Perit Dial Int
29(1):
111-114
2009
© 2009 International Society for Peritoneal Dialysis
Direct Measurement of Nitric Oxide Concentration in CAPD Dialysate
Seiichi Mochizuki1,2,*,
Aya Takayama2,4,
Tamaki Sasaki3,
Hideyuki Horike3,
Naoki Kashihara3,
Yasuo Ogasawara1,2,
Fumihiko Kajiya1,2,
Tomoki Kitawaki4 and
Hisao Oka4
Department of Medical Engineering1 Kawasaki
Medical School Department of Medical Engineering2
Kawasaki University of Medical Welfare Division of
Nephrology3 Department of Internal Medicine Kawasaki
Medical School Kurashiki City Graduate School of Health
Sciences4 Okayama University Medical School Okayama
City, Japan
*
e-mail:
mochi{at}me.kawasaki-m.ac.jp
In long-term continuous ambulatory peritoneal dialysis (CAPD) patients, a
gradual increase in peritoneal permeability to small solutes and water and
consequent failure of ultrafiltration capacity are often observed
(1). Endogenously produced
nitric oxide (NO) may regulate ultrafiltration capacity during CAPD, given its
crucial role in the regulation of vascular tone and permeability and its
interactions with angiogenic growth factors
(2–4).
Some previous studies suggested that abnormal production and/or oxidation of
NO, that is, changes in NO bioavailability, induce peritoneal injuries
(3,5).
Thus, it is highly important to directly measure NO to assess the
bioavailability of NO and its relevant role in CAPD. However, there have been
no studies on direct measurement of NO in CAPD. Recently, we reported the
basic performance and applicability of a newly developed NO sensor
(6) and have developed
catheter-type NO sensors for measurement in the aorta
(7–9)
and coronary circulation (10).
In the present study, we investigated the applicability of this type of NO
sensor to direct measurement of NO concentration in CAPD dialysate.
 |
MATERIALS AND METHODS
|
|---|
Subjects: 32 CAPD patients (22 males, 10 females; age 31 –
79 years) volunteered for this study. CAPD duration ranged from 2 months to 4
years and 7 months. Three types of dialysates, Dianeal-N PD4 1.5 (19
patients), 2.5 (6 patients) (Baxter Healthcare, Tokyo, Japan), and Midpeliq
L135 (7 patients) (Terumo, Tokyo, Japan) were used. Dialysate volume ranged
from 1000 to 2000 mL. This study protocol was approved by the Ethics Committee
of Kawasaki Medical School. Written informed consent was obtained from each
patient.
NO Sensor: An NO sensor (model: amiNO-700) and an NO monitor
(model: inNO-T; Innovative Instruments, Tampa, FL, USA) were used for
measurement of NO concentration in dialysates. Each sensor was calibrated with
NO-saturated pure water at the beginning and the end of the day of the
experiments (6). We recently
reported the details and basic performance of this NO sensor system as well as
its applicability to in vivo measurement of plasma NO concentration
in animal models
(6,7,10).
Both working and reference electrodes are encased in a cylindrical
gas-permeable membrane to isolate the electrodes from a sample solution to
assure the selectivity and stability of the sensor. Thus, only gas-phase
molecules such as NO can permeate the membrane. Compared with O2,
this sensor shows much higher selectivity to NO (about 25000 times).
Therefore, this sensor is not influenced by changes in solution contents, such
as glucose, proteins, pH, or dissolved O2, all of which may be
observed in dialysates.
Sample Preparation and Measurement of NO: To minimize sampling
frequency and determine if there is a difference in NO concentration in
relation to sampling timing, the following two types of spent dialysates were
collected at the time of dialysate exchange: "pre-exchange," which
dwelled in the peritoneal cavity for about 5 hours before dialysate exchange,
and "post-exchange," which dwelled in the peritoneal cavity for
about 1 minute after dialysate exchange.
Before sample measurement, an NO sensor was calibrated using NO-saturated
pure water (6) and immersed in
fresh (unused) dialysate for stabilization. The sensor was then immersed in
the spent dialysate. Concentration of NO was obtained as the maximum
difference between the fresh (unused; zero level) and the spent dialysates
(see Figure 1). We normalized
the NO concentration, named "NO amount," based on body size and
dialysate volume (Eq. 1):
 | (1) |
View larger version (5K):
[in this window]
[in a new window]
|
Figure 1 — Representative tracing of measured nitric oxide (NO) concentration
in both fresh and spent dialysates. Broken-line arrow indicates zero-to-peak
change in NO concentration between fresh and spent dialysates (measured NO
concentration).
|
|
Statistical Analysis: All data are expressed as mean ± SEM.
