Inhibitory Effect of S1PR2 Antagonist JTE-013 on Proliferation of Chronic Myeloid Leukemia Cells / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of sphingosine-1-phosphate receptor 2 (S1PR2) specific antagonist JTE-013 on the proliferation of human chronic myeloid leukemia (CML) cell line K562.@*METHODS@#K562 cells were treated with JTE-013 (0, 0.5, 1, 5, 10, 20 μmol/L) for 24 and 48 hours respectively, CCK8 assay was used to detect the cell viability. K562 cells were treated with JTE-013 (0, 5, 10, 20 μmol/L) for 24 hours, propidium iodide (PI) DNA staining was used to analyze the cell cycle, Western blot was used to determine the levels of P21 and Cyclin D1 protein expression.@*RESULTS@#JTE-013 inhibited the proliferation of CML cell line K562 in a dose dependent manner (r=-0.971). The proliferation rate of CML cells showed that the activity of CML cells decreased gradually with the increase of JTE-013 concentration (r=-0.971). The detection demonstrated that JTE-013 suppressed tumor cell proliferation through cell cycle arrest in G/G phase. Further detection of the protein expressions of G phase regulators showed that level of P21 increased, and expression of Cyclin D1 decreased.@*CONCLUSION@#JTE-013, a S1PR2 antagonist, can inhibit the proliferation of human CML K562 cells, which may be achieved by arresting the cells in G/G phase.

Sphingolipid Mediators of Myocardial Pathology

Cardiomyopathy is the leading cause of mortality worldwide. While the causes of cardiomyopathy continue to be elucidated, current evidence suggests that aberrant bioactive lipid signaling plays a crucial role as a component of cardiac pathophysiology. Sphingolipids have been implicated in the pathophysiology of cardiovascular disease, as they regulate numerous cellular processes that occur in primary and secondary cardiomyopathies. Experimental evidence gathered over the last few decades from both in vitro and in vivo model systems indicates that inhibitors of sphingolipid synthesis attenuate a variety of cardiomyopathic symptoms. In this review, we focus on various cardiomyopathies in which sphingolipids have been implicated and the potential therapeutic benefits that could be gained by targeting sphingolipid metabolism.

Altered expressions of SphK1 and S1PR2 in hippocampus of epileptic rats / 中国应用生理学杂志

OBJECTIVE@#To observe the expressions of sphingosine kinase 1 (SphK1) and sphingosine-1-phosphate receptor 2 (S1PR2) in hippocampus of epileptic rats and to investigate the pathogenesis of SphK1 and S1PR2 in epilepsy.@*METHODS@#One hundred and eight male Sprague-Dawley (SD) rats were randomly divided into control group (n=48) and pilocarpine (PILO) group (n=60). A robust convulsive status epilepticus (SE) was induced in PILO group rats by the application of pilocarpine. Control group rats were injected with respective of physiological saline. Pilocarpine group was randomly divided into 6 subgroups (n=8): acute group (E6 h, E1 d, E3 d), latent group (E7 d) and chronic group (E30 d, E56 d). Each subgroup has 8 control rats and 8 epileptic rats. Hippocampal tissue and brain slices were obtained from control rats and rats subjected to the Li-PILO model of epilepsy at 6 h, 1 d, 3 d,7 d,30 d and 56 d after status epilepticus (SE). Western blot technique was used to determine the expressions of SphK1 and S1PR2 in hippocampus at different point of time after pilocarpine treatment. Immunofluorescence was applied to detect the activation and proliferation of hippocampal astrocytes and the localization of SphK1 and S1PR2 in rat hippocampal astrocytes.@*RESULTS@#Compared with control group, the levels of SphK1 in acute phase (E3 d), latent phase (E7 d) and chronic phase (E30 d, E56 d) were significantly increased while the expressions of S1PR2 were decreased in acute phase (E3 d), latent phase (E7 d) and chronic phase (E30 d, E56 d)(P＜0.05 or P＜0.01). Immunofluorescence results showed astrocyte activation and proliferation in hippocampus of epileptic (E7 d) rats (P＜0.05). Confocal microscopy confirmed the preferential expressions of SphK1 and S1PR2 in epileptic rat(E7 d)hippocampal astrocytes.@*CONCLUSION@#The results indicate that SphK1 and S1PR2 may play an important role in the pathogenesis of epilepsy by regulating the activation and proliferation of hippocampal astrocytes and altering neuronal excitability.

