were also different. Yokozawa et al used a needle-type
bipolar electrode electrolytic organ rheometer, whereas we used
a transit-time ultrasonic-Doppler flow meter with a much
higher precision (±5%).
The effect of MLB on renal microcirculation may be
attributed to several factors. The primary contributor may be
its potent antioxidant properties. Recent studies suggest
that free radicals may play a key role in regulating renal
microvascular tone. Although studies investigating the roles
of oxygen radicals in the physiological regulation of renal
microcirculation have only recently begun, it is evident that
oxygen radicals have important direct and indirect actions in
both cortical and medullary
microcirculation[22_25]. Because
O2_ and NO both contain unpaired electrons in their outer
orbits, they undergo extremely rapid, diffusion-limited
radical-radical reactions, leading to the formation of peroxynitrite
anions (ONOO_ ), strong oxidants that could prompt the
generation of hydroxyl radicals (OH_). In the renal
microva-sculature, free radicals can cause vasoconstriction, mediate
the vasoconstriction of other agonists, and modulate the
action of vasodilators (inactivate nitric oxide and blunt
endothelium-dependent
vasodilation)[26_28]. These findings have led to the idea that antioxidants might be used
therapeutically as part of a nephroprotective
strategy[29_31]. Our current study supports this idea: MLB was proven to be a
potent inhibitor of the production of superoxides, hydrogen
peroxide, and hydroxyl radicals, the three most common
oxygen radicals in the renal microvasculature. Here we propose
that MLB, as a potent antioxidant, scavenges free radicals,
blocks the
O2__ONOO__OH_
cascade, promotes NO bioavail-ability and thus ameliorates renal microcirculation.
Some other factors may play a role too. MLB has been
reported to improve the renal circulatory state through
activation of kallikrein and promotion of prostaglandin
E2 production[7,9,13]. Tissue kallikrein cleaves the kininogen sub
strate to release the vasoactive peptide kinin, which binds to
endothelial bradykinin B2 receptors and stimulates the
release of potent vasodilators, including prostacyclin, nitric
oxide, and endothelium-derived hyperpolarizing
factor[32,33]. The paracrine agent
PGE2 is the predominant cyclooxygenase metabolite of arachidonic acid in the
kidney[34]. PGE2 plays an important role in tubular reabsorption of salt and water as
well as in the control of renal vascular resistance and the
maintenance of glomerular hemodynamics. Despite several
reports of PGE2-induced
vasoconstriction[35,36], there is convincing evidence that
PGE2 acts primarily on the preglo-merular vasculature to counteract the effects of the
vasocons-tricting hormones and protect the kidney from excessive
vasoconstriction[37_41]. Although the effect of MLB on
cyclo-oxygenase has not been studied, it is reported to be a potent
inhibitor of 5-lipoxygenase, and such inhibition of
lipoxy-gengase causes a shift of arachidonic acid from the
lipoxy-genase to the cyclooxygenase pathway, which is thought to
result in increased formation of cyclooxygenase metabo-
lites[3]. Indeed, Yokozawa et
al reported that MLB increased urinary excretion of prostaglandin
E2 (PGE2) and
6-keto-PGF1a, while thromboxane B2
(TXB2) remained unchanged or decreased in rats with renal
failure[3]. Activation of kallikrein, promotion of
PGE2 production, and scavenging of radicals
could act simultaneously to increase the bioavailability of
NO and prostacyclin, the major vasodilators in the kidney.
These three effects are suggested to be the major
contributors to increased renal microcirculation after MLB
administra-tion. Nonetheless, the exact mechanism by which MLB
ameliorates renal cortical microperfusion is not completely clear,
and should be evaluated further.
In conclusion, the major finding of this study is that
intravenously administered MLB dose-dependently
ameliorates renal microcirculation. This finding suggests that the
renal protective properties of MLB may be mediated in part
by vasodilation of the renal microvasculature.
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