BACKGROUND:Bone tissue transplantation or osteogenic material fil ing is after used for bone defect repair. To remove autologous bone tissues can lead to additional damage and secondary deformity, therefore, it is extremely urgent to search for a new osteogenic material. OBJECTIVE:To construct the porousβ-tricalcium phosphate (β-TCP)/col agen scaffold modified with human bone morphogenetic protein 2 (hBMP2) gene, and to observe its effects on differentiation of MC3T3-E1 cel lines. METHODS:The porousβ-TCP/col agen scaffold modified with hBMP2 gene was prepared. Then in vitro culture system of MC3T3-E1 cel lines with composite scaffold was established. There were scaffold and plate groups, and each group was divided into two subgroups according to the different concentrations of plasmid. Samples were col ected and observed morphological y by scanning electron microscope and light microscope after complex culture. After 1, 3, 7 and 14 days of induction, calcium nodules were observed through alizarin red staining, the cel cycle was detected by real-time PCR, and expressions ofαI-chain col agen type I gene, Osterix and bone sialoprotein were observed. RESULTS AND CONCLUSION:The number of cel s adhered, differentated and distributed on the composite scaffold was significantly higher than that of the single scaffold (P<0.05). Alizarin red staining and real-time PCR detection showed that the osteogenesis ability of MC3T3-E1 cel lines in the scaffold group was stronger than that in the plate group. To conclude, the porousβ-TCP/col agen scaffold modified with hBMP2 gene is an appropriate candidate for bone defect repair.

Objective To investigate the relationship between CYP2J2*7 mutation(G-76T) and coronary heart disease (CHD) in Chinese Hanpopulation and to study the effects of CYP2J2 geneover-expressionon the proliferation and migrationof aortic smooth muscle cells of ApoE-/- mice. Methods CYP2J2*7 genotype was detectedin 500 patients with CHD and 478 controlsubjects by the Polymerase Chain Reaction-Restriction Frag-ment Length Polymorphism (PCR-RFLP). Culturedaortic smooth muscle cells of ApoE-/- mice were divided into control group, sham transfectiongroup and CYP2J2 over-expression group. Cell proliferation and migration were investigated after CYP2J2 over-expressionby MTS and Transwell assay. Results The frequency of CYP2J2*7 in CHD group was significantly higher than that incontrol group (10.00% vs. 6.49%, P = 0.046). Same is the case in female cases(P = 0.026). Compared with these of aortic smooth muscle cells incontrol group and sham trans-fectiongroup, the cell proliferation in 24, 48, 72 h, and the cell migration in 48 h after CYP2J2 over-expression in CYP2J2 group were significantly suppressed. Conclusions CYP2J2*7 mutation might increase the risk of CHD in Chinese Han population. CYP2J2 over-expression can suppress the proliferation and migration of aortic smooth muscle cells and CYP2J2 might have the effect of anti-atherosclerosis.

OBJECTIVETo investigate the clinical characteristics of newly diagnosed acute myeloid leukemia (AML) with NPM1 mutation.

METHODSNPM1 mutation (including A, B, D mutation type) was detected in 206 patients with newly diagnosed AML by real-time quantitative RT-PCR.

RESULTSThe incidence of NPM1 mutation was 15.5% in total AML patients and 32.5% in normal karyotypes AML patients. The characteristics of 174 NPM1 wild type patients v.s. that of 32 NPM1 mutation patients was as follow, median age (46 vs 35 years old, P < 0.01), WBC counts (27 × 10(9)/L vs 8 × 10(9)/L, P < 0.01), BPC (82 × 10(9)/L vs 36 × 10(9)/L, P < 0.01), proportion of AML-M(5) (31.2% vs 5.8%, P = 0.01), incidence of normal karyotypes (92.6% vs 40.8%, P < 0.01), incidence of FLT3-ITD-positive (25.0% vs 7.5%, P < 0.01), CD34-positvie (23.3% vs 69.5%, P < 0.01), cases with fusion gene (0 vs 47.1%, P < 0.01). No statistic difference was found in sex, percentage of blasts in bone marrow, complete remission rate, overall survival between the two groups. Relapse-free survival in AML patients with NPM1-mutation and FLT3-ITD-negative tended to be higher than in those with NPM1-mutation and FLT3-ITD-positive.

CONCLUSIONIt is necessary to detect NPM1 mutation and FLT3-ITD in newly diagnosed AML patients, especially in patients with high WBC and BPC, CD34-negative, normal karyotype, which might help to molecular classification and treatment.

