Adrenomedullin inhibits the endothelin production induced by urotensin II in rat vascular smooth muscle cells

The aim of this study was to observe the effects of adrenomedullin (ADM) on endothelin (ET) production induced by urotensin Ⅱ (UⅡ) in rat vascular smooth muscle cells (VSMCs). Cultured VSMCs which were incubated with UⅡ (10^-8 mol/L) and with various concentrations of ADM were used to measure the VSMCs ³H-TdR incorpora- tion, the activity of extracellular signal-regulated kinase (ERK), the amount of ET mRNA and ET production in VSMCs. In this work we found that incubation with UⅡ(10^-8 mol/L) increased obviously the amount of ET mRNA in VSMCs and ET production in medium, however, coincubation with ADM (10^-10—10^-8 mol/L) and UⅡ(10^-8 mol/L) reduced the amount of ET mRNA by 15%, 24% and 45% (P< 0.01) respectively, compared with UⅡ alone. The content of ET in medium was 14.13, 11.38 and 11.00 pg/mL. ADM alone (10^-8 mol/L) had no effect on ET production in VSMCs. UⅡ (10^-8 mol/L) promoted the 3H-TdR incorpo- ration and activity of ERK in VSMCs. ADM inhibited VSMCs 3H-TdR incorporation and activation of ERK in a concentration-dependent manner. Compared with UⅡ group, after coincubation with ADM (10^-10—10^-8 mol/L) and UⅡ (10^-8 mol/L) the VSMCs 3H-TdR incorporation was decreased by 7% (P > 0.05), 32% (P < 0.05) and 41% (P < 0.01), respectively, and the activity of ERK was decreased by 24% (P > 0.05), 32% (P < 0.05) and 36% (P < 0.05), re- spectively, in a concentration-dependent manner. The results show that in cultured VSMCs ADM inhibits ET mRNA expression, ET production and proliferation stimulated by UⅡ, and that inhibitory effect of ADM on UⅡ bioaction could be mediated through inhibiting MAPK pathway.     

Regulation of angiotensinⅡon Gαq/11 protein of vascular smooth muscle cell and its underlying mechanism

To study the regulation of angiotensin Ⅱ(Ang Ⅱ) on Gαq/11 protein of vascular smooth muscle cell (VSMC) and its underlying mechanism, the protein synthesis was detected by [³H]-leucine incorporation. Gαq/11 expression was measured by Western blot in cultured VSMC of rat aorta. The results showed that the level of Gαq/11 was downregulated after stimulated by AngⅡ for 1—6 h, while it was upregulated significantly by 12—24 h stimulation (P < 0.01) in VSMC. The [³H]-leucine incorporation of VSMC was increased after 24 h Ang Ⅱ stimulation. The biphase regulation of Ang Ⅱ on Gαq/11 protein was blocked by the Ang Ⅱ type Ⅰ receptor (AT1) specific antagnist losartan or PLC inhibitor U73122, while PD98059 did not have this effect. These data indicated that Ang Ⅱ contributed to VSMC hypertrophy by regulating the level of Gαq/11, and this effect was mediated mainly through AT1 receptor-PLC signal transduction pathway.     

Expression, purification and characterization of the soluble CuA domain of cytochrome c oxidase ofParacoccus versutus

The key subunit Ⅱ of cytochrome c oxidase (CcO) contains a soluble binuclear copper center (Cu_A) domain. The Cu_A domain of Paracoccus versutus was cloned, expressed, purified and characterized. The gene encoding the Cu_A domain in pET11d vector was expressed in E. coli BL21 (DE3). The results showed that the Cu_A domain was expressed mostly in inclusion bodies and the Cu_A domain protein synthesized in E. coli cells represents approximately 10 percent of the total cellular proteins. Dissolved in urea, dialyzed and recombined with Cu⁺/Cu²⁺ and purified by the Q-sepharose fast flow anion-exchange column and Sephadex G-75 gel filtration column, the soluble purple-colored protein, which shows a single band in electrophoresis, was obtained. The UV-visible absorption spectrum of Cu_A domain showed that there are intense band at 478 nm and a shoulder peak at 530 nm, and two weak bands at 360 and 806 nm respectively, which can be assigned to the charge transfer and the interactions of obitals of Cu—S and Cu——Cu in the mixed-valence binuclear metal center (Cu₂S₂R₂). The far-UV CD spectrum indicated that this domain is predominantly in β-sheet structure. The fluorescence spectra showed that its maximal excitation wavelength and maximal emission wavelength are at 280 and 345 nm, respectively.     

