[Cytogenetic and Clinical Characteristics of Newly Diagnosed Elderly Patients with Acute Myeloid Leukemia].

OBJECTIVE: To investigate the cytogenetic characteristics and prognostic risk factors for elderly patients with newly diagnosed elderly acute myeloid leukemia(AML). METHODS: Cytogenetic test results of 76 elderly patients with AML admitted to the First Affiliated Hospital of the University of Science and Technology of China (Anhui Provincial Hospital) from April 2015 to December 2021 were retrospectively analyzed, and analyzed clinical characteristics of patients and risk factors influencing prognosis. RESULTS: According to cytogenetic risk stratification, 76 newly treated elderly AML patients were divided into the favorable, intermediate, and unfavorable groups with 6(7.9%), 58(76.3%), and 12(15.8%) cases, respectively. There was no significant difference in the patient's clinical characteristics and prognosis with the cytogenetics-risk classification groups. Correlation analysis showed that patients' objective response rate (ORR) was related to the age of onset and the mutation status of the CEBPA gene. Logistic regression analysis found that age ≥70 years was an independent risk factor for patients' ORR (OR=0.110, P=0.005). Remission determined the 1-year OS rate (OR=0.049, P=0.005). CONCLUSION: There is no significant difference in clinical characteristics among aged AML patients treated at initial treatment in different cytogenetic risk groups. The age of onset ≥70 years is the determinant of whether patients can obtain ORR, and the rate of ORR is closely related to the 1-year OS rate.

The Driving Force for 2014 Dengue Outbreak in Guangdong, China.

Dengue fever has rapidly spread in recent decades to become the most globally expansive viral vector-borne disease. In mainland China, a number of dengue outbreaks have been reported since 1978, but the worst epidemic in decades, involving 45230 cases and 76 imported cases, resulting in six deaths in Guangdong province, emerged in 2014. Reasons for this ongoing surge in dengue, both imported and autochthonous, are currently unclear and demand urgent investigation. Here, a seasonally-driven dynamic epidemiological model was used to simulate dengue transmission data recorded from the unprecedented outbreak. Sensitivity analysis demonstrate that delayed mosquito control, the continuous importations between the end of April to the early of July, the transmission of asymptomatic dengue infections, and the abnormally high precipitation from May to August might be the causal factors for the unprecedented outbreak. Our results suggested that the earlier and more frequent control measures in targeting immature and adult mosquitoes were effective in preventing larger outbreaks, and enhanced frontier health and quarantine from the end of April to the early of July for international communications and travelers.

[Effects of small interfering RNA specific for Her-2 gene on biological behavior of ovarian carcinoma cell].

OBJECTIVE: To explore the effects of small interfering RNA (siRNA) specific for Her-2 gene on biological behavior of ovarian carcinoma cell. METHODS: Her-2 siRNA recombinant plasmid and negative control plasmid were transfected into packing cell line PT67 by liposome, and PT67 was selected by puromycin later. SKOV3 was infected by the virus supernatant of stably transfected PT67 cell lines, and the stably transfected SKOV3 cell lines (SKOV3/siRNA, SKOV3/siRNA-negative) established by selection with puromycin were investigated in terms of the reduction levels of Her-2 mRNA and p185 by RT-PCR and immunohistochemistry. Cell proliferation was assayed with methyl thiazolyl tetrazolium, and cell cycle distribution and cell apoptosis were assayed with flow cytometry. The tumor growth of the null mice was analyzed after injection of SKOV3/siRNA and SKOV3/siRNA-negative into the skin. RESULTS: (1) The stable SKOV3 cell lines with a persistent silence of Her-2 gene were established. (2) The percentages of SKOV3/siRNA in G(0)/G(1) phase and S phase were 68.6%, 15.1% respectively; while the percentages of SKOV3/siRNA-negative in G(0)/G(1) phase and S phase were 55.8%, 23.3%. (3) The percentage of SKOV3/siRNA in early apoptosis was (10.500 +/- 0.250)%, while the percentage of SKOV3/siRNA-negative was (0.340 +/- 0.010)% (P < 0.01). (4) Compared with SKOV3/siRNA-negative, the proliferation of SKOV3/siRNA was delayed obviously (P < 0.05), and the growth of the corresponding implanted tumor slowed down significantly (P < 0.01). CONCLUSION: siRNA can inhibit the expression of Her-2 gene effectively, which restrains the biological behavior of ovarian carcinoma cell.

[Effect of natural killer cell line NK-92 against human ovarian carcinoma cells in vitro and in vivo].

OBJECTIVE: To study the cytotoxic activity of NK-92 cells irradiated against human ovarian cancer. METHODS: NK-92 cells were exposed to different doses of radiation and assayed for proliferation by a standard (3)H-thymidine incorporation assay and cell count by using trypan blue exclusion. The cytotoxic activity of NK-92 cells against targets was measured in a standard (51)Cr-release assay in vitro. The effectiveness of irradiated NK-92 cells on ovarian cancer was compared with the control group of cancers (without injection of irradiated NK-92 cells). RESULTS: (1) In vitro:The proliferation of NK-92 cells was inhibited by radiation of 4, 8 and 16 Gy, respectively. From the (3)H-thymidine incorporation data, irradiation by 4 Gy reduced cell proliferation to 29% of control, while 8 Gy reduced proliferation to 6%. The cytotoxicity of NK-92 cells at 4 Gy 2 days following irradiation was approximately 42%-62% for ovarian cancer cell HO-8910, while it was 33%-58% at 8 Gy. (2) In vivo: Tumor size in treatment group was (0.047 +/- 0.019) cm(3) on day 30 after inoculation, and (0.167 +/- 0.021) cm(3) on day 40 and (0.343 +/- 0.022) cm(3) on day 50, while the sizes were smaller in treatment group (P < 0.01). In addition, the tumor group animals died between 74-82 days after injection of HO-8910 cells, while the treatment group animals were alive over 120 days (P < 0.01). CONCLUSION: Our study indicates that injection of irradiated NK-92 cells may be a potentially effective treatment for human ovarian carcinoma.

