Decreased expression of Na,K-ATPase in the kidneys of rats with two-kidney, one-clip hypertension / 대한내과학회지

BACKGROUND/AIMS: This study investigated the role of Na,K-ATPase, the local renin-angiotensin-aldosterone system (RAAS), and atrial natriuretic peptide (ANP) system in the pathogenesis of renal tubular dysfunction and hypertension in rats with two-kidney, one-clip (2K1C) hypertension. METHODS: Adult male Sprague-Dawley rats were made 2K1C hypertensive for 4 weeks. The renal expression of Na,K-ATPase was determined by immunoblotting. The mRNA expression of renin, angiotensin-converting enzyme (ACE), aldosterone synthase (CYP11B2), mineralocorticoid receptor (MR), and the ANP system were determined in the kidney using real-time polymerase chain reaction. RESULTS: The blood pressure was increased in the 2K1C rats, compared with controls. The plasma renin activity and serum aldosterone concentrations were increased, as were the urine output and fractional excretion of sodium. The expression of Na,K-ATPase protein was decreased in the clipped kidney, as compared with the control kidney, while it remained unchanged in the contralateral kidney. The mRNA expression of renin, ACE1, CYP11B2, and MR was increased in the clipped kidney, but unchanged in the non-clipped kidney. The mRNA expression of ACE2 did not differ between the groups. The expression of ANP mRNA was increased in both clipped and non-clipped kidneys, as compared with control kidneys. CONCLUSIONS: The enhanced activity of the local RAAS may result in to ischemic tubular injury and the development of hypertension in 2K1C rats. The downregulation of Na,K-ATPase associated with tubular injury in the clipped kidney may account for the impaired tubular sodium reabsorption in 2K1C hypertension.

Increased Expression of Endothelin-1 and CYP11B2 in Gentamicin-Induced Nephropathy in Rat Kidney / 대한신장학회지

PURPOSE: An altered activity of vasoactive hormones as well as aldosterone synthase (CYP11B2) in the kidney may involve the pathogenesis of gentamicin-induced nephropathy. The present study was designed to investigate whether there are changes of local renin-angiotensin-aldosterone system (RAAS) and endothelin (ET) in the kidney of gentamicin-induced nephropathy in rats. METHODS: Male Sprague-Dawley rats (180-200 g) were intramuscularly injected with gentamicin (100 mg/kg per day) for 5 days. Vehicle was given for the control rats. The mRNA expression of local renin-angiotensin system, aldosterone synthase (CYP11B2), ET system and transforming grow factor-beta1 (TGF-beta1) was determined in the kidney by real-time polymerase chain reaction. The protein expression of TGF-beta in the kidney was determined by immunoblotting and immunohistochemistry. RESULTS: Following the gentamicin treatment, a renal failure was noted as evidenced by increased serum concentrations of creatinine along with a decrease of its clearance. TGF-beta1 expression was significantly increased in the kidney in gentamicin treated rats compared with that in controls. The abundance of ET-1 mRNA was significantly increased. The endothelin type A receptor expression was decreased while endothelin type B receptor was not changed. The expression of angiotensin converting enzyme 1 (ACE1) and ACE2 was decreased, whereas renin expression was not changed. The CYP11B2 expression was significantly increased in gentamicin treated rats, while mineralocorticoid receptor expression was not changed. CONCLUSION: The expression of ET-1 and CYP11B2 was up-regulated which may play a role in the pathogenesis of gentamicin-induced nephropathy.

Blockade of Nitric Oxide Synthesis Further Diminishes Aquaporin Water Channels in Rat Kidney Subjected to Ischemia/reperfusion Injury / 대한신장학회잡지

BACKGOUND: The present study examined whether a blockade of nitric oxide (NO) synthesis affects the regulation of aquaporin (AQP) water channels in rats subjected to renal ischemia/reperfusion (I/R). METHODS: Renal I/R was experimentally induced by clamping the left renal artery for 60 minutes in rats. The rats were kept for 7 days thereafter, during which they were supplied with tap water containing NG-nitro-L-arginine methyl ester (L-NAME, 100 mg/L). The expression of AQP1-3 was determined in the kidney by Western blot analysis. RESULTS: In renal I/R injury, the expression of AQP2 was significantly decreased. The treatment with L-NAME further diminished the expression of AQP2. Although the expression of either AQP1 or AQP3 was not significantly altered in the kidney subjected to I/R, it was also significantly decreased by the treatment with L-NAME. CONCLUSION: It is suggested that endogenous NO system should play a role in the regulation of AQP water channels in rat kidney subjected to I/R injury.

