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【摘要】 Transient administration of ANG II causes persistent salt-sensitive hypertension associated with arteriolopathy, interstitial inflammation, and cortical vasoconstriction; blocking the vascular and inflammatory changes with mycophenolate mofetil (MMF) prevents vasoconstriction. While infiltrating leukocytes during the salt-sensitive hypertension phase express ANG II, the functional role of ANG II during this phase is not known. We examined the acute effect of candesartan on renal hemodynamics during the established salt-sensitive hypertensive phase and related these findings to direct measurement of intrarenal ANG II and inflammatory cells in rats previously exposed to ANG II with or without MMF treatment. Sham controls were also examined. The administration of ANG II, followed by exposure to high-salt diet, resulted in hypertension, cortical vasoconstriction, an increase in interstitial inflammatory cells (44.8 ± 1.3 lymphocytes/mm 2, and 30.8 ± 1.2 macrophages/mm 2 ANG II vs. 19.6 ± 2 lymphocytes/mm 2, and 22 ± 0.7 macrophages/mm 2 Sham), and increase in renal ANG II levels (1,358 ± 74.6 pg/ml ANG II vs. 194 ± 9.28 pg/ml Sham). Treatment with MMF during the administration of exogenous ANG II resulted in reduction in renal interstitial inflammation (19.7 ± 0.9 lymphocytes/mm 2 and 15.9 ± 0.8 machophages/mm 2 ), ANG II levels (436.9 ± 52.29 pg/ml), cortical vasoconstriction, and stable blood pressure levels during the subsequent challenge with a high-salt diet. Acute administration of candesartan similarly reduced renal vasoconstriction and blood pressure. We conclude that the cortical vasoconstriction occurring with salt-sensitive hypertension following exposure to ANG II is mediated by intrarenal ANG II, related, at least in part, to the interstitial inflammation.
【关键词】 renal hemodynamics candesartan AT angiotensin II receptor blockade
RECENT STUDIES HAVE SUGGESTED that salt-sensitive hypertension may result from a subtle acquired renal injury that alters the ability to excrete salt ( 13 ). One experimental model in which this has been demonstrated is the postangiotensin II model ( 15, 24 ). In this model the transient infusion of ANG II results in hypertension with renal vasoconstriction and the development of microvascular and tubulointerstitial injury ( 15 ). Interestingly, after the exposure to ANG II, the rats become susceptible to the development of salt-sensitive hypertension despite maintaining a normal glomerular filtration rate (GFR) ( 9, 24 ). Specifically, when rats are placed on a high-salt diet (HSD) they rapidly become hypertensive in association with the development of cortical vasoconstriction ( 9 ). The hemodynamic changes are associated with the presence of tubulointerstitial inflammation consisting of T cells and macrophages and local oxidative stress ( 22, 24, 25, 34 ). Importantly, many of the infiltrating cells express ANG II ( 9, 21 ). The administration of the immunosuppressant mycophenolate mofetil (MMF) during the initial ANG II infusion can block these histological changes ( 21, 24 ) and prevent the cortical vasoconstriction and hypertensive response to the HSD ( 21, 26 ).
An important issue is whether there is a functional increase in ANG II in this model. Classically, HSDs are associated with the expansion of blood volume and a reduction in the activation of the renin-angiotensin axis. However, intrarenal angiotensin is not suppressed by plasma expansion and in fact, it may function as a separate compartment ( 10, 18 ). Furthermore, Ruan et al. ( 27 ) reported that HSD increases the renal vasoconstrictive response to ANG II by increasing intrarenal expression of AT 1 receptors. Hence, we postulated that an increase in intrarenal ANG II during salt loading, likely related to the renal interstitial inflammation, could be responsible for the cortical vasoconstriction in this model. These experiments were addressed to test this hypothesis.
METHODS
Experimental Protocol
Male Sprague-Dawley rats (350 g) were administered ANG II (Sigma, St. Louis, MO; 435 ng·kg -1 ·min -1 in Ringer lactate) for 2 wk via subcutaneous miniosmotic pumps (Alzet model 2002, Alza, Palo Alto, CA) as previously reported ( 9 ). At day 14, the minipump was removed and the rats were allowed to recover for 5 days on a normal-salt diet (0.4% NaCl). The rats were then placed on a HSD (Harlan, 4% NaCl) for 5 additional wk and micropuncture studies were performed ( Fig. 1 ). Care, use, and treatment of all animals in this study were in strict agreement with the guidelines set for by the Instituto Nacional de Cardiología I. Ch. Two sets of experiments were done.
