Comparative Proteomic Analysis of Mature and Immature Oocytes of the Swamp Buffalo (Bubalus bubalis).

Maternal protein components change markedly during mammalian oogenesis. Many of these proteins have yet to be characterized and verified. In this study, a proteomics approach was used to evaluate changes in proteins during oogenesis in the Swamp Buffalo (Bubalus bubalis). Proteins from 500 immature oocytes and 500 in vitro matured oocytes were subjected to two-dimensional electrophoresis, and more than 400 spots were detected. Image analysis indicated that 17 proteins were differentially expressed between the two groups. Eight proteins were identified by mass spectrometry. In mature oocytes, three proteins were down-regulated: major vault protein (MVP), N-acetyllactosaminide ß-1,6-N-acetylglucosaminyl-transferase (GCNT-2), and gem-associated protein (GEMIN)8, whereas five other proteins, heat shock protein (HSP)60, Ras-responsive element-binding protein 1 (RREB-1), heat shock cognate 71 kDa protein (HSC71), hemoglobin subunit α (HBA), and BMP-2-inducible protein kinase (BMP-2K), were up-regulated. The expression profiles of HSP60 and GEMIN8 were further verified by Western blotting. The changes in HSP60 protein expression demonstrate the increasing need for mitochondrial protein importation to facilitate macromolecular assembly during oocyte maturation. The down-regulation of GEMIN8 production implies that RNA splicing is impaired in mature oocytes.

Favorable prognosis of biallelic CEBPA gene mutations in acute myeloid leukemia patients: a meta-analysis.

OBJECTIVES: Increasing number of studies suggested that biallelic CEBPA (bi CEBPA) mutations were associated with favorable prognosis in patients with acute myeloid leukemia (AML), but the results remain inconclusive. We therefore present a meta-analysis to evaluate the prognostic value of bi CEBPA mutations in patients with AML. METHODS: A comprehensive literature search was undertaken through August 2014 looking for eligible studies. Pooled hazard ratios (HRs) estimates and 95% confidence intervals (95% CIs) in overall survival (OS) and event-free survival (EFS) were used to calculate estimated effect. RESULTS: Ten studies covering a total of 6219 subjects were included in this analysis. Overall, bi CEBPA mutations were associated with favorable clinical outcome in patients with AML (HR for EFS: 0.41, 95% CI: 0.32-0.52; for OS: 0.37, 95% CI: 0.27-0.50), in cytogenetically normal (CN)-AML (HR for EFS: 0.38, 95% CI: 0.29-0.49; for OS: 0.32, 95% CI: 0.23-0.43). When took the cohort of monoallelic CEBPA (mo CEBPA) mutated and wild-type CEBPA (wt CEBPA) AML as a reference group, bi CEBPA mutated AML also shown beneficial outcomes (HR for OS: 0.52, 95% CI: 0.37-0.72). No significant difference was found between mo CEBPA mutation and wt CEBPA in patients with AML or CN-AML (P > 0.05). CONCLUSION: Bi CEBPA mutations in patients with AML are strongly associated with a favorable prognosis, which suggested that bi CEBPA mutations would potentially serve as a novel prognostic marker in AML.

[Effects of Paclitaxel and Quizartinib Alone and in Combination on AML Cell Line MV4-11 and Its STAT5 Signal Pathway].

OBJECTIVE: To investigate the effects of paclitaxel, quizartinib and their combination on proliferation, apoptosis and FLT3/STAT5 pathway of human leukemia cell line MV4-11 (FLT3-ITD+). METHODS: MV4-11 cells were treated with paclitaxel and quizartinib at different concentrations for 24 h, 48 h and 72 h, respectively, and then the two drugs were combined at 48 h to compare the inhibition of proliferation, the apoptosis rate was detected by flow cytometry, the expression of FLT3 and STAT5 mRNA was determined by fluorescence quantitative PCR, and the protein expression of FLT3, p-FLT3, STAT5 and p-STAT5 was determined by Western blot. RESULTS: Different combination groups of paclitaxel and quizartinib had synergistic inhibitory effect. The cell survival rate in the combination group was significantly lower than that in the single drug group (P<0.05). The cell apoptosis rate in the combination group was significantly higher than that in the single drug group (P<0.001). The expression of FLT3 mRNA in combination group was significantly higher than that in two single drugs (P<0.01). The expression of STAT5 mRNA in combination group was significantly higher than that in quizartinib group (P<0.001); increased compared with paclitaxel group, but there was no statistical significance. The expression level of p-FLT3、p-STAT5 protein in the combination group was significantly lower than that in the single drug group (P<0.05, P<0.05). CONCLUSION: Paclitaxel combined with quizartinib can synergistically inhibit the proliferation of MV4-11 cell line and promote the apoptosis of MV4-11 cell line by inhibiting the activity of FLT3/STAT5 pathway.

[Effects of mangiferin on cell cycle status and CDC2/Cyclin B1 expression of HL-60 cells].