Linear regression and correlation analyses were conducted to evaluate the
correlation between the amount of NO produced before and after dialysate
exchange. Statistical analyses were conducted by paired t-test for comparison
between before and after dialysate exchange. A p value < 0.05 was
considered statistically significant.
|
RESULTS
|
|---|
Determination of NO Concentration in Dialysate:
Figure 1 shows a representative
tracing of NO concentration in dialysate. A stable baseline (zero level) was
measured in fresh dialysate. When switched to the spent dialysate, measured NO
concentration increased rapidly and reached its peak level. This trend was
observed in all samples studied. We used the zero-to-peak change in NO
concentration between fresh and spent dialysates as "measured NO
concentration" (indicated as a broken-line arrow in
Figure 1).
Comparison Between Before and After Dialysate Exchange: The
measured NO concentration in spent dialysate was significantly higher in the
pre-exchange samples than in the post-exchange samples (5.0 ± 0.7 vs
1.9 ± 0.3 nmol/L, p < 0.001). The "NO amount"
in spent dialysate was also significantly higher in the pre-exchange samples
than in the post-exchange samples (Figure
2; p < 0.001). This result suggests that NO is
released from peritoneal and surrounding tissues.
Correlation Between Before and After Dialysate Exchange: There was
a good positive correlation in the amount of NO between pre-exchange and
post-exchange dialysates (Figure
3; p < 0.001). This result suggests that, when NO is
produced more significantly in one subject than in another subject at one time
point, the same trend may be observed at another time point.
When compared among the three types of dialysates [Dianeal-N PD4 1.5 (n =
19), 2.5 (n = 6), and Midpeliq L135 (n = 7)], there seemed no significant
differences in the correlations between pre-exchange and post-exchange
dialysates among these three groups (p = NS). To test possible
effects of dialysate components on the lifetime of NO, we performed a
calibration with NO-saturated pure water in each fresh dialysate. There were
no significant differences in the measured NO current between the three
dialysates and saline [about 270 pA/(nmol NO/L) in all solutions].
|
DISCUSSION
|
|---|
The NO sensor demonstrated satisfactory stability and performance in the
measurement of NO in spent dialysates. It was demonstrated that NO remains at
a measurable level in dialysate and the interindividual difference in NO
bioavailability among patients can be evaluated quantitatively.
The longer a dialysate dwelled in the peritoneal cavity, the more NO tended
to be detected (Figure 2). This
suggests that NO remains stable in spent dialysate. In dwelling dialysates and
the surrounding tissues, there may be various sources of NO production,
including macrophages, mesothelial cells, fibroblasts, and vascular
endothelium
(11,12).
Our previous studies showed that inducible-type nitric oxide synthase (iNOS)
and neuronal-type NO synthase (nNOS) mRNAs can be detected in cultured rat
mesothelial cells, while endothelial NOS (eNOS) mRNAs are not detected. In
hyperglycemia, the expression of both iNOS and nNOS mRNAs is increased
(13). Combet et al.
reported enhanced expression and activity of eNOS in peritoneal biopsies of
long-term CAPD patients
(4).
A wide range in the NO amount was observed among the patients studied
(Figure 3). The fate of NO
(bioavailability of NO) in dialysates is determined by multiple factors,
including production rate (NOS expression and activity), transport phenomena
(diffusion and convection), and balance between oxidants and antioxidants
(extent of oxidative stress). Thus, the reasons for different NO amounts may
vary from patient to patient. Further studies including plasma NO measurement
will be required to clarify the detailed mechanisms.
In conclusion, direct measurement of NO may provide us with new information
enabling clarification of the causes of peritoneal dysfunction in long-term
CAPD. It may also be possible to quantitatively evaluate peritoneal functions
as well as biocompatibility of dialysis solutions and to develop therapeutics
to preserve peritoneal functions.
|
ACKNOWLEDGMENTS
|
|---|
This study was supported in part by Grant-in-Aid for Science Research,
Japan Society for the Promotion of Science.
