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OBJECTIVE:To explore the effects of extracts from Siegesbeckiae herba (HS) on doxorubicin (DOX)-induced chronic myocardial injury in mice. METHODS:48 mice were randomly divided into 3 groups. Mice in blank control(Con)group received distilled water once every day,ig,and normal saline(2 mL/100 g)once every day,ip,for 8 weeks;mice in DOX mice received distilled water(2 mL/100 g)once every day,ig,and DOX(3 mg/kg)once every week,ip,for 8 weeks;mice in DOX+HS group received HS(340 mg/kg)once every day,ig,and DOX(3 mg/kg)once every week,ip,for 8 weeks. After administra-tion,body mass,heart coefficient,cardiac function changes,serum biochemical index levels [alanine aminotransferase(ALT),as-partate aminotransferase (AST), creatine phosphokinase (CK), creatine kinase isoenzyme (CK-MB), lactate dehydrogenase (LDH)and total cholesterol(TC)] of mice were determined. RESULTS:Compared with Con group,body mass of mice in DOX group was decreased(P<0.01);heart coefficient was increased(P<0.05);heart rate slowed down,R-wave was increased(P<0.05 or P<0.01);serum biochemical indexed were increased,there was significant difference in AST(P<0.01). Compared with DOX group,heart coefficient of mice in DOX+HS group was decreased (P<0.01);heart rate was increased (P<0.01);serum biochemical indexes were decreased,there was significant differences in CK,LDH,TC(P<0.01). CONCLUSIONS:HS has cer-tain protective effects on DOX-induced chronic myocardial injury in mice.
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This paper is to report the exploration of the activation of Rho/ROCK signal pathway in 5-HT-induced proliferation of rat pulmonary artery smooth muscle cells (PASMCs) and the inhibitory effect of m-Nis on this pathway. PASMCs were cultured with the explant technique. MTT assay was used to explore the proliferation of PASMCs after 5-HT treated for different time and the intervening effect of m-Nis. RT-PCR and Western blot were used respectively to explore the mRNA expression of RhoA, ROCK1 and the protein expression of p-MYPT1 in 5-HT-treated PASMCs and intervening effect of m-Nis. The results of MTT assay suggested that 5-HT (1 µmol · L(-1)) treatment for 12-72 h significantly induced the proliferation of rat PASMCs (P<0.05 or P < 0.01), which were inhibited by m-Nis (1 x 10(-5), 1 x 10(-6), l x 10(-7), 1 x10(-8) mol · L(-1)) in dose-dependent manners (P < 0.05 or P < 0.01). Similarly, the mRNA expression of RhoA, ROCK1 and the protein expression of p-MYPT1 were also inhibited by m-Nis in different degrees (P < 0.05 or P < 0.01). Thus, the results of this study suggested that Rho/ROCK pathway played an important role in 5-HT-induced proliferation of rat PASMCs, m-Nis inhibited 5-HT-induced proliferation obviously, which may be related to the blockage of Rho/ROCK signal pathway.
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Aim To explore the changes of Apelin/APJ system in LPS-induced injury of rat pulmonary mi-crovascular endothelial cells( PMVECs) , and the effect and mechanism of Apelin. Methods PMVECs were cultured with the explant technique, and the identifica-tion of rat PMVECs was carried out by immunocyto-chemical staining of factorⅧrelated antigen. MTT as-say was used to evaluate the viability of PMVECs. The mRNA expression of Apelin and APJ was detected by RT-PCR. The protein expression of PCNA and the phosphorylation of Akt was analyzed by Western blot. Results The mRNA expression of Apelin and APJ showed a compensatory increase after LPS treatment for a short period of time ( P<0. 01 ) , but with the exten-sion of time, which was significantly inhibited, even lower than the control group ( P<0. 05 or P<0. 01 ) , suggesting that Apelin/ APJ system might be involved in LPS-induced PMVECs injury. MTT results showed that 10 -6 ~10 -9 mol · L-1 Apelin obviously promoted the proliferation of rat PMVECs ( P <0. 05 or P <0. 01 ) , and with certain concentration and time de-pendence. Moreover, Apelin also improved the LPS-induced PMVECs injury in different degrees ( P<0. 05 or P < 0. 01 ) . In addition, Western blot analysis showed that Apelin significantly reversed the decrease of the protein expression of PCNA and the Akt phos-phorylation level induced by LPS ( P <0. 05 or P <0. 01 ) . Conclusions The Apelin/APJ system is in-volved in LPS-induced PMVECs injury. Apelin plays an important role in protecting the pulmonary microvas-cular endothelial function and reversing the LPS-in-duced PMVECs injury, which might be related to the activation of Akt phosphorylation pathway.