OBJECTIVE: To study the effects of mangiferin on cell cycle status and CDC2/Cyclin B1 expression in HL-60 cells, and its molecular mechanism for treating leukemia. METHODS: The effect of Mangiferin on HL-60 cells proliferation was determined by MTT assay; The change of cell cycle status in HL-60 cells treated with mangiferin was performed by the flow cytometry; The expressions of CDC2 mRNA and Cyclin B1 mRNA were detected by semiquantitative RT-PCR. RESULTS: The growth inhibition effects of mangiferin in HL-60 cells were enhanced as the mangiferin concentration increased and exposure time prolonged; HL-60 cells in G2/M phase increased in a dose-dependent way 24 hours after mangiferin administration, indicating G2/M phase blockage; the expressions of CDC2 mRNA and Cyclin B1 mRNA enhanced in a dose-dependent way and came to the peak at 80 micromol/L mangiferin. CONCLUSION: Mangiferin can inhibit the proliferation of HL-60, block the cell cycle progression in G2/M phase, and it can significantly increase the expressions of CDC2 mRNA and Cyclin B1 mRNA of HL-60 cells. G2/M phase blockage may be one of its molecular mechanism for treating leukemia.

[Expression and Function of miR-99a-5p in Bone Marrow of Patients with MDS].

OBJECTIVE: To detect the expression of miR-99a-5p in myelodysplastic syndrome (MDS), to predict the target genes and to analyze its function by using bioinformatics. METHODS: The expression levels of bone marrow miR-99a-5p in MDS patients were detected by qRT-PCR, and the correlation of miR-99a-5p expression with clinical pathological characteristics, percentage of marrow blasts , chromosome karyotype and peripheral blood hemogram were analyzed. The target genes of miR-99a-5p were predicted by Targetscan, Miranda and Microcosm, and the intersection of the predicted results of 3 softwares was used as a potential target gene for miR-99a-5p. RESULTS: The expression level of bone marrow miR-99a-5p in MDS patients was significantly higher than that of healthy controls (P<0.01); According to the prognostic score of IPSS, MDS patients were divided into relatively low risk group (including low risk group and intermediate risk group 1) and relatively high risk group (including intermediate risk group 2 and high risk group). Analysis showed that the expression level of bone marrow miR-99a-5p in relatively low risk group was 2.40 times higher than that in healthy control group (P<0.01), the expression level of bone marrow miR-99a-5p in relatively high risk group was 6.66 times higher than that in healthy control group (P<0.01), the expression level of miR-99a-5p in relatively high risk group was 2.80 times higher than that in the relatively low risk group ( P<0.01) ; the expression level of bone marrow miR-99a-5p in t-MDS group was 6.35 times higher than that in healthy control group (P<0.001). There was a significant positive correlation between the expression level of miR-99a-5p and the percentage of marrow blasts (P<0.01), but the expression of miR-99a-5p did not correlate with chromosome karyotype and peripheral blood hemogram; In this study it was obtained that ST5 might participate in pathological mechanism of MDS by bioinformatic analyses and references. CONCLUSION: The expression levels of miR-99a-5p is up-regulated in MDS patients, and its expression showed a rising tendency which may be involved in the pathogenesis of MDS by regulating target gene ST5.

Increased serum concentrations of asymmetric dimethylarginine (ADMA) in patients with early-onset coronary artery disease.

BACKGROUND: Asymmetric dimethylarginine (ADMA) has been associated with an increased risk of cardiovascular disease. We investigated the role of serum ADMA concentrations in early-onset coronary artery disease (EOCAD). METHODS: Candidates for coronary artery angiography (age<50y for men and <55y for women) who met the inclusion criteria were enrolled in this study. Serum concentrations of ADMA were determined using ELISA. Severity of coronary atherosclerosis was estimated by number of diseased vessels. RESULTS: A total of 601 subjects (286 with EOCAD patients and 315 controls) were included in the study. ADMA concentrations were found to be significantly higher in the EOCAD group (0.480±0.110µmol/l) than in the control group (0.457±0.091, P=0.007). ADMA concentrations significantly increased with the number of diseased vessels (P<0.001). In addition, serum ADMA concentrations were affected by diabetes mellitus and smoking status, and were positively correlated with serum creatinine and body mass index (BMI). CONCLUSIONS: Our results show that serum ADMA concentrations were associated with the presence and severity of EOCAD, suggesting that ADMA may be involved in the progression of EOCAD.

An early warning scoring system for the prevention of acute heart failure.

