[Show abstract][Hide abstract] ABSTRACT: Fear extinction has been extensively studied, but little is known about the molecular processes that underlie the persistence of extinction long-term memory (LTM). We found that microinfusion of norepinephrine (NE) into the CA1 area of the dorsal hippocampus during the early phase (0 h) post-extinction enhanced extinction LTM 2 and 14 days after extinction. Intra-CA1 infusion of NE during the late phase (12 h) post-extinction selectively promoted extinction LTM 14 days after extinction, which was blocked by the β-receptor antagonist propranolol, protein kinase A (PKA) inhibitor Rp-cAMPS, and protein synthesis inhibitors anisomycin and emetine. The phosphorylation levels of PKA, cyclic adenosine monophosphate response element binding protein (CREB) and GluR1, and the membrane GluR1 level was increased by NE during the late phase post-extinction, which was also blocked by propranolol and Rp-cAMPS. These results suggest that the enhancement of extinction LTM persistence induced by NE requires the activation of the β-receptor/PKA/CREB signaling pathway and membrane GluR1 trafficking. Moreover, extinction increased the phosphorylation levels of Erk1/2, CREB, and GluR1, and the membrane GluR1 level during the late phase, and anisomycin/ emetine alone disrupted the persistence of extinction LTM, indicating that the persistence of extinction LTM requires late-phase protein synthesis in the CA1. Propranolol and Rp-cAMPS did not completely disrupt the persistence of extinction LTM, suggesting that another β-receptor/ PKA-independent mechanism underlies the persistence of extinction LTM. Altogether, our results showed that enhancing hippocampal noradrenergic activity during the late phase after extinction selectively promotes the persistence of extinction LTM.Neuropsychopharmacology accepted article preview online, 19 February 2014; doi:10.1038/npp.2014.42.
Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology 02/2014; · 8.68 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The effects of stress on emotional memory are distinct and depend on the stages of memory. Memory undergoes consolidation and reconsolidation after acquisition and retrieval, respectively. Stress facilitates the consolidation but disrupts the reconsolidation of emotional memory. Previous research on the effects of stress on memory have focused on long-term memory (LTM) formation (tested 24 h later), but the effects of stress on the persistence of LTM (tested at least 1 week later) are unclear. Recent findings indicated that the persistence of LTM requires late-phase protein synthesis in the dorsal hippocampus. The present study investigated the effect of stress (i.e., cold water stress) during the late phase after the acquisition and retrieval of contextual fear memory in rats. We found that stress and corticosterone administration during the late phase (12 h) after acquisition, referred to as late consolidation, selectively enhanced the persistence of LTM, whereas stress during the late phase (12 h) after retrieval, referred to as late reconsolidation, selectively disrupted the restabilized persistence of LTM. Moreover, the effects of stress on the persistence of LTM were blocked by the corticosterone synthesis inhibitor metyrapone, which was administered before stress, suggesting that the glucocorticoid system is involved in the effects of stress on the persistence of LTM. We conclude that stress within a restricted time window after acquisition or retrieval selectively affects the persistence of LTM and depends on the glucocorticoid system.
PLoS ONE 03/2013; 8(3):e59075. · 3.53 Impact Factor
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[Show abstract][Hide abstract] ABSTRACT: To establish a bcr-abl(+) cell line resistance to nilotinib, and to investigate the possible mechanisms of resistance.
K562 cells were treated with gradually increasing concentrations of nilotinib to generate resistance cell line K562-RN. The folder of drug-resistance was evaluated by MTT assay. Cells apoptosis rate was detected by flow cytometry, the mRNA level of bcr-abl fusion gene by FISH, and the expression of apoptosis relative gene mRNA and protein (such as bcr-abl, HO-1, mdr1, Bcl-2 and caspase-3) by RQ-PCR and western blot.
The resistant cell line K562-RN was successfully established, with 2.01 fold resistant to nilotinib compared with K562 cell line \[the IC(50) value of nilotinib to K562 and K562-RN were (12.320 ± 1.720) µmol/L and (24.742 ± 2.310) µmol/L, respectively\]. It also had the cross resistance to adriamycin, homoharringtonine, etoposide and imatinib. Treated with different concentrations of nilotinib, cell apoptosis rate of K562-RN was significantly lower than that of K562 cells. The rate of bcr-abl gene positive cells was 92% in K562-RN by FISH assay. The mRNA and protein levels of bcr-abl, HO-1 and mdr1 expression up-regulated in K562-RN cells, while those of caspase-3 expression down-regulated, being significantly statistical difference when compared with K562 cells (P < 0.05).