Fig. 1. Effect of ANG II infusion in the development of salt sensitivity during 7 wk of follow up. Rats receiving ANG II developed salt sensitivity, whereas concomitant treatment with mycophenolate mofetil (MMF) prevented salt-induced increase in blood pressure. Sham control rats on high-salt diet remained normotensive. Values are means ± SD. + P < 0.01 vs. ANG II+MMF. Values in the Sham-operated group are significantly (* P < 0.001) lower than the rest.
Micropuncture experiments. These included the following groups. ANG II group ( n = 10) rats received vehicle (0.5 ml of saline solution) 30-40 min before glomerular hemodynamic measurements during micropuncture. ANG II + Cand group ( n = 9) rats received single dose of 0.25 mg/kg of candesartan diluted in 0.5 ml of saline solution, 30-40 min before the glomerular hemodynamic measurements, during the micropuncture studies ( 5 ). ANG II + MMF ( n = 9) rats received 30 mg/kg body wt of MMF by gastric gavage during the 2 wk in which exogenous ANG II infusion was administered and during the 5-day washout period until the initiation of a HSD. Micropuncture studies were done as in the ANG II group above. ANG II+ MMF+Cand ( n = 9) rats received MMF (30 mg·kg -1 ·day -1 by gastric gavage) during the period of ANG II infusion and during the 5-day washout period until the initiation of a HSD. Rats received a single dose of 0.25 mg/kg of candesartan diluted in 0.5 ml of saline solution 30-40 min before glomerular hemodynamics were measured during the micropuncture studies. Sham-operated ( n = 10) rats had a surgical procedure for insertion of subcutaneous pumps but pumps were not inserted and received normal-salt diet for 2 wk followed by 5 wk of HSD. At the time of micropuncture, they received vehicle as described above. Sham + Cand ( n = 11) included 11 sham-operated rats as mentioned above; they received a normal-salt diet for 2 wk followed by 5 wk of HSD. Candesartan was administered before micropuncture experiments.
Determination of interstitial ANG II. These experiments were done to evaluate the interstitial ANG II concentrations during the salt-driven hypertension that follows 2 wk of exogenous ANG II infusion, with and without concomitant administration of MMF. For this purpose, we studied rats that received exogenous ANG II ( n = 10) or ANG II and MMF ( n = 12) for 2 wk, as described above, and then were given 5 wk of HSD. Sham-operated ( n = 12) rats were used as controls. Previously, the procedure was validated to obtain values from normal rats in our hands.
Blood Pressure Measurements
Systolic blood pressure (SBP) measurements were performed in conscious, restrained rats by tail-cuff plethysmography (Narco Biosystems, Austin, TX). Rats were conditioned twice before the blood pressure was measured at a basal period, every week during the first 2 wk and every 2 wk for the rest of the study.
Micropuncture Experiments
For micropuncture studies, the rats were anesthetized with pentobarbital sodium (30 mg/kg ip, and supplementary doses were administered as required). The rats were placed on a thermo-regulated table, and the temperature was maintained at 37°C. Polyethylene tubing was used to catheterize the trachea (PE-240), both jugular veins and femoral arteries (PE-50), and the left ureter (PE-10). The left kidney was exposed, placed in a Lucite holder and sealed, covering the kidney surface with Ringer solution. Mean arterial pressure (MAP) was continuously monitored with a pressure transducer (model p23 LX, Gould Hato Rey, Puerto Rico) and recorded on a polygraph (Grass Instruments, Quincy, MA). Blood samples were taken periodically every 45-60 min and replaced with blood from a normal donor rat.
Rats were maintained euvolemic by infusion of 10 ml/kg body wt of isotonic rat plasma during surgery, followed by an infusion of 10% polyfructosan (Inutest, Laevosan-Gesellschafft, Austria) in 0.9% sodium saline solution, at a 2.5-ml/h rate. After 60 min, seven timed samples of proximal tubular fluid were obtained to determine flow rate and polyfructosan concentration; intratubular hydrostatic pressure under free flow and stop-flow conditions, and peritubular capillary pressures were measured in other proximal tubules with a servo-null device (Servo-Nulling Pressure System, Instrumentation for Physiology and Medicine, San Diego, CA), as previously described ( 3 ). Polyfructosan was measured in plasma samples. Glomerular colloid osmotic pressure was estimated in protein from blood taken from the femoral artery and from the surface of the efferent arterioles.
