Taking advantage of the special Hg2+-promoted deprotection reaction of dithioacetals and the well-known coumarin chromophore, novel reactive fluorescent probes (C3 and C4) towards mercury ions were designed and synthesized. Upon the addition of mercury ions, probe C3 displayed remarkable fluorescence quenching, which was beneficial to the production of a high signal output during sensing process. Differently, compound C4 displayed dramatic red-shift in both fluorescence (60 nm) and absorption (60 nm) spectra, with the detection limit of 90 nM. By virtue of this unique chemical reaction, compound C4 had selective response towards Hg2+ over other metal ions and only the addition of Hg2+ could lead to apparent emission and absorption spectra changes. Moreover, C4 was successfully applied to microscopic imaging for the detection of Hg2+ in HeLa cells with ratiometric fluorescent methods.

Chorionic NAD-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) plays a pivotal role in controlling the amount of prostaglandins in the uterus and has been implicated in the process of labor. Prior studies identified hydrogen sulfide-generating enzymes cystathionine-beta-synthetase (CBS) and cystathionine-gamma-lyase (CSE) in fetal membranes. We investigated whether hydrogen sulfide is involved in the regulation of PGDH expression in the chorion during Labor. The chorionic tissues were obtained from pregnant women at preterm in Labor and at term in labor or not in Labor at term. Levels of CSE and CBS and hydrogen sulfide production rate were down-regulated in term in Labor and preterm in labor groups compared with not in labor at term group. The CBS level correlated to PGDH expression in the chorion. Hydrogen sulfide donor NaHS and precursor L-cysteine dose-dependently stimulated PGDH expression and activity in cultured chorionic trophoblasts. The effect of L-cysteine was blocked by CBS inhibitor and CBS siRNA but not by CSE inhibitor and CSE siRNA. Hydrogen sulfide treatment suppressed miR-26b and miR-199a expression in chorionic trophoblasts. miR-26b and miR-199a mimics blocked hydrogen sulfide upregulation of PGDH expression. Our results indicate that hydrogen sulfide plays pivotal roles in maintenance of PGDH expression in the chorion during human pregnancy. Reduced expression of hydrogen sulfide-generating enzymes contributes to an increased amount of prostaglandins in the uterus during Labor.

A new benzothiazole-derived fluorescence probe 2-(4-azidobenzyl)oxy-3-(benzo[d]thiazol-2-yl)-5-methylbenzaldehyde (HBTA) was synthesized from 3-(benzo[d] thiazol-2-yl)-2-hydroxy-5-methylbenzaldehyde, and the structure of HBTA was characterized. The recognition behaviors of HBTA to H2S were investigated and the results show that HBTA exhibits good selectivity and sensitivity to H2S with fast response and good anti-interference ability. The probe can be applied to detect H2S in a broad pH range, and the detection limit of HBTA for H2S was estimated to be 9.09x10(-7) mol.L (-1). Cell imaging studies reveal that HBTA is capable of detect H2S in live cells.

Recent evidence suggests that uterine activation for labor is associated with inflammation within uterine tissues. Hydrogen sulfide (H2S) plays a critical role in inflammatory responses in various tissues. Our previous study has shown that human myometrium produces H2S via its generating enzymes cystathionine-γ-lyase (CSE) and cystathionine-β-synthetase (CBS) during pregnancy. We therefore explored whether H2S plays a role in the maintenance of uterine quiescence during pregnancy. Human myometrial biopsies were obtained from pregnant women at term. Uterine smooth muscle cells (UMSCs) isolated from myometrial tissues were treated with various reagents including H2S. The protein expression of CSE, CBS and contraction-associated proteins (CAPs) including connexin 43, oxytocin receptor and prostaglandin F2αreceptor determined by Western blot. The levels of cytokines were measured by ELISA. The results showed that CSE and CBS expression inversely correlated to the levels of CAPs and activated NF-κB in pregnant myometrial tissues. H2S inhibited the expression of CAPs, NF-κB activation and the production of interleukin (IL)-1β, IL-6 and tumor necrosis factor α (TNFα) in cultured USMCs. IL-1β treatment reversed H2S inhibition of CAPs. Knockdown of CSE and CBS prevented H2S suppression of inflammation. H2S modulation of inflammation is through KATPchannels and phosphoinositide 3-kinase (PI3K) and extracellular signal-regulated kinase (ERK) signaling pathways. H2S activation of PI3K and ERK signaling is dependent on KATPchannels. Our data suggest that H2S suppresses the expression of CAPs via inhibition of inflammation in myometrium. Endogenous H2S is one of the key factors in maintenance of uterine quiescence during pregnancy.

