Gossypol acetic acid (GAA) has been shown to have male antifertility effects, but there are pronounced differences among animal species. In the search of endogenous effector molecules, which interfere with the functions of GAA, we have studied the in vitro effect of various amino acids on the inhibition of the purified LDH-X by GAA. Histidine, cysteine and glycine were shown to block the effect of GAA. The effects of these amino acids were concentration dependent. Histidine and glycine protection was found to be complex type in which both the Km and Vmax were decreased compared to control. Arginine, glutamic acid, phenylalanine and valine were found to be ineffective against the inhibitory action of GAA.
Amino Acids/pharmacology* ; Animal ; Enzyme Inhibitors/pharmacology* ; Goats ; Gossypol/pharmacology ; Gossypol/analogs & derivatives* ; Isoenzymes ; Lactate Dehydrogenase/antagonists & inhibitors* ; Male ; Spermatocidal Agents/pharmacology* ; Testis/enzymology

AIMTo study the effect of 3,4-oxo-isopropylidene-shikimic acid (ISA) on H2O2 (200 mol.L-1, 4 h) injured human umbilical vein endothelial cells (HUVEC).

METHODSMorphological change was observed under microscop. Cell viability was assessed by MTT assay. The release of intracellular lactate dehydrogenase (LDH) and NO was assessed by colorimetry. Radioimmunoassay was used to assess 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha).

RESULTSPretreatment with ISA for 6 h alleviated the morphological damage of H2O2 induced HUVECs. At the concentration of 1-100 mumol.L-1, ISA prevented the inhibitory effect on cell viability induced by H2O2 in dose-dependent manner, but increased the ratio of cell viability from 60.4% to 84.3%. ISA reduced LDH release and increased the level of NO and 6-keto-PGF1 alpha in H2O2 induced HUVECs.

CONCLUSIONISA exerted protective effect on H2O2 injured HUVEC.

6-Ketoprostaglandin F1 alpha ; metabolism ; Cell Survival ; drug effects ; Cells, Cultured ; Endothelial Cells ; metabolism ; pathology ; Humans ; Hydrogen Peroxide ; antagonists & inhibitors ; Infant, Newborn ; L-Lactate Dehydrogenase ; metabolism ; Nitric Oxide ; metabolism ; Protective Agents ; pharmacology ; Shikimic Acid ; analogs & derivatives ; pharmacology ; Umbilical Veins ; cytology

OBJECTIVETo investigate the effect of mitochondrial permeability transition pore inhibitor cyclosporine A (CsA) on lipopolysaccharide (LPS)-induced acute lung injury in mice.

METHODSAll male ICR mice were randomly divided into five groups (n = 24): control group, LPS group, dexamethasone group, cyclosporine A(CsA) group and CsA + atractyloside(Atr) group. Six hours after treatment with LPS, the activity of lactate dehydrogenlase (LDH) in bronchoalveolar lavage fluid (BALF) and level of tumor necrosis factor-alpha (TNF-alpha) in lung tissue were detected. The lung wet weight/dry weight ratio and the pulmonary capillary permeability index were also detected.

RESULTSIn contrast to LPS group, the mitochondrial permeability transition pore inhibitor CsA induced a decrease in LDH activity in the BALF and TNF-alpha level in lung tissue, lung wet weight/dry weight ratio and the pulmonary capillary permeability index were declined. Atractyloside, the activator of mitochondrial permeability transition pore, almost abolished the role of CsA on LPS-induced lung injury.

CONCLUSIONThese results suggested that CsA plays the protective effect on LPS-induced lung injury in mice, it is likely through inhibiting the opening of mitochondrial permeability transition pore.

Acute Lung Injury ; chemically induced ; physiopathology ; prevention & control ; Animals ; Cyclosporine ; pharmacology ; L-Lactate Dehydrogenase ; metabolism ; Lipopolysaccharides ; Male ; Mice ; Mice, Inbred ICR ; Mitochondrial Membrane Transport Proteins ; antagonists & inhibitors ; Protective Agents ; pharmacology ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo investigate the role of sodium-calcium exchanger (NCX) on ischemic preconditioning and pharmacological preconditioning.

METHODSCultured rat neonatal cardiomyocytes were randomly divided into 6 groups: (1) ischemia/reperfusion group (9 h ischemia followed by 1 h reperfusion, I/R), (2) ischemic preconditioning group (1.5 h ischemia/1 h reperfusion + I/R), (3) pharmacologic preconditioning group, adenosine (10 micromol/L) pretreated for 1 h + I/R, (4) calmodulin-dependent protein kinase II (CaMKII) inhibitor KN-93 (0.5 micromol/L for 0.5 h) + ischemic preconditioning group, (5) KN-93 + pharmacologic preconditioning group, (6) control group. The leakage of intracellular lactate dehydrogenase (LDH) in various groups was determined by biochemical autoanalyzer. Semi-quantitative RT-PCR was employed to measure the mRNA levels of sodium-calcium exchanger. Activity of sodium-calcium exchanger (Na(+)-dependent (45)Ca(2+) uptake) was measured by liquid scintillation counting.