Increased Expression of Sodium Transporters in Rats Chronically Inhibited of Nitric Oxide Synthesis

The present study was done to determine whether endogenous nitric oxide (NO) plays a role in the regulation of sodium transporters in the kidney. Male Sprague-Dawley rats were treated with NG-nitro-L-arginine methyl ester (L-NAME, 100 mg/L drinking water) for 4 weeks. Control rats were supplied with tap water without drugs. Expression of Na, K-ATPase, type 3 Na/H exchanger (NHE3), Na/K/2Cl cotransporter (BSC1), and thiazide-sensitive Na/Cl cotransporter (TSC) proteins was determined in the kidney by Western blot analysis. Catalytic activity of Na,K-ATPase was also determined. The treatment with L-NAME significantly and steadily increased the systemic blood pressure. Total and fractional excretion of urinary sodium decreased significantly, while creatinine clearance remained unaltered. Neither plasma renin activity nor aldosterone concentration was significantly altered. The alpha1 subunit expression and the catalytic activity of Na, K-ATPase were increased in the kidney. The expression of NHE3, BSC1 and TSC was also increased significantly. These results suggest that endogenously-derived NO exerts a tonic inhibitory effect on the expression of sodium transporters, including Na, K-ATPase, NHE3, BSC1, and TSC, in the kidney.

Effects of Indomethacin on Aquaporin Water Channels in Rat Kidney / 대한신장학회잡지

BACKGROUND: The present study was aimed to determine whether non-steroidal anti-inflammatory drugs (NSAID) alter the regulation of aquaporin (AQP) water channels in the kidney. METHODS: Male Sprague-Dawley rats were injected with indomethacin (5 mg/kg, twice a day, i.p.) for two days. Control group was injected with vehicle. The expression of AQP1-4 proteins was determined in the kidney by Western blot analysis. RESULTS: Following the treatment with indomethacin (5 mg/kg), the expression of AQP1 was not significantly affected in the cortex and outer medulla, but was decreased and inner medulla. The expression of AQP2 was significantly decreased in the cortex, outer medulla and inner medulla. The expression of AQP3 was not significantly affected in the cortex, but was decreased in the outer medulla and inner medulla. The expression of AQP4 was detected only in the inner medulla, which was also significantly decreased. A lower dose of indomethacin (2.5 mg/kg) or diclofenac (100 mg/kg, single dose) similarly decreased the expression of AQP2. Despite the changes in AQP proteins, there were no significant changes in the expression of heat shock proteins-25 and -70. CONCLUSION: These results suggest that NSAID specifically decrease the expression of AQP water channels in the kidney.

Upregulation of Renal Renin-Angiotensin System in Rats with Adriamycin-Induced Nephrosis

The present study was aimed to investigate whether the adriamycin-induced nephrosis is associated with an altered regulation of local renin-angiotensin system (RAS) in the kidney. Rats were subjected to a single injection of adriamycin (2 mg/kg body weight, IV) and kept for 6 weeks to allow the development of nephrosis. They were then divided into two groups, and supplied with and without cilazapril, an angiotensin converting enzyme (ACE) inhibitor, in drinking water (100 mg/l) for additional 6 weeks. Another group without adriamycin-treatment served as control. The mRNA expression of renin, ACE, type 1 and type 2 angiotensin II receptors (AT1R, AT2R), and transforming growth factor (TGF) -beta1 was determined in the cortex of the kidney by reverse transcription-polymerase chain reaction. Adriamycin treatment resulted in heavy proteinuria. Accordingly, the mRNA expression of renin, ACE, and AT1R was increased in the renal cortex, while that of AT2R was decreased. Co-treatment with cilazapril attenuated the degree of proteinuria. While not affecting the altered expression of renin, cilazapril decreased the expression of ACE to the control level. Cilazapril further increased the expression of AT1R, while it restored the decreased expression of AT2R. The expression of TGF-beta1 was increased by the treatment with adriamycin, which was abolished by cilazapril. An altered expression of local RAS components may be causally related with the development of adriamycin-induced nephrosis, in which AT1R is for and AT2R is against the development of nephrosis.