Humans ; Leukemia, Myeloid, Acute ; genetics ; Mutation ; Nuclear Proteins ; genetics ; Prognosis ; fms-Like Tyrosine Kinase 3 ; genetics

Background: Atherosclerosis (AS) is an inflammatory disease. Inflammation was considered to play a role in the whole process of AS. This study aimed to analyze the relationships of inflammatory factors and risk factors with different target organ damages (TOD) in essential hypertension (EH) patients and to explore its clinical significance. Methods: A total of 294 EH patients were selected and divided into four groups according to their conditions of TOD. Forty?eight healthy subjects were selected as control. The clinical biochemical parameters, serum amyloid A, serum tryptase, and lipoprotein?associated phospholipase A2 (Lp?PLA2) in each group were detected, and the related risk factors were also statistically analyzed. Results: Fibrinogen (Fbg) was the most significant independent risk factor in acute coronary syndrome (ACS) group (odds ratio [OR]:22.242, 95% confidence interval [CI]: 6.458–76.609, P < 0.001) with the largest absolute value of the standardized partial regression coefficient B''(b'': 1.079). Lp?PLA2 was the most significant independent risk factor in stroke group (OR: 13.699, 95% CI: 5.236–35.837, P < 0.001) with b'' = 0.708. Uric acid (UA) was the most significant independent risk factor in renal damage group (OR: 15.307, 95%CI: 4.022–58.250, P < 0.001) with b'' = 1.026. Conclusions: Fbg, Lp?PLA2, and UA are the strongest independent risk factors toward the occurrence of ACS, ischemic stroke, and renal damage in EH patients, thus exhibiting the greatest impacts on the occurrence of ACS, ischemic stroke, and renal damage in EH patients, respectively.

Objective:To evaluate the gene mutation profile and prognostic significance of adult cytogenetically normal acute myeloid leukemia (CN-AML) with CEBPA-bZIP mutation. Methods:Targeted sequencing was implemented on the diagnostic bone marrow DNA samples of 141 adult CN-AML subjects with CEBPA-bZIP mutation. The nomogram model for leukemia-free survival (LFS) rate was generated by combining genetic abnormalities and clinical data. Risk stratification was conducted based on prognostic variables and the effect of risk-adjusted consolidation therapy was investigated by Kaplan-Meier method. Results:Four variables were finally included in our nomogram model after multivariate Cox analysis,and an equation for risk score calculation was obtained,risk score=1.3002×white blood cell (WBC) (≥18.77×109/L)+1.4065×CSF3R mutation positive+2.6489×KMT2A mutation positive+1.0128×DNA methylation-related genes mutation positive. According to the nomogram model,patients were further divided into low-risk group (score=0,n=46) and high-risk group (score>0,n=95). Prognostic analysis showed that the 5-year LFS rate,5-year overall survival (OS) rate,and 5-year cumulative incidence of relapse (CIR) of patients who received allogeneic hematopoietic stem cell transplantation (allo-HSCT) in the high-risk group were 93.5％,97.1％,and 3.5％,while those in patients who received maintenance chemotherapy were 32.9％,70.5％,and 63.4％,respectively. The differences were statistically significant (all P<0.05). Allo-HSCT could significantly improve the prognosis of patients in high-risk group. However,no corresponding benefit was observed in the low-risk group. Conclusion:Adult CN-AML with CEBPA-bZIP mutation has a complex co-mutation pattern. The nomogram model based on mutations of CFS3R,KMT2A and DNA methylation-related genes together with WBC count can further divide this subset of patients into a relatively low-risk group and a relatively high-risk group. For individuals in the high-risk group,allo-HSCT is proposed as post-remission therapy. The above data will benefit the prognosis estimation and treatment decision for adult CN-AML with CEBPA-bZIP mutation.