RESULTS(1) Compared to the I/R group, the LDH leakages in both ischemic preconditioning group and pharmacologic preconditioning group were significantly reduced (P < 0.05) while significantly increased in the KN-93 + pharmacologic preconditioning group and the KN-93 + ischemic preconditioning group (P < 0.05). (2) The Na(+)-dependent (45)Ca(2+) uptake was significantly increased in the I/R group (P < 0.05) compared to control group and this increase could be significantly attenuated in ischemic preconditioning group and adenosine pretreatment group (P < 0.05). (3) The expression of NCX mRNA in I/R group was also significantly increased (P < 0.05) in the I/R group (P < 0.05) compared to control group and this increase could be significantly attenuated in ischemic preconditioning group and adenosine pretreatment group (P < 0.05), CaMKII inhibitor KN-93 significantly abolished these effects in preconditioning group (P < 0.05) and in adenosine pretreated group (P < 0.05).

CONCLUSIONNCX mediated the cardioprotective effects of ischemic preconditioning and pharmacological preconditioning in the neonatal cardiomyocytes I/R model.

Animals ; Calcium ; metabolism ; Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; antagonists & inhibitors ; Cells, Cultured ; Ischemic Preconditioning, Myocardial ; L-Lactate Dehydrogenase ; metabolism ; Myocardial Reperfusion Injury ; metabolism ; therapy ; Myocytes, Cardiac ; metabolism ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Sodium-Calcium Exchanger ; metabolism

AIMTo investigate the effects of NPPB, a chloride channel blocker, on proliferation of mesangial cells.

METHODSCell proliferation was determined by measuring cell number and 3H-thymidine incorporation. The LDH activity released from these cells was measured as evaluation of cell viability. The phase of cell cycle was detected by flow cytometry.

RESULTSCell proliferation assays showed that treatment with both NPPB (50 and 25 micromol x L(-1)) and in hypertonic media (100% increased osmolarity with D-mannitol ) significantly reduced the number of human MC and 3H-thymidine incorporation in a dose-dependent manner. But the LDH activity was not significantly altered in the treatment with 50 micromol x L(-1) NPPB. Flow cytometry experiments showed that 50 and 25 micromol x L(-1) NPPB arrested (84.2 +/- 2.4) % and (80.8 +/- 2.9) % of cells at G0/G1 stage, versus (70.5 +/- 1.4) % of control cells. Conclusion NPPB suppresses cell proliferation and produces growth arrest at G0/G1 phase in human MC by a mechanism probably associated with changes in cell volume.

Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Chloride Channels ; antagonists & inhibitors ; Dose-Response Relationship, Drug ; Humans ; L-Lactate Dehydrogenase ; metabolism ; Mesangial Cells ; cytology ; metabolism ; Nitrobenzoates ; administration & dosage ; pharmacology

AIMTo observe the effect of angiotensin-converting enzyme inhibitors (ACEI) and aldosterone receptor blockers on cardiac function to explore the mechanism of cardiac function descending and myocardial injury in calcium-overload rats.

METHODSCalcium-overload in rat was induced by administration of Vitamin D3 plus nicotine. To Estimate the extent of calcium-overload by calcium content. Angiotension II and aldosterone levels in the myocardia were measured by radioimmunoassay. Cardiac function (+/- LVdp/dt, LVESP and LVEDP) were measured by Powerlab. The malondialdehyde (MDA) content, activities of lactate dehydrogenase (LDH) and creatine kinase (CPK) were measured by biochemistry.

RESULTSCalcium content increased by 3.2-, 5.8 -fold in myocardial and artery, compared with controls. VDN-treated survivors showed lower + LVdp/dt(max) and -LVdp/dt(max) values, by 27% and 34%, respectively (both P < 0.01). Higher LVESP, and LVEDP by 42 % and 32% (P < 0.01); heart rate and mean arterial pressure were not significantly altered (P > 0.05). The lipid peroxidation products MDA and conjugated diene in myocardia were increased 22% (P < 0.01), 68% (P < 0.05) (P < 0.05), respectively. The plasma activity of CPK and LDH was greatly increased by 4.5-and 3.1-fold (P < 0.01), respectively. ACEI and spironolactone obviously relieved degree of calcium-overload and improved cardiac function and myocardial injury(P < 0.01). Calcium content in myocardia and artery was lower 44%, 39% and 57%, 34%. Lower MDA by 20%, 30%, lower conjugated diene by 44%, 35% than calcium-overload group. The plasma activity of CPK and LDH were obviously decreased 28%, 34% and 20%, 27%, compared with calcium-overload group.