Synthesis mechanism of LiNiVO4 by wet chemical method and mixed valence of nickel and vanadium

The single phase LiNiVO₄ has been successfully synthesized by adopting a new mild liquid route with oxalic acid as both complexant and precipitant, and this method is named the CPG method. The products were obtained by sintering the dry gel precursor which was prepared by the CPG method at 200—850℃ for 2—10 h in air. The products were tested by XRD, XPS, ESR and TGA-DTA, and the results indicate that the single phase LiNiVO₄ could be obtained at 450℃ for 2—3 h in air and LiNiVO₄ was still steady at 850℃ for 10 h. The valence analyses show that in LiNiVO₄ the valence of lithium is +1, both nickel and vanadium have the mixed valence, namely +2, +3 for nickel and +4, +5 for vanadium respectively. The LiNiVO₄ can be expressed as LiNi³⁺_xNi²⁺_1-xV⁴⁺_xV⁵⁺_1-xO₄ (0≤x<1). The pyrolysis mechanism of the dry gel is also discussed.     

Adrenomedullin inhibits the endothelin production in-duced by urotensin Ⅱ in rat vascular smooth muscle cells

The aim of this study was to observe the effects of adrenomedullin (ADM) on endothelin (ET) production induced by urotensin Ⅱ (UⅡ) in rat vascular smooth muscle cells (VSMCs). Cultured VSMCs which were incubated with UⅡ (10-8 mol/L) and with various concentrations of ADM were used to measure the VSMCs 3H-TdR incorpora-tion, the activity of extracellular signal-regulated kinase(ERK), the amount of ET mRNA and ET production inVSMCs. In this work we found that incubation with UⅡ(10-8 mol/L) increased obviously the amount of ET mRNA inVSMCs and ET production in medium, however,co-incubation with ADM (10-10-10-8 mol/L) and UⅡ(10-8mol/L) reduced the amount of ET mRNA by 15%, 24% and45% (P< 0.01) respectively, compared with UⅡ alone. Thecontent of ET in medium was 14.13, 11.38 and 11.00 pg/mL. ADM alone (10-8 mol/L) had no effect on ET production inVSMCs. UⅡ (10-8 mol/L) promoted the 3H-TdR incorpo-ration and activity of ERK in VSMCs. ADM inhibited VSMCs 3H-TdR incorporation and activation of ERK in aconcentration-dependent manner. Compared with UⅡgroup, after co-incubation with ADM (10-10-10-8 mol/L)and UⅡ (10-8 mol/L) the VSMCs 3H-TdR incorporation wasdecreased by 7% (P > 0.05), 32% (P < 0.05) and 41% (P <0.01), respectively, and the activity of ERK was decreasedby 24% (P > 0.05), 32% (P < 0.05) and 36% (P < 0.05), re-spectively, in a concentration-dependent manner. The resultsshow that in cultured VSMCs ADM inhibits ET mRNA ex-pression, ET production and proliferation stimulated by UⅡ, and that inhibitory effect of ADM on UⅡ bioaction could be mediated through inhibiting MAPK pathway.     