Human leukemic cell line resistance to nilotinib, K562-RN is established successfully by gradually increasing concentrations of drug. The mechanisms of resistance in K562-RN is probably associated with increasing expression of bcr-abl, HO-1, mdr1 and decreasing expression of caspase-3 mRNA and protein levels.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 11/2012; 33(11):906-10.
[Show abstract][Hide abstract] ABSTRACT: This study was aimed to investigate the effect of AMN107 (nilotinib) combined with heme oxygenase-1 (HO-1) inhibitor zinc protoporphyrin IX (ZnPPIX) on chronic myeloid leukemia (CML) cells and its mechanism. Proliferative rate of cells treated with AMN107 (10 µmol/L) and ZnPPIX (10 µmol/L) alone or both for different time was observed by MTT and trypan blue methods; the expression of HO-1 in the control group, ZnPPIX (10 µmol/L) group, AMN107 (10 µmol/L) group, AMN107 (10 µmol/L) combined with ZnPPIX (10 µmol/L) group was evaluated by semi-quantitative RT-PCR and Western blot at 48 h. Cell apoptosis was detected by flow cytometry with Annexin V/PI double staining at 48 h. The results showed that the strongest inhibition of cell proliferation was detected in combined group, and in a time-dependent manner; the expression level of HO-1 was lowest in combined group; the cell apoptosis rates were (11.38 ± 0.02)%, (17.44 ± 0.08)%, (39.81 ± 0.07)% and (56.46 ± 0.19)% in the control group, ZnPPIX group, AMN107 group, AMN107 combined with ZnPPIX group at 48 h respectively. It is concluded that the second-generation tyrosine kinase inhibitor AMN107 can induce the apoptosis in CML cells. Inhibition of HO-1 expression can enhance the killing effect of AMN107 on CML cells, which provides experimental evidence to further improve the clinical efficacy of CML treatment.
Zhongguo shi yan xue ye xue za zhi / Zhongguo bing li sheng li xue hui = Journal of experimental hematology / Chinese Association of Pathophysiology 07/2012; 20(4):867-71.
[Show abstract][Hide abstract] ABSTRACT: To investigate the effect of retrovirus mediated heme oxygenase (HO)-1 gene on chronic myeloid leukemia (CML) resistance cell apoptosis induced by nilotinib (AMN107).
High titer viral particles of pQCXIP-EGFP-HO-1 were prepared, and K562/A02 cells stably transfected with HO-1 gene was established. The expression of HO-1 in K562/A02 cells was detected by RT-PCR. After treated with AMN107 for 24 h, HO-1 mRNA and protein expression by RT-PCR and Western blot, respectively; Cell proliferation by MTT assay; bcr-abl fusion gene by RQ-PCR, and the apoptosis and cell cycle by flow cytometry.
Recombinant retrovirus vector was constructed successfully and K562/A02/HO-1 cells were successfully set up. The expression of HO-1 in K562/A02 cells was expressed clearly. After three groups cells treated with AMN107 for 24 h, the expression of HO-1 mRNA and protein was significantly higher in gene-transfected group than in either empty vector or no-transfected group. The difference was statisitically significant (\%P\% < 0.05). The cell proliferation ofs was inhibited, but the cell viability was significantly higher in gene-transfected group than in other two groups. The difference was statistically significant(\%P\% < 0.05); After treated with 10 µmol/L AMN107 for 24 h, the CT values of bcr-abl fusion gene were (18.15 ± 0.18) in K562/A02/HO-1 group, being significantly higher than that in K562/A02/LXSN (20.32 ± 0.20) and K562/A02 (20.51 ± 0.21) group, the difference was statistically significant (\%P\% < 0.05); the apoptosis rate were (17.26 ± 0.23)%, (39.47 ± 0.17)%, and (41.84 ± 0.09)%, respectively in three groups, and were (3.74 ± 0.03)%, (5.91 ± 0.08)% in K563/A02/HO-1 untreated with drug and K562/A02 untreated with drug group. The number of G(0)/G(1) phase and S phase cells markedly decreased. The cells were arrested in G(2)/M phase. But cell cycle in gene-transfected group did not change significantly.
AMN107 inhibits proliferation of CML resistance cells and induces cell apoptosis. HO-1 gene can protect CML resistance cells to apoptosis. There was a relationship between HO-1 gene and the growth of CML resistance cells.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 05/2012; 33(5):383-7.
