Anatomical observation and transcriptome analysis of buds reveal the association between the AP2 gene family and reproductive induction in hybrid larch (Larix kaempferi × Larix olgensis).

Hybrid larch is an excellent afforestation species in northern China. The instability of seed yield is an urgent problem to be solved. The biological characteristics related to seed setting in larch are different from those in angiosperms and other gymnosperms. Studying the developmental mechanism of the larch sporophyll can deepen our understanding of conifer reproductive development and help to ensure an adequate supply of seeds in the seed orchard. The results showed that the formation of microstrobilus primordia in hybrid larch could be observed in anatomical sections collected in the middle of July. The contents of endogenous gibberellin 3 (GA3) and abscisic acid (ABA) were higher and the contents of GA4, GA7, jasmonic acid and salicylic acid were lower in multiseeded larch. Transcriptome analysis showed that transcription factors were significantly enriched in the AP2 family. There were 23 differentially expressed genes in the buds of the multiseeded and less-seeded types, and the expression of most of these genes was higher in the buds than in the needles. We conclude that mid-July is the early stage of reproductive organ development in hybrid larch and is suitable for the study of reproductive development. GA3 and ABA may be helpful for improving seed setting in larch, and 23 AP2/EREBP family genes are involved in the regulation of reproductive development in larch.

[Retrospective study on forty cases of medical legal dispute].

OBJECTIVE: To analyze the main causes of medical dispute and the main types of medical malpractice. The related problems were discussed in forensic expertise. METHODS: Forty cases of medical dispute from 2006 to 2008 in our institute were analyzed retrospectively. RESULTS: In 40 cases of medical dispute, city-level hospitals, county-level hospitals, town-level hospitals and private clinics were 11 (27.5%), 24 (60.0%), 2 (5.0%) and 3 (7.5%) cases respectively. The internal medicine departments, surgical departments, gynaecological and obstetric departments, pediatric departments and outpatient center were 16 (40.0%),10 (25.0%), 9 (22.5%), 2 (5.0%) and 3 (7.5%) cases, respectively. The amount of cases from city-level hospitals, county-level hospitals diagnosed by the medical experts as the medical malpractice showed less than that from town-level hospitals and private clinics. CONCLUSION: The amount of cases of medical dispute from city-level and county-level hospitals were more than that of town-level hospitals. But the amount of cases diagnosed by the medical experts as medical malpractice from city-level and county-level hospitals were less than that of town-level hospital and private clinics.

Improving Ethyl Acetate Production in Baijiu Manufacture by Wickerhamomyces anomalus and Saccharomyces cerevisiae Mixed Culture Fermentations.

Ethyl acetate content has strong influence on the style and quality of Baijiu. Therefore, this study investigated the effect of Saccharomyces cerevisiae Y3401 on the production of ethyl acetate by Wickerhamomyces anomalus Y3604. Analysis of cell growth showed that Y3401 influences Y3604 by nutrient competition and inhibition by metabolites, while the effect of Y3604 on Y3401 was mainly competition for nutrients. Mixed fermentation with two yeasts was found to produce more ethyl acetate than a single fermentation. The highest yield of ethyl acetate was 2.99 g/L when the inoculation ratio of Y3401:Y3604 was 1:2. Synergistic fermentation of both yeasts improved ethyl acetate production and increased the content of other flavor compounds in liquid and simulated solid-state fermentation for Baijiu. Saccharomyces cerevisiae had a positive effect on ethyl acetate production in mixed culture and provides opportunities to alter the aroma and flavor perception of Baijiu.

Enhanced production of ethyl acetate using co-culture of Wickerhamomyces anomalus and Saccharomyces cerevisiae.

The style and quality of Baijiu is greatly influenced by ethyl acetate. Therefore, improving and controlling the ethyl acetate levels in Baijiu is important. This study investigated ethyl acetate production using a co-culture of Saccharomyces cerevisiae Y3401 and Wickerhamomyces anomalus Y3604. More ethyl acetate was produced in mixed fermentations using both yeasts than in single fermentations. The highest ethyl acetate yield was 6.41 g/L using a Y3401/Y3604 ratio of 3:1. Synergistic fermentation using both yeasts not only improved ethyl acetate production, but also increased the contents of other flavor compounds, such as ß-phenethyl alcohol and phenethyl acetate. Therefore, the co-culture of S. cerevisiae and W. anomalus had a positive effect on ethyl acetate production and provides opportunities for altering the aroma and flavor perception of Baijiu.