The ability of candesartan to block the effects of ANG II on blood pressure was tested in five rats. Administration of intravenous 50 ng ANG II produced a rise in MAP of 60 ± 8.7 mmHg; this effect was completely blocked with 0.25 mg candesartan. In addition, at the end of the micropuncture studies in groups that received candesartan, a 50-ng bolus of ANG II did not produce any change in blood pressure.
Polyfructosan concentrations were determined by the Davidson and Sackner ( 7 ) technique. The volume of fluid collected from an individual proximal tubule was estimated from the length of the column of fluid in a capillary tube of uniform bore and known internal diameter. The concentration of tubular polyfructosan was measured by the method of Vurek and Pegram ( 37 ). The protein concentration in the efferent samples was determined by the Viets et al. ( 36 ) method.
Proximal single nephron glomerular filtration rate (SNGFR), intratubular pressure during free flow conditions (FF) and under stopped flow conditions after blocking the tubular lumen with a long oil column (SFP, mmHg), glomerular capillary hydrostatic pressure (PGC), peritubular capillary pressure (PTCP), afferent oncotic pressure ( A), efferent oncotic pressure ( E), glomerular capillary hydrostatic pressure gradient ( P), single-nephron filtration fraction, single nephron glomerular blood (Qa) and plasma flow (GPF), afferent (AR) and efferent (ER) resistances, and ultrafiltration coefficient ( K f, nl·s -1 ·mmHg -1 ) were all calculated according to equations given elsewhere ( 2, 8 ). We estimated PGC by the stopped flow method according to the following equation: PGC = SFP + A ( 2, 8 ).
It should be mentioned that, in micropuncture studies, proximal collections were taken to measure SNGFR, since this is the only available method to evaluate whole glomerular hemodynamics. However, this procedure impairs the tubuloglomerular feedback mechanism (TGF) and results in an overestimation of SNGFR; if this is so, calculated values of glomerular plasma flow are higher and AR and RE lower than the real values. However, the method is generally accepted ( 3, 21a, 25a ). Since the increase of afferent resistance with ANG II is mainly mediated through TGF, and the chronic high intake of salt induces low TGF sensitivity ( 28 ), the absence of TGF-mediated effects, on one hand, is a limitation in this study, but on the other hand, it allowed the evaluation of glomerular hemodynamics without the interference of TGF ( 3, 29 ). The evaluation of TGF by microperfusion is beyond the scope of this study.
Renal ANG II concentration was determined in the renal cortex through microdialysis, according to the method described by Nishiyama et al., Fox et al., and Siragy et al. ( 10, 18, 30 ). The efficiency of the microdialysis probe was determined in vitro, testing ANG II concentrations from 2 to 10 nM, the regression line obtained with the data was Y = 20.17x + 13.20, r 2 = 0.94. At a perfusion rate of 1.5 µl/min, the relative equilibrium rate was 66 ± 2% which did not deteriorate with time (data not shown). The in vitro recovery may not be equivalent to the in vivo recovery, but it has been generally accepted as a reasonable estimate for the interstitial concentrations of endogenous compounds ( 18, 30 ). The high correlation coefficient ( r 2 = 0.94) of the linear regression obtained with four different concentrations of ANG II indicates the adequacy of the microdialysis probe. The 66 ± 2% recovery attained may introduce some error for the calculations of recovery in vivo but, at any rate, similar for all the groups studied ( 18, 30 ).
In vivo calibrations were performed from 0 to 180 min to test the stability of the ANG II concentrations according to the method described by Nishiyama et al. ( 18 ). Thirty minutes after implantation of the probe, the concentrations of ANG II fell from 151.6 ± 6.3 pg/ml to reach a steady state (106.5 ± 9.0 pg/ml), and remained stable up to 180 min (nearly 100 pg/ml). Previous studies have determined that ANG II derived from tubular fluid does not contaminate the dialysate ( 18 ).