[Show abstract][Hide abstract] ABSTRACT: Exposure to cues previously associated with drug intake leads to relapse by activating previously acquired memories. Based on previous findings, in which cannabinoid CB(1) receptors were found to be critically involved in specific aspects of learning and memory, we investigated the role of CB(1) receptors in nicotine reward memory using a rat conditioned place preference (CPP) model. In Experiment 1, rats were trained for CPP with alternating injections of nicotine (0.5mg/kg, s.c.) and saline to acquire the nicotine-conditioned memory. To examine the effects of rimonabant on the reconsolidation of nicotine reward memory, rats were administered rimonabant (0, 0.3, and 3.0mg/kg, i.p.) immediately after reexposure to the drug-paired context. In Experiment 2, rats were trained for CPP similarly to Experiment 1. To examine the effects of rimonabant on the reinstatement of nicotine reward memory, rimonabant (0, 0.3, and 3.0mg/kg, i.p.) was administered before the test of nicotine-induced CPP reinstatement. In Experiment 3, to evaluate whether rimonabant itself produces a reward memory, rats were trained for CPP with alternating injections of different doses of rimonabant (0, 0.3, and 3.0mg/kg) and saline. Rimonabant at a dose of 3.0mg/kg significantly disrupted the reconsolidation of nicotine memory and significantly blocked the reinstatement of nicotine-induced CPP. However, rimonabant itself did not produce CPP. These findings provide clear evidence that CB(1) receptors play a role in nicotine reward memory, suggesting that CB(1) receptor antagonists may be a potential target for managing nicotine addiction.
Pharmacology Biochemistry and Behavior 06/2011; 99(4):738-42. · 2.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the effect of heme oxygenase-1 (HO-1) expression on cell growth and apoptosis in imatinib resistant chronic myeloid leukemia (CML) cells (K562/A02-IM), and explore the relationship between HO-1 gene and CML.
The expression of HO-1 in 20 drug-resistant CML patients was detected by RT-PCR. Different concentrations of hemin were used to induce HO-1 expression of K562/A02-IM, HO-1 expression at different time was detected by RT-PCR and Western blot analysis. Cell apoptosis was detected by Annexin V/PI staining, and MTT assay was used to detect viability of K562/A02-IM cells after induction or inhibition of HO-1 gene by hemin and zinc protoporphyrin (ZPP).
RT-PCR showed that HO-1 was expressed in the bone marrow mononuclear cells (BMMNCs). When treated with hemin at different concentrations (0, 10, 20, 40 µmol/L) for 16 h, the expression of HO-1 in K562/A02-IM was increased in a dose-dependent manner, and peaked at 20 µmol/L of hemin for 16 h. The apoptosis rates were (17.61 ± 0.01)%, (12.13 ± 0.11)%, (7.94 ± 0.03)% and (4.62 ± 0.15)% at 0,10, 20 and 40 µmol/L of hemin respectively for 16 h and were (14.7 ± 0.05)%, (8.1 ± 0.07)% and (16.3 ± 0.13)% at 20 µmol/L of hemin treatment for 8,16, and 24 h respectively. Hemin induced apoptosis of K562/A02-IM cells in a dose-dependent manner. The expression of HO-1 was induced in K562/A02-IM cells in a dose-dependent manner, and the survival of K562/A02-IM cells was significantly increased as compared to that of control group. When HO-1 was inhibited by ZPP, the cells survival was sharply decreased compared to that of the control group (P < 0.05).
HO-1 was expressed in the BMMNCs. It is a kind of molecules whose expression can be induced and can promote the growth of drug-resistant cells. Inhibition of HO-1 expression probably be used for the treatment of drug-resistant CML.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 06/2011; 32(6):388-91.
[Show abstract][Hide abstract] ABSTRACT: The intense associative memories that develop between drug-paired contextual cues and rewarding stimuli or the drug withdrawal-associated aversive feeling have been suggested to contribute to the high rate of relapse. Various studies have elucidated the mechanisms underlying the formation and expression of drug-related cue memories, but how this mechanism is maintained is unknown. Protein kinase M ζ (PKMζ) was recently shown to be necessary and sufficient for long-term potentiation maintenance and memory storage. In the present study, we used conditioned place preference (CPP) and aversion (CPA) to examine whether PKMζ maintains both morphine-associated reward memory and morphine withdrawal-associated aversive memory in the basolateral amygdala (BLA). We also investigate the role of PKMζ in the infralimbic cortex in the extinction memory of morphine reward-related cues and morphine withdrawal-related aversive cues. We found that intra-BLA but not central nucleus of the amygdala injection of the selective PKMζ inhibitor ZIP 1 day after CPP and CPA training impaired the expression of CPP and CPA 1 day later, and the effect of ZIP on memory lasted at least 2 weeks. Inhibiting PKMζ activity in the infralimbic cortex, but not prelimbic cortex, disrupted the expression of the extinction memory of CPP and CPA. These results indicate that PKMζ in the BLA is required for the maintenance of associative morphine reward memory and morphine withdrawal-associated aversion memory, and PKMζ in the infralimbic cortex is required for the maintenance of extinction memory of morphine reward-related cues and morphine withdrawal-related aversive cues.