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AIM To investigate effect of dipfluzine on sodium current (INa+) in isolated single guinea-pig ventricular myocytes. METHODS INa+ was measured by whole cell patch-clamp technique in single isolated guinea-pig ventricular myocytes which were prepared by enzymatic dissociation method. RESULTSCardiac INa+ was elicited by 50-ms pulses to +50 mV from holding potential at -80 mV with a step of +10 mV, which could be blocked completely by tetrodotoxin 10 μmol·L-1. The peak INa+ occurred at about -20 mV and the reversal potential for INa+ was about +30 mV. Dipfluzine inhibited cardiac INa+ in a concentration-dependent manner. The blocking effect of dipfluzine on INa+ was reversible. Dipfluzine suppressed cardiac INa+, without modifying maximum activation potential and reversal potential. The peak of INa+ was decreased by about 46% at -20 mV and shape of I-V curve was not altered by dipfluzine 40 μmol·L-1. Dipfluzine shifted the steady-state inactivation curve of INa+ towards more negative without changing the slope factor and produced very little change in the steady-state activation curve towards more positive. The mean half activation voltage was (-34.9±5.1) mV and slope factor was (6.0±4.8) mV under control condition and (-33.7±3.6) mV and (5.6±2.4) mV following exposure to dipfluzine 40 μmol·L-1. The half inactivation voltage was (-73.0±4.6)mV and slope factor was (4.8±1.8)mV under control condition and (-82.8±7.2)mV and (4.8±1.8)mV following dipfluzine 40 μmol·L-1 treatment. Dipfluzine delayed recovery of cardiac INa+ from inactivation state. The time course of recovery was (36±11) ms in control group and (79±28) ms in dipfluzine 40 μmol·L-1 group. CONCLUSION Dipfluzine inhi- bits cardiac INa+ in a concentration-dependent manner and has higher affinity for the inactivated state than that for resting state of Na+ channels.
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Effect of strophanthidin (Str) on intracellular calcium concentration ([Ca2+]i) was investigated on isolated ventricular myocytes of guinea pig. Single ventricular myocytes were obtained by enzymatic dissociation technique. Fluorescent signal of [Ca2+]i was detected with confocal microscopy after incubation of cardiomycytes in Tyrode's solution with Fluo3-AM. The result showed that Str increased [Ca2+]i in a concentration-dependent manner. The ventricular myocytes began to round-up into a contracture state once the peak level of [Ca2+]i was achieved in the presence of Str (10 μmol·L-1), but remained no change in the presence of Str (1 and 100 nmol·L-1). Tetrodotoxin (TTX), nisodipine, and high concentration of extracellular Ca2+ changed the response of cardiomycytes to Str (1 and 100 nmol·L-1), but had no obvious effects on the action of Str (10 μmol·L-1). The elevation of [Ca2+]i caused by Str at all of the detected concentrations was partially antagonized by rynodine (10 μmol·L-1) or the removal of Ca2+ from Tyrode's solution. In Na+, K+-free Tyrode's solution, the response of cardiomycytes in [Ca2+]i elevation to Str (10 μmol·L-1) was attenuated, while remained no change to Str (1 and 100 nmol·L-1). TTX, nisodipine, and high concentration of extracellular Ca2+ changed the response of cardiomycytes to Str at all of the detected concentrations in Na+, K+-free Tyrode's solution. The study suggests that the elevation of [Ca2+]i by Str at the low (nomomolar) concentrations is partially mediated by the extracellular calcium influx through Ca2+ channel or a "slip mode conductance" of TTX sensitive Na+ channel. While the effect of Str at high (micromolar) concentrations was mainly due to the inhibition of Na+, K+-ATPase. Directly triggering the release of intracellular Ca2+ from sarcoplasmic reticulum (SR) by Str may be also involved in the mechanism of [Ca2+]i elevation.
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AIM To investigate whether dipfluzine (Dip) possesses antiarrhythmic effect on experimental arrhythmias and effect on cytosolic calcium in ventricular myocytes of guinea-pig. METHODS Experimental arrhythmias were induced by strophanthin G infusion through jugular vein in guinea-pigs and by myocardial ischemia-reperfusion (I-R) in rats respectively. Cytosolic calcium concentration ([Ca2+]i) of isolated guinea-pig ventricular myocytes was examined with laser confocal scanning microscope. RESULTSIn guinea-pigs pretreatment with Dip 20 mg·kg-1 increased the dosages of strophanthin G required to induce ventricular premature contraction (VP), ventricular tachycardia (VT), ventricular fibrillation (VF) and cardiac arrest (CA), pretreatment with Dip 10 mg·kg-1 increased the dosages of strophanthin G required to induce VP. In the I-R-induced arrhythmic model of rats, Dip 20 mg·kg-1 decreased the number of rats exhibiting VT, VF and CA, and the number of rats exhibiting VF and CA was decreased by Dip 10 mg·kg-1. Both Dip and verapamil (Ver) decreased [Ca2+]i of the ventricular myocytes in normal Tyrode′s solution. The Ca2+ overload evoked by high extracellular Ca2+ levels was inhibited by Dip and Ver, and the prophylactic effect of Dip was less than that of Ver, while the curative effect of Dip was more obvious than that of Ver. CONCLUSION Dip has antiarrhythmic effect, which is likely related to the modulation on the intracellular calcium homeostasis.
