curcumin and sphingosine-kinase

Our previous studies have confirmed that the sphingosine kinase 1 (SphK1)-sphingosine 1-phosphate (S1P) signaling pathway in the kidney under diabetic conditions is closely correlated with the pathogenesis of diabetic nephropathy (DN). The activation of SphK1-S1P pathway by high glucose (HG) can increase the expression of fibronectin (FN), an important fibrotic component, in glomerular mesangial cells (GMCs) by promoting the DNA-binding activity of transcription factor AP-1. However, the mechanism responsible for the sustained activation of SphK1-S1P pathway remains unclear. Given the binding motifs for AP-1 within the first intron of the SphK1 gene, we speculated that the activated AP-1 in the kidney under HG condition possibly regulates SphK1 expression in a positive feedback manner, thereby promoting the sustained activation of SphK1-S1P pathway and mediating the pathological progression of DN. Here, we observed the effect of AP-1 on SphK1 expression in GMCs and explored the molecular mechanism involved in the sustained activation of SphK1-S1P pathway. We found two consensus binding motifs for AP-1 in the promoter sequences and non-coding region downstream of the transcriptional initiation of the rat SphK1 gene by chromatin immunoprecipitation assay. The treatment of GMCs with both HG and S1P significantly increased the protein expression of c-Jun and c-Fos, and obviously enhanced the phosphorylation of c-Jun at Ser63 and Ser73, and c-Fos at Ser32. Knockdown of c-Jun and c-Fos with siRNAs substantially inhibited the expression of SphK1 and FN, whereas overexpression of c-Jun and c-Fos significantly increased the expression of SphK1 and FN. Curcumin treatment greatly decreased the levels of c-Jun, c-Fos, SphK1, and FN in the kidney tissues of diabetic rats. SiRNAs targeting SphK1 and S1P2 receptor respectively inhibited the phosphorylation of c-Jun (ser63 and ser73) and c-Fos (ser32), as well as FN expression under both normal and HG conditions. Our data demonstrated that the activated SphK1-S1P signaling pathway in GMCs under diabetic conditions is closely associated with AP-1 to form a positive feedback loop. This positive feedback loop functions as an important molecular basis for the sustained activation of SphK1-S1P pathway and increased FN expression that lead to the initiation and progression of DN.

Topics: Animals; Binding Sites; Cells, Cultured; Curcumin; Diabetes Mellitus; Diabetic Nephropathies; DNA-Binding Proteins; Enzyme Inhibitors; Fibronectins; Glucose; JNK Mitogen-Activated Protein Kinases; Lysophospholipids; Male; Mesangial Cells; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Promoter Regions, Genetic; Protein Binding; Proto-Oncogene Proteins c-fos; Rats; Receptors, Lysosphingolipid; RNA Interference; RNA, Small Interfering; Signal Transduction; Sphingosine; Sweetening Agents; Transcription Factor AP-1

Curcumin, a major polyphenol from the golden spice Curcuma longa commonly known as turmeric, has been recently discovered to have renoprotective effects on diabetic nephropathy (DN). However, the mechanisms underlying these effects remain unclear. We previously demonstrated that the sphingosine kinase 1-sphingosine 1-phosphate (SphK1-S1P) signaling pathway plays a pivotal role in the pathogenesis of DN. This study aims to investigate whether the renoprotective effects of curcumin on DN are associated with its inhibitory effects on the SphK1-S1P signaling pathway. Our results demonstrated that the expression and activity of SphK1 and the production of S1P were significantly down-regulated by curcumin in diabetic rat kidneys and glomerular mesangial cells (GMCs) exposed to high glucose (HG). Simultaneously, SphK1-S1P-mediated fibronectin (FN) and transforming growth factor-beta 1 (TGF-β1) overproduction were inhibited. In addition, curcumin dose dependently reduced SphK1 expression and activity in GMCs transfected with SphK(WT) and significantly suppressed the increase in SphK1-mediated FN levels. Furthermore, curcumin inhibited the DNA-binding activity of activator protein 1 (AP-1), and c-Jun small interference RNA (c-Jun-siRNA) reversed the HG-induced up-regulation of SphK1. These findings suggested that down-regulation of the SphK1-S1P pathway is probably a novel mechanism by which curcumin improves the progression of DN. Inhibiting AP-1 activation is one of the therapeutic targets of curcumin to modulate the SphK1-S1P signaling pathway, thereby preventing diabetic renal fibrosis.

Topics: Animals; Antioxidants; Cells, Cultured; Curcumin; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Drug Evaluation, Preclinical; Fibronectins; Gene Expression; Glucose; JNK Mitogen-Activated Protein Kinases; Kidney; Lysophospholipids; Male; Mesangial Cells; Phosphotransferases (Alcohol Group Acceptor); Rats; Rats, Sprague-Dawley; Signal Transduction; Sphingosine; Transcription Factor AP-1; Transforming Growth Factor beta1

Recent published studies suggest that increasing levels of ceramides enhance the chemo-sensitivity of curcumin. Using in vitro approaches, we analyzed the impact of sphingosine kinase-1 (SphK-1) inhibition on ceramide production, and evaluated SphK1 inhibitor II (SKI-II) as a potential curcumin chemo-sensitizer in ovarian cancer cells. We found that SphK1 is overexpressed in ovarian cancer patients' tumor tissues and in cultured ovarian cancer cell lines. Inhibition of SphK1 by SKI-II or by RNA interference (RNAi) knockdown dramatically enhanced curcumin-induced apoptosis and growth inhibition in ovarian cancer cells. SKI-II facilitated curcumin-induced ceramide production, p38 activation and Akt inhibition. Inhibition of p38 by the pharmacological inhibitor (SB 203580), a dominant-negative expression vector, or by RNAi diminished curcumin and SKI-II co-administration-induced ovarian cancer cell apoptosis. In addition, restoring Akt activation introducing a constitutively active Akt, or inhibiting ceramide production by fumonisin B1 also inhibited the curcumin plus SKI-II co-administration-induced in vitro anti-ovarian cancer effect, suggesting that ceramide accumulation, p38 activation and Akt inhibition are downstream effectors. Our findings suggest that low, well-tolerated doses of SKI-II may offer significant improvement to the clinical curcumin treatment of ovarian cancer.

Topics: Apoptosis; Apoptosis Regulatory Proteins; Blotting, Western; Cell Line, Tumor; Cell Proliferation; Ceramides; Curcumin; Enzyme-Linked Immunosorbent Assay; Female; Growth Inhibitors; Humans; Ovarian Neoplasms; Phosphotransferases (Alcohol Group Acceptor); RNA Interference; RNA, Small Interfering; Transfection