Deficiency of Sphingosine-1-Phosphate Receptor 2 (S1P₂) Attenuates Bleomycin-Induced Pulmonary Fibrosis

Sphingosine 1-phosphate (S1P) levels are often found to be elevated in serum, bronchoalveolar lavage, and lung tissue of idiopathic pulmonary fibrosis patients and experimental mouse models. Although the roles of sphingosine kinase 1 and S1P receptors have been implicated in fibrosis, the underlying mechanism of fibrosis via Sphingosine 1-phosphate receptor 2 (S1P₂) has not been fully investigated. Therefore, in this study, the roles of S1P₂ in lung inflammation and fibrosis was investigated by means of a bleomycin-induced lung fibrosis model and lung epithelial cells. Bleomycin was found to induce lung inflammation on day 7 and fibrosis on day 28 of treatment. On the 7(th) day after bleomycin administration, S1P₂ deficient mice exhibited significantly less pulmonary inflammation, including cell infiltration and pro-inflammatory cytokine induction, than the wild type mice. On the 28(th) day after bleomycin treatment, severe inflammation and fibrosis were observed in lung tissues from wild type mice, while lung tissues from S1P₂ deficient mice showed less inflammation and fibrosis. Increase in TGF-β1-induced extracellular matrix accumulation and epithelial-mesenchymal transition were inhibited by JTE-013, a S1P₂ antagonist, in A549 lung epithelial cells. Taken together, pro-inflammatory and pro-fibrotic functions of S1P₂ were elucidated using a bleomycin-induced fibrosis model. Notably, S1P₂ was found to mediate epithelial-mesenchymal transition in fibrotic responses. Therefore, the results of this study indicate that S1P₂ could be a promising therapeutic target for the treatment of pulmonary fibrosis.

Decreased Expression of Sphingosine-1-Phosphate Receptor 1 in the Blood Leukocyte of Rheumatoid Arthritis Patients

Sphingosine-1-phosphate (S1P) plays an important role in trafficking leukocytes and developing immune disorders including autoimmunity. In the synovium of rheumatoid arthritis (RA) patients, increased expression of S1P was reported, and the interaction between S1P and S1P receptor 1 (S1P1) has been suggested to regulate the expression of inflammatory genes and the proliferation of synovial cells. In this study, we investigated the level of S1P1 mRNA expression in the blood leukocytes of RA patients. In contrast to the previous reports, the expression level of this gene was not correlated to their clinical scores, disease durations and ages. However, S1P1 was transcribed at a significantly lower level in the circulating leukocytes of RA patients when compared to age-, and sex-matched healthy controls. Since these data may suggest the participation of S1P1, further studies are needed to determine the role of this receptor in the pathogenesis of RA.

Lentiviral vectors carrying siRNA inhibit S1PR3 gene expression in the corpus cavernosum smooth muscle cells of rats with spontaneous hypertension / 中华男科学杂志

Objective@#To screen lentiviral vectors carrying siRNA which can specifically down-regulate the gene expression of the sphingosine-1-phosphate receptor 3 (S1PR3) in the corpus cavernosum smooth muscle (CCSM) cells of rats with spontaneous hypertension (SHT) and investigate the influence of the vectors on the signaling pathways of ROCK1, ROCK2 and eNOS in the CCSM cells of SHT rats.@*METHODS@#Using the S1PR3 mRNA sequence of the rat as an interfering target, we designed and synthesized three pairs of siRNA sequences (siRNA1, 2 and 3) targeting S1PR3 and one pair of negative control, and then constructed and packaged them into lentiviral vectors. We cultured the CCSM cells of SHT and Wistar-Kyoto (WKY) rats in vitro and randomly divided them into groups A (SHT untransfected control), B (SHT transfected and carrying negative control virus), C (SHT transfected and carrying siRNA1 targeting S1PR3), D (SHT transfected and carrying siRNA2 targeting S1PR3), E (SHT transfected and carrying siRNA3 targeting S1PR3), and F (WKY untransfected control). With the multiplicity of infection (MOI) = 60, we transfected the CCSM cells of the SHT rats with the lentiviral vector and then determined the expression of the green fluorescent protein (GFP) as well as the mRNA and protein expressions of S1PR3, ROCK1, ROCK2 and eNOS in the CCSM cells of the SHT and WKY rats by RT-PCR and Western blot.@*RESULTS@#Gene sequencing proved the successful construction of the lentiviral vector. The transfection efficiency of the CCSM cells of the rats was >80% in groups B, C, D and E. Compared with group A, the mRNA and protein expressions of S1PR3, ROCK1 and ROCK2 exhibited no significant difference in group B but were remarkably decreased in groups C, D, E and F (P0.05) but remarkably lower than those in group F (P0.05) but markedly increased in groups A, B, C and D (P< 0.05), while those of eNOS remarkably decreased in groups A, B, C, D and E (P< 0.05).@*CONCLUSIONS@#The three constructed lentiviral vectors carrying siRNA targeting different loci of the S1PR3 gene could significantly inhibit the expression of S1P3 as well as RhoA/Rho kinase signaling pathways in the CCSM cells of SHT rats, and the vector carrying siRNA3 exhibited the highest inhibitory effect.

