OBJECTIVETo investigate whether the cardioprotective effect of hemin against ischemia/reperfusion (I/R) injury is through the inhibition of calpain activity, and to explore its underlying mechanism.

METHODSSixty-four SD rats were randomly divided into eight groups (n = 8): sham, I/R, MDL+ I/R, MDL, hemin + I/R, hemin, and ZnPP + hemin+ I/R, ZnPP. Iangendorff isolated rat heart perfusion model was used. The rat hearts were suffered from 40 min of ischemia followed by 30 min of reperfusion. After that, left ventricular developed pressure (LVDP) was recorded. Infarct size and release of lactate dehydrogenase (LDH) were measured. Calpain, heme oxygenase (HO), and caspase 3 activities were evaluated. Expression of calpastatin protein was detected by Western blot.

RESULTS(1) After suffered from ischemia/reperfusion, the calpain activity and caspase 3 activity increased. MDL28170, an inhibitor of calpain, prevented ischemia/reperfusion induced increases in LDH and infarct size, improved the LVDP recovery. (2) Compared with ischema/reperfusion rat hearts, pretreatment of hemin enhanced the HO-1 activity, decreased the calpain and caspase 3 activities, declined LDH release and infarct size, and improved LVDP recovery. (3) Ischemia/reperfusion reduced the expression of calpastatin protein in rat hearts, which was inhibited by hemin pretreatment. And HO-1 inhibitor could abolish the cardioprotection of hemin.

CONCLUSIONCardioprotective effect of hemin against ischemia/reperfusion injury is through the inhibition of calpain activity, the mechanism might be involved in the increase in calpastatin protein expression.

Animals ; Calpain ; metabolism ; Cardiotonic Agents ; pharmacology ; Caspase 3 ; metabolism ; Heme Oxygenase-1 ; metabolism ; Hemin ; pharmacology ; L-Lactate Dehydrogenase ; metabolism ; Myocardial Reperfusion Injury ; drug therapy ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo determine whether U50488H, a selective agonist of kappa-opioid receptor, could induce biphasic (early and late) cardioprotection against myocardial ischemia/reperfusion injury and to explore the underlying mechanisms.

METHODSIsolated perfused rat hearts were subjected to 30 min of ischemia followed by 120 min reperfusion and the cardiac function was evaluated.

RESULTLeft ventricular end-diastolic pressure (LVEDP), left ventricular developed pressure (LVDP) and maximal velocity of contraction and relaxation (+/-dP/dtmax) were improved when U50488H was administered 1 or 24 h before ischemia (P<0.05). Myocardial infarct size, activities of creatine kinase (CK) and lactate dehydrogenase (LDH) in the coronary effluent were lower in the U50488H pretreatment group than those in the control group. Administration of a selective cyclooxygenase-2 (COX-2) inhibitor, celecoxib abolished the late phase of cardioprotection produced by administration of U50488H 24 h before ischemia. Activities of CK and LDH in the coronary effluent were higher in U50488H and celecoxib co-pretreatment group than those in U50488H group. However, administration of celecoxib did not block the early phase of cardioprotection by 1 h treatment of U50488H before ischemia.

CONCLUSIONThe late (but not the early) phase of cardioprotection induced by kappa-opioid receptor agonist might be mediated by COX-2.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer ; pharmacology ; Animals ; Cardiotonic Agents ; pharmacology ; Creatine Kinase ; metabolism ; Cyclooxygenase 2 ; physiology ; In Vitro Techniques ; Ischemic Preconditioning, Myocardial ; L-Lactate Dehydrogenase ; metabolism ; Male ; Myocardial Infarction ; enzymology ; pathology ; Myocardial Reperfusion Injury ; prevention & control ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; agonists

To approach the effect of CCAAT/enhancer binding proteins (C/EBPs) on un-terminal differentiation of HL-60 cells after treatment with Arsenic Trioxide ( As_2O_3) . Methods The changes of cell morphology were observed by Wright staining, the alteration in the cell proliferation was determined by WST1 experiment and the NBT reduction assay was used to detect the differentiation condition of cells, determination and analysis cell cycle. The expressions of C/EBP? and C/EBP? mRNA in HL-60 cells exposed to ATRA and As_2O_3 were assayed by semi-quantitative RT-PCR. Results It was found that ATRA could up- regulate the mRNA expression of C/EBP? obviously, but down-regulate the mRNA expression of C/EBP?. As_2O_3 could up-regulate the mRNA expression of C/EBP? lightly, down-regulate the expression of C/EBP?. Conclusion Both of ATRA and As_2O_3 can down-regulate the mRNA expression of C/EBP?,but there is no significant difference between these two groups,ATRA and As2O3 can up- regulate the mRNA expression of C/EBP?, with significant differences (P

OBJECTIVETo investigate the reversal effect of dihydromyricetin(DMY) on drug resistance of K562/A02 cells to adriamycin and explore its possible mechanism.

