Kihito Takahashi

Experimental and/or genetically controlled alterations of the renal microsomal cytochrome P450 epoxygenase induce hypertension in rats fed a high salt diet.

Excess dietary salt induces a cytochrome P450 arachidonic acid epoxygenase isoform in rat kidneys (Capdevila, J. H., S. Wei, J. Yang, A. Karara, H. R. Jacobson, J. R. Falck, F. P. Guengerich, and R. N. Dubois. 1992. J. Biol. Chem. 267:21720-21726). Treatment of rats on a high salt diet with the epoxygenase inhibitor, clotrimazole, produces significant increases in mean arterial blood pressure (122 +/- 2 and 145 +/- 4 mmHg for salt and salt- and clotrimazole-treated rats, respectively). The salt- and clotrimazole-dependent hypertension is accompanied by reductions in the urinary excretion of epoxygenase metabolites and by a selective inhibition of the renal microsomal epoxygenase reaction. The prohypertensive effects of clotrimazole are readily reversed when either the salt or clotrimazole treatment is discontinued. The indication that a salt-inducible renal epoxygenase protects against hypertension, are supported by studies with the Dahl rat model of genetic salt-sensitive hypertension. Dahl resistant animals responded to excess dietary salt by inducing the activity of their kidney microsomal epoxygenase(s) (0.102 +/- 0.01 and 0.240 +/- 0.04 nmol of products formed/min per mg of microsomal protein for control and salt-treated rats, respectively). Despite severe hypertension during excess dietary salt intake (200 +/- 20 mmHg), Dahl salt-sensitive rats demonstrated no increase in renal epoxygenase activity. These studies indicate that acquired or inherited abnormalities in renal epoxygenase activities and/or regulation can be related to salt-sensitive hypertension in rodents. Studies on the human renal epoxygenase and its relationship to salt hypertension may prove useful.

Predominant functional roles for thromboxane A2 and prostaglandin E2 during late nephrotoxic serum glomerulonephritis in the rat.

While much is known regarding acute nephrotoxic serum (NTS)-induced glomerular injury, the glomerular dynamics and pathophysiologic mediators of the more relevant chronic autologous phase remain poorly defined. Studies were performed in rats 14 d after injection of rabbit serum (n = 6), NTS in the absence (n = 6), or presence, of a cyclooxygenase inhibitor, ibuprofen (n = 6) or a thromboxane A2 (TxA2) receptor antagonist, L-670,596 (n = 5). A mesangial macrophage/monocyte infiltrate was noted with equal intensity in all NTS-treated rats. Glomerular generation rates of prostaglandin (PG) E2, PGF2a, and TxA2 in nephritic kidneys were dramatically increased as compared to controls. 2 wk after NTS, there was an increase in glomerular plasma flow rate (SNPF), attainment of filtration pressure disequilibrium, and augmentation of net transcapillary hydraulic pressure difference (delta P). Glomerular filtration rate (GFR), however, was reduced, due to a marked fall in the glomerular capillary ultrafiltration coefficient (Kf). Cyclooxygenase inhibition resulted in normalization of glomerular eicosanoid generation rates, amelioration of proteinuria, afferent vasoconstriction, and normalization of SNPF, delta P, Kf, and GFR. Selective antagonism of TxA2 also led to preservation of Kf, but was without effect on SNPF, thereby leading to elevated values for GFR. Thus, in contrast to the pathophysiologic role of arachidonate-lipoxygenase products in the early heterologous phase, PG-mediated vasodilatation and TxA2-induced reductions in Kf and GFR underlie glomerular functional changes during autologous mesangioproliferative glomerulonephritis.

Leukotriene D4 is a mediator of proteinuria and glomerular hemodynamic abnormalities in passive Heymann nephritis.

We assessed the role of leukotrienes (LTs) in Munich-Wistar rats with passive Heymann nephritis (PHN), an animal model of human membranous nephropathy. 10 d after injection of anti-Fx1A antibody, urinary protein excretion rate (Upr) in PHN was significantly higher than that of control. Micropuncture studies demonstrated reduced single nephron plasma flow and glomerular filtration rates, increased transcapillary hydraulic pressure difference, pre- and postglomerular resistances, and decreased ultrafiltration coefficient in PHN rats. Glomerular LTB4 generation from PHN rats was increased. Administration of the 5-LO activating protein inhibitor MK886 for 10 d markedly blunted proteinuria and normalized glomerular hemodynamic abnormalities in PHN rats. An LTD4 receptor antagonist SK&F 104353 led to an immediate reduction in Upr and to reversal of glomerular hemodynamic impairment. Ia(+) cells/glomerulus were increased in PHN rats. In x-irradiated PHN rats, which developed glomerular macrophage depletion, augmented glomerular LT synthesis was abolished. Thus, in the autologous phase of PHN, LTD4 mediates glomerular hemodynamic abnormalities and a hemodynamic component of the accompanying proteinuria. The synthesis of LTD4 likely occurs directly from macrophages or from macrophage-derived LTA4, through LTC4 synthase in glomerular cells.

