20-HETE in acute kidney injury.

Ischemia/reperfusion injury (IRI) is a common cause of acute kidney injury (AKI) that is associated with a patient mortality of up to 50%. Currently there are not effective pharmacologic therapies for AKI. This Commentary highlights recent evidence indicating that 20-HETE plays an important role in IRI and that drugs that target this pathway have potential as therapeutic agents for AKI.

Activator of G protein signaling 3 promotes epithelial cell proliferation in PKD.

The activation of heterotrimeric G protein signaling is a key feature in the pathophysiology of polycystic kidney diseases (PKD). In this study, we report abnormal overexpression of activator of G protein signaling 3 (AGS3), a receptor-independent regulator of heterotrimeric G proteins, in rodents and humans with both autosomal recessive and autosomal dominant PKD. Increased AGS3 expression correlated with kidney size, which is an index of severity of cystic kidney disease. AGS3 expression localized exclusively to distal tubular segments in both normal and cystic kidneys. Short hairpin RNA-induced knockdown of endogenous AGS3 protein significantly reduced proliferation of cystic renal epithelial cells by 26 +/- 2% (P < 0.001) compared with vehicle-treated and control short hairpin RNA-expressing epithelial cells. In summary, this study suggests a relationship between aberrantly increased AGS3 expression in renal tubular epithelia affected by PKD and epithelial cell proliferation. AGS3 may play a receptor-independent role to regulate Galpha subunit function and control epithelial cell function in PKD.

20-HETE activates the Raf/MEK/ERK pathway in renal epithelial cells through an EGFR- and c-Src-dependent mechanism.

20-Hydroxyeicosatetraenoic acid (20-HETE) has been reported to promote mitogenicity in a variety of cell types, including renal epithelial cells. However, the signal transduction pathways activated by 20-HETE have not been fully defined. The present study evaluated the effects of 20-HETE and its more stable agonist analogs 20-hydroxyeicosa-5(Z),14(Z)-dienoic acid (5,14-20-HEDE) and N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-20-HEDGE) on the Raf/MEK/ERK and phosphatidylinositol 3-kinase (PI3K)-Akt pathway in LLC-PK(1) renal epithelial cells. 20-HETE (20 microM) increased phosphorylation of Raf-1 (2.5 +/- 0.2-fold), MEK1/2 (6.3 +/- 1.6-fold), and ERK1/2 (5.8 +/- 0.3-fold) compared with vehicle-treated cells. Similarly, the 20-HETE analogs also strongly activated ERK1/2 in a Raf-1- and MEK1/2-dependent manner. Moreover, 5,14-20-HEDE increased Akt phosphorylation by 2.2 +/- 0.3-fold. 20-HETE and 5,14-20-HEDE also promoted activation (Y1086) of epidermal growth factor receptor (EGFR; Y1086) by 1.9 +/- 0.2- and 2.5 +/- 0.2-fold, respectively. These effects were completely blocked by the EGFR inhibitor EKB-569 (0.1 microM). Moreover, EKB-569 (0.1 microM), as well as a c-Src inhibitor, SKI-606 (0.05 microM), completely abolished the 20-HETE-mediated activation of the Raf/MEK/ERK and PI3K-Akt pathways. Blockade of PKC with bisindolylmaleimide I had no effect on 20-HETE-induced ERK1/2 activation. This study demonstrated that 20-HETE activated the Raf/MEK/ERK and Akt pathways in renal epithelial cells secondary to the activation of c-Src and EGFR.

Chronic blockade of 20-HETE synthesis reduces polycystic kidney disease in an orthologous rat model of ARPKD.

20-Hydroxyeicosatetraenoic acid (20-HETE) has been implicated as a potential mediator in epithelial cell proliferation and cyst formation in polycystic kidney disease (PKD). In the present study, we studied the effects of chronic blockade of 20-HETE synthesis in an orthologous rodent model of autosomal recessive polycystic kidney disease (ARPKD), the PCK rat. RT-PCR analysis indicated that the expression of CYP4A1, CYP4A2, CYP4A3, and CYP4A8 mRNA was increased two- to fourfold in cystic PCK compared with noncystic Sprague-Dawley rat kidneys. Daily administration of a 20-HETE synthesis inhibitor, HET-0016 (10 mg x kg(-1) x day(-1) ip) for 4-7 wk significantly reduced kidney size by 24% from 4.95 +/- 0.19 g in vehicle-treated PCK rats to 3.76 +/- 0.15 g (n = 4). Collecting tubule morphometric cystic indices were reduced in HET-0016-treated PCK rats (2.1 +/- 0.2; n = 4) compared with vehicle-treated PCK rats (4.4 +/- 0.1; n = 4). The cellular mechanism by which 20-HETE may play a role in cyst formation has not been well characterized, but there was a significantly lower (P < 0.05) level of intracellular cAMP and decreased phosphorylation (activation) of ERK1/2 protein in PCK rat kidneys (n = 3) treated with HET-0016 . These studies indicate a potential role of 20-HETE in cyst formation in the orthologous rodent PCK model of ARPKD.

Gene therapy to the kidney using viral vectors.

Many pediatric diseases have reached a therapeutic plateau using currently available surgical and pharmacological approaches. Gene therapy has emerged as an exciting new technology to manipulate cells in the mammalian system, and in some cases, this method has achieved amazing therapeutic benefits. Compared to other organs, such as the brain, liver and lung, methods to genetically modify renal cells have received relatively little attention. The current review will discuss the challenges and important developments regarding gene therapy to the kidney, and relate the recent successes and failures to the future potential of gene therapy as a treatment modality in the context of pediatric disease.

Correlation between decreased sensitivity of the Daudi lymphoma cells to VP-16-induced apoptosis and deficiency in DNAS1L3 expression.

A functional relationship between the apoptotic endonuclease DNAS1L3 and the chemotherapeutic drug VP-16 was established. The lymphoma cell line, Daudi, exhibited a significant resistance to VP-16 treatment in comparison to the lymphoma/leukemia cell line, U-937. While U-937 cells degraded their DNA into internucleosomal fragments, Daudi cells failed to undergo such fragmentation in response to the drug. Activation of both caspase-3 and DNA fragmentation factor was not sufficient to trigger internucleosomal DNA fragmentation in Daudi cells. No correlation was found between expression levels of topoisomerase-II, Pgp, Bcl-2, Bax, or Bad and decreased sensitivity of Daudi cells to VP-16. Daudi cells failed to express DNAS1L3 and ectopic expression of this protein significantly sensitized the cells to VP-16. An enhancement of caspase-3 activity and collapse of mitochondrial membrane potential underlie DNAS1L3-mediated sensitization of Daudi cells to VP-16, which may be a direct result of DNAS1L3-mediated increase in PARP-1-activating DNA breaks after VP-16 treatment. Our results suggest that DNAS1L3 plays an active role in lymphoma cell sensitization to VP-16 and that its deficiency may constitute a novel mechanism of drug resistance in these cells.