A new benzothizole-based fluorescent probe 1 for Hg2+ recognition utilizing "ESIPT+AIE" strategy has been developed. In THE/H2O (1:1, v/v, PBS 20 mM, pH = 8.5) mixed solution, probe 1 displays rapid fluorescence responses to Hg2+ ions with high selectivity and sensitivity through Hg2+-triggered releasing of a compound possessing "ESIPT+AIE" characteristics. Cell imaging investigations indicate that probe I is cell permeable with low toxicity to MCF-7 cells, and applicable to detect Hg2+ ions in living MCF-7 cells. (C) 2017 Elsevier Ltd. All rights reserved.

A fluorescent probe (DBT) based on conjugation of 2,3-dimethylbenzo[d] thiazol-3-ium iodide to 3-(benzo[d] thiazol-2-yl)-2-hydroxy-5-methylbenzaldehyde for the detection of sulfur dioxide derivatives(HSO3/SO32) has been developed. DBT exhibits highly selective colorimetric and ratiometric fluorescence dual response to HSO3- with a fluorescence detection limit of 3.50 x 10(-7) M. The HSO3 - sensing mechanism was confirmed to undergo nucleophilic addition of HSO3- to alkene double bond of DBT as evidenced by H-1 NMR and HRMS analysis. The probe is MCF-7 cell permeable and applicable for fluorescence imaging of exogenous and intrinsically generated intracellular SO2 derivatives in the living cells. Fluorescence co-localization studies reveal that DBT is a mitochondria-targetable fluorescent probe. Thus, DBT has potential application for exploring the role played by SO2 derivatives in biology. (C) 2017 Elsevier B.V. All rights reserved.

Increased production of soluble fms-like tyrosine kinase-1 (sFlt-1) from placenta is one of the major contributors to the development of preeclampsia. Our previous study has shown that hydrogen sulfide (H2S) inhibits sFlt-1 release in placenta. In the present study, we sought to investigate whether endogenous H2S affects sFlt-1 production and elucidate which H2S-producing enzyme is responsible for its effect in placenta. It was found that, besides cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE), 3-mercaptopyruvatesulfurtransferase (3-MST) was identified in human placenta and mainly localized in syncytiotrophoblasts. There was no significant difference in expression level of 3-MST among preeclamptic and normal placentas. Treatment of cultured syncytiotrophoblasts with NaHS and l-cysteine suppressed sFlt-1 mRNA expression and caused a decrease in sFlt-1 protein content in culture media of the cells. Transfection of syncytiotrophoblasts with CBS siRNA and CSE siRNA reversed the above effects of l-cysteine. Furthermore, NaHS and l-cysteine treatment decreased the half-life of sFlt-1 mRNA and increased the expression of miR-133b targeting sFlt-1. MiR-133b expression was downregulated in preeclamptic placentas and correlated with the level of CBS and CSE. These results indicate that H2S is an important regulatory factor in sFlt-1 production in placenta. Reduced H2S generation in placenta contributes to development of preeclampsia by enhancing sFlt-1 production.

Hydrogen sulfide (H2S) has been implicated to angiogenesis in various tissues. We sought to investigate the role of hydrogen sulfide (H2S) in regulating production of vascular endothelial growth factor (VEGF) proteins, the key factors of angiogenesis and vasculogenesis, in placenta.Methods: Placental tissues were obtained from pregnant women with preeclampsia and healthy pregnant women who underwent elective cesarean section. Explants and trophoblasts were isolated from healthy placentas and treated with H2S donor and precursor. Western blotting was used to determine the levels of cystathionine beta-synthase (CBS) and cystathionine gamma-lyase (CSE). The levels of VEGF mRNA, miR miR-200c,-20a and -20b were determined by quantitative real time PCR.Results: NaHS and L-cysteine increased VEGF but not placenta growth factor (PIGF) production in cultured explants and trophoblasts. Transfection of CBS and CSE siRNA reversed the stimulatory effect of L-cysteine on VEGF production in placental cells. H2S prolonged the half-life of VEGF mRNA and decreased the expression of miR-200c,-20a and -20b in placental cells. MiR-200c mimic and inhibitor affected VEGF mRNA and protein level, whereas miR-20a or -20b mimic and inhibitor affect VEGF protein release but not mRNA expression. The expression level of miR-200c,-20a and -20b as well as the level of CBS, CSE and VEGF were downregulated in preeclamptic placentas.Conclusion: H2S produced via CSE and CBS plays a critical role in VEGF production in human placenta. Reduced expression of CSE and CBS may contribute to the abnormal production of angiogenic factors in preeclamptic placenta. (C) 2016 Elsevier Ltd. All rights reserved.