CONCLUSIONCalcium-overload could lead to cardiac function descending and myocardial injury in calcium-overload rats by VDN. ACEI and spironolactone could reduce calcium-overload in myocardial and ameliorate cardiac function and decrease myocardial injury.

Angiotensin-Converting Enzyme Inhibitors ; pharmacology ; Animals ; Calcium ; adverse effects ; Creatine Kinase ; metabolism ; L-Lactate Dehydrogenase ; metabolism ; Lipid Peroxidation ; Male ; Malondialdehyde ; analysis ; Mineralocorticoid Receptor Antagonists ; Myocardium ; metabolism ; Nicotine ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Spironolactone ; pharmacology ; Vitamin D ; pharmacology

OBJECTIVE: To investigate the protective effect of a non-specific NMDA receptor antagonist memantine on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. METHODS: Healthy male mice were divided into 4 groups: a normal group, a memantine group, an ALI group and a memantine+ALI group. The ALI group was induced by intraperitoneal injection of LPS (10 mg/kg). Memantine (10 mg/kg) was injected intraperitoneally before the injection of LPS to determine the effect of blockade of NMDA receptor in the memantine+ALI group. The lung wet/dry ratio was detected. HE staining was preformed to show the morphological changes in the lung tissue. Myeloperoxidase enzyme (MPO) activity and malondialdehyde (MDA) content in the lung tissue were detected. ELISA was used to detect the tumor necrosis factor-α (TNF-α) content and lactate dehydrogenase (LDH) activity in the bronchoalveolar lavage fluid (BALF). RESULTS: Memantine pretreatment improved the LPS-induced ALI lung tissue morphological changes, reduced their lung wet/dry ratio, the levels of TNF-α and LDH activity in BALF, and also reduced the MPO and MDA content in the lung tissue. CONCLUSION Blockade of NMDA receptors can ameliorate LPS-induced mice ALI.
Acute Lung Injury ; drug therapy ; Animals ; Bronchoalveolar Lavage Fluid ; L-Lactate Dehydrogenase ; metabolism ; Lipopolysaccharides ; Lung ; metabolism ; Male ; Malondialdehyde ; metabolism ; Memantine ; pharmacology ; Mice ; Peroxidase ; metabolism ; Receptors, N-Methyl-D-Aspartate ; antagonists & inhibitors ; Tumor Necrosis Factor-alpha ; metabolism

AIMTo investigate the cardio-protective effect of crocetin on primary culture of cardiac myocyte treated with noradrenaline.

METHODSAfter adding crocetin, the primary culture of cardiac myocyte was injured by 1.0 micromol x L(-1) noradrenaline. The activity of lactic dehydrogenase (LDH), mitochondrion succinic dehydrogenase (MSDH) and ATPase were assayed. The mitochondrion membrane potential was detected by Rh123. The percentage of cardiac myocyte apoptosis was observed by flow cytometry.

RESULTSCrocetin significantly decreased the activity of LDH in culture supernatant, increased the activity of MSDH, ATPase (Na+-K+ ATPase, Ca2+ ATPase) and mitochondrion membrane potential.

CONCLUSIONCrocetin could alleviate the disturbance of energy metabolism and decrease the percentage of apoptosis of cardiac myocyte treated with noradrenaline.

Animals ; Animals, Newborn ; Apoptosis ; drug effects ; Calcium-Transporting ATPases ; metabolism ; Carotenoids ; isolation & purification ; pharmacology ; Cells, Cultured ; Crocus ; chemistry ; L-Lactate Dehydrogenase ; metabolism ; Membrane Potentials ; drug effects ; Mitochondria ; physiology ; Myocytes, Cardiac ; cytology ; enzymology ; Norepinephrine ; antagonists & inhibitors ; Plants, Medicinal ; chemistry ; Protective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Sodium-Potassium-Exchanging ATPase ; metabolism ; Succinate Dehydrogenase ; metabolism