Sphingosine 1-Phosphate Receptor Modulators and Drug Discovery

Initial discovery on sphingosine 1-phosphate (S1P) as an intracellular second messenger was faced unexpectedly with roles of S1P as a first messenger, which subsequently resulted in cloning of its G protein-coupled receptors, S1P₁₋₅. The molecular identification of S1P receptors opened up a new avenue for pathophysiological research on this lipid mediator. Cellular and molecular in vitro studies and in vivo studies on gene deficient mice have elucidated cellular signaling pathways and the pathophysiological meanings of S1P receptors. Another unexpected finding that fingolimod (FTY720) modulates S1P receptors accelerated drug discovery in this field. Fingolimod was approved as a first-in-class, orally active drug for relapsing multiple sclerosis in 2010, and its applications in other disease conditions are currently under clinical trials. In addition, more selective S1P receptor modulators with better pharmacokinetic profiles and fewer side effects are under development. Some of them are being clinically tested in the contexts of multiple sclerosis and other autoimmune and inflammatory disorders, such as, psoriasis, Crohn’s disease, ulcerative colitis, polymyositis, dermatomyositis, liver failure, renal failure, acute stroke, and transplant rejection. In this review, the authors discuss the state of the art regarding the status of drug discovery efforts targeting S1P receptors and place emphasis on potential clinical applications.

Effect of rhG-CSF Mobilization on S1P5 Expression in T Lymphocyte Subsets of Allo-HSCT Donors / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To explore the effect of recombinant human granulocyte colony-stimulating factor (rhG-CSF) mobilization on S1P5 expression in T lymphocyte subsets of allo-HSCT donors.</p><p><b>METHODS</b>The peripheral blood was collected from 10 allo-hematopoietic stem cell transplantation (allo-HSCT) donors before and after mobilization with rhG-CSF for 4 days. The flow cytometry was used to detect S1P5 expression in T lymphocyte subsets.</p><p><b>RESULTS</b>There was no S1P5 expression on the surface of T-lymphocytes both before and after rhG-CSF mobilization. After fixation with permeabilization agent, S1P5 expression could be detected in lymphocytes after rhG-CSF mobilization, which indicates S1P5 may be located in cells. Compared with level before rhG-CSF mobilization, S1P5 expression was significantly increased in T lymphocyte subsets after rhG-CSF mobilization, CD3(+)T cells (57.92±2.32)% vs (7.94±1.47)%(P<0.05）, CD4(+)T cells (72.58±1.73)% vs （5.48±0.82）%(P<0.05）, CD8(+)T cells（51.79±3.57）% vs （6.46±1.01）%（P<0.05）,CD3-/CD56(+)NK cells（40.00±1.47）% vs（4.97±0.74）%（P<0.05）. The up-regulated level of S1P5 expression in CD4(+)T cells was most high(P<0.05).</p><p><b>CONCLUSION</b>S1P5 expression significantly increases in T lymphocyte subsets after rhG-CSF mobilization, and the up-regulated level of S1P5 expression in CD4(+)T cells is the most high.</p>