A new reactive probe, 1-(benzo[d]thiazol-2-yl)naphthalen-2-yl-picolinate (BTNP), was designed and synthesized. BTNP acts as a highly selective probe to Cu(2+) in DMSO/H2O (7/3, v/v, Tris-HCl 10mM, pH=7.4) solution based on Cu(2+) catalyzed hydrolysis of the picolinate ester moiety in BTNP, which leads to the formation of an ESIPT active product with dual wavelength emission enhancement. The probe also possesses the advantages of simple synthesis, rapid response and high sensitivity. The pseudo-first-order reaction rate constant was calculated to be 0.205min(-1). Moreover, application of BTNP to Cu(2+) detection in living cells and real water samples was also explored.

Placental amino acid transporters and peroxisome proliferator-activated receptors (PPARs) have been implicated to placental development and therefore regulation of fetal growth. We analyzed the correlation between the expression of amino acid transporters and PPARs and investigated whether PPARs control the expression of amino acid transporters in placentas. It was found that protein expression of PPARγ and L-type amino acid transporter 1(LAT1) and 2 (LAT2) was decreased in small-for-gestational-age (SGA) placentas. LAT1, LAT2 and taurine transporter (TAUT) expression correlated to PPARγ level and birth weight. In cultured placental cells, PPARγ agonist stimulated LAT1 and LAT2 and TAUT, which was reversed by PPARγ siRNA. PPARγ up-regulation of LAT1 and TAUT was through specificity protein 1 (Sp-1) while stimulation of LAT2 expression was via induction of gene transcription. Our data suggest that PPARγ, SP-1, LAT1 and LAT2 in placentas are involved in control of fetal growth. PPARγ signaling pathway may be the therapeutic target for intrauterine growth restriction.

Chorionic NAD-dependent 15-hydroxy prostaglandin dehydrogenase (PGDH) plays a pivotal role in controlling the amount of prostaglandins in the uterus. Peroxisome proliferator-activated receptors (PPARs) are implicated to be involved in parturition. In this study, we investigated whether PPARs are involved in control of PGDH expression in chorion. The chorionic tissues were collected from the following groups of the women with singleton pregnancy: term no Labor (TNL), term labor (TL) and preterm labor (PTL). Chorionic trophoblasts were isolated and cultured in vitro. Immunocytochemistry analysis showed that PPAR alpha, PPAR beta, and PPAR gamma were localized to trophoblasts in chorion. The protein levels of PGDH, PPAR beta, and PPAR gamma were localized to trophoblasts in chorion. The protein Levels of PPAR alpha, PPAR beta, and PPAR gamma were reduced in TL tissues compared to that of TNL group. PPAR alpha, PPAR beta, and PPAR gamma expression correlated to PGDH in TNL tissues, whereas only PPAR gamma expression correlated to PGDH in TL chorion tissues. PGDH expression was decreased in PTL tissues compared with TL group, whereas the expression of PPARs was not significantly different between TL and PTL groups. The agonists of three PPARs dose-dependently stimulated PGDH activity, mRNA, and protein expression in cultured chorionic cells. PPARs did not affect the stability of PGDH mRNA but stimulated the transcriptional activity of HPGD gene. Our results suggest that PPARs play pivotal roles in maintenance of PGDH expression in chorion during human pregnancy.

