Peroxisome proliferator-activated receptor-␥ (PPAR␥) is a nuclear hormone receptor that is critical for adipogenesis and insulin sensitivity. Ligands for PPAR␥ include some polyunsaturated fatty acids and prostanoids and the synthetic high affinity antidiabetic agents thiazolidinediones. However, the identity of a biologically relevant endogenous PPAR␥ ligand is unknown, and limited insight exists into the factors that may regulate production of endogenous PPAR␥ ligands during adipocyte development. To address this question, we created a line of 3T3-L1 preadipocytes that carry a ␤-galactosidase-based PPAR␥ ligand-sensing vector system. In this system, induction of adipogenesis resulted in elevated ␤-galactosidase activity that signifies activation of PPAR␥ via its ligand-binding domain (LBD) and suggests generation and/or accumulation of a ligand moiety. The putative endogenous ligand appeared early in adipogenesis in response to increases in cAMP, accumulated in the medium, and dissipated later in adipogenesis. Organically extracted and high pressure liquid chromatography-fractionated conditioned media from differentiating cells, but not from mature adipocytes, were enriched in this activity. One or more components within the organic extract activated PPAR␥ through interaction with its LBD, induced lipid accumulation in 3T3-L1 cells as efficiently as the differentiation mixture, and competed for binding of rosiglitazone to the LBD of PPAR␥. The active species appears to be different from other PPAR␥ ligands identified previously. Our findings suggest that a novel biologically relevant PPAR␥ ligand is transiently produced in 3T3-L1 cells during adipogenesis. Peroxisome proliferator-activated receptor-␥ (PPAR␥)1 is a nuclear hormone receptor that forms obligate heterodimers with the retinoid X receptor (RXR) (1) and binds to direct repeat 1-type motifs found in the promoter sites of target genes (2). PPAR␥ is a major modulator of several aspects of development and homeostasis. It is expressed in breast, colon, prostate, macrophages, and adipose tissue (3-6) and has been shown to play a critical role in glucose and lipid metabolism (7), macrophage function (8), and adipogenesis (9).As a member of the nuclear receptor superfamily, PPAR␥ is activated through ligand binding, which results in allosteric changes in receptor conformation, recruitment of coactivators, assembly of a transcriptional complex, and regulated transcription of target genes (10). Among known PPAR␥ agonists are the synthetic high affinity antidiabetic drugs thiazolidinediones (11) and a number of natural substances. These include the dehydration product of prostaglandin D 2 , 15-deoxy-⌬ 12,14 -prostaglandin J 2 (15-dPGJ 2 ) (12, 13); derivatives of linoleate, 9-and 13-hydroxyoctadeca-9Z,11E-dienoic acids, both found in oxidized low density lipoprotein (14); certain polyunsaturated fatty acids (15,16); and oxidized alkyl phospholipids such as lysophosphatidic acid (17). Most of these natural ligands bind PPAR␥ with relatively low affinity compared...

Within the kidney, the (pro)renin receptor (PRR) is predominantly expressed in the collecting duct (CD), particularly in intercalated cells, and it is regulated by the PGE receptor EP Notably, EP also controls urinary concentration through regulation of aquaporin 2 (AQP2). Here, we tested the hypothesis that sequential activation of EP and PRR determines AQP2 expression in the CD, thus mediating the antidiuretic action of vasopressin (AVP). Water deprivation (WD) elevated renal PRR expression and urinary soluble PRR excretion in rats. Intrarenal infusion of a PRR decoy peptide, PRO20, or an EP antagonist partially prevented the decrease in urine volume and the increase in urine osmolality and AQP2 expression induced by 48-hour WD. In primary cultures of rat inner medullary CD cells, AQP2 expression induced by AVP treatment for 24 hours depended on sequential activation of the EP receptor and PRR. Additionally, mice lacking PRR in the CD exhibited increased urine volume and decreased urine osmolality under basal conditions and impaired urine concentrating capability accompanied by severe volume loss and a dangerous level of plasma hyperosmolality after WD. Together, these results suggest a previously undescribed linear AVP/PGE/EP/PRR pathway in the CD for regulation of AQP2 expression and urine concentrating capability.

