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Autophagy is an important homoeostatic mechanism for the lysosomal degradation of protein aggregates and damaged cytoplasmic components. Recent studies suggest that autophagy which is induced by TGF-β1 suppresses kidney fibrosis in renal tubular epithelial cells (RTECs) of obstructed kidneys. Sphingosine kinase 1(SK1), converting sphingosine into endogenous sphingosine-1-phosphate (S1P), was shown to modulate autophagy and involved in the processes of fibrotic diseases. Since SK1 activity is also up-regulated by TGF-β1, we explored its effect on the induction of autophagy and development of renal fibrosis in this study. In vitro, SK1 expression and activity were markedly increased by TGF-β1 stimulation in a time and concentration dependent manner, and concomitant changes in autophagic response were observed in HK-2 cells. Further, knockdown of SK-1 led to a decrease of autophagy whereas overexpression of SK1 caused a greater induction of autophagy. In addition, overexpression of SK1 resulted in decreased of mature TGF-β levels through autophagic degradation. In vivo, SK1 enzymatic activity and autophagic response were both up-regulated in a mouse model of kidney fibrosis induced by unilateral ureteral obstruction (UUO); meanwhile, increased of mature TGF-β1 and deposition of extracellular matrix (ECM) were observed in tubulointerstitial areas compared with sham-operated mice. However, aggravation of renal fibrosis was detected when SK1 inhibitor PF-543 was applied to suppress SK1 enzymatic activity in UUO mice. At the same time, autophagy was also inhibited by PF-543. Thus, our findings suggest that SK1 activation is renoprotective via induction of autophagy in the fibrotic process. [ABSTRACT FROM AUTHOR]

Abstract: Sphingosine-1-phosphate (S1P) is a pleiotropic lipid mediator that acts either on G protein-coupled S1P receptors on the cell surface or via intracellular target sites. In addition to the well established effects of S1P in angiogenesis, carcinogenesis and immunity, evidence is now continuously accumulating which demonstrates that S1P is an important regulator of fibrosis. The contribution of S1P to fibrosis is of a Janus-faced nature as S1P exhibits both pro- and anti-fibrotic effects depending on its site of action. Extracellular S1P promotes fibrotic processes in a S1P receptor-dependent manner, whereas intracellular S1P has an opposite effect and dampens a fibrotic reaction by yet unidentified mechanisms. Fibrosis is a result of chronic irritation by various factors and is defined by an excess production of extracellular matrix leading to tissue scarring and organ dysfunction. In this review, we highlight the general effects of extracellular and intracellular S1P on the multistep cascade of pathological fibrogenesis including tissue injury, inflammation and the action of pro-fibrotic cytokines that stimulate ECM production and deposition. In a second part we summarize the current knowledge about the involvement of S1P signaling in the development of organ fibrosis of the lung, kidney, liver, heart and skin. Altogether, it is becoming clear that targeting the sphingosine kinase-1/S1P signaling pathway offers therapeutic potential in the treatment of various fibrotic processes. This article is part of a Special Issue entitled Advances in Lysophospholipid Research. [Copyright &y& Elsevier]

Purpose: Sphingosine kinase-1 (SphK1) was shown in preclinical models and non-genitourinary cancers to be instrumental in cancer progression, adaptation to hypoxia and in tumour angiogenesis. No data were available in human prostate cancer. The present study was designed to assess SphK1 expression and activity in radical prostatectomy specimens and to research correlations with clinical features. Materials and methods: Transverse section of fresh tissue was obtained from 30 consecutive patients undergoing laparoscopic prostatectomy. SphK1 enzymatic activities of tumour and normal counterpart were determined. Relationships with PSA, Gleason sum, pathological stage, resection margin status and treatment failure were researched. SphK1 pattern of expression was then assessed on tissue microarray. Results: A significant 2-fold increase in SphK1 enzymatic activity(11.1±8.4 versus 5.9±3.2 (P <0.04)) was observed in cancer. The upper quartile of SphK1 activity was associated with higher PSA (16.7 versus 6.4ng/ml, P =0.04), higher tumor volumes (20.7 versus 9.8, P =0.002), higher rates of positive margins (85.7% versus 28.6%, P =0.01) and surgical failure (71.4% versus 9.5%, P =0.003) than the lower three quartiles. Odds ratios (OR) for treatment failure showed a strong relationship with SphK1 activity (OR: 23.7, P =0.001), positive resection margins (OR: 15.0, P =0.007) and Gleason sum (⩾4+3, OR: 8.0, P =0.003). Tissue microarrays showed discrete epithelial expression that varied with Gleason sum with significant relationship between SphK1 expression and higher Gleason sum. Conclusion: In complement to preclinical literature, the demonstrated relationships between SphK1-increased activity in cancer and relevant clinical features confirm a central role for SphK1 in prostate cancer that herald promising avenues in risk-assessment and treatment. [Copyright &y& Elsevier]