Development and evaluation of real-time loop-mediated isothermal amplification for hepatitis B virus DNA quantification: a new tool for HBV management.

BACKGROUND: Accurate quantitation of hepatitis B viral load is a critical aspect in screening and monitoring HBV infection. OBJECTIVES: We used loop-mediated isothermal amplification (LAMP) to develop a real-time fluorogenic (RtF-LAMP) protocol to quantitate HBV DNA. Quantitative analysis was obtained by measuring time-to-positive (TTP), a biomarker similar to cycle threshold (Ct) in real-time PCR. STUDY DESIGN: Sensitivity, specificity, reproducibility, and dynamic range for RtF-LAMP were evaluated using molecular and biological standards. Four hundred and two patient samples were then used to compare the performance of RtF-LAMP to a commercial real-time PCR assay (DaAn Gene Co, China). RESULTS: The lower detection limit (LDL) of RtF-LAMP by Probit analysis at the 95% detection level was 210 copies/ml, and the dynamic range was 8 orders of magnitude. The conversion factor for results obtained with the RtF-LAMP assay was 1 IU/ml equals to 4.4 copies/ml. Coefficients of variation (CV) reflected low intra-assay and inter-assay variability (4.24-12.11%). In a large number of serum samples, there was excellent an correlation between RtF-LAMP and real-time PCR (R(2)=0.96). There was a good agreement between the two tests except at the detection cutoff of the real-time PCR assay. CONCLUSION: Our RtF-LAMP protocol appears to be precise, accurate and rapid. It could be a valuable tool for the detection of HBV in large clinical and epidemiological studies.

[Characteristics of death cases caused by road traffic accident in Shenzhen].

OBJECTIVE: By summarize the characteristics of death cases caused by road traffic accident, to provide information and data for prevention of traffic accident. METHODS: To retrospectively analyze 4148 death cases caused by road traffic accident in Shenzhen. The characteristics studied include the age and sex of the dead, the cause of death, time and place of the accidents and vehicle types, etc. RESULTS: The death was mainly male and the proportion of male to female is 2.45:1; the accidents mainly occured in 6:00-8:00 and 18:00-2:00; 72% of the accidents took place on the main suburban roads. The proportions for each traffic modes of the death were: 44% of walking, 19% of bike, 15% of motorbike. Most traffic accidents were induced by truck, 83.2% of the death causations were severe head injury and 13.3% were complex injuries. CONCLUSION: The death cases of road-traffic accident in Shenzhen has obvious characteristic and maybe is preventible.

Mindset switching increases the use of 'want-based' over 'should-based' behaviors.

This study examines the consequences of mindset switching on behavioral choices in want/should conflicts. Building on the insights of the ego depletion literature, we propose that mindset switching depletes individuals' self-control resources and therefore prompts the choice of want behavior, which provides immediate pleasure, over should behavior, which provides long-term utility. Four laboratory experiments with university students that stimulated individuals to switch mindsets were conducted to test our hypotheses. Experiment 1 demonstrated that switching between individualist and collectivist mindsets increased the subjects' tendency to prefer popular magazines over scientific journals. Experiment 2 replicated the results by testing the relationship between an abstract/concrete mindset-switching task and want/should online behavioral choices. The mediating effect of ego depletion was also supported. Experiment 3 retested the main effect of language-switching on reading choices, and the mediating effect of ego-depletion. Experiment 3 also tested the moderating effect of the Need for Cognition, and eliminated the alternative explanation of cognitive fatigue. In Experiment 4, actual food choices were used as the direct measure of want/should behaviors to test the robustness of our findings. The results consistently supported our hypotheses that mindset switching has significant effects on behavioral choices in terms of overindulgence, such as increasing want behavior and thus foregoing should behavior.

[Experimental Research of Circular RNA hsa_circ_0000254 in Acute Myeloid Leukemia].