Differential Regulation of Atrial Natriuretic Peptide in Clipped and Contralateral Kidneys in Two-Kidney, One Clip Hypertension / 대한신장학회잡지

BACKGROUND: The present study was aimed to determine the pathophysiological implications of local atrial natriuretic peptide (ANP) system in the kidney in two- kidney, one clip (2K1C) hypertension. METHODS: Rats were made 2K1C hypertensive, and their mRNA expressions of ANP and natriuretic peptide receptors (NPR) were determined in the clipped and contralateral kidneys by reverse transcription-polymerase chain reaction. RESULTS: The expression of ANP was decreased in the clipped kidney and increased in the contralateral kidney. Similarly, the expression of both NPR-A and NPR-C was decreased in the clipped kidney and increased in the contralateral kidney. CONCLUSION: These findings indicate a differentially-altered ANP system in the clipped and the contralateral kidneys in 2K1C hypertension.

Decreased Expression of Aquaporin-2 Water Channels in the Kidney in Rats Treated with Chlorpropamide / 대한신장학회잡지

BACKGROUND: Whether blood glucose levels may change the regulation of aquaporin(AQP) water channels in the kidney was investigated. METHODS: Male Sprague-Dawley rats were treated with chlorpropamide(40 mg/100 g body weight per day, per oral, for 7 days), and their expression of AQP1-3 and type VI adenylyl cyclase proteins was determined in the kidney. RESULTS: Following the treatment with chlorpropamide, the blood glucose level was significantly decreased compared with that in the control(64+/-8 vs 106+/-7 mg/dL, n=6 each, p < 0.01). Accordingly, the expression of AQP2 proteins was decreased in the cortex, outer medulla, and inner medulla. The AQP2 targeting was not significantly altered, as evidenced by parallel decreases of its expression in the membrane and the cytoplasmic fractions. No significant changes were observed in the expression of either AQP1 or of AQP3. The protein expression of type VI adenylyl cyclase was not significantly altered. CONCLUSION: These results suggest that hypoglycemia attenuates the expression of AQP2 water channels in the kidney.

Influence of Nitric Oxide Synthesis Inhibition on Endothelin-1 and Its Receptor Expression in Rat Kidney / 대한신장학회잡지

BACKGROUND: The present study was aimed to evaluate the influence of nitric oxide(NO) synthesis inhibition on endothelin(ET) expression in rat kidney. METHODS: Male Sprague-Dawley rats were treated with N(G)-nitro-L-arginine methyl ester(L-NAME, 100 mg/L drinking water) for 4 weeks to inhibit the endogenous synthesis of NO. The tissue expression of ET-1, ET(A) receptor, and ET(B) receptor mRNA in the kidney was determined by reverse transcription-polymerase chain reaction. RESULTS: Tissue levels of NO metabolites were significantly decreased in the plasma and the kidney, along with the increased blood pressure. The expression of ET-1 mRNA was increased in the cortex, but not in the medulla. The expression of ET(A) and ET(B) receptor mRNA was not significantly altered either in the cortex or in the medulla. The plasma level of ET-1 peptide was significantly increased, along with the increased blood pressure, when L-NAME(200 microgram/kg per min, iv) was administered in an acute preparation of animals. Accordingly, the expression of ET-1 mRNA was increased in the cortex, whereas that of ET(A) and ET(B) receptor mRNA was not altered. CONCLUSION: These results suggest that enhanced activity of ET system induced by NO synthesis inhibition may be associated with hypertension although direct association between two factors is not confirmed.