Studies on the glomerular microcirculatory actions of manidipine and its modulation of the systemic and renal effects of endothelin

We examined the actions of intravenously administered manidipine on systemic and renal microcirculatory hemodynamics and its efficacy in antagonizing endothelin-1 (ET-1)-evoked responses. Manidipine was a potent vasodilator with preferential activity in the renal vasculature. Its administration in optimal doses resulted in decreases in systemic arterial pressures accompanied by increases in renal perfusion and filtration rates. Its primary sites of action in the kidney were at both pre- and postglomerular arteriolar sites. Manidipine was capable of near-total reversal of the sustained elevations in arterial pressure and the progressive reductions in renal blood flow and glomerular filtration rates induced by intravenously administered ET-1. In the presence of prolonged calcium channel blockade, subsequent administration of ET-1 led to paradoxic hypotensive responses, which could be profound and blocked by an inhibitor of nitric oxide synthesis. These unexpected vasorelaxant actions of ET-1 in the presence of manidipine were likely caused by the dual effects of antagonism of its own intrinsic vasoconstrictor action (through calcium channel blockade), as well as ET-1-evoked release of the endothelium-derived relaxing factor, nitric oxide.

The effect of thromboxane A2 receptor antagonism on amphotericin B-induced renal vasoconstriction in the rat

Amphotericin B (AmB) is the drug of choice for most systemic fungal infections, but doses are frequently reduced because of nephrotoxicity. We investigated the role of thromboxane as a mediator for this nephrotoxicity. Vehicle or amphotericin (0.60 mg/kg) was infused into the left renal artery in four groups of rats, and renal plasma flow (RPF) and glomerular filtration rate (GFR) were measured. Group 1 received vehicle for 90 min. Group 2 received vehicle followed by a 30 minute AmB infusion which caused a significant and reversible fall in the RPF and GFR. Group 3 received vehicle followed by AmB infusion, but were infused with a bolus of ibuprofen (20 mg/kg) 45 minutes before AmB. This group exhibited an insignificant attenuation in the fall in RPF and GFR. Group 4 received vehicle followed by AmB, but were infused with a bolus and continuous infusion of the thromboxane receptor antagonist SQ29,548. This group demonstrated an attenuation in the fall in RPF and a significant decrease in GFR compared to AmB control rats. In addition, the rat glomeruli were incubated with AmB (4 ug/ml). Supernatant levels of thromboxane B2 were significantly elevated in the presence of AmB vs buffer alone. We conclude that the reduction in RPF and GFR observed with AmB infusion in the rat is partially mediated by release of thromboxane.

Regulation of MAP-Kinase Activation by 8-Iso-Prostaglandin F2a in Cultured Rat Aortic Smooth Muscle Cells

8-iso-prostaglandin F2α (8-iso-PGF2α), an F2-isoprostane, is produced in vivo by a cyclooxygenase-independent, free radical-catalyzed mechanism involving peroxidation of arachidonic acid1. This isoprostane is a highly potent renal vasoconstrictor2 and stimulates proliferation of cultured rat aortic smooth muscle cells (AoSMC) through enhancement of phosphoinositide turnover3. AoSMC have distinct F2-isoprostane binding sites, although these putative receptors bear homology to the thromboxane A2 (TxA2) receptor4. In this study, we report the activation of mitogen activated protein kinases (MAP-kinases)5 by 8-iso-PGF2α in AoSMC and its regulating mechanism involving protein kinase C (PKC) and pertussis toxin (PT)-sensitive GTP binding proteins.

Endothelin and Endothelium-Derived Relaxing Factor in the Control of Glomerular Filtration and Renal Blood Flow

The functional importance of the vascular endothelium in the regulation of local vascular smooth muscle and organ blood flow is now well-established. Since the discovery of endothelial-derived relaxing factor (EDRF) (1), a variety of biochemical and mechanical stimuli have been shown to induce the release of endothlial-derived relaxing or contracting factors. It seems likely that EDRF is nitric oxide (2) and it now appears that endothelial-derived contracting factor (EDCF) is primarily the peptides in the endothelin family (3–5).

The Functional Significance of Lipoxygenase Products of Arachidonic Acid in the Glomerular Microcirculation

Intrarenal administration of Lipoxin A4 (LXA4) results in vasodilatory responses, but reduces the ultrafiltration coefficient, Kf. Lipoxin A4 competes for the 3H-Leukotriene D4 (LTD4) binding to rat mesangial cells and stimulates inositol trisphosphate (INsP3) formation from these cells. Reversal of stereochemical orientation of the polar substituents at C5 and C6 from S,R to R,S in both LTD4 and LXA4 abolishes both homologous and heterologous competition for 3H-LTD4 binding to mesangial cells and leads to total loss of biologic activity. Taken together, these data provided evidence for the involvement of a common recognition site mediating the actions of these two eicosanoids on mesangial cells. Cyclooxygenase inhibition reversed the vasodilatory action of LXA4, suggesting that LXA4-mediated increases in RPF were due to the secondary release of vasodilatory cyclooxygenase products. Intrarenal arterial administration of other lipoxins, LXB4 and 7-cis, 11-trans LXA4, induced vasoconstriction. Administration of the last named lipoxin which has a configuration at C5 and C6 identical with that of LTD4, resulted in systemic and renal effects similar to those of LTD4, suggesting that these actions resulted from activation of LTD4 receptor-coupled responses.