OBJECTIVE: To clone the circular RNA hsa_circ_0000254 and construct its lentiviral over-expression vector. METHODS: The sequence of hsa_circ_0000254 (a total of 524 bp long) was synthesized and cloned by using pGH vector. The vector was cut by EcoR I and BamH I, and artificial hsa_circ_0000254 was obtained, then inserted in pLCDH-ciR to construct the recombinant expression vector pLCDH-circ254(C254), which was confirmed by DNA sequencing. The lentiviral expression vectors pLCDH-circ254(C254) and NC(pLCDH-ciR) were cotransfected into 293T cells by lipofectamineTM 2000(lipo2000). After transfection for 40 hours, the cells were collected and verified by PCR and sequencing. RESULTS: Restriction analysis and DNA sequencing demonstrated that the lentiviral vector pLCDH-circ254(C254) was constructed successfully, the expression efficiency increased 10000 times after transfection of cells. CONCLUSION: The successful construction of the lentiviral expression vector pLCDH-circ254(C254) results in the production of high-titer lentivirus, so as to facilitate further study of the molecular functions of hsa_circ_0000254.

Microvascular Myocardial Ischemia in Patients With Diabetes Without Obstructive Coronary Stenosis and Its Association With Angina

Objective@#To investigate the incidence of microvascular myocardial ischemia in diabetic patients without obstructive coronary artery disease (CAD) and its relationship with angina. @*Materials and Methods@#Diabetic patients and an intermediate-to-high pretest probability of CAD were prospectively enrolled. Non-diabetic patients but with an intermediate-to-high pretest probability of CAD were retrospectively included as controls. The patients underwent dynamic computed tomography-myocardial perfusion imaging (CT-MPI) and coronary computed tomography angiography (CCTA) to quantify coronary stenosis, myocardial blood flow (MBF), and extracellular volume (ECV). The proportion of patients with microvascular myocardial ischemia, defined as any myocardial segment with a mean MBF ≤ of 100 mL/min/100 mL, in patients without obstructive CAD (Coronary Artery Disease–Reporting and Data System [CAD-RADS] grade 0–2 on CCTA) was determined. Various quantitative parameters of the patients with and without diabetes without obstructive CAD were compared. Multivariable analysis was used to determine the association between microvascular myocardial ischemia and angina symptoms in diabetic patients without obstructive CAD. @*Results@#One hundred and fifty-two diabetic patients (mean age: 59.7 ± 10.7; 77 males) and 266 non-diabetic patients (62.0 ± 12.3; 167 males) were enrolled; CCTA revealed 113 and 155 patients without obstructive CAD, respectively. For patients without obstructive CAD, the mean global MBF was significantly lower for those with diabetes than for those without (152.8 mL/min/100 mL vs. 170.4 mL/min/100 mL, P < 0.001). The mean ECV was significantly higher for diabetic patients (27.2% vs. 25.8%, P = 0.009). Among the patients without obstructive CAD, the incidence of microvascular myocardial ischemia (36.3% [41/113] vs. 10.3% [16/155], P < 0.001) and interstitial fibrosis (69.9% [79/113] vs. 33.3% [8/24], P = 0.001) were significantly higher in diabetic patients than in the controls. The presence of microvascular myocardial ischemia was independently associated with angina symptoms (adjusted odds ratio = 3.439, P = 0.037) in diabetic patients but without obstructive CAD. @*Conclusion@#Dynamic CT-MPI + CCTA revealed a high incidence of microvascular myocardial ischemia in diabetic patients without obstructive CAD. Microvascular myocardial ischemia is strongly associated with angina.

The formula of ω-3 polyunsaturated fatty acids improving cognitive impairment in patients with depres-sion:a clinical randomized double-blind controlled trial / 中国神经精神疾病杂志