Coordinate expression of renin and cyclooxygenase-2 in rats with two-kidney, one clip and deoxycorticosterone acetate-salt hypertension

The present study was aimed to examine whether the expression of renin is associated with that of cyclooxygenase-2 (COX-2) in the kidney. Male Sprague-Dawley rats were made two-kidney, one clip (2K1C) or deoxycorticosterone acetate (DOCA)-salt hypertensive, to stimulate or to inhibit the endogenous renin-angiotensin system, respectively. The expression of renin and COX-2 mRNA was determined in the cortex of the kidney by reverse transcription-polymerase chain reaction. 2K1C hypertensive rats showed an increased expression of renin as well as of COX-2 in the clipped kidney. The expression of renin was decreased in parallel with that of COX-2 in the contralateral non-clipped kidney. Removal of the renal arterial clip reversed the expression of both genes, along with the blood pressure, to the control level. On the other hand, DOCA-salt hypertension was associated with parallel decreases of renin and COX-2 expression. These results indicate that renin and COX-2 genes are coordinately expressed in the kidney.

Decreased expression of aquaporin-2 water channels in the kidney in rats treated with reserpine

Whether there exists a Sympathetic neural mechanism regulating the expression of aquaporin (AQP) water channels in the kidney was investigated. Male Sprague-Dawley rats were treated with reserpine (1 mg/kg, IP), and the expression of AQP1-4 proteins was determined in the kidney one day thereafter. Following the treatment with reserpine, the systolic blood pressure measured in a conscious state was significantly decreased in the experimental group compared with that in the control (83+/-8 vs 124+/-6 mmHg n=6 each, P<0.05). The expression of AQP2 proteins was decreased in the cortex, outer medulla, and inner medulla. The decrease of AQP2 proteins was in parallel in the membrane and the cytoplasmic fractions, suggesting a preserved AQP2 targeting. No significant changes were observed in the expression of AQP1, AQP3, or AQP4. Neither basal nor AVP-stimulated formation of cAMP was significantly altered. These results suggest that the sympathetic nervous system has a tonic stimulatory effect specifically on the expression of AQP2 water channels in the kidney.

Increased expression of nitric oxide synthases in left and right ventricular hypertrophy

The present study was aimed to explore pathophysiological implications of nitric oxide in the development of left and right ventricular hypertrophy. To induce selective left and right ventricular hypertrophy, rats were made two-kidney, one clip (2K1C) hypertensive and treated with monocrotaline (MCT), respectively. Six weeks later, the hearts were taken and their ventricular tissue mRNA and protein expression of endothelial constitutive isoform of nitric oxide synthase (NOS) were determined by reverse transcription-polymerase chain reaction and Western blot analysis, respectively. In 2K1C hypertensive rats, the expression of NOS mRNA was increased in parallel with its proteins in the left ventricle, but not in the right ventricle. In MCT-treated rats, the expression of NOS mRNA and proteins were proportionally increased in the right ventricle, but not in the left ventricle. These results suggest that the expression of NOS is specifically increased in association with the ventricular hypertrophy, which may be a mechanism counteracting the hypertrophy.

Altered Renal Expression of Aquaporin-2 Water Channels in Rats with Experimental Two-Kidney, One Clip Hypertension

The present study was aimed at examining the regulation of aquaporin (AQP)-2 water channels in the kidney in two-kidney, one clip (2K1C) hypertension. Rats were made 2K1C hypertensive for 6 weeks, and their expression of AQP2 channel proteins was determined in the clipped and contralateral kidneys. To examine the upstream affecting AQP2 channels, adenylyl cyclase activity was also determined. Along with the hypertension, in the clipped kidney, the abundance of AQP2 proteins was significantly decreased in the cortex, outer and inner medulla, while their trafficking remained unaltered. Concomitantly with the reversal of the blood pressure at 24 hours following removal of the clip, the AQP2 abundance also returned to the control level. The arginine vasopressin-evoked generation of cAMP was decreased in the clipped kidney, which again was reversed to the control level following removal of the clip. In contrast, the expression of AQP2 channels as well as the activity of adenylyl cyclase remained unaltered in the contralateral kidney. These results indicate an altered regulation of AQP2 water channels in the clipped kidney in 2K1C hypertension.