This study is to investigate the protective effect of (-) clausenamide against the neurotoxicity of okadaic acid in SH-SY5Y cell line, and injection beta-amyloid peptide25-35 (Abeta25-35) to the cerebral ventricle in ovariectomy (OVX) rats. MTT assay, LDH assay, and Hoechst 33258 staining were used to detect the effect of (-) clausenamide on the toxicity of okadaic acid in SH-SY5Y cell line. The animal model was induced by ovariectomized and injection of Abeta25-35 in the cerebroventricle of rats. The effect of (-) clausenamide on learning and memory deficiency was observed by step-through test. Electron microscope, Nissl body staining, and HE staining were used to examine the morphological changes in hippocampus and cerebral cortex neurons. Pretreatment of (-) clausenamide and LiCl decreased the rate of cell death from MTT, LDH release, and apoptosis from Hoechst 33258 staining in SH-SY5Y cell line. The step-through tests showed (-) clausenamide could improve the ability of learning and memory. The Nissl body staining and HE staining experiments also showed the neuroprotective effects of (-) clausenamide on the neurons of hippocampus and cerebral cortex. (-) Clausenamide has the protective effects against the neurotoxicity induced by okadaic acid and Abeta25-35.
Amyloid beta-Peptides ; antagonists & inhibitors ; toxicity ; Animals ; Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Survival ; drug effects ; Cerebral Cortex ; cytology ; Clausena ; chemistry ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Female ; Hippocampus ; cytology ; Humans ; L-Lactate Dehydrogenase ; metabolism ; Lactams ; isolation & purification ; pharmacology ; Learning ; drug effects ; Lignans ; isolation & purification ; pharmacology ; Memory Disorders ; physiopathology ; Neuroblastoma ; metabolism ; pathology ; Neurons ; drug effects ; Neuroprotective Agents ; isolation & purification ; pharmacology ; Okadaic Acid ; antagonists & inhibitors ; toxicity ; Ovariectomy ; Peptide Fragments ; antagonists & inhibitors ; toxicity ; Plants, Medicinal ; chemistry ; Rats ; Rats, Sprague-Dawley

The present study was aimed to investigate whether calreticulin (CRT) was involved in the protective effect of hypoxic preconditioning (HPC) against oxidative stress injury in rat cardiomyocytes. Neonatal cardiomyocytes were prepared from Sprague-Dawley rats aged 24 h, and cultured in DMEM medium containing 10% fetal bovine serum. The cultured cardiomyocytes were randomly divided into 8 groups as follows: (1) hydrogen peroxide stress (H(2)O(2) group); (2) brief hypoxic exposure for 20 min to simulate HPC (HPC group); (3) hypoxic exposure for 20 min followed by normoxic reoxygenation for 24 h before hydrogen peroxide stress (HPC + H(2)O(2) group); (4) SB203580 (a specific inhibitor of p38 MAPK) + HPC + H(2)O(2) group; (5) CRT antisense oligonucleotide transfection (AS group); (6) AS + H(2)O(2) group; (7) AS + HPC + H(2)O(2) group; (8) control group. Morphological observation, lactate dehydrogenase (LDH) leakage and flow cytometry were employed to assess cell apoptosis and necrosis. RT-PCR and Western blot were used to detect CRT expression and activity of p38 MAPK. All experiments were repeated at least four separate times. The results obtained were as follows: (1) HPC relieved cell injury caused by H(2)O(2). Compared with that in H(2)O(2) group, the cell survival rate increased by 18.0% (P<0.05), apoptotic rate and LDH leakage in culture medium decreased by 19.4% and 53.0%, respectively (P<0.05) in HPC + H(2)O(2) group. (2) H(2)O(2) induced CRT over-expression (7.1-fold increase compared with control, P<0.05), while HPC resulted in mild CRT up-regulation (2.4-fold increase compared with control, P<0.05), suggesting that HPC can relieve the over-expression of CRT induced by H(2)O(2). (3) CRT AS transfection weakened the protection of HPC. Compared with that in HPC + H(2)O(2) group, the cell survival rate decreased by 4% (P<0.05), and apoptotic rate and LDH leakage in culture medium increased by 2.6% and 39.0%, respectively (P< 0.05) in AS + HPC + H(2)O(2) group. (4) The protection of HPC and HPC-induced upregulation of CRT were almost eliminated when SB203580 was administered before HPC. These results suggest that HPC up-regulates CRT expression through the p38 MAPK signaling pathway and protects cardiomyocytes from oxidative stress injury.
Animals ; Apoptosis ; Calreticulin ; metabolism ; Cell Hypoxia ; Cell Survival ; Cells, Cultured ; Hydrogen Peroxide ; Imidazoles ; pharmacology ; Ischemic Preconditioning, Myocardial ; L-Lactate Dehydrogenase ; metabolism ; Myocytes, Cardiac ; pathology ; Oxidative Stress ; Pyridines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Up-Regulation ; p38 Mitogen-Activated Protein Kinases ; antagonists & inhibitors