Activation of Phospholipase D in Rat Thymocytes by Sphingosine

Lee, Young-kyun,Choi, Myung-Un Korean Chemical Society 2002 Bulletin of the Korean Chemical Society Vol.23 No.10

Sphingosine is known to regulate a wide range of cell physiology including growth, differentiation, and apoptosis. In this study, we examined the effect of sphingosine on the phospholipase D (PLD) activity in rat thymocytes. Sphingosine potently stimulated PLD in the absence of extracellular calcium, while depletion of intracellular calcium by BAPTA/AM treatment completely blocked activation of PLD by sphingosine. Sphingosine-induced increase of the intracellular calcium concentration was confirmed using a fluorescent calcium indicator Fluo-3/AM. A phosphoinositide-specific phospholipase C inhibitor U73122 partially inhibited the stimulation of PLD by sphingosine. When mouse PLD2 gene was transfected into mouse thymoma EL4 cells, which lack intrinsic PLD activity, sphingosine could stimulate PLD2 significantly while overexpression of human PLD1 had no effect. Taken together, the sphingosine-stimulated PLD activity in rat thymocytes is dependent on the mobilization of intracellular calcium and appears to be due to the PLD2 isoform.

Effects of Sphingosine-1-phosphate on Vestibular Nuclear Neurons

Jae-Hyuk Lee,Sujeong Jang,Song-Hee Kim,Han-Seong Jeong,Jong-Seong Park(박종성) 대한의생명과학회 2010 Biomedical Science Letters Vol.16 No.1

This study was designed to investigate the effects of sphingosine-1-phosphate on the neuronal activity of rat medial vestibular nuclear neurons. Sprague-Dawley rats aged 14 to 16 days were decapitated under ether anesthesia. After treatment with pronase and thermolysin, the dissociated medial vestibular nuclear neurons were transferred into a chamber on an inverted microscope. Spontaneous action potentials and potassium currents were recorded by standard patch-clamp techniques under current and voltage-clamp modes respectively. 15 medial vestibular nuclear neurons revealed excitatory responses to 1 and 5 μM of sphingosine-1-phosphate. The spike frequency and resting membrane potential of these cells were increased by sphingosine-1-phosphate. The amplitude of afterhyperpolarization was decreased by sphingosine-1-phosphate. Whole potassium currents of medial vestibular nuclear neurons were decreased by sphingosine-1-phosphate (n=12). Sphingosine-1-phosphate did not affect the charybdotoxin-treated potassium currents. These experimental results suggest that sphingosine-1-phosphate increases the neuronal activity of the medial vestibular nuclear neurons by altering the resting membrane potential and afterhyperpolarization.

Sphingosine Kinase: Biochemical and Cellular Regulation and Role in Disease

Taha, Tarek Assad,Hannun, Yusuf Awni,Obeid, Lina Marie Korean Society for Biochemistry and Molecular Biol 2006 Journal of biochemistry and molecular biology Vol.39 No.2

Sphingolipids have emerged as molecules whose metabolism is regulated leading to generation of bioactive products including ceramide, sphingosine, and sphingosine-1-phosphate. The balance between cellular levels of these bioactive products is increasingly recognized to be critical to cell regulation; whereby, ceramide and sphingosine cause apoptosis and growth arrest phenotypes, and sphingosine-1-phosphate mediates proliferative and angiogenic responses. Sphingosine kinase is a key enzyme in modulating the levels of these lipids and is emerging as an important and regulated enzyme. This review is geared at mechanisms of regulation of sphingosine kinase and the coming to light of its role in disease.

Deficiency of Sphingosine-1-Phosphate Reduces the Expression of Prohibitin and Causes β-Cell Impairment via Mitochondrial Dysregulation

홍석우,이진미,권혜미,박세은,이은정,박철영,오기원,박성우,이원영 대한내분비학회 2018 Endocrinology and metabolism Vol.33 No.3