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AIM To establish an Endothelin 1 induced focal cerebral ischemia and reperfusion model. METHODS Endothelin 1(ET 1), a potent vasoconstrictor, was injected near the middle cerebral artery to induce reduction in cerebral blood flow and ischemic neuronal damage. The changes of cerebral blood flow in striatum were characterised using hydrogen clearance technique. The neurologic scores were performed and the infarct volume was identified by TTC staining at 6 h and 24 h after ET 1 application, respectively. RESULTS ET 1 induced a dose dependent reduction of cerebral blood flow in striatum and the CBF at 10 min after ET 1 injection were the lowerest. CBF at 10 min post injection was (27 1?2 9)% in group 300 pmol, (12 7?2 1) % in group 360 pmol, (11 9?1 8)% in group 400 pmol and (9 5?1 6)% in group 500 pmol , respectively. Neurologic score showed that ET 1 could induce variable grade neurologic deficit. The infarct volume were increased with the increment of ET 1 concentration and showed a close correlation, which were (3 9?0 3)% in group 300 pmol, (7 4?0 5)% in group 360 pmol, (11 3?1 3)% in group 400 pmol, and (16 2?1 8)% in group 500 pmol respectively, r =0 992 6 ( P
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AIM To study the effects of strophanthidin (Str) on cardiac function and Na +,K +-ATPase activity in isolated guinea pig heart failure model. METHODS Langendorff isolated heart failure models made by perfusing heart with K-H solution containing sodium pentobarbital. Eight-channel physiological recording instrument was used to determine cardiac function. Colorimetry method was used to determine cardiac sarcolemmal Na +,K +-ATPase activity. RESULTS Str increased the heart rate, left ventricular systolic pressure and the maximum rise or decline rate of left ventricular pressure in a concentration-dependent manner at 1?10 -9～1?10 -7 mol?L -1. But Str caused first a rise, then a reduction of contractility and arrhythmia accompanied by inhibition of Na +,K +-ATPase when the concentration of Str was higher than 1?10 -6 mol?L -1. Str had no obvious effect on Na +,K +-ATPase activity at 1?10 -7 mol?L -1, but increased cardiac activity at 1?10 -10～1?10 -8 mol?L -1. CONCLUSION The inotropic effect and heart toxicity of Str at higher concentration is due to inhibition of Na +,K +-ATPase, but the inotropic effect of Str at lower concentration is not the result of inhibition of Na +,K +-ATPase activity.
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AIM To study the effect and mechanism of valdecoxib, a selective COX 2 inhibitor, on human gastric cancer BGC 823 cells. METHODS MTT assay and flow cytometry were used to observe the effect of valdecoxib on proliferation, apoptosis and the cell cycle distribution of BGC 823 cells. Laser confocal microscopy, transmission electron microscope and DNA fragmentation assay were further used to detect the apoptosis. The content of LDH was examined using LDH kit. RESULTS Valdecoxib in 25～400 ?mol?L -1 significantly inhibited the proliferation of BGC 823 cell in a time and dose dependent fashion, the inhibition rate of proliferation was 24 0%～92 0% after 72 h, and the rate of apoptosis was increased from (2 6?0 7)% to (7 6?1 5) %～(16 5?1 5)%. 100～400 ?mol?L -1 valdecoxib also decreased the proliferation index and the proportion of cells in the S phase, increased the proportion of cells in the G 0/G 1 phase, but had no effect on the proportion of cell in the G 2/M phase. CONCLUSION Valdecoxib inhibits human gastric cancer BGC 823 cells growth by inducing apoptosis and cell cycle arrest. The growth inhibitory effect of 400 ?mol?L -1 valdecoxib is also associated with cell necrosis.
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Objective To evaluate the inhibitory effect of valdecoxib on the growth of the cancer cell lines and involvement of COX-2 in this inhibition. Methods Western blotting and immunocytochemistry were used to detect the expression of COX-2. MTT assay was used to determine inhibitory effect of the drugs on the cell growth. The content of PGE_ 2 in cell medium was determined with PGE_ 2 ELISA kit. Results ①Clone 26 cells expressed high levels of COX-2, whereas BGC-823,HGC-27 and SK-OV-3 cell had no COX-2 expression. ②Valdecoxib inhibited the growth of BGC-823, HGC-27, SK-OV-3 and clone 26 cells, with a IC_ 50 of 110.7, 99.2, 113.3, 117.6 ?mol/L, respectively. ③The inhibitory effect of these drugs on BGC-823 and clone 26 cell was in the descending order of valdecoxib, SC-560 and indomethacin. ④PGE_ 2 did not antagonize the effect of valdecoxib, SC-560 and indomethacin on BGC-823 and clone 26 cells. ⑤The inhibitory effect of valdecoxib and indomethacin on the growth of clone 26 cells was not compatible with that on PGE_ 2 . Conclusion The inhibitory effect of valdecoxib on cell growth is not related to its effect on COX-2.