For this procedure, additional groups of 10-12 rats each were studied at the end of the follow-up (7 wk); normal rats under standard chow were included to determine the values of ANG II in our laboratory. The surgical preparation of the animals was identical to that described previously for the micropuncture studies. A 10-mm polysulfone microdialysis probe (Fresenius Medical Care) with a 33,000-D transmembrane diffusion cut-off was implanted into the renal superficial cortex ( 17, 25 ); steel needles were inserted in both ends of the fiber. The inflow of the probe was connected with a polyethylene tubing (PE-10) to a microinfusion pump (Harvard apparatus). The probe was perfused with Ringer solution containing 1% bovine serum albumin (pH 7.4), at a rate of 1.5 µl/min. The dialysate was collected directly from the outflow steel tubing into a small tube containing a solution of inhibitors (30 µl of 500 mM EDTA, 15 µl of 1 mM enalaprilat and 30 µl of 125 M o -phenantrolene and 0.2 mM pepstatin A in 95% ethanol), to allow the dialysate effluent to be immediately mixed with the inhibitors. After 90 min of the microdialysis probe implantation, three 60-min sample collections were taken. Immediately thereafter, each sample was vortex-mixed, and 100 µl of the solution were transferred to a tube containing 1 ml chilled 100% methanol. The samples were stored at -70°C until analyzed.
Determination of ANG II was performed by enzyme-linked competitive immunoassay using a commercial Kit (SPI Bio, Bertin Group, France), following the procedure recommended by the manufacturer, and according to the method described by Nishiyama et al. and Fox et al. ( 10, 18 ). The dialysate samples in methanol were evaporated to dryness with nitrogen, reconstituted in assay buffer, and assayed after extraction in phenyl cartridges; an 8-points standard curve (0 to 125 pg/ml) was used. The intra- and interassay variations were 3.93 ± 0.56 and 5.10.70 ± 0.58%, respectively. Results are expressed in picogram per milliliter. No cross-reactivity was found with ANG I, ANG 1-7, ANG 3-8, or ANG III.
Histological Studies
In six rats from each group, coronal sections of the left kidney were obtained at the time of death. The fragments were fixed in 15% formalin and embedded in paraffin. Lymphocyte and macrophage infiltration was studied with the immunoperoxidase technique described before ( 23, 25 ). Briefly, tissues were successively washed and incubated with 20 ml of ExtraAvidin (2.5 mg/ml; Sigma) and 30 ml of 0.001% biotin in PBS. Afterwards, tissues were incubated for 2 h at 37°C with 50 ml of the corresponding primary monoclonal antibody, diluted 1:30 in Tris saline buffer (TSB), pH 7.8. After being washed in TSB for 15 min, tissues were incubated 1 h with 30 ml of rat antimouse IgG (ab') biotin-conjugated fragments with minimal cross-reactivity with human, horse and rat serum proteins (Accurate Chemical Westbury), and finally for 30 min with 60 ml of peroxidase-conjugated Extravidin. After the final wash, tissues were incubated for 15 min in diaminobenzidine and H 2 O 2 in TBS.
The primary antibodies used were anti-CD5 (mouse monoclonal anti-rat thymocytes and lymphocytes) and anti-ED-1 (mouse monoclonal anti-rat monocytes and macrophages), purchased from Biosource International (Camarillo, CA). Cellular infiltration was evaluated in tubulointerstitial areas and expressed as positive cells per glomerular cross-section or positive cells per square millimeter (mm 2 ).
Statistical Analysis
Groups were evaluated by one-way ANOVA followed by a Tukey-Kramer posttest. Statistical significance was defined at P < 0.05. Results in tables and figures are expressed as means ± SE.
RESULTS
Salt-Sensitive Hypertension Is Associated with Increased Intrarenal ANG II and Tubulointerstitial Inflammation
As previously reported ( 9, 15 ), rats transiently infused with ANG II developed hypertension when placed subsequently on a HSD. Control (sham-operated) rats and rats that received MMF during the ANG II infusion did not develop salt-sensitive hypertension ( Fig. 1 ). We previously reported that the administration of MMF blocks ANG II-induced tubulointerstitial inflammation and tubular injury and that this correlates with the prevention of the subsequent salt-sensitive hypertension ( 9, 21 ). In the current study similar findings were observed, as noted by a significant reduction in T cells (CD5+ cells) and macrophages (ED-1+ cells) in MMF-treated rats compared with ANG II alone (see Table 2 ).
Table 1. Renal hemodynamics in rats with salt-sensitive hypertension, rats with simultaneous MMF treatment, and normotensive sham rats on a high-salt diet after 7 wk of follow up, and effect of a bolus of candesarten on glomerular dynamics of similar groups at the end of the study
Table 2. Glomerular and tubulointerstitial inflammatory infiltration observed after 5 wk of a high-salt diet in rats with salt-sensitive hypertension, rats with concomitant MMF treatment, and normotensive sham rats, and inflammatory infiltration during acute treatment with candesarten
ANG II concentration in renal interstitial fluid after 5 wk of HSD are shown in Fig. 2. Intrarenal ANG II levels in MMF-treated rats were significantly reduced, although they remained higher than sham controls. The intrarenal ANG II levels observed correlated with the degree of salt-sensitive hypertension ( Fig. 3 ) as well as with the total number of CD5+ cells ( Fig. 4 ) when all groups were combined.