[Expression of platelet collagen receptor-glycoprotein VI fragment in E. coli and its biological activities].

This study was aimed to further investigate the function of platelet collagen receptor-glycoprotein VI and to screen its specific inhibitor. The extracellular domain of platelet glycoprotein VI (GPVI) in E. coli was expressed by recombinant technology, the extracellular domain cDNA of GPVI was amplified from pBluescript KS(-)-GPVI plasmid by PCR. Proved by sequencing, the expression vector pET-20b(+)-GPVI was constructed, which was then transformed into E. coli (BL21(DE3)pLysS) and induced by IPTG. The recombinant GPVI was purified on Ni-NTA resin column and renatured in PBS containing GSH and GSSG. The anti-penta His McAb and anti-GPVI polyclonal antibody were used to identify the recombinant GPVI in Western blotting. Collagen binding test was conducted to investigate the biological activity of recombinant GPVI. The results showed that the recombinant GPVI was expressed in E. coli and successfully purified, which was confirmed to be similar to the native GPVI in Western blotting. The recombinant GPVI can bind the type I collagen in dose-dependent manner. In conclusion, the recombinant GPVI can be achieved in E. coli and restore its native characteristics after renaturation.

[Expression of von willebrand factor-A3 domain in E coli and its biological function].

The interaction among collagen, von Willebrand factor (vWF) and glycoprotein Ib axis is the first step in hemostasis and thrombosis, especially under high shear condition. To develop a new remedy of anti-thrombosis, mRNA from endothelial cells was extracted, and reverse transcription PCR was adopted to amplify DNA of interest. After sequencing, recombinant expression vector was constructed. The amplified DNA fragment of vWF domain A3 was inserted into expression vector with 6 x his taq, pET20b(+), the recombinant was transformed into E coli (strain DE3) and induced by IPTG. Recombinant vWF-A3 was designated as a recombinant fragment comprising residues 918 - 1114 of mature vWF subunit. It was purified through Ni-NTA resin column and refolded in Tris buffer containing GSH and GSSG. The results showed that rvWF-A3 was expressed successfully in E coli (strain DE3), accounting for 46% of total bacterial protein with its purity of over 95%. It was identified that rvWF-A3 is capable to bind collagen and inhibit the wild vWF binding to collagen by competition. It is concluded that rvWF-A3 fragment might be an effective antithrombotic agent for preventing arterial thrombosis.

[Construction and expression of mouse-human chimeric Fab fragment gene of monoclonal antibody SZ-2 against platelet].

AIM: To reduce immunogenicity of a monoclonal antibody SZ-2 specific for human platelet. METHODS: Reverse transcription and polymerize chain reaction were used to amplify the variable region genes of monoclonal antibody SZ-2. The cloned V(H) and V(L) genes were sequenced and fused to human IgG1 constant region gene CH1 and Ckappa in plasmid pSW1. The recombinant plasmid were transformed into E. coli. The expressed recombinant proteins were analysed. RESULTS: The V(H) and V(L) genes were homologous with the published gene sequences of mouse antibody variable region. The concentration of chimeric Fab fragment in expression supernatant was about 180 microg/L detected by ELISA. Western blot analysis showed that SZ-2 Fab/Hu maintained the binding activity to human platelet GPIb. The recombinant proteins could suppress platelet aggregation induced by Ristocatin. CONCLUSION: The variable region genes of SZ-2 are cloned and the mouse-human chimeric Fab fragment is expressed successfully in E. coli.

[Molecular cloning of human vWF/A1 gene and its expression].

To study the mechanism of thrombogenesis and search new anti-thrombotic agent, the cDNA for human vWF A1 domain was high-level expressed in E. coli and recombinant protein of vWF A1 with biologic activity was obtained. The gene encoding A1 domain was amplified by PCR from plasmid containing full length cDNA of human vWF. After confirming by DNA sequencing analysis, the recombinant expression plasmid pQE31-vWF/A1 was constructed and introduced into E. coli M15 strain, then induced by IPTG; the expressed protein was purified with Ni-NTA agarose, identified by Western blotting. The results showed that the 854 bp DNA fragment was obtained by PCR from the plasmid containing full length cDNA for human vWF and its sequence was identical to the published sequence. High level expression of A1 protein was yielded after 5 hour-induction, which amounted to 30% of total bacteria protein in inclusion body. Western blot demonstrated it possessed good antigenicity and high specificity. It is concluded that cDNA for vWF/A1 had been cloned successfully, high level expression of A1 protein was achieved in E. oli. This study will provide a basis for the further clinical and basic research on the role of vWF in thrombosis and hemostasis.