Synthesis and structure of Cu5(BTA)6(TTA)4 · 5DMF, a copper-nitrogen cluster containing η Π3-benzotriazolate

The title cluster compound—Cu₅(BTA)₆(TTA)₄·5DMF was prepared using thenyltrifluoroacetone and benzotriazolate ligands. The crystal structure indicates that a tetrahedral array of Cu(1), Cu(2), Cu(3) and Cu(4) ions surrounding a central Cu(5) ion are held together by bridging tridentate BTA^- and terminated by TTA^- bond cap. The three nitrogen atoms of a BTA^- bond three different copper ions to form a η³-benzotriazolate. The central Cu ion has a distorted octahedral structure and the surrounding Cu ions are 5 coordinated forming distorted tetragonal structures. The distances between the surrounding Cu(Ⅱ) ions and the central Cu(Ⅱ) ion are in the range of 0.3561—0.3755 nm and those between the surrounding Cu(Ⅱ) ions are in the range of 0.5785—0.6301 nm.     

Research situation and progress of non-equilibrium plasma chemistry

Generally, the non-equilibrium plasma is produced at low pressure by a glow discharge (1.33 Pa—1.33 kPa) including the radio frequency (13.56 MHz), microwave (2450 MHz), AC or DC high voltage discharges. As a method to directly apply energy to a reaction system, some successful applications have been obtained in the fields such as chemical synthesis and decomposition at plasma, sputtering and filming, deposition at the gas state, polymerization, modification on the material surface, etching, ashing at low temperature and so on. For example, in 1999, Zhang et al.^[1] got a high conversion rate of 98.2% for CH₄ synthesis by a glow discharge at the condition of 850℃ with the catalyst of Ni/a-Al₂O₃. In 1990, Matsumoto et al.^[2] reduced the concentration of CO₂ from 10% to 8% by DC corona torch discharge with Ar catalyst at 0.08 kPa. In 1988, Boenig et al.^[3] synthesized rare N₂H₄ and H₂O₂ with a glow discharge, in which N₂H₄ is a kind of pureness fuel and propellant of rocket. Tanaka et al.^[4] investigated the synthesis of ammonia from nitrogen-hydrogen plasmas in radiofrequency and microwave discharges with catalysts of iron and molybdenum wires of 100 pieces at 620 K and 650 Pa. After 2-h synthesis, the amounts of synthesized NH₃ and N₂H₄ were only 1.5 mmol/g and 2.5 mol/g, respectively. Because it is only at low pressure that electrons can obtain enough energy to carry out chemical reaction, the vacuum system is necessary, which results in the complicated technology and low production efficiency.......     

Iron fractions in the apoplast of intact root tips of Zea mays L. seedlings affected by nitrogen form

The effects of ammonium (NH₄⁺-N ) and nitrate ( NO₃^--N ) were examined on Fe fractions and FeCN (ferricyanide) reductase activity in intact root tips (0—3 cm) of young maize (Zea mays L. cv. Lenz) in solution culture by using short-term experiment under controlled Fe deficiency conditions (containing high HCO₃^- concentration in preculture solution). The results showed that Fe(Ⅱ) concentrations in root tip apoplast of maize were only 20—40 nmol/g FW which accounted for 7%—13% of total Fe. Most of Fe in root tips existed as Fe(Ⅲ) compounds. Imposition of the roots to NH₄⁺-N or NO₃^--N for 60 min led to an increase of Fe(Ⅱ) in root tip apoplast. NH₄⁺-N led to an increased concentration of Fe(Ⅱ) and exchangeable Fe (Fe(Ⅱ) and Fe (Ⅲ)) in root tips, while NO₃^--N increased FeCN reductase activity. The relationship between pH and Fe fractions, FeCN reductase activity was also discussed.     