Fig. 2. ANG II interstitial concentrations in the cortex of rats that developed salt sensitivity (ANG II), as well as rats which received ANG II but salt-sensitive hypertension was prevented with MMF treatment (ANG II+MMF), and normotensive rats (Sham rats on high-salt diet and normal rats). Values are means ± SE.
Fig. 3. Correlation between ANG II concentration and development of salt sensitivity ( ). MMF prevented the development of salt sensitivity in rats which received concomitant ANG II infusion (ANG II+MMF; ).
Fig. 4. Correlations among ANG II interstitial concentrations in the cortex of rats with salt-sensitive hypertension (ANG II), rats in which salt sensitivity was prevented (ANG II+MMF) with ANG II-positive cells, lymphocytes (CD5-positive cells) and machophages (EDI-positive cells).
Effects of Candesartan on Renal Hemodynamics
To evaluate the functional role of ANG II in salt-sensitive hypertension phase, glomerular hemodynamics were assessed following the acute administration of candesartan. As previously reported, the administration of ANG II followed by exposure to a HSD resulted in profound cortical vasoconstriction with an increase in afferent and efferent arteriolar resistances, in association with a reduction in SNGFR and blood flow. These abnormal findings were significantly ameliorated in the rats treated with MMF ( Fig. 5 ). The acute administration of candesartan also completely blocked the arteriolar vasoconstriction (afferent and efferent) in the postangiotensin II salt-sensitive hypertension model ( Fig. 5 ). Similarly, the ultrafiltration coefficient ( K f ) was reduced in the ANG II group (ANG II group = 0.027 ± 0.001, vs. sham = 0.034 ± 0.001, P < 0.05) but was significantly improved in MMF-treated rats (0.033 ± 0.002) and the ANG II group that received an acute injection of candesartan (0.033 ± 0.001). Indeed, the increase in K f in these latter two groups was similar to that observed in sham-operated rats (0.034 ± 0.001).
Fig. 5. Effects of MMF and candesartan on glomerular hemodynamics of rats with salt-sensitive hypertension.
In contrast with rats treated with ANG II alone, in ANG II+MMF rats, candesartan administration was associated with no effects on vascular tone; in fact, afferent and efferent resistances were not different when compared with ANG II+MMF rats receiving vehicle. However, since MAP was decreased, glomerular plasma flow, PGC, SNGFR and K f were 26, 13, 22, and 20% lower, respectively, than in ANG II+MMF group that received vehicle. Similar effects were observed in sham rats, i.e., no effects on vascular resistance, associated with a slight fall in glomerular plasma flow, PGC, SNGFR and K f. ( Table 1 and Fig. 5 ). Candesartan treatment 1 h before death did not alter the cellular infiltration in any group ( Table 2 ).
DISCUSSION
We previously proposed that many forms of salt-sensitive hypertension are due to subtle renal microvascular and interstitial inflammatory injury ( 13, 15 ). We have been able to demonstrate the engagement of this pathway in both genetic and experimentally induced models of hypertension ( 15, 23, 24 - 26 ). A key finding in these models is the presence of renal ischemia and the infiltration of immune cells that express oxidants and ANG II ( 5, 6, 17, 19, 21a, 22 ). Indeed, the blockade of the inflammatory cell infiltration with agents such as MMF ( 9, 21 ), or the inhibition of local oxidants ( 35 ), can ameliorate the salt-sensitive hypertension. While we have also proposed a role for intrarenal ANG II in this process based on the presence of ANG II-positive cells, a formal testing of this hypothesis has not been performed.
In this paper, we examined the role of ANG II in a model of salt-sensitive hypertension in which renal injury is induced by a short-term exposure to ANG II. Specifically, the infusion of ANG II for 2 wk results in the acute development of hypertension with renal microvascular and tubulointerstitial injury. While stopping the ANG II results in rapid resolution of the hypertension, the subsequent exposure of the rat to a HSD results in the rapid redevelopment of hypertension. The mechanism was shown to be associated with cortical vasoconstriction ( 9 ) and to result in the infiltration of inflammatory cells ( 9, 24 ). The administration of MMF was able to prevent the salt-sensitive hypertension, cortical vasoconstriction, and renal interstitial inflammatory changes ( 9, 22 ).