This study was purposed to investigate the effect of adenovirus-mediated transfer of PDCD5 gene on apoptosis of K562 cells induced by etoposide. Recombinant adenovirus PDCD5 (Ad-PDCD5), control vectors Ad-null and Ad-eGFP were constructed by AdMax vector system respectively. After K562 cells were transfected by Ad-PDCD5, Ad-null or Ad-eGFP with different multiplicity of infection (MOI), the expression level of the PDCD5 gene was examined by RQ-RT-PCR assay. The effects of etoposide in combination with Ad-PDCD5 on the proliferation and apoptosis of K562 cells were measured by using MTT assay and flow cytometry with Annexin-V-FITC/PI dual labeling technique, respectively. The results showed that the transfection efficiencies of Ad-eGFP in K562, Jurkat and CEM cells ranged from 60% to 86%. Expression level of PDCD5 gene in K562 cells was evidently increased following transfection with Ad-PDCD5. The Ad-PDCD5 synergistically enhanced the apoptotic percentage of K562 cells induced by VP-16, as compared with that of Ad-null + VP16 and VP-16 alone respectively. It is concluded that Ad-PDCD5 may be a potential agent for enhancing the chemotherapy effect.
Adenoviridae ; genetics ; metabolism ; Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; genetics ; metabolism ; Etoposide ; pharmacology ; Humans ; K562 Cells ; Neoplasm Proteins ; genetics ; metabolism ; RNA, Messenger ; metabolism ; Recombinant Proteins ; genetics ; metabolism ; Transfection

OBJECTIVETo evaluate levels of common specific fusion transcripts M-bcr-abl, m-bcr-abl, TEL-AML1, AML1-ETO, PML-RAR alpha, CBF beta-MYH11 in untreated leukemia patients.

METHODSSpecific fusion transcript levels were detected by TaqMan-based real-time quantitative RT-PCR technique in a total of 208 samples, including 195 bone marrow samples from 50 M-bcr-abl(+) chronic phase-chronic myeloid leukemia (CML-CP), 10 M-bcr-abl(+) acute lymphoblastic leukemia (ALL), 19 m-bcr-abl(+) ALL, 11 TEL-AML1(+) ALL, 30 AML1-ETO(+) acute myeloid leukemia (AML), 58 PML-RAR alpha(+) acute promyelocytic leukemia (APL) and 17 CBF beta-MYH11(+) AML patients and 13 peripheral blood samples from 13 M-bcr-abl(+) CML-CP patients. abl was chosen as internal control gene. Fusion transcript level was calculated as fusion transcript copies/abl transcript copies in percentage.

RESULTSBone marrow and peripheral blood samples of CML-CP patients had similar M-bcr-abl fusion transcript levels (median 30% vs 35%, P > 0.05). M- and m-bcr-abl (median 64% vs 54%) levels were similar in ALL patients (P > 0.05), M-bcr-abl level was significantly higher in ALL than CML-CP patients(P < 0.001). Median TEL-AML1 level was 228% in ALL patients. Among AML patients, AML1-ETO level was significantly higher than CBF beta-MYH11 and PML-RAR alpha levels (median 388% vs 145%, 388% vs 47%, all P < 0.001), CBF beta-MYH11 level was significantly higher than PML-RAR alpha level (P < 0.001). Fusion transcript levels of L-, V- and S-type PML-RAR alpha were 45%, 44% and 55%, respectively. L-type was significantly lower than S-type (P = 0.04).

CONCLUSIONSFusion transcript levels in untreated leukemia patients were different and patient-to-patient variations did exist. Detection of fusion transcript levels in untreated leukemia patients not only provides baseline for minimal residual disease monitoring and treatment evaluation but also enable the comparison in inter-laboratory data.

Adolescent ; Adult ; Bone Marrow Cells ; metabolism ; Child ; Child, Preschool ; Core Binding Factor Alpha 2 Subunit ; genetics ; Female ; Fusion Proteins, bcr-abl ; genetics ; Humans ; Leukemia ; genetics ; Male ; Middle Aged ; Oncogene Proteins, Fusion ; genetics ; RUNX1 Translocation Partner 1 Protein ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Transcription, Genetic