Application of CRISPR/Cas9 Gene Editing Technique to Establish S1PR5 Gene Knockout Mice / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To establish the S1PR5 gene knockout mouse model by using CRISPR/Cas9 gene editing technique so as to provide the tool for studying the regulating role of sphingosine-1-phosphate receptor 5 (S1PR5) in allogeneic hematopoietic stem cell transplantation.</p><p><b>METHODS</b>Single guide RNA (sgRNA) plasmids against the exon 3 of S1PR5 were designed and constructed. Then the sgRNA and hCas9 were transcribed by T7 RNA polymerase in vitro. Cas9 mRNA and sgRNA were mixed and microinjected into fertilized eggs of C57BL/6 mice. T7E1 digestion and gene sequencing were used to detect the mutations of S1PR5. Quantitative PCR (qPCR) and Western blot were used to detect the expression of S1PR5.</p><p><b>RESULTS</b>Finally 2 kinds of F2 generation of homozygous S1PR5 deficent mice (S1PR5-170/-170 mice and S1PR5-215/-215 mice) were gained, and in these 2 model mice the S1PR5 did not express at mRNA and protein levels.</p><p><b>CONCLUSION</b>A mouse model with S1PR5 dificiency has been successfully established, which shall lay a foundation for future investigation of S1PR5.</p>

Influence of S1PR5 Defect on the Lymphocyte Distribution in Mice / 中国实验血液学杂志

<p><b>BACKGROUND</b>The sphingosine 1-phosphate (S1P) receptors (S1PRs) are a group of G protein-coupled receptors expressed on the surface of lymphocytes. The interaction between S1P and S1PRs plays a significant role in the migration and distribution of lymphocytes.</p><p><b>OBJECTIVE</b>To investigate the influence of S1PR5 defect on the lymphocytes distribution in mice.</p><p><b>METHODS</b>The distribution of different subsets of lymphocyte in the mice with S1PR5 defect was examined by flow cytometry.</p><p><b>RESULTS</b>Compared with wild type mice, the number of NK cells in the peripheral blood (PB) and spleen (SP) from the mice with S1PR5 defect decreased very significantly 〔PB: 6.4±0.45% vs 2.2±0.47(P<0.01,n=3)；SP: 3.0±0.91% vs 0.68±0.14%(P<0.05,n=3)〕. However, the NK cell number in the bone marrow (BM) and lymphonodes (LN) of the mice with S1PR5 defect increased very significantly 〔BM: 0.97±0.20 % vs 2.6±0.35% (P<0.01, n=3); LN: 0.35±0.16% vs 1.7±0.15% (P<0.01, n=3)〕. The percentages of CD3(+) lymphocyte in peripheral blood, spleen and lymph node were not statistically significantly different between these 2 types of mice 〔PB: 17.3±7.9% vs 17.0±4.6% (P>0.05, n=3); SP: 33.0±6.0% vs 27.4±1.8% (P>0.05, n=3); LN: 42.3±10.7% vs 51.2±2.7% (P>0.05, n=3)〕.</p><p><b>CONCLUSION</b>S1PR5 defect can significantly influence the NK cell distribution.</p>