A high-fructose (HF) diet is shown to induce salt-sensitive hypertension but the underlying mechanism largely remains unknown. The major goal of the present study was to test the role of renal (pro)renin receptor (PRR) in this model. In Sprague-Dawley rats, HF intake increased renal expression of full-length PRR (fPRR), which were attenuated by allopurinol. HF intake also upregulated renal mRNA and protein expression of sodium/hydrogen exchanger 3 (NHE3) and Na/K/2Cl cotransporter (NKCC2) as well as in vivo NKCC2 activity, all of which were nearly completely blocked by a PRR decoy inhibitor PRO20 or allopurinol treatment. Parallel changes were observed for indices of intrarenal renin-angiotensin-system (RAS) including renal and urinary renin and angiotensin II levels. Radiotelemetry demonstrated that HF or a high-salt diet alone did not affect mean arterial pressure (MAP), but the combination of the two maneuvers induced a ~10 mm Hg increase of MAP, which was blunted by PRO20 or allopurinol treatment. In cultured human kidney 2 cells, both fructose and uric acid (UA) increased protein expression of soluble PRR (sPRR) in a time- and dose-dependent manner; fructose-induced PRR upregulation was inhibited by allopurinol. Taken together, our data suggest that fructose via UA stimulates renal expression of PRR/sPRR that stimulate NHE3 and NKCC2 expression and intrarenal RAS to induce salt-sensitive hypertension.

Proteinuria is a characteristic of chronic kidney disease and also a causative factor that promotes the disease progression, in part, via activation of the intrarenal renin-angiotensin system (RAS). (Pro)renin receptor (PRR), a newly discovered component of the RAS, binds renin and (pro)renin to promote angiotensin I generation. The present study was performed to test the role of soluble PRR (sPRR) in albumin overload-induced responses in cultured human renal proximal tubular cell line human kidney 2 (HK-2) cells. Bovine serum albmuin (BSA) treatment for 24 h at 20 mg/ml induced renin activity and inflammation, both of which were attenuated by a PRR decoy inhibitor PRO20. BSA treatment induced a more than fivefold increase in medium sPRR due to enhanced cleavage of PRR. Surprisingly, this cleavage event was unaffected by inhibition of furin or a disintegrin and metalloproteinase 19. Screening for a novel cleavage enzyme led to the identification of site-1 protease (S1P). Inhibition of S1P with PF-429242 or siRNA remarkably suppressed BSA-induced sPRR production, renin activity, and inflammatory response. Administration of a recombinant sPRR, termed sPRR-His, reversed the effects of S1P inhibition. In HK-2 cells overexpressing PRR, mutagenesis of the S1P, but not furin cleavage site, reduced sPRR levels. Together, these results suggest that PRR mediates albumin-induced cellular responses through S1P-derived sPRR.

Proteinuria is not only a common feature of chronic kidney diseases (CKD) but also an independent risk factor promoting CKD progression to end-stage renal failure. However, the underlying molecular mechanisms for protein overload-induced renal injury remain elusive. The present study examined the role of (pro)renin receptor (PRR) in pathogenesis of albumin overload (AO)-induced nephropathy and activation of intrarenal renin-angiotensin system (RAS) in rats. Wistar rats underwent unilateral nephrectomy and were treated for 7 weeks with vehicle, bovine serum albumin (5 g/kg/d via a single i.p. injection) alone or in conjunction with a PRR decoy inhibitor PRO20 (500 μg/kg/d via 3 s.c. injections). The AO rat model exhibited severe proteinuria, tubular necrosis, and interstitial fibrosis, oxidative stress, inflammation, accompanied by elevated urinary N-acetyl-beta-D-glucosaminidase activity and urinary β2-microglobulin secretion, all of which were significantly attenuated by PRO20. Urinary and renal levels of renin, angiotensinogen (AGT), and Ang II were elevated by AO and suppressed by PRO20, contrasting to largely unaltered plasma levels of the RAS parameters. The AO model also showed increased renal expression of full-length PRR and soluble PRR (sPRR) and urinary excretion of sPRR. Taken together, we conclude that PRR antagonism with PRO20 alleviates AO-induced nephropathy via inhibition of intrarenal RAS.