Neuropsychopharmacology: official publication of the American College of Neuropsychopharmacology 06/2011; 36(10):1972-81. · 8.68 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To construct an eukaryotic expression vector containing the aldehyde dehydrogenase-2 (ALDH2) gene, and determine whether transfection with the ALDH2 gene can provide protection against hydrogen peroxide-induced oxidative damage, as well as attenuate apoptosis or cell death in human peripheral blood mononuclear cells (PBMCs).
The ALDH2 gene was cloned from human hepatocytes by RT-PCR. The eukaryotic expression vector containing the gene was constructed and then transfected into PBMCs via liposomes. RT-PCR, indirect immunofluorescence assay, and Western blot were used to evaluate the expression of the transgene in target cells. MTT assay and flow cytometry were used to detect the effects of ALDH2 on PBMCs damaged by hydrogen peroxide (H₂O₂). The level of intracellular reactive oxygen species (ROS) was determined by fluorescence spectrophotometry.
The eukaryotic expression vector pcDNA3.1/myc-His-ALDH2 was successfully constructed and transfected into PBMCs. RT-PCR results showed higher mRNA expression of ALDH2 in the gene-transfected group than in the two control groups (empty vector-transfected group and negative control). Indirect immunofluorescence assay and Western blot indicated distinct higher protein expression of ALDH2 in the gene-transfected group. The cell survival rate against H₂O₂-induced oxidative damage was higher in the ALDH2 gene-transfected group. Moreover, apoptosis rates in gene-transfected PBMCs incubated with 50 and 75 μmol/L H₂O₂ decreased by 7% and 6%, respectively. The generation of intracellular ROS was also markedly downregulated.
ALDH2 gene transfection can protect PBMCs against H₂O₂-induced damage and attenuate apoptosis, accompanied with a downregulation of intracellular ROS. ALDH2 functions as a protector against oxidative stress.
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE: To construct a eukaryotic expression vector containing aldehyde dehydrogenase-2 (ALDH2) gene and investigate the effects and its possible mechanisms of ALDH2 gene on cell proliferation and anti-oxidative damage in the K562 cells. METHODS: An eukaryotic expression vector containing the ALDH2 gene cloned from human hepatocytes was constructed and transfected into K562 cells by liposome. RT-PCR and Western blot were used to evaluate the expression of ALDH2. MTT assay was used to check the cell proliferation and trypan blue exclusion to check K562 cells damage induced by hydrogen peroxide (H2O2). RT-PCR and fluorescence spectrophotometry were used to determine the expression of heme oxygenase-1 (HO-1) and the generation of intracellular reactive oxygen species (ROS) respectively. RESULTS: RT-PCR and Western blot analysis showed distinct higher ALDH2 protein expression in gene transfected group. The latter group had a higher cell proliferation (P < 0.05) and survival rate against H2O2 induced-oxidative damage, being increased by 7.8 times (IC(50) was 12.3 µmol/L and 1.4 µmol/L for K562-pcDNA3.1-ALDH2 and control cells, respectively, P < 0.01). The HO-1 mRNA expression and the generation of intracellular ROS were downregulated at a specific concentration of H2O2 in the ALDH2 gene transfected group. CONCLUSION: ALDH2 gene transfection can protect K562 cells against oxidative damage, and the downregulation of HO-1 expression and intracellular ROS may be involved in this process.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 11/2010; 31(11):721-725.