Expressions of S1P1-3 in the corpus cavernosum of castrated male rats / 中华男科学杂志

<p><b>OBJECTIVE</b>To investigate the expressions of sphingosine-1-phosphate receptors 1-3 (S1P1- 3) in the corpus cavernosum of castrated male rats and its relationship with the NOS/NO/cGMP and RhoA/Rho kinase signaling pathways.</p><p><b>METHODS</b>We equally randomized 18 eight-week-old healthy male SD rats into a sham-operation control, a castration, and a testosterone replacement (TR) group and harvested the bilateral testes and epididymides from the rats in the latter two groups, followed by 4 weeks of subcutaneous injection of testosterone propionate at 3 mg per kilogram of the body weight per day for those in the TR group and that of plant oil for those in the control and castration groups. At the age of 12 weeks, we measured the serum testosterone (T) level and maximum intracavernous pressure/mean arterial pressure (ICPmax/MAP) of the animals and determined the expressions of SlP1-3, eNOS, P-eNOS, ROCK1, and ROCK2 in the corpus cavernosum by Western blot and immunohistochemistry.</p><p><b>RESULTS</b>The serum T level was significantly decreased in the rats of the castration group as compared with those of the control and TR groups ([0.41 ± 0.04] vs [16.01 ± 1.02] and [15.84 ± 1.32] nmol/L, P < 0.01), with no statistically significant difference between the latter two groups. The ICPmax/MAP at 0 V, 3 V, and 5 V electric stimulation was remarkably lower in the rats of the castration group (0.088 ± 0.014, 0.323 ± 0.014, and 0.432 ± 0.012) than in those of the control group (0.155 ± 0.011, 0.711 ± 0. 010, and 0.819 ± 0.024) and TR group (0.153 ± 0.012, 0.696 ± 0.017, and 0.763 ± 0.027) (P < 0.01), with no significant difference between the latter two groups. With GAPDH as internal control, the animals of the castration group showed markedly reduced expressions of S1P1 ([49.99 ± 3.39]%), eNOS ([46.82 ± 3.81]%) , and P-eNOS ([45.42 ± 4.35]%) in comparison with those in the control group ([72.57 ± 3.06], [89.76 ± 3.98], and [82.53 ± 8.92] and TR group ([71.77 ± 4.43], [87.19 ± 4.23], and [79.82 ± 7.38]%) (P < 0.01) , while the expressions of S1P2, S1P3, ROCK1, and ROCK2 were significantly upregulated in the castration group ([82.35 ± 4.13], [61.03 ± 5.14], [74.50 ± 4.02], and [69.83 ± 5.75]%) as compared with those in the control group ([41.67 ± 1.68], [31.66 ± 2.67], [35.69 ± 5.56], and [39.85 ± 7.17]%) and TR group ([42.80 ± 3.87], [32.25 ± 4.22], 38.06 ± 5.21], and [42.36 ± 4.44]%) (P < 0.01).</p><p><b>CONCLUSION</b>Androgen deficiency induces significant reduction of ICPmax/ MAP in male rats, which is possibly associated with the decline of S1P1 in the corpus cavernosum, inhibition of the eNOS/NO/cGMP signaling pathway, increased expressions of S1P2 and S1P3, and activation of the RhoA/Rho kinase signaling pathway.</p>

Effect of a novel selective S1P1 agonist, Syl948, on mouse skin transplantation / 药学学报

Syl948 is a synthesized selective S1P1 agonist with novel structure. HTRF-IP1 test indicated that Syl948-P, the active form of Syl948 in vitro, has strong activity against S1P1 (EC50: 83 +/- 16 nmol x L(-1)), but its effect on S1P3 was very weak (EC50: 1 026 +/- 90 nmol x L(-1)). In SD rats, oral administration of Syl948 10 mg x kg(-1) significantly decreased the peripheral blood lymphocytes (PBL), with the maximal PBL inhibition rate of 63%, which was as similar as equal dose of fingolimod (FTY720). Oral administration of Syl948 10 mg x kg(-1) had no effect on heart rate of SD rats, which was better than FTY720. Daily oral administration with Syl948 (2 or 4 mg x kg(-1)) significantly prolonged the survival time of the allografts of skin slice on mice. In summary, the above results demonstrated that Syl948 has great selectivity in vitro and good activity in vivo, which indicated its potential use as an anti-rejection drug in skin transplantation.

Sphingosine-1-phosphate receptors respond differently to early myocardial ischemia and ischemia-reperfusion in vivo / 生理学报

Sphingosine-1-phosphate (S1P) has been demonstrated to be a mediator and marker of heart diseases. We hypothesized that the expression of S1P receptors is involved in the S1P-mediated cardioprotection in vivo and may serve as a biomarker of ischemic heart disease. In vivo models of myocardial ischemia (MI) and ischemia-reperfusion (IR) were established by ligation of the left anterior descending artery (LAD) of rat heart, the mRNA expressions of S1PR1-3 were detected using real time PCR at different time intervals after ischemia (LAD for 15 min, 30 min, and 1 h) and IR. The results showed that mRNA expression of S1PR3, but not S1PR1 and S1PR2, increased greatly after IR. No statistical difference was found in any of the three S1P receptors after MI within 1 h. Regarding the studies of lipid concentration changes in myocardiopathy, we conclude that S1P receptors are not early response biomarkers for MI. There are different mechanisms when S1P plays a protection role in heart during MI and IR. The cooperation of lipid content and S1P receptor expression appears to form a regulation network during MI and IR.