Objective To investigate the effects of different ratios of ω-3 polyunsaturated fatty acids(ω-3 PUFA)on depression and cognitive impairment in patients with major depression.Methods A randomized,double-blinded controlled trial was used to randomly assign patients with depression to a cognitive improvement group,a depression improvement group,and a placebo group.The cognitive improvement group took 1388 mg of docosahexaenoic acid(DHA)and 692 mg of eicosapentaenoic acid(EPA)every day and the depression improvement group took 1248 mg of EPA and 832 mg of DHA every day.The placebo group took the same dose of soybean oil for 12 weeks,during which psychiatric medication was maintained.The 24-item Hamilton depression scale(HAMD-24)was used to evaluate depressive symptoms,and the standardized MATRICS consensus cognitive battery(MCCB)was used to evaluate cognitive function after 6 weeks and 12 weeks,respectively.Results The study recruited a total of 46 patients with depression including 22 in the cognitive improvement group,12 in the depression improvement group,and 12 in the placebo group.After 6 weeks of treatment,the HAMD-24 scores were significantly lower in the depression improvement group(19.00±10.70)and cognitive improvement group(16.58±9.39)than in the placebo group(31.10±10.03)(P<0.01).After 12 weeks of treatment,HAMD-24 scores were significantly lower in the depression improvement group(13.58±8.43)than in the placebo group(28.10±15.04)(P=0.02).No significant interaction effect was found on the cognitive assessment scores in any dimension after 6 weeks and 12 weeks of treatment(P>0.05).The incidence rate of adverse events in the depression improvement group was 16.7%(2/12),and no adverse events were reported in the other two groups.There was no significant difference in the incidence of adverse events among the three groups(P=0.13).Conclusion Treatment with ω-3PUFA for 6 weeks can improve the depressive symptoms of patients with depression.The formula with a higher ratio of EPA exhibits higher effectiveness while the two groups of ω-3PUFA formulas with different ratios do not improve cognitive function.

CT Fractional Flow Reserve for the Diagnosis of Myocardial Bridging-Related Ischemia: A Study Using Dynamic CT Myocardial Perfusion Imaging as a Reference Standard

Objective@#To investigate the diagnostic performance of CT fractional flow reserve (CT-FFR) for myocardial bridging-related ischemia using dynamic CT myocardial perfusion imaging (CT-MPI) as a reference standard. @*Materials and Methods@#Dynamic CT-MPI and coronary CT angiography (CCTA) data obtained from 498 symptomatic patients were retrospectively reviewed. Seventy-five patients (mean age ± standard deviation, 62.7 ± 13.2 years; 48 males) who showed myocardial bridging in the left anterior descending artery without concomitant obstructive stenosis on the imaging were included. The change in CT-FFR across myocardial bridging (ΔCT-FFR, defined as the difference in CT-FFR values between the proximal and distal ends of the myocardial bridging) in different cardiac phases, as well as other anatomical parameters, were measured to evaluate their performance for diagnosing myocardial bridging-related myocardial ischemia using dynamic CT-MPI as the reference standard (myocardial blood flow < 100 mL/100 mL/min or myocardial blood flow ratio ≤ 0.8). @*Results@#ΔCT-FFRsystolic (ΔCT-FFR calculated in the best systolic phase) was higher in patients with vs. without myocardial bridging-related myocardial ischemia (median [interquartile range], 0.12 [0.08–0.17] vs. 0.04 [0.01–0.07], p < 0.001), while CT-FFR systolic (CT-FFR distal to the myocardial bridging calculated in the best systolic phase) was lower (0.85 [0.81–0.89] vs. 0.91 [0.88–0.96], p = 0.043). In contrast, ΔCT-FFRdiastolic (ΔCT-FFR calculated in the best diastolic phase) and CT-FFR diastolic (CT-FFR distal to the myocardial bridging calculated in the best diastolic phase) did not differ significantly. Receiver operating characteristic curve analysis showed that ΔCT-FFRBsystolic had largest area under the curve (0.822; 95% confidence interval, 0.717–0.901) for identifying myocardial bridging-related ischemia. ΔCT-FFRsystolic had the highest sensitivity (91.7%) and negative predictive value (NPV) (97.8%). ΔCT-FFRdiastolic had the highest specificity (85.7%) for diagnosing myocardial bridging-related ischemia. The positive predictive values of all CT-related parameters were low. @*Conclusion@#ΔCT-FFR systolic reliably excluded myocardial bridging-related ischemia with high sensitivity and NPV. Myocardial bridging showing positive CT-FFR results requires further evaluation.

Clinical application of WeChat MDT in the perioperative period of secondary hyperparathyroidism / 中华内分泌外科杂志