Background: Emerging evidence suggests that sphingolipids may be involved in type 2 diabetes. However, the exact signaling defectthrough which disordered sphingolipid metabolism induces β-cell dysfunction remains unknown. The current study demonstratedthat sphingosine-1-phosphate (S1P), the product of sphingosine kinase (SphK), is an essential factor for maintaining β-cell functionand survival via regulation of mitochondrial action, as mediated by prohibitin (PHB). Methods: We examined β-cell function and viability, as measured by mitochondrial function, in mouse insulinoma 6 (MIN6) cellsin response to manipulation of cellular S1P and PHB levels. Results: Lack of S1P induced by sphingosine kinase inhibitor (SphKi) treatment caused β-cell dysfunction and apoptosis, with repressionof mitochondrial function shown by decreases in cellular adenosine triphosphate content, the oxygen consumption rate, theexpression of oxidative phosphorylation complexes, the mitochondrial membrane potential, and the expression of key regulators ofmitochondrial dynamics (mitochondrial dynamin-like GTPase [OPA1] and mitofusin 1 [MFN1]). Supplementation of S1P led to therecovery of mitochondrial function and greatly improved β-cell function and viability. Knockdown of SphK2 using small interferingRNA induced mitochondrial dysfunction, decreased glucose-stimulated insulin secretion (GSIS), and reduced the expression ofPHB, an essential regulator of mitochondrial metabolism. PHB deficiency significantly reduced GSIS and induced mitochondrialdysfunction, and co-treatment with S1P did not reverse these trends. Conclusion: Altogether, these data suggest that S1P is an essential factor in the maintenance of β-cell function and survival throughits regulation of mitochondrial action and PHB expression.

Sphingosine-1-phosphate의 HaCaT 세포의 증식에 대한 이중적 효과에 대한 연구

김형훈,강숙경,장성은,이미우,최지호,성경제,문기찬,고재경 大韓皮膚硏究學會 2003 大韓皮膚硏究學會誌 Vol.10 No.4

초고성능액체크로마토그래피-텐덤질량분석법에 의한 혈청 중스핑고신과 스핑고신-1-포스페이트의 동시 분석

나승훈(Seonghoon Na),윤혜란(Hye-Ran Yoon) 대한약학회 2021 약학회지 Vol.65 No.1

Sphingosine (SPH) and sphingosine-1-phosphate (S1P) are emerging as key players in asthma metabolism and in numerous cellular inflammation processes. To identify potential biomarkers of asthma and inflammatory therapeutics, it is essential to determine their levels. Herein, we developed a rapid and sensitive UHPLC-MS/MS method to simultaneously quantify SPH and S1P in human serum using C17-SPH and C17-S1P as internal standards. After methanol precipitation of serum proteins, the supernatants were analyzed by MS/MS performed in the positive ion mode by multiple reaction monitoring. UHPLC analysis (C18 column) was performed using two mobile phase systems (water containing 0.1% formic acid, and 85% acetonitrile containing 0.1% formic acid) within 5 min of the short run. The calibration curves were linear in the range of 0.002-1.5 µg/mL for S1P and SPH with an R 2 greater than 0.9999. The LOD and LOQ were 0.0002 and 0.0004 µg/mL for S1P, and 0.0005 and 0.001 µg/mL for SPH, respectively. The accuracy and precision of the method were in the range of 89.8-100.7% (RSD, 1.5-2.8%) for both SPH and S1P species. We were able to quantify both molecules in serum from healthy and asthmatic patients. These results suggest that SPH and S1P are promising potential biomarkers, and also contribute to the basic data for the construction of an omics-based platform for preventive index prior to asthma diagnosis.

The role of sphingosine-metabolizing enzymes in viral immunopathogenesis

Young-Jin Seo 한국실험동물학회 2021 한국실험동물학회 학술발표대회 논문집 Vol.2021 No.7

Diverse viruses continue to pose a threat to public health by causing illness and mortality in humans. Discovering host factors that regulate viral propagation and pathogenesis is vital for the development of novel drugs. In this study, we investigated the role of sphingosine-metabolizing enzymes in acute and chronic viral infections. We demonstrated that the inhibition of sphingosine kinase (SphK) 1 or SphK2 enhances the host defense against acute and chronic viruses. The temporal inhibition of SphK1 or SphK2 protected mice from influenza virus-induced morbidity and mortality. Furthermore, we found that SphK2 functions during chronic viral (Clone 13 strain of lymphocytic choriomeningitis virus) infection to limit T cell immune pathology, which subsequently aids in the establishment of virusinduced immunosuppression and the resultant viral persistence. Notably, oral instillation of an SphK2- selective inhibitor promoted protective T cell responses and accelerated the termination of LCMV Clone 13 persistence in mice. Collectively, our results indicate that sphingosine-metabolizing enzymes function as pro-viral factors during viral infection. Therefore, SphK1 and SphK2 represent potential host targets for therapeutics against acute and chronic viral infections.