The expression levels of programmed cell death 5 (PDCD5) are down-regulated in many malignancies. SG611-pdcd5, a recombinant conditionally replicative adenovirus carrying pdcd5 gene expression cassette, can evidently kill the leukemic cells and protect selectively the normal cells. The purpose of this study was to investigate the synergistic killing effect of SG611-pdcd5 and low-dose etoposide (VP-16) on K562 cells. K562 cells were treated with different concentrations of VP-16 or different multiplicities of infection (MOI) of SG611-pdcd5. After 48 hours of incubation the cell viability was determined by using MTT assay. The results showed that the cell viability of SG611-pdcd5 (MOI = 40) plus VP-16 (0.5 µg/ml) group significantly decreased as compared with single SG611-pdcd5 (MOI = 40) treatment group or single VP-16(0.5 µg/ml) treatment group (42.00 ± 5.75% vs 59.45 ± 4.12%; 42.00 ± 5.75% vs 82.91 ± 3.41%, respectively, both p < 0.05). The synergistic killing effect of SG611-pdcd5 plus VP-16 was higher than that of PDCD5 protein plus VP-16 or that of non-replicating adenovirus carrying pdcd5 (Ad-pdcd5) plus VP-16 (both p < 0.05). The cell viability of VP-16 (4.0 µg/ml) plus SG611-pdcd5 (MOI = 40) group, VP-16 (4.0 µg/ml) plus proPDCD5 (40 µg/ml) group and VP-16 (4.0 µg/ml) plus Ad-pdcd5 (MOI = 80) group was 37.09 ± 1.89%, 52.36 ± 1.64% and 73.64 ± 4.33%, respectively. It is concluded that SG611-pdcd5 can promote K562 cell death induced by low-dose VP-16. The combination of SG611-pdcd5 and VP-16 can enhance the killing effect on leukemic cells.
Adenoviridae ; genetics ; Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; Apoptosis Regulatory Proteins ; genetics ; Cell Survival ; Etoposide ; pharmacology ; Genetic Vectors ; Humans ; K562 Cells ; Neoplasm Proteins ; genetics

The purpose of this study was to construct a recombinant conditionally replicating adenovirus (CRAd) expressing programmed cell death 5 (pdcd5). Pdcd5 gene was inserted in the E3 region of SG600-a CRAd in which the key genes for virus replication E1a and E1b were controlled under the human telomerase reverse transcriptase promoter (hTERTp) and the hypoxia response element (HRE) respectively, and with a deletion of 24 nucleotides within CR2 region of E1a. The insertion and orientation of all recombined plasmids were confirmed by restriction enzyme digestion and polymerase chain reaction (PCR). The infection efficiencies of a recombined virus carrying enhanced green fluorescent protein (EGFP) in leukemic cell lines were observed by using fluorescence microscope. The relative pdcd5 expression levels of K562 after being infected with SG611-pdcd5 were detected by real-time quantitative PCR. The results showed that the construction of SG611-pdcd5 was completed and confirmed. Pdcd5, hTERTp, HRE, skeleton and fiber11 of recombinant adenovirus SG611-pdcd5 were successfully amplified. The infection efficiencies of SG611-EGFP were all above 70% in both leukemic K562 and MEG-01 cell lines. SG611-pdcd5 expressed pdcd5 with high efficiency in leukemic cells as compared with Ad-pdcd5 or SG611 (p < 0.001). The expression level of pdcd5 increased gradually along with the increase of MOI. It is concluded that the triple-regulated adenovirus of SG611-pdcd5 containing the pro-apopro-tic gene pdcd5 has been successfully established with high pdcd5 expression level in leukemic cells, indicating that the recombinant adenovirus, SG611-pdcd5, promises further development of targeted tumor gene therapy.
Adenoviridae ; genetics ; Apoptosis Regulatory Proteins ; genetics ; Genetic Therapy ; methods ; Neoplasm Proteins ; genetics ; Oncolytic Viruses ; genetics ; Promoter Regions, Genetic ; Telomerase ; genetics

OBJECTIVETo explore the effect of one dressing of porcine acellular dermal matrix on deep partial thickness burns.

METHODSFrom January 1997 to January 2004, sixty-seven cases of deep partial thickness total burned surface area (TBSA) from 50% to 90% burn wound were treated by a single dressing of porcine acellular dermal matrix (the porcine acellular dermal matrix group). Ten cases of deep partial thickness burned patients with the same TBSA treated by exposure method served as the exposure method group. The healing time of the wound was observed. The patients were followed up for 3 months to 2 years, and the scar proliferation was observed.

RESULTSThe deep partial-thickness wound would be healed without dressing change in the porcine acellular dermal matrix group, and the average healing time was (12.2 +/- 2.6) days. The average healing time of the exposure method group was (27.4 +/- 3.5) days. Follow up of the patients within 3 months to 2 years showed that scar proliferation in the porcine acellular dermal matrix group was much less than that in the exposure method group, even no scar proliferation was observed in some patients.

CONCLUSIONWithout tangential excision, autografting and dressing change, a single dressing of porcine acellular dermal matrix on deep partial thickness burn wound could shorten the healing time and inhibit scar proliferation.

Animals ; Biological Dressings ; Burns ; pathology ; therapy ; Cicatrix ; prevention & control ; Female ; Follow-Up Studies ; Humans ; Male ; Swine ; Treatment Outcome ; Wound Healing