High perfusion pressure accelerates renal injury in salt-sensitive hypertension

Abstract

Renal injury in the Dahl salt-sensitive rat mimics human salt-sensitive forms of hypertension that are particularly prevalent in black individuals, but the mechanisms that lead to the development of this injury are incompletely understood. We studied the impact of renal perfusion pressure (RPP) on the development of renal injury in this model. During the development of salt-induced hypertension over 2 wk, the RPP to the left kidney was maintained at control levels (125 +/- 2 mmHg) by continuous servocontrol inflation of an aortic balloon implanted between the renal arteries; during the same period, the RPP to the right kidney rose to 164 +/- 8 mmHg. After 2 wk of a 4% salt diet, DNA microarray and real-time PCR identified genes related to fibrosis and epithelial-to-mesenchymal transition in the kidneys exposed to hypertension. The increased RPP to the right kidney accounted for differences in renal injury between the two kidneys, measured by percentage of injured cortical and juxtamedullary glomeruli, quantified proteinaceous casts, number of ED-1-positive cells per glomerular tuft area, and interstitial fibrosis. Interlobular arteriolar injury was not increased in the kidney exposed to elevated pressure but was reduced in the control kidney. We conclude that elevations of RPP contribute significantly to the fibrosis and epithelial-to-mesenchymal transition found in the early phases of hypertension in the salt-sensitive rat.

Figure 1.

(A) In SS rats (n = 6), the femoral arterial pressure (○) was monitored to reflect the perfusion pressure of the left kidney, which was continuously servocontrolled at the baseline level after initiation of the 4.0% NaCl diet. The carotid arterial pressure of the same rats (•), reflecting the perfusion pressure of the right kidney, was allowed to increase in response to the increased salt diet. A separate sham group of SS rats (n = 6; ▴) were maintained on 0.4% salt diet throughout the study. Means ± SEM of 24 h averages are summarized for each day of study. (B) A representative pressure tracing of 3-min averages is shown from a single rat.

Figure 2.

Eight of the 57 genes found by microarray to be differentially expressed between the outer medulla of the uncontrolled right kidney (high pressure) and controlled left kidney (normal pressure) were analyzed using real-time PCR. Ratios of higher pressure kidneys (right kidneys) over lower pressure kidneys (left kidneys) for mRNA expression level are shown for the microarray (▪) and real-time PCR methods (); n = 6 for microarray, n = 6 for real-time PCR; *P < 0.05. C4, complement C4; Col IVα1, collagen type IV α1; EGR1, early growth response 1; GPSP2, regulator of G-protein signaling protein 2; IGFBP5, insulin-like growth factor-binding protein 5; MM2, matrix metalloproteinase 2; Myl9, myosin regulatory light chain Myl9; TIMP-1, tissue inhibitor of metalloproteinase 1.

Figure 3.

Positive immunostaining of SMA (▪), a marker of epithelial transdifferentiation, and collagen IV () was greater in the outer medulla of the uncontrolled right kidney than the controlled left kidney (n = 6; *P < 0.05).

Figure 4.

Representative images demonstrating the renal injury in uncontrolled right kidney and the servocontrolled left kidney. (A) Casts demonstrated with Gomori's trichrome staining. (B) Interstitial fibrosis in the renal outer medulla determined by immunostaining with SMA antibody. (C) Tubular injury in the renal juxtamedullary region determined by immunostaining with OPN antibody.

Figure 5.

(A) The percentage of the outer medulla containing blocked tubules filled with protein (percentage of cast region) was determined from Gomori trichrome–stained kidney sections. The uncontrolled right kidney tended to be higher than the servocontrolled left kidney (P = 0.052). (B) Positive immunostaining of OPN was quantified in both cortex and outer medulla. OPN expression was significantly increased in the cortex of uncontrolled high-pressure right kidney compared with the servocontrolled left kidney (n = 6; *P < 0.05). No sham data were obtained for this measurement.

Figure 6.

A glomerulus visualized by Gomori trichrome stain was considered injured when its injury score was >2 on a scale of 0 to 4. The percentage of glomeruli with an injury score >2 was determined for the cortical (▪) and the juxtamedullary () glomeruli. The protected, servocontrolled left kidney had significantly fewer injured glomeruli in both regions than in the uncontrolled, high-pressure right kidney (n = 6; *P < 0.05). Sham left kidney was not significantly different from the servocontrolled left kidney (Table 2).

Figure 7.

Kidneys were immunostained with ED-1 antibody, and the number of ED-1–positive cells in the glomerular tuft was determined as an indication of macrophage infiltration. Consistent with the glomerular injury shown in Figure 6, there were fewer ED-1 cells observed in the servocontrolled left kidney compared with those of the uncontrolled right kidney (n = 6; *P < 0.05). ED-1–positive cells of sham left kidneys (Table 2) were not different from those of controlled left kidney.

Figure 8.

Interlobular arterial injury was determined by tracing the inner and outer circumferences with image analysis software and determining the ratio of the area of the inner lumen to the area of the outer circumference of the vessel. Median wall thickness ratio of the servocontrolled left kidney was significantly less than uncontrolled right kidney (n = 6; *P < 0.05). The ratio for the sham kidney (Table 2) was significantly higher than the uncontrolled left kidney.