An explanation to the high efficiency ofm-THPC (temporfin) used in photodynamic therapy

Meso-tetrahydroxylphenyl chlorin (m-THPC) is one of the most efficient prospective sensitizers used in photodynamic therapy (PDT). ESR spectroscopy, fluorescence quenching experiments and cyclic voltammogram measurement were used to study its redox properties. The results showed that the ability of m-THPC generating superoxide radical anions was very strong, and the rate constant of m-THPC fluorescence quenching by oxygen k_q (O₂)=1.46×10¹⁰ mol^-1·s^-1. The values of fluorescence quen- ching rate constant of m-THPC by some other electron acceptors, such as methyl viologen (MV²⁺) and anthraquinone (An), were also measured. And they were k_q (MV²⁺)=5.51×10⁹ mol^-1·s^-1, k_q (An)=7.81×10⁹ mol^-1·s^-1. The oxidation potential of m-THPC was examined to be +0.62 V (vs. NHE) in acetonitrile. All these suggested that m-THPC should be a much stronger electron donor than photofrin, the currently used in clinical photodrug, and may react easily through electron transfer with biological matter to yield various radicals. So it seemed reasonable that the type Ⅰ reaction may play an important role in the high activity of m-THPC-PDT.     

Sources of cumulus expansion enabling factor (CEEF) in porcine follicles

It was shown that expansion of porcine cumulus did not depend on oocyte-secreted factor(s), and it is therefore presumed that porcine CEEF may not be produced exclusively by the oocyte. In this experiment, we used mouse oocytectomized complexes (OOX), which were incapable of CEEF production, to assess the secretion of CEEF by evacuated zona, oocytes of different quality and somatic cells in the porcine follicles. The results showed that: (ⅰ) Evacuated zonae from both porcine and mouse oocytes did not produce CEEF. (ⅱ) Porcine oocytes of A, B and C types from 3—6 mm follicles were not significantly different in both production and activity of CEEF. (ⅲ) Both porcine OOX from 3—6 mm follicles and granulose cells from < 1 mm follicles secreted CEEF in a large quantity, independent of gonadotropins; mural granulose cells from 3—6 mm follicles, however, produced neglectable amount of CEEF. (ⅳ) The follicular fluid from 3—6 mm porcine follicles contained CEEF activity that was concentration-dependent, and thus it enabled cumulus expansion in 60% mouse OOX when used at 10% of concentration, but the expansion rate of mouse OOX decreased to 9% when the concentration was increased to 50%. (ⅴ) Mouse OOX cultured in porcine CEEF-containing M199 expanded only in the presence of gonadotropins, suggesting that the activity of porcine CEEF is hormone-dependent.     

Urotensin Ⅱ increases endothelin production by vascular smooth muscle cells in rats

The aim of this study was to observe the effects of urotensin Ⅱ (UII) on production of endothelin (ET) in rat aortic vascular smooth muscle cells (VSMC). Cultured VSMCs incubated with various concentrations of UII were used to measure the VSMC 3H-TdR incorporation, the amount of ET mRNA and ET production in VSMCs. In this work we found that UII (10-10-10-8 mol/L) promoted VSMC 3H-TdR incorporation (47%-83%, P < 0.01) and increased the amount of ET mRNA by 17.1% (P < 0.05) to 112.8% (P < 0.01), respectively, in a concentration dependent manner compared with control. After 4 and 8 h incubation, 10-10-10-8 mol/L of UII elevated the ET synthesis and release in a concentration dependent manner. After 4 h incubation, the content of ET in medium was 4.9, 5.36 and 7.12 pg/mL (P < 0.01). After 8 h incubation, the ET content released from VSMCs was 12.6, 12.07 and 17.17 pg/mL (P < 0.01). In addition, it was found that BQ123, a specific ETA receptor antagonist, obviously decreased the VSMC DNA synthesis induced by UII. The results of this study showed that UII could stimulate the ET mRNA expression and ET production in VSMC. The effects of UII on VSMC DNA synthesis were partly mediated by ET autocrine pathway. It suggests that the interaction between UII and ET plays an important biological regulating role as endogenous active peptides.     