Objective:To observe the application effect of the multi-disciplinary team (MDT) process and program supported by WeChat group in the perioperative period of secondary hyperparathyroidism (SHPT) .Methods:A total of 80 SHPT patients who were hospitalized in Hunan Provincial People’s Hospital and Qidong County People’s Hospital from Jul. 2017 to Oct. 2019 were selected and divided into MDT group (40 cases) and control group (40 cases) according to the principle of complete randomization. In MDT group, there were 21 females and 19 males, aged (48.80±9.08) years old, ranging from 26 to 74 years; in the control group, there were 23 females and 17 males, aged (47.90±8.89) years old, ranging from 24 to 74 years. The control group were given a conventional treatment plan, and the MDT group were implemented with the WeChat MDT process on this basis. The perioperative preparation time, operation time, intraoperative blood loss, postoperative extubation time, and continuous full parathyroid were compared between the two groups. The levels of intact parathyroid hormone (iPTH) and blood calcium and phosphorus were compared with the postoperative complications and patient satisfaction in the two groups. The data in this study were analyzed using SPSS 23.0 statistical software.Results:The preoperative preparation time (4.35±1.12) d, operation time (130.00±32.58) min, intraoperative blood loss (15.75±7.89) ml, and postoperative extubation time (3.80±0.82) d in the MDT group were significantly lower than the preoperative preparation time of the control group (6.86±1.85) d, operation time (162.57±41.65) min, intraoperative blood loss (60.75±11.5) ml, postoperative extubation time (5.97±1.25) d ( P<0.05) 1 week after operation, the iPTH (20.86±1.52) pg/ml and blood calcium level (2.23±0.24) mmol/L of the MDT group were significantly lower than those of the control group (103.47±8.27) pg/ml and blood calcium level (2.87±0.21) mmol/L ( P<0.05) , meanwhile the blood phosphorus level of the MDT group (1.52±0.56) mmol/L was significantly higher than the blood phosphorus level of the control group (1.18±0.25) mmol/L ( P<0.05) . The number of complications in the MDT group (20 cases) was significantly lower than the number of complications in the control group (48 cases) ( P<0.05) ; and the satisfaction of treatment in the MDT group (100.00%) was significantly better than that of the control group (80.00%) ( P<0.05) . Conclusion:The WeChat MDT diagnosis and treatment process and treatment plan are safe and effective, which can effectively shorten the operation time, reduce the operation risk, reduce postoperative complications, and increase patient satisfaction. It can be further promoted in the clinic.

Dynamic CT Myocardial Perfusion Imaging in Patients without Obstructive Coronary Artery Disease: Quantification of Myocardial Blood Flow according to Varied Heart Rate Increments after Stress

Objective@#The present study aimed to investigate the association between myocardial blood flow (MBF) quantified by dynamic CT myocardial perfusion imaging (CT-MPI) and the increments in heart rate (HR) after stress in patients without obstructive coronary artery disease. @*Materials and Methods@#We retrospectively included 204 subjects who underwent both dynamic CT-MPI and coronary CT angiography (CCTA). Patients with more than minimal coronary stenosis (diameter ≥ 25%), history of myocardial infarction/ revascularization, cardiomyopathy, and microvascular dysfunction were excluded. Global MBF at stress was measured using hybrid deconvolution and maximum slope model. Furthermore, the HR increments after stress were recorded. @*Results@#The median radiation dose of dynamic CT-MPI plus CCTA was 5.5 (4.5–6.8) mSv. The median global MBF of all subjects was 156.4 (139.8–180.4) mL/100 mL/min. In subjects with HR increment between 10 to 19 beats per minute (bpm), the global MBF was significantly lower than that of subjects with increment between 20 to 29 bpm (153.3 mL/100 mL/min vs. 171.3 mL/100 mL/min, p = 0.027). This difference became insignificant when the HR increment further increased to ≥ 30 bpm. @*Conclusion@#The global MBF value was associated with the extent of increase in HR after stress. Significantly higher global MBF was seen in subjects with HR increment of ≥ 20 bpm.

High diversity of endophytic fungi from the pharmaceutical plant, Heterosmilax japonica Kunth revealed by cultivation-independent approach.

Heterosmilax japonica Kunth is well recognized for its diuretic effects in China. However, little is known about its endophytic fungi. In this study, microbial communities inhabiting the stems of H. japonica in spring and summer were investigated by light microscopy and cultivation-independent approaches, such as RFLP analysis and sequencing of rDNA ITS library. Molecular phylogenetic analysis showed that a broad spectrum of fungi, including Mycosphaerella, Phomopsis, Aureobasidium, Cladosporium, Glomerella, Botryosphaeria, Guignardia, is able to colonize the plants internally. Particularly, several rDNA sequences determined in this study like YJ4-61 are not specifically affiliated with any currently documented fungal sequences in the public database. Several sequence types, such as YJ4-9 and YJ4-70, are significantly similar to some uncultured environmental samples. Furthermore, our result also showed that the samples collected in spring harbored more abundant endophytic populations than that in summer, implying a seasonal fluctuation for the endophytes in H. japonica.