We demonstrate that this model is associated with an increase in renal ANG II levels. Since these animals are received a HSD that is known to suppress the systemic renin angiotensin system, the findings represent an example of how intrarenal angiotensin may be dissociated from plasma angiotensin ( 18 ). This study also provides evidence that intrarenal ANG II may have a role in maintaining cortical vasoconstriction and sodium retention. Similar findings have been shown in experimental models of volume expansion ( 10 ) and in interstitial nephritis ( 34 ).
The possibility that the ANG II was mediating the renal vasoconstriction levels was supported by the presence of a strong correlation between the level of ANG II and blood pressure ( Fig. 3 ) and the interstitial inflammatory response. While potential stimulation of ANG II production in tubular cells by the inflammatory reaction has not been studied, one likely source of the intrarenal ANG II is the infiltrating immune cells, as previous studies have shown that many of these cells are ANG II-positive by immunostaining ( 21, 24 ). The observation that the renal ANG II levels were reduced by MMF (even though the rats received the same dose of ANG II) strongly argues against the possibility that intrarenal ANG II represents exogenous ANG II given nearly 6 wk earlier.
To investigate the functional role of ANG II in the salt-sensitive hypertension model, we examined the effects of acute administration of candesartan in this model. As shown in Table 1, AT 1 blockade decreased blood pressure in all groups, but the magnitude of the fall was larger in the ANG II group; candesartan was able to reverse the cortical vasoconstriction in ANG II-treated rats, suggesting that this pathophysiological state is mediated by the AT 1 receptor. Similar effects were seen with K f, in which there was a near complete correction to that observed in controls ( Fig. 5 ). Such a vasodilator response was not observed in the ANG II + MMF + Cand, or in Sham + Cand groups, in which the lack of a vasodilatory renal response can be attributed to an increased renal sympathetic nerve activity in the afferent and efferent arterioles, induced by the fall in blood pressure ( 4 ). In this regard, since SNGFR was taken from proximal collections, the TGF was prevented ( 3, 29 ). Nevertheless, the site(s) at which ANG II interacts with glomerular function is still unclear, since AT 1 receptors are located in macula densa cells, afferent and efferent arterioles and mesangial cells ( 16, 20 ). Thus this limitation is partially balanced by the fact that it allows evaluation of glomerular hemodynamics without interference of TGF ( 3 ). Thus we can only suggest a defective autoregulatory response to the fall in blood pressure in Sham and ANG II+MMF groups, since TGF is the main component of the autoregulatory response. In addition, it should be recognized that while our results suggest that renal ANG II is responsible for the glomerular hemodynamic findings in the salt-driven hypertension that follows exogenous angiotensin administration, the potential role of incomplete suppression of circulating angiotensin levels ( 12 ) was not addressed in this investigation.
In conclusion, we demonstrated that intrarenal ANG II levels are elevated in a model of salt-sensitive hypertension and that this correlates with the presence of intrarenal inflammation and cortical vasoconstriction. Importantly, blocking the AT 1 receptor can ameliorate the renal vasoconstriction. These studies are consistent with the hypothesis that the intrarenal angiotensin system in association with interstitial immune infiltration plays a pivotal role in this model of salt-sensitive hypertension.
GRANTS
This study was supported by Grants 30605M and 40934M (to M. Franco) from the National Council of Science and Technology (CONACYT), Mexico, FONACYT, Grant F-2005000283 and Asociación de Amigos del Riñón, Venezuela (B. Rodríguez-Iturbe), and National Institutes of Health (NIH) Grants DK-52121 and HL-68607. Dr. Johnson was supported by NIH Grant RO-1-HL-68607.
ACKNOWLEDGMENTS
We thank AstraZeneca for the kind donation of candesartan. We thank B. Chávez-Renteria for technical assistance.
J. Herrera-Acosta is deceased.
【参考文献】
Alvarez V, Quiroz Y, Nava M, Pons H, and Rodriguez-Iturbe B. Overload proteinuria is followed by salt-sensitive hypertension caused by renal infiltration of immune cells. Am J Physiol Renal Physiol 283: F1132-F1141, 2002.
Azar S, Tobian L, and Johnson MA. Glomerular, efferent arteriolar, peritubular capillary, and tubular pressures in hypertension. Am J Physiol 227: 1045-1050, 1974.