Bone marrow stem/progenitor cell mobilization in C57BL/6J and BALB/c mice / 한국실험동물학회지

Bone marrow (BM) has been considered as a reservoir of stem/progenitor cells which are able to differentiate into ectodermal, endodermal, and mesodermal origins in vitro as well as in vivo. Following adequate stimulation, such as granulocyte stimulating factor (G-CSF) or AMD3100, BM resident stem/progenitor cells (BMSPCs) can be mobilized to peripheral blood. Several host-related factors are known to participate in this mobilization process. In fact, a significant number of donors are resistant to G-CSF induced mobilization protocols. AMD3100 is currently used in combination with G-CSF. However, information regarding host-related factors which may influence the AMD3100 directed mobilization is extremely limited. In this study, we were to get some more knowledge on the host-related factors that affect the efficiency of AMD3100 induced mobilization by employing in vivo mobilization experiments. As a result, we found that C57BL/6J mice are more sensitive to AMD3100 but less sensitive to G-CSF which promotes the proliferation of BMSPCs. We excluded S1P as one of the host related factor which influences AMD3100 directed mobilization because pre-treatment of S1P receptor antagonist FTY720 did not inhibit BMSPC mobilization. Further in vitro experiments revealed that BALB/c mice, compared to C57BL/6J mice, have less BMSPCs which migrate in response to host related factors such as sphingosine-1-phosphate (S1P) and to CXCL12. We conclude that AMD3100-directed mobilization depends on the number of BMSPCs rather than on the host-related factors. These results suggest that the combination of AMD3100 and G-CSF is co-operative and is optimal for the mobilization of BMSPCs.

[ effect of resistance training on plasma and skeletal muscles sphingosine-1-phosphate levels of male Wistar rat]

Sphingosine 1-phosphate [S1P] is a bioactive platelet-derived sphingolipid that is involved in regulation of proliferation, differentiation, hypertrophy and anti-apoptosis of cells and activation of satellite cells. The purpose of present study was to examine the effect of resistance training on S1P levels of plasma and skeletal muscles in male Wistar rats. Twenty four 8-week-old male Wistar rats were used in this study. The initial body weight of rats was 190 to 250 gr. All animals were maintained in pairs in an environmentally controlled room at 22°C, 12:12-h photoperiod cycle and allowed normal cage activity. The animals were fed standard rat chow and water ad libitum. After a week of acclimation to the animal facility, the rats were assigned randomly to a control [N=12] or training [N=12] group. Resistance training was done using a 1 meter height ladder with 2 cm grid with an 85 degree incline, and weights attached to rat's tails. The content of sphingosine-1- phosphate [S1P] present in the chloroform layer was determined by means of high performance liquid chromatography [HPLC]. Resistance exercise training increased the total content of S1P in FHL [fast-twitch] [P=0.003] and soleus [slow-twitch] [P=0.008] muscles and plasma [P=0.001] in comparison with control group. It is concluded that resistance exercise training strongly affects the S1P content in fast and slow twitch muscles and plasma

Senescent endothelial dysfunctions were mediated by S1P2 receptor in cultured human umbilical vein endothelial cells / 中南大学学报(医学版)

OBJECTIVE@#To investigate the variation of senescent endothelial function by regulating the sphingosine-1-phosphate receptor type 2 (S1P2) expression in cultured human umbilical vein endothelial cells (HUVECs).@*METHODS@#The S1P2 receptor expression was regulated by transfecting the cDNA or shRNA of S1P2 in cultured HUVECs. The expression levels of S1P2 receptor in HUVECs were detected by RT-PCR and Western blot. EC chemotaxis was measured by the transwell migration assay. The wound healing assay was performed by a scratch wound model on EC monolayer. Matrigel morphogenesis assay was employed to assess the in vitro angiogenic responses.@*RESULTS@#After up-regulating the S1P2 expression in young ECs, the S1P-stimulated formation of a tubular-like network in Matrigel was dramatically diminished in transfected ECs (P<0.05). Quantification of the wound healing assay showed that transfected ECs grew much slower than young ECs (P<0.05). The chemotactic capability was significantly decreased in transfected ECs (P<0.05). Furthermore, the senescent-associated impairments were revoked by the downregulation of S1P2 receptor in senescent HUVECs.@*CONCLUSION@#The impaired functions (chemotactic, wound-healing and morphogenetic responses) in senescent HUVECs in vitro are mediated by S1P2 receptor.