[Show abstract][Hide abstract] ABSTRACT: Cocaine use and relapse involves learned associations between cocaine-associated environmental contexts and discrete stimuli and cocaine effects. Initially, these contextual and discrete cues undergo memory consolidation after being paired with cocaine exposure. During abstinence, cocaine cue memories can undergo memory reconsolidation after cue exposure without the drug. We used a conditioned place preference (CPP) procedure in rats to study the role of neuronal protein kinase cyclin-dependent kinase 5 (Cdk5) in consolidation and reconsolidation of cocaine cue memories. We found that the expression of cocaine CPP in drug-free tests 1 d after CPP training (four pairings of 10 mg/kg cocaine with one context and four pairings of saline with a different context) increased Cdk5 activity, and levels of the Cdk5 activator p35 in basolateral but not central amygdala. We also found that basolateral (but not central) amygdala injections of the Cdk5 inhibitor beta-butyrolactone (100 ng/side) immediately (but not 6 h) after cocaine-context pairings during training prevented subsequent cocaine CPP expression. After training, acute basolateral (but not central) amygdala beta-butyrolactone injections immediately before testing prevented the expression of cocaine CPP; this effect was also observed on a second test performed 1 d later, suggesting an effect on reconsolidation of cocaine cue memories. In support, basolateral beta-butyrolactone injections, given immediately (but not 6 h) after a single exposure to the cocaine-paired context, prevented cocaine CPP expression 1 and 14 d after the injections. Results indicate that basolateral amygdala Cdk5 activity is critical for consolidation and reconsolidation of the memories of cocaine-associated environmental cues.
Journal of Neuroscience 08/2010; 30(31):10351-9. · 6.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To construct the human cytochrome P-450 CYP2E1 (CYP2E1) recombinant adenovirus vector and detect its anti-tumor effects in combination with chemotherapeutic drugs so as to provide rationales for gene directed enzyme prodrug therapy (GDEPT).
CYP2E1 cDNA was cloned from human liver and a recombinant adenovirus vector constructed at a titer of 1 × 10(12) pfu/ml. Human bone marrow-derived mesenchymal stem cells (BMSCs) were separated, cultured, purified and detected. The tropism of BMSCs for cancer cells was detected by Transwell technique. These recombinant vectors were transferred into BMSCs and A375 cells and the expressions of EGFP and CYP2E1 detected by fluorescence microscope, RT-PCR and Western blot respectively. Inverted microscope, MTT and Annexin V-FITC/PI were employed to detect the anti-tumor effect of CYP2E1 recombinant adenovirus vectors in combination with chemotherapeutic prodrug dacarbazine (DTIC).
The recombinant adenovirus vectors pAd5CMV-NpA-CYP2E1 and pAd5CMV-NpA-EGFP were constructed successfully. BMSCs were successfully separated and they could migrate in vitro through a polycarbonate filter toward K562 and A375 cells in the lower chamber. Fluorescence microscope was used to detect the expression of EGFP while both RT-PCR and Western blot revealed a high expression of CYP2E1 in gene-transfected group cells. The dead cell counts of co-culture group of gene-transfected BMSCs and wild type A375 cells were significantly higher than those of the control under inverted microscope. The results of MTT showed that the growth inhibition of gene-transfected group cells was increased with DTIC concentration in a concentration dependent manner. IC(50) of group BMSCs-CYP2E1 + A375 was (0.17 ± 0.13) mmol/L, and that of the control group was (0.65 ± 0.20) mmol/L (P < 0.01). The DTIC concentration at which BMSCs were relatively safe might be selected at 0.05 mmol/L. Annexin V-FITC/PI confirmed that the apoptosis rate of BMSCs-CYP2E1 + A375 group was significantly higher than that of the control group after a 48-hour treatment of 0.05 mmol/L DTIC (26.8 ± 2.0 vs 8.7 ± 1.3, P < 0.01).
With an in vitro tropism for cancer cells, BMSCs transferred with CYP2E1 recombinant adenovirus vectors have anti-tumor effects probably synergistically with chemotherapeutic drugs.
[Show abstract][Hide abstract] ABSTRACT: To explore bone marrow-derived mesenchymal stem cells (BMSC) mediated gene directed enzyme prodrug targeting anti-tumor therapy (GDEPT).
CYP1A2 gene was cloned from human hepatocytes by RT-PCR, and the eukaryote expression vector was constructed and transferred into Raji cells and human BMSCs via liposome. The targeted anti-tumor effect of BMSC-CYP1A2 cooperated with dacarbazine (DTIC) was measured. RT-PCR and Western blot were used to detect the expression of CYP1A2. Migration assay was detected with Transwell Plates. MTT was used to evaluate the growth inhibitory effect. Cell apoptosis was determined with Annexin V-FITC/PI staining by flow cytometry(FCM).