Blantz RC and Tucker BJ. Measurements of glomerular dynamics. In: Methods in Pharmacology, edited by Martinez Maldonado M. New York: Plenum, 1978, p. 141-163.
Cervenka L, Wang CT, and Navar LG. Effects of acute AT 1 receptor blockade by candesartan on arterial pressure and renal function in rats. Am J Physiol Renal Physiol 274: F940-F945, 1998.
Chabrashvili T, Kitiyakara C, Blau J, Karber A, Aslam S, Welch WJ, and Wilcox CS. Effects of Ang II type 1 and 2 receptors on oxidative stress, renal NADPH oxidase and SOD expression. Am J Physiol Regul Integr Comp Physiol 285: R117-R124, 2003.
Costerousse O, Allegrini J, Lopez M, and Alhenc-Gelas F. Angiotensin I-converting enzyme in human circulating mononuclear cells: genetic polymorphism of expression in T-lymphocytes. Biochem J 290: 33-40, 1993.
Davidson WD and Sackner MA. Simplification of the anthrone method for the determination of inulin in clearance studies. J Lab Clin Med 62: 351-356, 1963.
Deen WM, Troy JL, Robertson CR, and Brenner BM. Dynamics of glomerular ultrafiltration in the rat. IV. Determination of the ultrafiltration coefficient. J Clin Invest 52: 1500-1508, 1973.
Franco M, Tapia E, Santamaria J, Zafra I, Garcia-Torres R, Gordon KL, Pons H, Rodríguez-Iturbe B, Johnson RJ, and Herrera-Acosta J. Renal cortical vasoconstriction contributes to development of salt sensitive hypertension after angiotensin II exposure. J Am Soc Nephrol 12: 2263-2271, 2001.
Fox J, Guan S, Hymel AA, and Navar LG. Dietary Na and ACE inhibition effects on renal tissue angiotensin I and II and ACE activity in rats. Am J Physiol Renal Fluid Electrolyte Physiol 262: F902-F909, 1992.
Henry RJ, Segalove M, and Sobel C. Turbidimetric determination of proteins with sulfosalicylic and trichloroacetic acids. Proc Soc Exp Biol Med 92: 748-751, 1956.
Hu L, Sealey JE, Chen R, Zhou Y, Merali C, Shi Y, Laragh JH, and Catanzaro DF. Nitric oxide synthase inhibition accelerates the pressor response to low-dose angiotensin II, exacerbates target organ damage, and induces renin escape. Am J Hypertens 17: 395-403, 2004.
Johnson RJ, Herrera-Acosta J, Schreiner GF, and Rodríguez-Iturbe B. Subtle acquired renal injury as a mechanism of salt-sensitive hypertension. N Engl J Med 346: 913-923, 2002.
Johnson RJ, Gordon KL, Suga S, Duijvestijn AM, Griffin K, and Bidani A. Renal injuy and salt-sensitive hypertension after exposure to catecholamines. Hypertension 34: 151-159, 1999.
Lombardi D, Gordon KL, Polinsky P, Suga S, Schwartz SM, and Johnson RJ. Salt-sensitive hypertension develops after short-term exposure to angiotensin II. Hypertension 33: 1013-1019, 1999.
Navar LG. Integrating multiple paracrine regulators of renal microvascular dynamics. Am J Physiol Renal Physiol 274: F433-F444, 1998.
Nakagawa T, Kang DH, Oashi R, Suga S, Herrera-Acosta J, Rodríguez Iturbe B, and Johnson RJ. Tubuloinerstitial disease: role of ischemia and microvascular disease. Curr Opin Nephrol Hypertens 12: 233-242, 2003.
Nishiyama A, Seth DM, and Navar LG. Renal interstitial fluid concentrations of angiotensin I and II in anesthetized rats. Hypertension 39: 129-134, 2002.
Okamura A, Rakugi H, Ohishi M, Yanagitani Y, Takiuchi S, Moriguchi K, Fennessy PA, Higaki J, and Ogihara T. Upregulation of renin-angiotensin system during differentiation of monocytes to macrophages. J Hypertens 17: 537-545, 1999.
Paxton WG, Runge M, Horaist C, Cohen C, Alexander RW, and Bernstein KE. Immunohistochemial localization of rat angiotensin II AT 1 receptor. Am J Physiol Renal Fluid Electrolyte Physiol 264: F989-F995, 1993.