[Effect of panaxadiol saponin and panaxtrol saponin on proliferation of human bone marrow hemopoietic progenitor cells].

OBJECTIVE: To observe the effect of panaxadiol saponin (PDS) and panaxtrol saponin (PTS) on proliferation of human bone marrow hemopoietic progenitor cells (HPC). METHODS: PDS and PTS were separated and purified from ginsenosides, and the effects on HPC were studied using in vitro hemopoietic progenitor cell colony-forming technique, by observing the proliferation of human burst forming unit-erythroid progenitor (BFU-E), colony-forming unit-erythroid (CFU-E), colony-forming unit-granulocyte/macrophage (CFU-GM) and colony-forming unit-pluripotent hemopoietic progenitor (CFU-Mix) in mice after PDS and PTS stimulation. RESULTS: Different concentration of PDS (2.5-200 micrograms/ml) could stimulate the proliferation of HPC obviously, showing increase of CFU-E, BFU-E, CFU-GM and CFU-Mix by 54.9 +/- 6.3%, 48.8 +/- 5.1%, 27.6 +/- 4.2% and 48.9 +/- 3.9% respectively, which was higher than that of the control group. While stimulated by PTS of the same concentration, the CFU-E and BFU-E was lower than that of control significantly (P < 0.05); when the terminal concentration of PTS was 200 micrograms/ml, CFU-E and BFU-E was zero respectively. In the CFU-GM culture, PTS in concentration of 12.5 micrograms/ml could cause the proliferation increased by 29.7 +/- 2.2% (P < 0.05), but in concentration of 100 micrograms/ml and 200 micrograms/ml, it showed inhibitory effect on CFU-GM, the inhibition rate being 48.6 +/- 3.9% and 100% respectively. CONCLUSION: PDS is the effective component of ginsenosides in stimulating proliferation of human bone marrow HPC. PTS is an component with inhibitory action on proliferation of CFU-E and BFU-E and its effect on CFU-GM was depending on its concentration.

Effects of sodium copper chlorophyllin on mesenchymal stem cell function in aplastic anemia mice.

OBJECTIVE: To investigate the effects of sodium copper chlorophyllin (SCC) on the proliferation, differentiation and immunomodulatory function of mesenchymal stem cells (MSCs) from mice with aplastic anemia. METHODS: A mouse model of aplastic anemia was established by exposure of BALB/c mice to sublethal doses of 5.0 Gy Co60 γ radiation, followed by transplantation of 2×10(6) lymph node cells from DBA/2 donor mice within 4 h after radiation. Aplastic anemic BALB/c mice were randomly divided into six groups: the treated groups, which received 25, 50, or 100 mg/kg/day SCC, respectively; a positive control group treated with cyclosporine A (CsA); and an untreated model control group (model group); while, the non-irradiated mice as the normal control group. SCC or CsA were administered by gastrogavage for 20 days, starting on day 4 after irradiation. Peripheral blood cells were counted and colony-forming fibroblasts (CFU-F) in the bone marrow were assayed. The ability of MSCs to form calcium nodes after culture in osteoinductive medium was also observed. The immunosuppressive effect of MSCs on T lymphocytes was analyzed by enzyme-linked immunosorbent assay and flow cytometry, to evaluate the efficacy of SCC in mice with aplastic anemia. RESULTS: Peripheral blood white cell and platelet counts were increased by medium and high SCC doses, compared with the untreated control. CFU-Fs were also increased compared with the untreated control, and the numbers of calcium nodes in MSCs in osteoinductive medium were elevated in response to SCC treatment. The percentage of Forkhead box protein 3 (FOXP3(+)) T cells was increased in T cell-MSC cocultures, and the cytokine transforming growth factor ß1 was up-regulated in SCC-treated groups. CONCLUSION: The results of this study suggest that SCC not only promotes the proliferation and differentiation of MSCs, but also improves their immunoregulatory capacity in mice with aplastic anemia.