Abstract: PDGF is known to be critically implicated in skeletal muscle repair; however its molecular mechanism of action has been only marginally investigated. In this study we show that in mouse myoblasts PDGF transactivates S1P1 receptor via sphingosine kinase (SK)-1 activation and that this molecular event exerts a negative regulation of the mitogenic effect elicited by this growth factor. Indeed, pharmacological inhibition of S1P1, or its specific silencing increased PDGF-dependent cell proliferation, whereas S1P1 overexpression diminished the biological effect. Moreover, the mitogenic response to PDGF was enhanced by pharmacological inhibition of SK activity as well as specific silencing of SK1 but not SK2. Furthermore, ERK1/2 signalling pathway was found to be upstream of the observed attenuation of PDGF-induced cell proliferation. Interestingly, PDGF-directed engagement of S1P1 exerted also a positive modulatory action of the growth factor-dependent cell motility. The here highlighted dual role of S1P1-mediated signalling in response to myoblast challenge with PDGF is likely important to guarantee the fine control of the biological response to this growth factor, finalized to efficient repopulation of skeletal muscle after damage, where a tight balance between proliferation and migration of tissue progenitor cells is required. [ABSTRACT FROM AUTHOR]

Transforming growth factor-β2 (TGF-β2) stimulates the expression of pro-fibrotic connective tissue growth factor (CTGF) during the course of renal disease. Because sphingosine kinase-1 (SK-1) activity is also upregulated by TGF-β, we studied its effect on CTGF expression and on the development of renal fibrosis. When TGF-β2 was added to an immortalized human podocyte cell line we found that it activated the promoter of SK-1, resulting in upregulation of its mRNA and protein expression. Further, depletion of SK-1 by small interfering RNA or its pharmacological inhibition led to accelerated CTGF expression in the podocytes. Over-expression of SK-1 reduced CTGF induction, an effect mediated by intracellular sphingosine-1-phosphate. In vivo, SK-1 expression was also increased in the podocytes of kidney sections of patients with diabetic nephropathy when compared to normal sections of kidney obtained from patients with renal cancer. Similarly, in a mouse model of streptozotocin-induced diabetic nephropathy, SK-1 and CTGF were upregulated in podocytes. In SK-1 deficient mice, exacerbation of disease was detected by increased albuminuria and CTGF expression when compared to wild-type mice. Thus, SK-1 activity has a protective role in the fibrotic process and its deletion or inhibition aggravates fibrotic disease. [ABSTRACT FROM AUTHOR]

Abstract: Sphingosine 1-phosphate (S1P) is a potent mitogenic signal generated from sphingosine by the action of sphingosine kinases (SKs). In this study, we show that in the human arterial endothelial cell line EA.hy 926 histamine induces a time-dependent upregulation of the SK-1 mRNA and protein expression which is followed by increased SK-1 activity. A similar upregulation of SK-1 is also observed with the direct protein kinase C activator 12-O-tetradecanoylphorbol-13-acetate (TPA). In contrast, SK-2 activity is not affected by neither histamine nor TPA. The increased SK-1 protein expression is due to stimulated de novo synthesis since cycloheximide inhibited the delayed SK-1 protein upregulation. Moreover, the increased SK-1 mRNA expression results from an increased promoter activation by histamine and TPA. In mechanistic terms, the transcriptional upregulation of SK-1 is dependent on PKC and the extracellular signal-regulated protein kinase (ERK) cascade since staurosporine and the MEK inhibitor U0126 abolish the TPA-induced SK-1 induction. Furthermore, the histamine effect is abolished by the H1-receptor antagonist diphenhydramine, but not by the H2-receptor antagonist cimetidine. Parallel to the induction of SK-1, histamine and TPA stimulate an increased migration of endothelial cells, which is prevented by depletion of the SK-1 by small interfering RNA (siRNA). To appoint this specific cell response to a specific PKC isoenzyme, siRNA of PKC-α, -δ, and -ε were used to selectively downregulate the respective isoforms. Interestingly, only depletion of PKC-α leads to a complete loss of TPA- and histamine-triggered SK-1 induction and cell migration. In summary, these data show that PKC-α activation in endothelial cells by histamine-activated H1-receptors, or by direct PKC activators leads to a sustained upregulation of the SK-1 protein expression and activity which, in turn, is critically involved in the mechanism of endothelial cell migration. [Copyright &y& Elsevier]