Quiroz Y, Pons H, Gordon KL, Rincon J, Chavez M, Parra G, Herrera-Acosta J, Gómez-Garre D, Largo R, Egido J, Johnson RJ, and Rodríguez-Iturbe B. Mycophenolate mofetil prevents salt-sensitive hypertension resulting from nitric oxide synthesis inhibition. Am J Physiol Renal Physiol 281: F38-F47, 2001.
Rajagopalan S, Kurz S, Munzel T, Tarpey M, Freeman BA, Griendling KK, and Harrison DG. Angiotensin II-mediated hypertension in the rat increases vascular superoxide production via membrane NADH/NADPH oxidase activation. Contribution to alterations of vasomotor tone. J Clin Invest 97: 1916-1923, 1996.
Reckelhoff JF and Romero JC. Role of oxidative stress in angiotensin-induced hypertension. Am J Physiol Regul Integr Comp Physiol 284: R893-R912, 2003.
Rincon J, Parra G, Quiroz Y, Benatuil L, and Rodriguez-Iturbe B. Cyclosporin A reduces expression of adhesion molecules in the kidney of rats with chronic serum sickness. Clin Exp Immunol 121: 391-398, 2000.
Rodríguez-Iturbe B, Pons H, Quiroz Y, Gordon K, Rincon J, Chávez M, Parra G, Herrera-Acosta J, Gomez-Garre D, Largo R, Egido J, and Johnson RJ. Mycophenolate mofetil prevents salt-sensitive hypertension resulting from angiotensin II exposure. Kidney Int 59: 2222-2232, 2001.
Rodríguez-Iturbe B, Quiroz Y, Nava M, Bonet L, Chávez M, Herrera-Acosta J, Johnson RJ, and Pons HA. Reduction of renal immune cell infiltration results in blood pressure control in genetically hypertensive rats. Am J Physiol Renal Physiol 282: F191-F201, 2002.
Romano G, Favret G, Edda F, and Ettore B. Measurements of nephron filtration from distal and proximal tubules. J Nephrol 11: 211-215, 1998.
Romero F, Rodríguez-Iturbe B, Parra G, González L, Herrera-Acosta J, and Tapia E. Mycophenolate mofetil prevents the progressive renal failure induced by 5/6 renal ablation in rats. Kidney Int 55: 945-955, 1999.
Ruan X, Wagner C, Chatziantoniou C, Kurtz A, and Arenshorts WJ. Regulation of angiotensin II receptor AT 1 subtypes in renal afferent arterioles during chronic changes in sodium diet. J Clin Invest 99: 1072-1081, 1997.
Schnermann J. Juxtaglomerular cell complex in the regulation of renal salt excretion. Am J Physiol Regul Integr Comp Physiol 274: R263-R279, 1998.
Schnermann J, Davis JM, Wunderlich P, Levine DZ, and Horster. Technical problems in the micropuncture determination of nephron filtration rate and their functional implications. Pflügers Arch 329: 307-320, 1971.
Siragy HM, Howell NL, Ragsdale NV, and Carey RM. Renal interstitial fluid angiotensin. Modulation by anesthesia, epinephrine, sodium depletion and renin inhibition. Hypertension 25: 1021-1024, 1995.
Siragy HM. Angiotensin II compartmentalization within the kidney: effects of salt diet and blood pressure alterations. Curr Opin Nephrol Hypertens 15: 50-53, 2006.
Vanegas V, Ferrebuz A, Quiroz Y, and Rodriguez-Iturbe B. Hypertension in page (cellophane-wrapped) kidney is due to interstitial nephritis. Kidney Int 68: 1161-1170, 2005.
Vaziri ND, Ni Z, Oveisi F, and Trnavsky-Hobbs DI. Effect of antioxidant therapy on blood pressure, and NO synthase expression in hypertensive rats. Hypertension 36: 957-964, 2000.
Viets JW, Deen WM, Troy JL, and Brenner BM. Determination of serum protein concentration in nanoliter blood samples using fluorescamine or o-phthalaldehyde. Anal Biochem 88: 513-521, 1978.
Vurek GG and Pegram SE. Fluorometric method for the determination of nanogram quantities of inulin. Anal Biochem 16: 409-419, 1966.
作者单位：1 Department of Nephrology, Instituto Nacional de Cardiología Ignacio Chávez, Mexico City; 2 Hospital Universitario, Instituto de Investigaciones Biomédicas, Universidad del Zulia, Maracaibo, Venezuela; 3 Division of Nephrology, Hypertension and Transplantation, University of Fl