Gene product expression of cyclin D<Subscript>2</Subscript> and p16 during the transition from cardiac myocyte hyperplasia to hypertrophy

The current study was to investigate mRNA expression of cyclin D₂ and p16 during the transition from cardiac myocyte hyperplasia to hypertrophy. Cultured cardiac myocytes (CM) and fibroblasts (FC) obtained from 1-day-old Sparague-Dawley rats were used in this study. We have determined (1) hyperplasia by cell growth curve and fluorescence activated cell sorting (FACS); and (2) ultrastructure by electron microscope observation; and (3) expresions of cyclin D₂ mRNA and p16 mRNA by using in situ hybridization and image analysis. The results were shown (1) Results of cell growth curve and FACS analysis showed CM could proliferate in the first 3 cultured days (4 days in postnatal development). But the ability decreased quickly, concomitant with the differentiation. (2) The ultrastructure of CM showed the large amount of myofilaments and mitochondrion and FC showed moderate amount of rough endoplasmic reticulum. (3) The expression of cyclin D₂ mRNA in 3−, 4−, 5−day CM group was 0.89 times (p<0.05), 0.80 times (p<0.05) and 0.56 times (p<0.01) of that in 1-day group respectively. P16 mRNA in 2−, 3−, 4−, 5−day CM group were 1.63 times (p<0.01), 1.72 times (p<0.01), 1.99 times (p<0.01) and 2.84 times (p<0.01) of that in 1−day group respectively. It can be concluded that cultured neonatal rat cardiac myocytes could proliferate during the first 3 cultured days, but the ability of proliferation decreased, from the fourth day, concomitant with differentiation. Cyclin D₂ and p16 have the key roles during the transition from myocyte hyperplasia to hypertrophy. Biography: Zhang Yu-xia (1974-), female, Master, research direction: cardiovascular pathology.     

Efficient transfer and expression of human clotting factor IX cDNA in neonatal hemophilia B mice mediated by VSV-G pseudotyped retrovirus

The feasibility of in vivo gene therapy for hemophilia B by VSV-G pseudotyped retroviral vector was introduced. The novel packaging cell line 293GPG was used to produce VSV-G/G1NaBAIX pseudotyped virus with the highest titers up to 8.5×10⁸ cfu·mL^-1. In contrast to the conventional retrovirus, VSV-G pseudotyped virus was more resistant to inactivation by serum complements (P＜0.001). Our results also demonstrated that VSV-G pseudotyped virus was more stable in neonatal mice serum than in adult mice serum (P＜0.01). After intraperitoneal injection of different doses of virus, hFIX antigen was detected and lasted for more than 120 d, the highest level reached (72.5±6.1) ng·mL^-1. Moreover, the functional activity was improved to some extent in all hFIX-treated mice, the most remarkable improvement was observed in the mice treated with higher dose of virus whose clotting activity increased to (3.4±1.5)% and APTT (activated partial thromboplastin time) reduced to (43.2±7.2) s. The anti-hFIX antibody was not detected by the method of Bethesda, no germ line transmission and any side effects associated with gene transfer were found. Our results indicated that neonatal gene therapy for hemophilia B mice by VSV-G pseudotyped retrovirus is promising.     

Cardiac protective role of a novel erythrocyte-derived depressing factor on rats and its Ca^2＋ mechanism