Imbalance release of proangiogenic and antiangiogenic factor from placenta causes maternal endothelial dysfunction which is main manifestation of preeclampsia(PE). Hydrogen sulfide (H2S) “the third endogenous gaseous signaling transmitter”, has been shown to have a pro‐angiogenic effect. We investigated whether H2S modulate the release of proangiogenic and antiangiogenic factors in human placentas. It was found that H2S synthesis enzymes cystathionine β‐synthase(CBS) and cystathionine γ‐lyase(CSE) were expressed in human placenta. The levels of CBS and CSE positively correlated with the levels of agiogenic factor vascular endothelial growth factor(VEGF) and inversely correlated to antiangiogenic factor soluble fms‐like tyrosine kinase 1(sFlt‐1) in placentas. H2S precursor L‐cysteine significantly increased VEGFA release whereas decreased sFlt‐1 release in cultured placental explants andd placental trophoblasts. The effects of H2S precursor could be reversed by siRNA of CBS and CSE. Moreover, H2S decreased miR‐200c which target VEGF and increased miR‐133b which target sFlt‐1. The level of miR‐200c was significantly increased whereas miR‐133b decreased in PE placentas. Our results suggest that H2S is an important factor that maintains the balance of proangiogenic factor VEGFA and antiangiogenic factor sFlt‐1 in placenta. Reduced H2S generation in placentas leads to imbalance release of proangiogenic and antiangiogenic factor from placenta.Grant Funding Source: Natural Science Fundation of China No.81370734

Placental 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) is reduced in pregnancies complicated with preeclampsia (PE). Peroxisome proliferator-activated receptors β/δ (PPARβ/δ) have been shown to suppress 11β-HSD2 expression in human placental cells. Our objectives were to investigate whether the reduced 11β-HSD2 expression is associated with the changes in PPARs in PE placentas, and whether PPARα and PPARγ affect 11β-HSD2 expression in placental cells. PPARα and PPARβ/δ mRNA and protein expression was increased, whereas PPARγ mRNA and protein expression was decreased in PE placentas. 11β-HSD2 protein expression was inversely correlated with PPARβ/δ in normal placentas but correlated positively with PPARγ and inversely to PPARα in PE placentas. In cultured placental cells, PPARα agonist inhibited, whereas PPARγ agonist stimulated, 11β-HSD2 mRNA and protein expression and activity in a dose-dependent manner. Knockdown of retinoid X nuclear receptor α (RXRα) resulted in a loss of PPARγ effect but not PPARα effect on11β-HSD2. The PPARα effect remained, but the PPARγ effect was lost in the presence of the translational inhibitor cycloheximide. PPARγ agonist dose-dependently stimulated specificity protein 1 (Sp-1) protein expression. Inhibition or knockdown of Sp-1 resulted in a loss of the effects of PPARα and PPARγ. The Sp-1 protein level was not correlated with 11β-HSD2 and PPARs in normal placentas, whereas Sp-1 expression was correlated with 11β-HSD2, PPARγ, and PPARβ/δ in PE placentas. Our data indicate that 11β-HSD2 expression can be modulated by PPARα and PPARγ in placental trophoblasts through Sp-1. Decreased 11β-HSD2 expression in PE placenta might be associated with decreased PPARγ but increased PPARα expression.

The chorion laeve controls the levels of active prostaglandins within the uterus by NAD-dependent 15-hydroxy prostaglandin dehydrogenase (PGDH). The expression of PGDH in chorion is modulated by glucocorticoids and progesterone. In this study, we investigated glucocorticoid receptor (GR) and progesterone receptor A and B (PRA and PRB) in the regulation of PGDH expression in chorion, and we determined whether reduced PGDH expression in chorion during labor is associated with the changes in GR and PR expression by real-time RT-PCR and Western blot analysis. Dexamethasone (DEX) inhibited PGDH expression whereas progesterone stimulated PGDH expression in chorionic trophoblasts. DEX suppressed PGDH expression in GR overexpression and PR knockdown cells. The inhibitory effect of DEX did not occur in GR knockdown cells. Progesterone inhibited PGDH in GR overexpression and PR knockdown cells and it stimulated PGDH in PRB overexpression cells whereas it suppressed PGDH in PRA overexpression cells. Knockdown of c-Jun resulted in a loss of progesterone- and DEX-induced effects. PGDH was down-regulated in chorion tissues during labor. PRB was decreased whereas PRA and GR were increased in chorion during labor. Glucocorticoids inhibit PGDH expression via GR in chorionic trophoblasts. Progesterone enhances PGDH expression through PRB, whereas it inhibits PGDH expression via GR and PRA. Decreased PGDH expression is associated with increased GR and PRA, although decreased PRB, in chorion during labor.

