Upregulation of Cyclooxygenase-2 Expression in Porcine Macula Densa With Chronic Nitric Oxide Synthase Inhibition

The macula densa (MD) is composed of specialized morphologically distinct tubular epithelial cells located on the glomerular side of the distal tubule. By sensing electrolyte concentration in the distal tubule, the MD provides feedback to control preglomerular resistance and renin secretion, thereby participating in control of electrolyte and water homeostasis. Activity of the Na-K-2Cl cotransporter in the MD is thought to initiate a series of reactions that control glomerular filtration rate and renin secretion. Cyclooxygenase-2 (COX-2) and neuronal nitric oxide synthase (nNOS) are colocalized in the MD. The products of these enzymes—prostaglandins (PGs) and nitric oxide (NO), respectively—have been reported to be modulators of this series of reactions, counteracting afferent arteriolar vasoconstriction mediated by tubuloglomerular feedback (TGF), as well as TGF-induced inhibition of renin secretion.7,12,21,23,28,31 In addition, both enzymes are upregulated with interventions, such as low salt diets, administration of loop diuretics, and renovascular hypertension. That these enzymes colocalize in the MD suggests that nNOS might be involved in mediating expression of COX-2 and vice versa.5 Interestingly, hemodynamic responses to COX-2 inhibition, as seen after nonselective NOS blockade, could not be duplicated by selective nNOS blockade, thus suggesting that other NOS isoforms could be involved.3 Castrop et al proposed that MD-dependent renin secretion does not require intact nNOS or endothelial NOS (eNOS) activity but, rather, NO generated by global NOS activity.4 In addition to eNOS and nNOS, the other isoform of NOS—inducible NOS (iNOS)—may participate in this process.30 The isoform is constitutively expressed in rodent kidney; its renal expression is less certain in large animal models.20 Initial data from in vitro and in vivo studies suggested that NO enhances production of COX-2-derived products and so acts as a modulator of the COX-2 pathway.5,9 These data led to the hypothesis that NO bioavailability is obligatory for COX-2 expression and/or activity (Fig. 1a). Studies to the contrary have been reported. There was no reduction in COX-2 immunoreactivity in the MD cells of nNOS knockout mice.27 Indeed, studies have demonstrated that COX-2-derived PGs play an important role in buffering renal vasoconstriction secondary to a reduction in NO bioavailability.3,24 Figure 1 Schematic illustration of cyclooxygenase-2 (COX-2) regulation: 1a and 1b, the obligatory nitric oxide (NO) hypothesis; 1c and 1d, the reciprocal regulation hypothesis. Abbreviations: nNOS, neural isoform of NO synthase; PG, prostaglandin; AA, afferent ... Constitutive expression of low levels of COX-2 in the MD of normal kidney has been documented in several laboratory animal species.14 Initial studies in humans failed to detect COX-2 in the MD;15 more recent studies have documented its expression in MD of normal human kidneys.1 Markedly increased COX-2 protein was detected in the MD of elderly humans, children with Bartter-like syndrome, and hypertensive patients.13,16,19 Decreased endogenous NO production is associated with aging and is known to contribute to hypertension.10,11 Thus, an alternative to the obligatory NO hypothesis noted above has emerged—namely, that COX-2 expression and NOS expression are reciprocally regulated in MD (Fig. 1c). This hypothesis suggests that in populations characterized by reduced NO bioavailability, COX-2 is upregulated in response to, and to compensate for, a relative lack of NO (Fig. 1d). We hypothesized that, with chronic inhibition of NOS, COX-2 in the MD would be upregulated to compensate for diminished production of endogenous NO. We used a large animal model—the Yucatan miniature porcine model—to test this hypothesis. We use this model in our laboratory because the anatomy and physiology of its cardiovascular system exhibit many similarities to those of the human. In several of our studies, we have noted that certain aspects of renal function (eg, renal blood flow) faithfully model human physiology.17