Effect of Sphingosine-1-Phosphate on Intracellular Free Ca²⁺ in Cat Esophageal Smooth Muscle Cells

( Dong Kyu Lee ),( Young Sil Min ),( Seong Su Yoo ),( Hyun Sub Shim ),( Sun Young Park ),( Uy Dong Sohn ) 한국응용약물학회 2018 Biomolecules & Therapeutics(구 응용약물학회지) Vol.26 No.6

A comprehensive collection of proteins senses local changes in intracellular Ca²⁺ concentrations ([Ca²⁺]_i) and transduces these signals into responses to agonists. In the present study, we examined the effect of sphingosine-1-phosphate (S1P) on modulation of intracellular Ca²⁺ concentrations in cat esophageal smooth muscle cells. To measure [Ca²⁺]_i levels in cat esophageal smooth muscle cells, we used a fluorescence microscopy with the Fura-2 loading method. S1P produced a concentration-dependent increase in [Ca²⁺]_i in the cells. Pretreatment with EGTA, an extracellular Ca²⁺ chelator, decreased the S1P-induced increase in [Ca²⁺]_i, and an L-type Ca²⁺-channel blocker, nimodipine, decreased the effect of S1P. This indicates that Ca²⁺ influx may be required for muscle contraction by S1P. When stimulated with thapsigargin, an intracellular calcium chelator, or 2-Aminoethoxydiphenyl borate (2-APB), an InsP₃ receptor blocker, the S1P-evoked increase in [Ca²⁺]_i was significantly decreased. Treatment with pertussis toxin (PTX), an inhibitor of Gi-protein, suppressed the increase in [Ca²⁺]_i evoked by S1P. These results suggest that the S1P-induced increase in [Ca²⁺]_i in cat esophageal smooth muscle cells occurs upon the activation of phospholipase C and subsequent release of Ca²⁺ from the InsP₃-sensitive Ca²⁺ pool in the sarcoplasmic reticulum. These results suggest that S1P utilized extracellular Ca²⁺ via the L type Ca²⁺ channel, which was dependent on activation of the S1P₄ receptor coupled to PTX-sensitive G_i protein, via phospholipase Cmediated Ca²⁺ release from the InsP₃-sensitive Ca²⁺ pool in cat esophageal smooth muscle cells.

Integrative roles of sphingosine kinase in liver pathophysiology

Kim Kyu Min,Shin Eun Jin,Yang Ji Hye,Ki Sung Hwan 한국독성학회 2023 Toxicological Research Vol.39 No.4

Bioactive sphingolipids and enzymes that metabolize sphingolipid-related substances have been considered as critical messengers in various signaling pathways. One such enzyme is the crucial lipid kinase, sphingosine kinase (SphK), which mediates the conversion of sphingosine to the potent signaling substance, sphingosine-1-phosphate. Several studies have demonstrated that SphK metabolism is strictly regulated to maintain the homeostatic balance of cells. Here, we summarize the role of SphK in the course of liver disease and illustrate its effects on both physiological and pathological conditions of the liver. SphK has been implicated in a variety of liver diseases, such as steatosis, liver fibrosis, hepatocellular carcinoma, and hepatic failure. This study may advance the understanding of the cellular and molecular foundations of liver disease and establish therapeutic approaches via SphK modulation.

Divergent Synthesis of Diastereomeric Sphingosines from a Chiral Aziridine

강온유,Mi-Ri Shin,강한영 대한화학회 2016 Bulletin of the Korean Chemical Society Vol.37 No.7

All four stereoisomers of sphingosines were synthesized starting from a single intermediate, chiral aziridine (2), which was efficiently prepared by enzymatic desymmetrization in an enatiopure form. Aziridine (2) was converted to 3, which was used for the synthesis of 4. Both the advanced key intermediates, vinylaziridines 3 and 4, were successfully converted to threo-sphingosines 1a and 1b, respectively. Ring-closing metathesis (RCM) using the Grubbs II catalyst was the key reaction in the synthesis. Two erythro-sphingosines 1c and 1d were synthesized by the ring-expansion reactions of vinylaziridines 3 and 4, followed by RCM reactions. The successful divergent synthesis confirmed that chiral vinylaziridine 2 can be used as a key intermediate for the synthesis of sphingosine-related natural products.