The cardiac protective role of a novel erythro-cyte-derived depressing factor (EDDF) on spontaneous hy-pertensive rats (SHR), calcium overload (CaO) rats and Wistar rats and its mechanism was evaluated. Mean artery pressure (MAP), heart rate (HR) and LVdp/dtmax were measured by physiological recorder. The effect of EDDF on the Ca2+-ATPase activity in myocardial sarcoplasmic reticu-lum (SR) of CaO rats was determined by inorganic phos-phate assay. Calcium transport in myocytes was measured by 45Ca2+ radioactive isotope measurement. The phosphoryla-tion levels of extracellular signal-regulated protein kinases (ERK1/2) in myocardial tissue of SHR and CaO rats were measured by Western blot method. And the ultrastructures of cardiac muscle cells were observed with the transmission electron microscope. The results indicated that EDDF could significantly decrease MAP, HR and LVdp/dtmax in a dose dependent manner (P < 0.05). It seems that the mechanism might relate with activating the Ca2+-APTase, enhancing the uptake and release of Ca2+ from SR (P < 0.05), decreasing the phosphorylation levels of ERK1/2 of myocytes (P < 0.01) and lightening the ultrastructural lesion of cardiac muscle cells. In CaO rats, the Ca2+-ATPase activity decreased clearly com-pared to control (64.99 7.16 vs 94.48 7.68 nmol·min-1 ·mg-1 protein, P < 0.01), while EDDF (100 mg/mL) could significantly increase the activity (87.93 ?9.54 vs 64.99 ?7.16, P < 0.05, n = 7). Both uptake and release rate of Ca2+ (祄ol 45Ca2+/g protein/min) from myocardial SR of CaO rats re-markably decreased compared to control (32.40 ?2.70 and 15.46 ?1.49 vs 61.09 ?10.89 and 25.47 ?4.29, P < 0.05); EDDF (100 mg/mL) could significantly stimulate their activi-ties (50.48 6.76 and 21.76 2.75 vs 32.40 2.70 and 15.46 1.49, P < 0.05). EDDF could evidently down-regulate the phosphorylation of ERK1/2 in myocardial tissue from SHR and CaO rats (P < 0.01), lighten the ultrastructural lesion of cardiac muscle cells of SHR as well. It is concluded that EDDF seems to play protective roles on both structure and function of heart, which closely related with amelioration of Ca2+ transport and inhibition of Ca2+-MAP kinase pathway.     

Foliar δ^13C and δ^15N values of C3 plants in the Ethiopia Rift Valley and their environmental controls

The foliar C and N stable isotopic compositions （δ^13C and δ^15N） and the relationships between these compositions and environmental factors of C3 plants in the Ethiopia Rift Valley were investigated. There were three distribution patterns for foliar δ^13C with mean values of -26.7‰±0.4‰, -29.7‰ ±0.6‰ , and -26.9‰± 1.2‰ in cold-moist, temperate-moist, and arid-hot environments, respectively. The δ^15N values ranged from -1.4‰ ±1.7‰ to 14.3‰ ± 0.1‰, with higher values under arid-hot conditions and the lowest values in plants growing at higher altitudes under cold-moist conditions. A strong negative relationship between mean annual precipitation and δ^15N explained more than half of the observed variation in the δ^15N values （r2= 0.54, P 〈 0.001）; a modest positive relationship was also found between δ^15N and temperature （r2 = 0.32, P 〈 0.01）. A weakly positive relationship existed between δ^13C and temperature, and changes in δ^13C values with precipitation and altitude followed quadratic curves. This suggests a shift in the effects of water and heat conditions caused by altitude on carbon isotopic discrimination.     

Compression behavior and equation of state of Ni77P23 amorphous alloy

The compression behavior of Ni77P23 amorphous alloy is investigated at room temperature in a diamond-anvil cell instrument using insitu high pressure energy dispersive X-ray diffraction with a syn- chrotron radiation source. The equation of state is determined by fitting the experimental data accord- ing to Birch-Murnaghan equation: -ΔV/V0=0.08606P-3.2×10-4P2 5.7×10-6P3. It is found that the structure of Ni77P23 amorphous alloy is stable under pressures up to 30.5 GPa.     