The authors thank Ms. Mayumi Ono for her superb technical assistance.
|
REFERENCES
|
|---|
- Churchill DN, Thorpe KE, Nolph KD, Keshaviah PR, Oreopoulos DG,
Pagé D. Increased peritoneal membrane transport is associated with
decreased patient and technique survival for continuous peritoneal dialysis
patients. J Am Soc Nephrol 1998;9
: 1285-92.[Abstract]
- Yang CW, Hwang TL, Wu CH, Lai PC, Huang JY, Yu CC, et al.
Peritoneal nitric oxide is a marker of peritonitis in patients on continuous
ambulatory peritoneal dialysis. Nephrol Dial
Transplant 1996; 11:2466
-71.[Abstract/Free Full Text]
- Douma CE, de Waart DR, Struijk DG, Krediet RT. The nitric oxide
donor nitroprusside intraperitoneally affects peritoneal permeability in CAPD.
Kidney Int 1997;51
: 1885-92.[Medline]
- Combet S, Miyata T, Moulin P, Pouthier D, Goffin E, Devuyst O.
Vascular proliferation and enhanced expression of endothelial nitric oxide
synthase in human peritoneum exposed to long-term peritoneal dialysis.
J Am Soc Nephrol 2000;11
: 717-28.[Abstract/Free Full Text]
- Ni J, Cnops Y, McLoughlin RM, Topley N, Devuyst O. Inhibition of
nitric oxide synthase reverses permeability changes in a mouse model of acute
peritonitis. Perit Dial Int 2005;25
(Suppl 3):S11
-14.[Abstract/Free Full Text]
- Mochizuki S, Himi N, Miyasaka T, Nakamoto H, Takemoto M, Hirano K,
et al. Evaluation of basic performance and applicability of a newly
developed in vivo nitric oxide sensor. Physiol
Meas 2002; 23:261
-8.[Medline]
- Mochizuki S, Miyasaka T, Goto M, Ogasawara Y, Yada T, Akiyama M,
et al. Measurement of acetylcholine-induced endothelium-derived
nitric oxide in aorta using a newly developed catheter-type nitric oxide
sensor. Biochem Biophys Res Commun 2003;306
: 505-8.[Medline]
- Imanishi T, Kobayshi K, Kuroi A, Mochizuki S, Goto M, Yoshida K,
et al. Effects of angiotensin II on NO bioavailability evaluated
using a catheter-type NO sensor. Hypertension2006; 48:1058
-65.[Abstract/Free Full Text]
- Imanishi T, Kuroi A, Ikejima H, Mochizuki S, Goto M, Akasaka T.
Evaluation of pharmacological modulation of nitroglycerin-induced impairment
of nitric oxide bioavailability by a catheter-type nitric oxide sensor.
Circ J 2007; 71:1473
-9.[Medline]
- Neishi Y, Mochizuki S, Miyasaka T, Kawamoto T, Kume T, Sukmawan R,
et al. Evaluation of bioavailability of nitric oxide in coronary
circulation by direct measurement of plasma nitric oxide concentration.
Proc Natl Acad Sci U S A 2005;102
: 11456-61.[Abstract/Free Full Text]
- Plum J, Tabatabaei MM, Lordnejad MR, Pipinika O, Razeghi P, Huang
C, et al. Nitric oxide production in peritoneal macrophages from
peritoneal dialysis patients with bacterial peritonitis. Perit Dial
Int 1999; 19(Suppl 2):S378
-83.[Abstract/Free Full Text]
- Davenport A, Fernando RL, Varghese Z. Intraperitoneal nitric oxide
production in patients treated by continuous ambulatory peritoneal dialysis.
Blood Purif 2004;22
: 216-23.[Medline]
- Ozeki M, Sasaki T, Fujimoto S, Satoh M, Kobayashi S, Haruna Y,
et al. The implication of NO and ROS their interaction in the
pathogenesis of peritoneal injury [Abstract]. J Am Soc
Nephrol 2003; 14:475A
.
This article has been cited by other articles:
|
|
|
|
 
D. Lang and P. E. James
FREE NITRIC OXIDE IN SPENT CAPD FLUID: A LEAP OF FAITH?
Perit. Dial. Int.,
September 1, 2009;
29(5):
505 - 507.
[Full Text]
[PDF]
|
|
|
TOP
MATERIALS AND METHODS
RESULTS