METHODSK562 and K562/A02 cells were treated with DMY (5, 10, 20, 40, 60, 80 and 100 mg/L) and ADM (100-0.05 mg/L) for 48 h. The viability of K562 cells and K562/A02 cells was tested and the reversal effect of DMY on drug resistance of K562/A02 cells to adriamycin was analyzed by MTT. The relative concentration of ADM in cells was measured by flow cytometry. Protein expressions of drug resistance related genes including P-glycoprotein (P-gp), multidrug resistance associated protein 1 (MRP1), glutathione transferase π (GSTπ) and BCL-2 were measured by Western Blot.

RESULTSThe proliferation of K562 and K562/A02 cells was significantly decreased by DMY in dose-dependent manner as compared with control group (r1=0.37, r2=0.38). The ICof ADM on K562 and K562/A02 cells were 71.23±6.51 and 72.88±5.49 mg/L respectively. DMY (5, 10 and 20 mg/L) was low cytotoxicity. DMY (5, 10 and 20 mg/L) enhanced the sensitivity of K562/A02 cells to ADM in dose-dependent manner (r1=-0.62, r2=-0.71) and the reversal multiples was from 1.38 to 28.591. The relative concentrations of ADM in K562/A02 of DMY (5, 10 and 20 mg/L) group cells were significantly increased in dose-dependent manner compared with the control group (r=0.34). Compared with the control group, the expressions of drug resistance related protein P-gp, MRP1, GSTπ and BCL-2 were significantly decreased in dose-dependent manner in DMY (5, 10 and 20 mg/L) group (r1=-0.41, r2=-0.37, r3=-0.58, r=-0.46). Compared with the ADM group, the protein expressions of drug resistance related genes P-gp, MRP1, GSTπ and BCL-2 in DMY (5, 10 and 20 mg/L)+ADM group were significantly decreased in dose-dependent manner (r1=-0.55, r2=-0.41, r3 =-0.38, r4=-0.44).

CONCLUSIONDMY enhances the sensitivity of K562/A02 cells to ADM, its mechanism may be related with decrease of P-gp, MRP1, GSTπ and BCL-2 expressions.

OBJECTIVE: To explore the appropriate procedures for preparing extracellular microvesicles (MV) derived from human mesenchymal stem cells (MSC). METHODS: Human MSCs from umbilical cords were cultured in a serum-free medium and maintained in a basal medium for 72 hours after the cell confluence reached to 80%. The supernatants of cultured cells were collected and MVs were enriched. MVs were identified by flow cytometry and electron microscopy. The total protein amount in MVs was used as a parameter for the content of MVs. The supernatants were adjusted to different pH values, and the output of MVs was detected. The supernatants were also collected for enriching the MV and detecting the protein content of MV after the cells were maintained in the basic medium for different time. RESULTS: Flow cytometric analysis showed that the MVs expressed CD9, CD63 and CD81, morphologically presented round under an electron microscope and the diameter of MV was around 100 nm. After enrichment of MV, the protein content of MVs in the supernatants was 416.8±128.1, 255.4±77.9 and 142.8±46.4 μg per 10 MSC，respectively at pH of supernatant 3, 7 and 9 (P＜0.05). The protein content of the supernatants per 10 MSC was 173.6±44.5, 262.4±49.6 and 364.2±37.8 μg respectively after starvation culture for 48, 72 and 96 hrs (P＜0.05). CONCLUSION MVs can be readily collected after MSCs were starved for 96 hours, and the pH of the supernatants is adjusted at 3.0.
Cell-Derived Microparticles ; Cells, Cultured ; Flow Cytometry ; Humans ; Mesenchymal Stem Cells ; Umbilical Cord