Sexual difference in gonadal development of embryonic chickens after treatment of polychlorinated biphenyls

Polychlorinated biphenyls (PCBs) are ubiquitous, persistent and lipophilic environmental endocrine disrupters that can be biomagnified in human and animals including birds and thus to affect reproductive functions. Poultry meat and eggs contain a great deal of fat and possibly concentrate higher PCBs and other environmental contaminants such as dichlorodiphenyl-trichloroethane (DDT). In order to identify the adverse effects of PCBs and possible sexual difference on chicken gonadal development, Aroclor 1254 was injected into fertilized Hyline chicken eggs before incubation. Four groups of eggs received yolk injection of either peanut oil as control, 1, 10 or 100 mg/egg of Aroclor 1254 as treatment group, respectively. After hatching the gonads were removed for histological examination. Testicular structure was apparently changed and exhibited dramatic decrease in area of the transverse sections (1 mg/egg, P＜0.01; 10 and 100 mg/egg, P＜0.001), diameter (10 and 100 mg/egg, P＜0.05) and relative area of the testicular tubules (10 mg/egg, P＜0.05; 100 mg/egg, P＜0.01). Most testicular tubules of the highest dose group degenerated and even disappeared. The differentiation of germ cells was retarded in all groups treated with Aroclor 1254. Some germ cells were irregular in shape, with vacuolated cytoplasm and hyperchromatism nucleus (pyknosis) in 1 and 10 mg/egg groups and almost all germ cells in 100 mg/egg group. In contrast, the area of the left ovarian transverse sections increased dramatically (10 and 100 mg/egg,P＜0.001), the ovarian cortex manifested a significant increment in thickness (1, 10 and 100 mg/egg, P＜0.001) and a higher number of oocytes was observed (1, 10 and 100 mg/egg, P＜0.001) in the female chickens treated with Aroclor 1254 compared with the control group. A few oocytes with vacuolated cytoplasm and hyperchromatic nucleus were also observed in ovarian cortex after PCBs exposure. These results showed that PCBs interfered with gonadal development with a sex-specific pattern and retarded differentiation of spermatogonia into spermatocytes but accelerated differentiation of oogonia into oocytes in the embryonic chickens.     

Action mechanisms of a new erythrocyte-derived depressing factor

To investigate the action mechanisms of a new erythrocyte-derived depressing factor (EDDF), the focus is placed on the effect of EDDF on both cytosolic and nuclear free calcium (Ca²⁺) transportation in vascular smooth muscle cell (VSMC), as well as the apoptosis and cell cycle of VSMC of rats. EDDF has been extracted from human erythrocytes. The changes of Ca²⁺ levels in cytoplasm ([Ca²⁺]_i) and nucleus ([Ca²⁺]_n) have been observed using a laser scanning confocal microscope together with fluo-3/AM as a calcium indicator. Flow cytometric technique was used to study the effect of EDDF on cell cycle and apoptosis of VSMC. [Ca²⁺], and [Ca²⁺]_n were significantly decreased through several different pathways: ( i ) it reduced the Ca²⁺ influx by blocking L-type voltage-dependent calcium channel (L-VDC) and R-type voltage-dependent calcium channel (R-VDC); (ii) it inhibited the Ca²⁺ release from inositol 1, 4, 5-trisphosphate (IP₃) sensitive calcium store; and (iii) activated Ca²⁺-ATPase of sarcoplasmic reticulum (SR) and promoted the transportation of Ca²⁺ from cytoplasm to SR. However, EDDF seemed to have little inhibitory effect on the Ca²⁺ release from ryonodine sensitive calcium pool. It was also found that EDDF (10⁻⁴ g/mL) significantly decreased the proportion of S phase of human umbilical vein (HUV) and inhibited the proliferation of VSMC induced by angiotensin II (Angll, 10⁻⁵ mol/L). The apopotosis did not occur when VSMC was cultured under normal condition. While VSMC apoptosis was induced by Angll (10⁻⁵ mol/L) and EDDF (10⁻⁴ g/mL) seemed to have little effect on it. The inhibitory effect of EDDF on the elevation of [Ca²⁺]_i and [Ca²⁺]_n of VSMC might play an essential role in its action mechanisms and the ways it affects the Ca²⁺ handling of VSMC demonstrate that EDDF was different from other endogenous blood pressure regulators and some known antihypertensive drugs. EDDF could inhibit the proliferation of VSMC, which indicated that it might be beneficial to the prevention and treatment of hypertension and arteriosclerosis.