The results of FCM and differentiation induction were in line with the characteristics of BMSC. The expression of CYP1A2 was confirmed by RT-PCR and Western blot. Growth inhibition of Raji-CYP1A2 cells was increased with DTIC concentration in a dose-dependent manner (IC(50) was 1.67 mmol/L). However, BMSC was less sensitive to the cytotoxic effects of DTIC (IC(50) of 9.26 mmol/L and 7.53 mmol/L for BMSC-pcDNA3.1 and BMSC-CYP1A2 cells, respectively) than Raji cells did (IC(50) of 5.62 mmol/L and 1.67 mmol/L for Raji-pcDNA3.1 and Raji-CYP1A2 cells, respectively). BMSC migrated toward Raji cells in Transwell plate. BMSC-CYP1A2 cells mediated a bystander killing effect for CYP1A2-negative Raji cells when they were co-cultured with BMSC-CYP1A2 cells.
DTIC can be catalyzed by CYP1A2 in vitro. BMSC-based enzyme prodrug system of CYP1A2 and DTIC can induce lymphoma cell apoptosis targetedly via bystander killing effect.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 10/2009; 30(10):667-71.
[Show abstract][Hide abstract] ABSTRACT: This study was aimed to investigate the frequency of FMS-like tyrosine kinase 3 (FLT3) mutations including internal tandem duplication (ITD) mutation of juxtamembrane region and point mutation of the second tyrosine kinase domain (TKD) in acute myeloid leukemia (AML) patients and its clinical significance. The ITD mutation in FLT3 exon 14, 15 of bone marrow mononuclear cells was detected by genomic DNA-PCR, the TKD point mutation in FLT3 exon 20 was detected by genomic DNA-PCR combined with restriction endonuclease digest. The results indicated that among 131 newly diagnosed AML patients, 21 patients (16.0%) showed FLT3-ITD positive, 3 patients (2.3%) showed FLT3-TKD positive. None was found harboring both mutations. The WBC and bone marrow blast counts in FLT3-ITD positive patients seemed both higher than those in patients with wild-type FLT3 (FLT3-wt), but there was significant difference only in WBC count (p<0.05). The complete remission (CR) rate in FLT3-ITD positive patients was 47.6%, which was significantly lower than that in FLT3-wt patients (88.1%, p<0.05). There was no statistical difference in CR rate between FLT3-ITD positive and negative patients in 20 cases of M3; the CR rate in FLT3-ITD positive patients with non M(3) was 37.5 (6/16) which was obviously lower than that in FLT3-wt patients with non M3 (90.6%, 48/53) (p<0.05). 3 FLT3-ITD positive patients with CR relapsed after CR for 14 (2-20) months with relapse rate 50% (3/6) which was higher than that in FLT3-wt patients (29.2%, 14/48). It is concluded that FLT3 mutation is common in AML patients, while FLT3-ITD mutation is more frequent than FLT3-TKD mutation. The AML patients with FLT3-ITD mutation have a poor prognosis, while FLT3-TKD point mutation does not significantly influences prognosis of the patients. Therefore early detection of FLT3 mutation may be important for targeting therapy and evaluating clinical prognosis of AML patients.
Zhongguo shi yan xue ye xue za zhi / Zhongguo bing li sheng li xue hui = Journal of experimental hematology / Chinese Association of Pathophysiology 10/2009; 17(5):1135-9.
[Show abstract][Hide abstract] ABSTRACT: This study was aimed to investigate the status of c-KIT, Fms-like tyrosine kinase 3 (FLT3) and Janus kinase 2 (JAK2) mutations in acute myeloid leukemia (AML) patients with t (8; 21) and to analyze their relation to clinical feature and prognosis. PCR, AS-PCR, restriction and sequencing methods were used respectively to detect the FLT3, JAK 2 and c-KIT mutations in 8 cases of de novo AML with t (8; 21) and 6 cases of relapsed AML with t (8; 21). The results showed that the c-KIT mutation was found in 2 cases out of 14 AML patients with t (8; 21) (14.3%), among them 1 case had c-KIT D816V mutation, the other had c-KIT D816Y mutation. The FLT3-ITD mutation was detect in 1 out of 14 patients (7.1%), but JAK2 mutation could not be detected in all 14 cases. In conclusion, tyrosine kinase mutation relates to AML with t (8; 21), patients with tyrosine kinase mutation may have higher relapse, extramedullary infiltration and poor prognosis. The screening c-KIT, FLT3 mutations may play an important role in evaluating prognosis and guiding treatment of t (8; 21) AML.
Zhongguo shi yan xue ye xue za zhi / Zhongguo bing li sheng li xue hui = Journal of experimental hematology / Chinese Association of Pathophysiology 09/2009; 17(4):866-9.