There is evidence that gonadal hormones may affect the perception of painful stimulation, although the underlying mechanisms remain unclear. This investigation was undertaken to determine whether the adenosine 5′-triphosphate (ATP) receptor subunit, P2X3, is involved in the modulatory action of estrogen in peripheral pain signal transduction in dorsal root ganglion (DRG). The mechanical pain behavior test, real-time quantitative reverse transcription–polymerase chain reaction analysis, and Western blot methods were used to determine the mean relative concentrations and functions of P2X3 receptors in DRG in sham, ovariectomized (OVX), and estradiol replacement (OVX+E2) female rats and in sham and orchiectomized male rats. The mechanical hyperalgesia appeared after ovariectomy, which was subsequently reversed after estradiol replacement, whereas it was not observed after orchiectomy in male rats. Plantar injection of 2′(3′)-O-(2,4,6-trinitrophenyl) ATP (TNP-ATP), a P2X3 and P2X2/3 receptor antagonist, resulted in an increase of the pain threshold force in OVX rats while had no effect on sham rats. Furthermore, A-317491, a selective P2X3/P2X2/3 receptor antagonist, significantly reversed the hyperalgesia of OVX rats. Injection of ATP into the plantars also caused a significant increase of the paw withdrawal duration in OVX rats compared with that seen in the sham group, which became substantially attenuated by TNP-ATP. P2X3 receptors expressed in DRG were significantly increased in both mRNA and protein levels after ovariectomy and then reversed after estrogen replacement, while a similar increase was not observed after orchiectomy in male rats. Furthermore, P2X3 mRNA was significantly decreased 24 h after the application of 17β-estradiol in a concentration-dependent manner in cultured DRG neurons. ICI 182,780, an estrogen receptor antagonist, blocked the reduction in the protein level. These results suggest that the female gonadal hormone, 17β-estradiol, might participate in the control of peripheral pain signal transduction by modulating P2X3 receptor-mediated events in primary sensory neurons, probably through genomic mechanisms.

Objective. Metabolites profiling provides global insights in characterisations and interactions of metabolites under specific physiological or pathologic states. The aim of this study was to examine serum metabolic changes in bilateral ovariectomised (OVX) rats and those exposed to 17β-oestradiol (E2, 30 μg/kg bw, sc.) or genistein (50 μg/kg bw, sc.) for 12 weeks. Methods. Serum samples were detected by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-q-TOF-MS). Significant variables were found by the orthogonal partial least-squares analysis and identified by matching their tandem MS fragmentation patterns with reference standards. Results. There were 54 variables changed in OVX group compared with sham group. Genistein might induce more intensive changes of metabolites than E2 in OVX rats. Among the 54 significant variables of OVX group, 8 and 20 were found to be affected by E2 and genistein, respectively. Finally, four of them were identified as arachidonic acid (AA), eicosapentaenoic acid (EPA), ergocalciferol and cholecalciferol. Both E2 and genistein could reverse levels of AA, EPA and ergocalciferol to sham levels. Only genistein treatment could reverse the level of cholecalciferol. Conclusions. This study provided a serum profile of ovary dysfunction and showed some similarities and differences between E2 and genistein treatments of the OVX model.

苦参碱(Matrine,MT)、氧化苦参碱(Oxymatrine,OMT)是从传统中药豆科槐属植物苦参(Sophora flavescens Air)根中提取分离出来的主要的生物碱,具有抗肝纤维、保肝、抗乙肝病毒等药理作用[1～3].

Corticotropin-releasing hormone (CRH) has been shown to exhibit various functions in hippocampus. In the present study, we examined the effect of CRH on the expression of serum/glucocorticoid-inducible protein kinase-1 (SGK-1), a novel protein kinase, in primary cultured hippocampal neurons. A dose-dependent increase in mRNA and protein levels of SGK-1 as well as frequency of SGK-1-positive neurons occurred upon exposure to CRH (1 pmol/l to 10 nmol/l). These effects can be reversed by the specific CRH-R1 antagonist antalarmin but not by the CRH-R2 antagonist astressin 2B. Blocking adenylate cyclase (AC) activity with SQ22536 and PKA with H89 completely prevented CRH-induced mRNA and protein expression of SGK-1. Blockage of PLC or PKC did not block CRH-induced SGK-1 expression. Our results suggest that CRH act on CRH-R1 to stimulate SGK-1 mRNA and protein expression in cultured hippocampal neurons via a mechanism that is involved in AC/PKA signaling pathways.