Abstract: Sphingosine 1-phosphate (S1P) the product of sphingosine kinase (SK) action plays an important role in various pathological conditions like inflammation and cancer. In this study, we show that in the human breast cancer cell line MCF7, epidermal growth factor (EGF) stimulates SK-1 activity in a biphasic manner with a first peak after 15 min and a second delayed activation occurring after 1 h up to 18 h and thereafter declining again. This delayed activation is accompanied by increased mRNA and protein expression of SK-1, but not SK-2. Mechanistically, the transcriptional upregulation is dependent on the classical mitogen-activated protein kinase, protein kinase C (PKC) and the phosphoinositide 3-kinase, since specific inhibitors of these enzymes all abolish the EGF-induced mRNA upregulation and activity of SK-1. Moreover, dexamethasone also suppressed EGF-induced SK-1 mRNA expression and activity which is reversed by the glucocorticoid receptor antagonist RU486. To see whether EGF-induced upregulation of SK-1 is of relevance for tumor progression, we investigated two hallmarks of carcinogenesis, i.e., cell proliferation and migration. Stimulation of cells with EGF leads to enhanced [3H]thymidine incorporation into DNA and also to stimulated migration in a modified Boyden chamber assay. When cells are depleted of SK-1, but not SK-2, by siRNA transfection or by dexamethasone treatment, EGF-induced proliferation and migration are drastically reduced. In summary, these data show that EGF causes an acute stimulation of SK-1 activity and, moreover, triggers a delayed SK activation which is due to increased gene transcription and de novo synthesis of SK-1, which in turn directs cells towards growth and increased motility. Thus, the sphingosine kinase-1 may represent a novel attractive target for cancer therapy. [Copyright &y& Elsevier]

Influence of quail egg on pathologies has increased research interests and series of investigations are currently being done on its influence against these pathologies. The influence of quail egg against 2-butoxyethanol induced hemolysis and disseminated thrombosis was investigated to determine the enzymatic regulations that ensue in the amelioration of deleterious hemolytic and disseminated thrombosis displayed in female Wistar rats. Quail egg was separated into three (3) components (extracts)-quail egg yolk water soluble (QYWS) and fat soluble (QYFS), and albumen extract (QA) and the inorganic and organic compositions were characterized. Depranocytotic assaults was achieved by 250 mg/kg of 2-Butoxyethanol administered for 4 days, the clinical observation revealed a dark purple-red discoloration on the distal tails of the rats and therapeutic applications followed with 1000 mg/kg BWT of QYWS, QYFS and QA, and 15 mg/kg BWT of hydroxyurea. Morphological evaluation, haematological estimations and biochemical evaluations of the influence on the activities of sphingosine kinase-1, RNase, red cell carbonic anhydrase, lactate dehydrogenase, glutathione peroxidase and caspase-3, vis a vis the concentrations of sphingosine-1 phosphate, selenium and zinc (plasma and urine). In vitro anti-inflammatory influence of quail egg components were investigated against hemolysis and key enzymes of inflammation-cycloxygenase, lipoxygenase and β-glucuronidase. The in vitro anti-inflammatory effects of QYWS, QYFS and QA were concentration dependent from 200 to 800 μg/ml against hemolysis and the key enzymes of inflammation. The characterization of inorganic and organic bioactive composition of the yolk and albumen revealed the presence of folic acid, cobalamin, pyridine, riboflavin, ascorbic acid as well as vitamins D and E, selenium, zinc, iron and calcium. These had reflected in the attenuation of the induced hemolytic and disseminated thrombosis by regulations of enzymes linked to the infarction, apoptosis and oxidative stress characterized in sickle cell index. [ABSTRACT FROM AUTHOR]

• 2% Isoflurane and 3% sevoflurane are neuroprotective equally. • Isoflurane and sevoflurane are effective against early brain injury after SAH. • Isoflurane and sevoflurane suppress post-SAH apoptosis and brain inflammation. • Effect of isoflurane and sevoflurane is mediated by sphingosine-related pathway. • Isoflurane and sevoflurane are good candidates for neuroanesthesia after SAH. The first step to treat aneurysmal subarachnoid hemorrhage (SAH) is aneurysmal obliteration under general anesthesia but not treat the SAH itself and the secondary effects. However, the identification of anesthetics with properties that help to attenuate post-SAH brain injury can be useful for improving outcomes of SAH patients. We examined whether 2% isoflurane and 3% sevoflurane posttreatment are protective against early brain injury (EBI) after SAH. This study used 87 8-week-old male CD-1 mice. We induced SAH by endovascular perforation in mice. Animals were randomly divided into 4 groups: sham-operated (n = 16), SAH + vehicle-medical air (n = 26), SAH + 2% isoflurane (n = 22), and SAH + 3% sevoflurane (n = 23). Neurobehavioral function, brain water content and Western blotting were evaluated at 24 h. The expression of sphingosine kinase (SphK), cleaved caspase-3 and cyclooxygenase-2 (COX2) was determined by Western blotting. Cell death was examined by terminal deoxynucleotidyl transferase-mediated uridine 5′-triphosphate-biotin nick end-labeling staining. Both 2% isoflurane and 3% sevoflurane significantly improved neurobehavioral function, and brain edema at 24 h after SAH and attenuated cell death, associated with an increase in SphK1, a decrease in cleaved caspase-3 and COX2. The neuroprotective effects were similar between 2% isoflurane and 3% sevoflurane. These findings suggest that both 2% isoflurane and 3% sevoflurane significantly inhibited EBI by suppressing post-SAH apoptosis and brain inflammation possibly via the SphK1-related pathway. [ABSTRACT FROM AUTHOR]