[Show abstract][Hide abstract] ABSTRACT: Cytokine interferon-alpha (IFN-alpha) is an immunomodulator and neuromodulator, which modulates central nervous system function partially by activating opioid receptors. However, the role that IFN-alpha plays in relapse to drug abuse is still largely unknown. Thus, we studied whether human recombinant INF-alpha (rIFN-alpha) would reinstate morphine-conditioned place preference (CPP) in rats. In Experiment 1, rats were trained for morphine-CPP with 8-day alternate subcutaneous (s.c.) injections of morphine and saline, and the effect of human rIFN-alpha (20 000 IU/5 microl, intracerebroventricularly) on CPP reinstatement was examined after extinction. In Experiment 2, rats underwent morphine (5 mg/kg, s.c.) unconditioned training with 8 daily alternate injections of morphine (5 mg/kg, s.c.) and saline. Then, the effect of human rIFN-alpha (20 000 IU/5 microl, intracerebroventricularly) on reinstatement of CPP was examined after extinction. In Experiment 3, the effect of opioid receptor antagonist naloxone (1 mg/kg, intraperitoneally) on human rIFN-alpha-induced reinstatement of morphine-CPP was investigated. We found that human rIFN-alpha reinstated morphine-CPP in rats trained under morphine conditioning after extinction, but did not affect CPP in rats that underwent unconditioned training. Naloxone significantly inhibited human rIFN-alpha-induced reinstatement of morphine-CPP. These results indicate that IFN-alpha is a stimulus for reinstatement of morphine-CPP by activation of opioid receptors, which extends our understanding on the high comorbidity of heroin relapse and viral infection.
[Show abstract][Hide abstract] ABSTRACT: Chronic lymphocytic leukemia (CLL) is infrequent in Chinese people. Conventional cytogenetic analysis underestimates the frequency of chromosome aberrations in CLL due to the low rate of spontaneous mitoses. The aim of this study was to prospectively explore the frequency of chromosomal abnormalities in Chinese patients with CLL using interphase fluorescence in situ hybridisation (FISH) and probes for 12 centromere, 13q14, 14q32, 17p13, 11q22 and 6q23 on 143 patients with CLL. Molecular cytogenetic aberrations were found in 104 patients (72.7%) and 40 patients (28.0%) with more than two abnormalities. The most frequent abnormality was del(13q14) (47.6%), followed by trisomy 12 (21.7%), 14q32 translocation (19.6%), del(17p13) (12.6%), del(11q22) (11.9%) and del(6q23) (4.9%), respectively. The percentages of patients with aberrations by FISH were 75.4%, 72.3% and 67.7% for Binet stages A, B and C, respectively. In early stage (Binet A), del(13q14) aberration was more frequent than in Binet B and C (61.5% vs. 31.9% and 41.9%) (P=0.021). Patients with advanced stage (Binet C) had more frequent del(17p13) aberration than in Binet A and B (32.3% vs. 9.2% and 4.3%) (P=0.008). It was showed that the frequencies of the chromosomal abnormalities in our study population were similar to the frequencies in Western countries.
[Show abstract][Hide abstract] ABSTRACT: The objective of this study was to investigate the relationship between the expressions of breast cancer resistance protein (BCRP) gene and drug resistance as well as prognosis in adult patients with acute lymphocytic leukemia (ALL). Semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) was used to detect the expression of BCRP gene in 97 adult patients with acute lymphocytic leukemia (ALL) and 30 normal subjects. Beta(2) microglobulin (beta(2)-MG) was used as an internal reference. BCRP/beta(2)-MG ratio >or= 0.48 was defined as BCRP positive. The results showed that the positive ratio of BCRP gene expression in untreated group was 28.7%, in contrast that in refractory and relapsed patients was 51.2%. The first complete remission rates in patients with negative and positive expression were 71.7% and 29.7% respectively. In ALL subtypes BCRP gene expression of L3 type was distinctly higher than that of L(1), L(2) types. In immune types the BCRP gene expression of B-ALL was higher than that of T cell type, especially in mature B cell type with obviously statistical significance (p<0.01). It is concluded that the high expression of BCRP gene may induce clinical drug resistance, and may be an unfavorable factor for prognosis in adult patients with acute lymphocytic leukemia.
Zhongguo shi yan xue ye xue za zhi / Zhongguo bing li sheng li xue hui = Journal of experimental hematology / Chinese Association of Pathophysiology 03/2008; 16(1):31-4.