Research progress of the selective sphingosine-1-phosphate receptor 1 agonists / 药学学报

Sphingosine-1-phosphate (S1P) is a lysophospholipid signaling molecule that regulates important biological functions in both intracellular and extracellular compartments. It interacts with five G protein-coupled receptors subtypes (S1PR(1-5)) to generate multiple downstream signaling. Activation of S1PR1 has been validated to be involved in the process of immune modulation. Fingolimod (FTY720), the novel S1PR1 agonist, has been approved for the treatment of multiple sclerosis in clinical trials. The study towards discovery of selective S1PR1 agonists has become hot spot for immunological diseases. This article summarized the research progress of S1PR1 agonists, emphasizing their structure types, structure-activity relationship and direction of development.

Immunosuppressive effect of S1P1 receptor agonist FTY720 / 药学学报

FTY720 is a synthetic compound derived from the metabolites of Isaria sinclairii. Its unique chemical structure and mechanism appear to be distinctive from other known immunosuppressors. In the present study, the effect of FTY720 on immunosuppression and toxicity to heart was evaluated by detection of lymphocytes count, heart rate in rats, the survival time of the allografts of skin slice in mice and binding to S1P1 and S1P3 receptors by confocal. The results showed that FTY720 could induce lymphopenia, reduce the heart rates in rats and prolong the survival time of the allografts of skin slice in mice. The assay results on confocal showed that FTY720 can bind with S1P1 and S1P3 on surface of CHO-S1P1 and CHO-S1P3 cells. FTY720 could be developed for wide application for organ transplantation and self-immunity diseases.

Influence of NK cell S1PR5 expression on graft versus host disease in allogeneic hematopoietic stem cell transplantation / 中国实验血液学杂志

Natural killer (NK) cells can suppress the development of graft vs host disease (GVHD) while retaining antitumor response in allogeneic hematopoietic stem cell transplantation (allo-HSCT). Sphingosine-1-phosphate receptor 5 (S1PR5) can regulate NK cell migration and distribution in vivo by interacting with sphingosine-1-phosphate (S1P). This study was aimed to investigate S1PR5 expression change of NK cells in allo-HSCT and to explore the relationship between S1PR5 change and frequency of acute/chronic graft-versus-host disease (aGVHD/cGVHD). The S1PR5 expression was detected by real time quantitative PCR in the RNA extracted from blood NK cells of 17 couples of donor and recipient one month after allo-HSCT. The results showed that S1PR5 mRNA level variations in NK cells of donors and recipients post-allo-HSCT were not statistically significant (0.235 ± 0.191 vs 0.330 ± 0.261, P > 0.05). S1PR5 expression of NK cells was significantly lower in patients with aGVHD than those in patient without aGVHD (0.973 ± 0.834 vs 6.166 ± 5.32, P < 0.05). Compared with the corresponding donor, S1PR5 expression levels of patient declined by more than 10 that caused the high incidence of aGVHD. No significant correlation was found between S1PR5 expression of NK cells and cGVHD (3.401 ± 2.324 vs 2.762 ± 1.972, P > 0.05). It is concluded that the decreased expression level of NK cells S1PR5 is associated with aGVHD occurrence. Possible mechanism is due to S1PR5 low expression affecting distribution of NK cells in vivo, so affecting the regulation of NK cells for aGVHD.

Effect of sphingosine 1-phosphate/sphingosine 1-phosphate receptor signal pathway on function of neutrophils / 中国实验血液学杂志

The aim of this study was to examine the priming effect of sphingosine 1-phosphate (S1P) on fMLP-activated neutrophils, mainly to detect the neutrophil respiratory burst products, and to investigate the signaling pathway involved in S1P activity. Flow cytometry was used to evaluate the new isolated neutrophil; the superoxide anion output was detected indirectly by cytochrome C reduction in respiratory burst; the dihydro-rhodamine 123 was used to detect the intensity of respiratory burst; the signal transduction pathways of neutrophil respiratory burst were explored by Western blot. The results showed that after pretreated with S1P, the level of superoxide anion released by fMLP-activated neutrophils significantly increased; the Rhodamine 123 mean fluorescence intensity in S1P primed fMLP-activated neutrophils group was significantly higher than that in fMLP treatment group; PI3K and Akt proteins involved in the signal pathway of neutrophil respiratory burst. It is concluded that S1P is a new priming reagent, which primes respiratory burst of fMLP-activated neutrophils; this signal pathway may be that S1P interacts with its receptor, activates PI3K, then activates Akt-transmitting signals through NADPH oxidase, finally results in the respiratory burst.