BACKGROUND: Early prediction and identification of the onset of acute heart failure (AHF) in high-risk patients are of great significance for preemptive treatment and a better prognosis. We sought to find a scoring system to predict the onset of AHF in patients in the acute heart failure unit (AHFU). METHODS: Data for 433 patients at of AHF in the AHFU were analyzed. We recorded sex, age, history of coronary artery disease, hypertension, diabetes, and primary percutaneous coronary intervention. We also reviewed temperature, pulse, SpO2, respiratory rate, urine volume, and emotional state every hour before the onset of AHF. All admission and follow-up data were retrieved from hospital charts. Factors were analyzed using a binary logistic regression model to create the SUPER (SpO2, urine volume, pulse, emotional state, and respiratory rate) scoring model. We divided the scoring system into four levels: low-, intermediate-, high-, and extremely high-risk. Patients fitting the four risk levels were followed up for 6 to 24 months. RESULTS: SpO2, hourly urine volume, pulse, emotional state and respiratory rates were associated with an independent increased risk for the onset of AHF. The SUPER score for the patients in the AHFU predicted the onset of AHF 3.90 ± 1.94 h (1-17 h) earlier. The areas under the ROC curve for the SUPER score and the modified early warning score were 0.811 and 0.662 (p<0.05), indicating that the SUPER score provided a better warning of the AHF. A low-, intermediate-, high-, and very high-risk SUPER predicted the likelihood of AHF at 17.3%, 61.3%, 84.4%, and 94.0%, respectively. The differences in mortality rates between the four levels were statistically significant (p<0.05). CONCLUSIONS: In patients at high risk of AHF, the SUPER scoring system could predict the onset of AHF 2 to 6h earlier. Preemptive treatment according to the SUPER score may prevent or delay AHF occurrence to improve quality of life and reduce mortality.

Neuroglobin protects cardiomyocytes against apoptosis and cardiac hypertrophy induced by isoproterenol in rats.

Neuroglobin (Ngb) is well known as a physiological role in oxygen homeostasis of neurons and perhaps a protective role against hypoxia and oxidative stress. In this study, we found that Ngb is expressed in rat heart tissues and it is related to isoproterenol induced cardiac hypertrophy. Moreover, overexpression or knock-down of Ngb influences the expression of hypertrophic markers ANP and BNP and the ratio of hypertrophic cells in rat H9c2 myoblasts when isoproterenol treatment. The Annexin V-FITC/PI Staining, Western blot and qPCR analysis showed that the involvement in p53-mediated apoptosis of cardiomyocytes of Ngb is might be the mechanism. This protein could prevent the cells against ROS and POS-induced apoptosis not only in nervous systems but also in cardiomyocytes. From the results, it is concluded that Ngb is a promising protectant in the cardiac hypertrophy, it may be a candidate target to cardiac hypertrophy for clinic treatment.

[Methylation status of γ-globin gene promoter in ß-thalassemia major].

This study was aimed to detect and identify the promoter CpG island methylation of γ-globin gene in peripheral blood mononuclear cells from patients with ß-thalassemia major and healthy adult in Guangxi province, as well as to analyze the difference of promoter methylation rate of each CpG sites between them, and then to screen the promoter CpG island main methylation sites which maybe influence γ-globin expression. The template DNA was modified by bisulfite genomic sequencing PCR; the promoter sequences of γ-globin gene were amplified by technique Touchdown PCR, and then the PCR products were cloned and sequenced for obtaining methylation status of each CpG sites in target fragments, and then the accurate methylation sites and levels were detected quantitatively. The results indicated that the 4 CpG methylation sites locating at 28, 122, 231 and 234 bp in sequences were hypermethylated. As compared with healthy adults, the DNA methylation rate of 122 and 231 bp CpG sites in patients with ß-thalassemia major was obviously lower, however, methylation rates of 28 and 234 bp sites were not significantly different between patients and healthy adults. It is concluded that the methylation sites 28, 122, 231 and 234 bp of γ-globin gene promoter are found both in patients with ß-thalassemia major and healthy adults. The 122 and 231 bp sites are identified preliminarily to be involved in the regulation of γ-globin expression. This study provides the experimental evidence for alleviating the clinical symptoms of ß-thalassemia major and targeting gene treatment through the regulation of γ-globin.

[Effects of ligustrazine on expression of PECAM-1/CD31 and hematopoietic reconstitution in syngenic bone marrow transplantation of mice].

To estimate the effect of ligustrazine on the expression of PECAM-1/CD31 and hematopoietic reconstitution in syngenic bone marrow transplanted mice, 56 BALB/c mice were divided into 3 groups: normal control group, BMT control group, ligustrazine treated group (BMT + Ligustrazine) and syngenic BMT mouse models were established according to the literatures. The BMT control group and the ligustrazine treated group were given orally 0.2 ml saline and 2 mg ligustrazine twice a day respectively. On days 7, 14, 21 after BMT, mice were killed and peripheral blood cells, bone marrow nucleated cells (BMNC) were detected. Histological observation of bone marrow was made and the CD31 expression was assayed by flow cytometry. The results showed that in ligustrazine treated group the peripheral blood cell, BMNC counts on days 7, 14, 21 after BMT were higher than in the BMT control group (P < 0.01 or P < 0.05). The expression of CD31 in ligustrazine treated group on days 7, 14, 21 after BMT was also higher than in the BMT control group (P < 0.01 or P < 0.05). In conclusion, ligustrazine enhances CD31 expression in bone marrow cells after syngenic bone marrow transplantation of mice, which may be related to the mechanisms underlying the ligustrazine accelerating hematopoietic reconstitution in syngeneic bone marrow transplantation.