[Show abstract][Hide abstract] ABSTRACT: To investigate the effect of lipopolysaccharide(LPS) on the expression and activity of Toll-like receptor 4(TLR-4) in mesenchymal stem cells (MSC).
MSCs were harvested from adult rats bone marrow cells by density gradient centrifugation and adhesive culture. The purity of MSC were identified with the cell morphology and osteogenic capacity. The phenotypes were assayed by flow cytometry. Cultured MSCs were treated by LPS with various concentration (1 microg/ml, 10 microg/ml or 100 microg/ml) for 24 hours. The relative expression levels of TLR-4 mRNA were detected by semiquantitative RT-PCR, and costimulatory molecules (CD80, CD86 and MHC-II) expressed on MSC were analyzed by flow cytometry. The levels of TNF-alpha in supernatants were determined by double-antibody sandwich ELISA.
The expression levels of CD80, CD86, MHC-II, TLR-4 mRNA and TNF-alpha in MSC were (9.56 +/- 0.69)%, (22.03 +/- 2.03)%, (2.51 +/- 0.97)%, relative magnitude (0.61 +/- 0.10), (4.97 +/- 2.98) pg/ml, respectively. After incubation with LPS, MSC expressed higher levels of TLR-4 mRNA and costimulatory molecules, and the levels of TNF-alpha were higher than that in untreated group. Among the various concentration of LPS, 10 microg/ml emerged as the most effective in increasing the levels of TLR-4 mRNA (relative magnitude 1.55 +/- 0.02), costimulatory molecules [CD80 (41.70 +/- 2.92)%, CD86 (59.72 +/- 2.00)%, MHC-II (24.56 +/- 2.19)%] and TNF-alpha [(213.12 +/- 69.08) pg/ml] (P < 0.01). The levels of TLR-4 mRNA, costimulatory molecules and TNF-alpha began to decrease when MSC exposed to 100 microg/ml LPS (P < 0.05). Except for the levels of TNF-alpha [(158.05 +/- 28. 05) pg/ml] and MHC-II [(5.62 +/- 2.31)%] (P > 0.05), the levels of CD80, CD86, MHC-II and TLR-4 mRNA were significantly lower than the 10 microg/ml treatment group (P < 0.01).
MSCs are able to express TLR-4 mRNA. LPS could activate the expression of TLR-4 in MSC obviously, but the activity is dependent on the specific concentration.
Zhonghua xue ye xue za zhi = Zhonghua xueyexue zazhi 12/2007; 28(12):828-31.
[Show abstract][Hide abstract] ABSTRACT: The coordinated change of haematopoietic supporting microenvironment in bone marrow (BM) is crucial for innate immunity and inflammation. As the precursors of marrow stroma, BM derived mesenchymal stem cells (MSCs) promote haematopoietic function, but their roles in innate immunity or inflammation have not been investigated. Here we investigated the expression of Toll like receptor 4 (TLR-4) and the effect of lipopolysaccharide (LPS) on its expression in BM MSCs in vitro.
MSCs were harvested from adult rat's BM cells by density gradient centrifugation and adhesive culture. The purity of MSCs were identified with the cell morphological feature and osteogenic capacity, the phenotypes were tested by flow cytometry. Cultured MSCs were treated by LPS (1 microg/ml, 10 microg/ml or 100 microg/ml) for 24 hours. The relative expression levels of TLR-4 mRNA were detected by semiquantitative reverse transcription polymerase chain reaction and costimulatory molecules (CD80, CD86 and MHC-II) expressed on MSCs were analyzed by flow cytometry. The levels of tumor necrosis factor-alpha (TNF-alpha) in supernatants were determined by enzyme linked immunosorbent assay.
After incubation with LPS, MSCs expressed the higher levels of TLR-4 mRNA, costimulatory molecules and TNF-alpha than the untreated group: LPS 10 microg/ml was the most effective (P < 0.01); the levels of TLR-4 mRNA, costimulatory molecules and TNF-alpha decreased when MSCs were exposed to 100 microg/ml LPS. Except for MHC-II and TNF-alpha (P > 0.05), the levels of CD80, CD86 and TLR-4 mRNA were significantly lower than that in the treated group of 10 microg/ml (P < 0.01).
MSCs expressed TLR-4 mRNA. LPS activated the functional expression levels of TLR-4 in MSCs although the activity may depend on the concentration of LPS.
Chinese medical journal 10/2007; 120(19):1685-8. · 1.02 Impact Factor