Experimental Research of Circular RNA hsa_circ_0000254 in Acute Myeloid Leukemia / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To clone the circular RNA hsa_circ_0000254 and construct its lentiviral over-expression vector.</p><p><b>METHODS</b>The sequence of hsa_circ_0000254 (a total of 524 bp long) was synthesized and cloned by using pGH vector. The vector was cut by EcoR I and BamH I, and artificial hsa_circ_0000254 was obtained, then inserted in pLCDH-ciR to construct the recombinant expression vector pLCDH-circ254(C254), which was confirmed by DNA sequencing. The lentiviral expression vectors pLCDH-circ254(C254) and NC(pLCDH-ciR) were cotransfected into 293T cells by lipofectamine 2000(lipo2000). After transfection for 40 hours, the cells were collected and verified by PCR and sequencing.</p><p><b>RESULTS</b>Restriction analysis and DNA sequencing demonstrated that the lentiviral vector pLCDH-circ254(C254) was constructed successfully, the expression efficiency increased 10000 times after transfection of cells.</p><p><b>CONCLUSION</b>The successful construction of the lentiviral expression vector pLCDH-circ254(C254) results in the production of high-titer lentivirus, so as to facilitate further study of the molecular functions of hsa_circ_0000254.</p>

Effects of Panax notoginseng saponins on proliferation and differentiation in NIH3T3 cells.

OBJECTIVE: To investigate the effects of Panax notoginseng saponins (PNS) on the proliferation and differentiation in NIH3T3 cells. METHODS: NIH3T3 cells were treated by various concentrations of PNS 0, 0.05, 0.10, 0.20, and 0.40 g/L. The vitality and proliferation potential of cells were detected by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, the alkaline phosphatase (ALP) activity was measured by p-nitrophenyl phosphate (pNPP) assay, and the mineralization formation ability was tested for the cellular differentiation toward osteoblast, as well as the expression level of phosphorylated extracellular signal-regulated kinase1/2(P-ERK1/2), extracellular signal-regulated kinase1/2 (ERK1/2) protein kinase was analyzed by Western blot with total cell lysate of NIH3T3 cells treated by PNS. RESULTS: Both MTT and pNPP assay showed that optical density (OD) values were increased in response to PNS treatment at a dose-dependent pattern. The mineralization formation ability was enhanced in PNS-treated NIH3T3 cells compared with untreated cells. Meanwhile, the expression level of P-ERK1/2 protein kinase was up-regulated in PNS-treated NIH3T3 cells, while, the expression level of ERK1/2 protein kinase revealed no obvious difference with or without PNS treated cells. CONCLUSION: PNS could pay a role to promote the proliferation and differentiation in NIH3T3 cells by means of up-regulation of P-ERK1/2 protein kinase.

p53 gene mutations of familial breast cancer and early-onset breast cancer in part population of southern China / 中南大学学报(医学版)

Objective:To investigate the site and characteristic ofp53 gene mutations in familial or early-onset breast cancer patients in part population of southern China.Methods:A total of 150 patients with familial and early-onset breast cancer in parts population of southern China were enrolled.Genomic DNA was isolated from each peripheral blood sample,and the entire coding sequence and exon and intron splicing region of p53 gene were amplificated by PCR in the 150 patients.The mutation analysis were detected by denaturing high performance liquid chromatography (DHPLC) and confirmed by DNA sequence analysis.Results:In the 150 patients with familial and early-onset breast cancer,6 mutations including one novel pathogenic mutation 869_888 ins20 (insert mutation) and 5 previously reported pathogenic mutations (deletion mutation 643_660de118 and 4 missense mutation 91G＞A,215C＞G,537T＞G,743G＞A) were identified in p53 gene encoding region in 9 patients of breast cancer.Moreover,one same sense mutation 141G＞A in exon 4,one 16 bases deletion in intron 3,and 9 single nucleotide polymorphisms in p53 gene introns were also identified.The total mutation frequency ofp53 gene in 150 patients with familial breast cancer and early-onset breast cancer from part population of southern China was 6.00％,and the mutation frequency of familial breast cancer and early-onset breast cancer was 6.81％ and 6.25％,respectively.Conclusion:The total mutation frequency ofp53 gene in 150 patients with familial breast cancer and early-onset breast cancer from partpopulation of southern China is higher than the frequency previously reported.The pathogenicity of the novel mutations (insert mutation) 869_888ins20 will be confirmed by function analysis in the future study.The deletion mutation 643_660de118 enriches the p53 gene mutation database among Chinese population,which is probably the specific mutation of breast cancer in Chinese population.