0.05). It is decrease greatly, than those in anti-VEGF control (P0.05).The toxicity of CRM9 control are markedly higher than those in blank control groups (P

To study the function and potential application of nkx2.5, a critical gene for heart development, we constructed a recombinant adenovirus overexpressing nkx2.5 gene (Ad-Nkx2.5) with the AdEasy system. To evaluate the effect and mechanism of Ad-Nkx2.5 against oxidative injury, the H9c2 myocardial cells were infected with the recombinant adenoviruses Ad-Nkx2.5 or Ad-EGFP, and subsequently exposed to H2O2 to induce apoptosis. The anti-apoptotic potential of Ad-Nkx2.5 was validated by MTT assay for cell viability, Hoechst33342 staining for cellular morphology, and immunoblotting for caspase-3 activity. Ad-Nkx2.5 infection led to an increased survival rate of H9c2 cells and decreased the amount of caspase-3 in an active form. Additionally, overexpression of Nkx2.5 inhibited the release of cytochrome C from the mitochondria into the cytosol. Mechanismic studies showed that Nkx2.5 upregulated bcl-2 gene expression and significantly repressed H2O2-induced expression of bax detected by Real-time PCR. Additionally, H2O2 treatment did not affect the nuclear localization of Nkx2.5. These findings indicate that adenovirus-mediated nkx2.5 gene transfer exerted a protective effect on H9c2 cells against H2O2-induced apoptosis via mitochondrial pathway, and the Nkx2.5-mediated expression modulation of apoptosis-associated genes could be involved in this event.
Adenoviridae ; genetics ; metabolism ; Animals ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Cell Line ; Genetic Vectors ; genetics ; Homeobox Protein Nkx-2.5 ; Homeodomain Proteins ; biosynthesis ; genetics ; Hydrogen Peroxide ; pharmacology ; Myocytes, Cardiac ; cytology ; Oxidative Stress ; drug effects ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rats ; Recombinant Proteins ; biosynthesis ; genetics ; Transcription Factors ; biosynthesis ; genetics ; bcl-2-Associated X Protein ; metabolism

Objective To investigate the influencing factors involved in the establishment of a C57BL/6 J model of metastatic melanoma in the lung,including the way of tumor inoculation,the number of inoculated cells and the time of tumor formation. Methods Mouse melanoma B16F10 cells were cultured in vitro. 1)Eighteen healthy male C57BL/6 J mice were randomly divided into three groups. Mice in each group received 100 μL cell suspension(including 3 ×106 melanoma cells)via intravenous,intraperitoneal and subcutaneous injection,respectively. After two weeks,the mice were killed and dissected,and the tumor growth and metastasis were observed. 2)Eighteen male mice were randomly divided into three groups. Mice in each group were injected with 3 ×106cells,1 ×106cells, and 3 ×105cells through the tail vein,respectively. After two weeks,mice were killed and dissected,and the tumor growth and metastasis were observed. 3)Eighteen male mice were randomly divided into three groups. Mice in each group were injected with 1×106cells though the tail vein. Mice were killed and dissected after one week, two weeks and three weeks, respectively. The growth and metastasis of tumor were observed. Results 1)The success rate of lung metastasis was 100% in the mice with intravenous injection,but not in mice receiving intraperitoneal injection and subcutaneous injection. 2)The size of metastatic melanoma nodules were moderate in mice inoculated by 1 ×106cells. The number of melanoma metastatic foci was too high in the mice inoculated with 3 ×106cells,but too low in the mice inoculated with 3 ×105cells. 3)Significant metastatic melanoma foci were observed in the mice killed and dissected after two weeks with no death. The number of melanoma foci in the lung was too high in the mice killed after three weeks,while was too low in the mice killed at one week after tumor cell inoculation. Conclusions Intravenous injection of 1×106mouse melanoma cells into C57BL/6 J mice and killed after two weeks is an optimal method for establishment of a mouse model of metastatic melanoma in the lung, and is worth of recommendation.

Objective:To compare the therapeutic effects and safety of dexmedetomidine and midazolam on patients with severe coronavirus disease 2019 (COVID-19) who received non-invasive ventilation.Methods:Patients with COVID-19 who needed non-invasive ventilation in one critical care medicine ward of Wuhan Jinyintan Hospital during the team support period from the department of critical care medicine of Renmin Hospital of Wuhan University from January 23rd to February 15th in 2020 were investigated retrospectively. Ramsay score, mean arterial pressure (MAP), heart rate (HR), respiratory rate (RR), arterial oxygen partial pressure (PaO 2) before sedation and at 1, 12, 24 hours after sedation, sleep time were collected, and the side effects such as excessive sedation, fall of tongue, abdominal distension, aspiration, bradycardia, escalation to invasive mechanical ventilation during 24 hours were also collected. According to different sedative drugs, patients were divided into the control group (without sedative drugs), dexmedetomidine group and midazolam group. The changes of indicators among the three groups were compared. Results:Fourteen patients were injected with dexmedetomidine (loading dose of 1 μg/kg for 10 minutes, maintained at 0.2-0.7 μg·kg -1·h -1); 9 patients were injected with midazolam (loading dose of 0.05 mg/kg for 2 minutes, maintained at 0.02-0.10 mg·kg -1·h -1); 12 patients didn't use sedative drugs due to limitations of previous hospital or patients' rejection. In dexmedetomidine group and midazolam group, the Ramsay score was maintained at 2-3 points after sedation, which were higher than those of control group at different time points after sedation, and there was no significant difference between dexmedetomidine group and midazolam group. MAP of dexmedetomidine group and midazolam group decreased gradually after sedation. MAP after 1-hour sedation was significantly lower than that before sedation, and MAP after 24 hours sedation was significantly lower than that in the control group [mmHg (1 mmHg = 0.133 kPa): 109.7±11.5, 107.1±12.3 vs. 121.1±13.3, both P < 0.05]. HR decreased gradually after sedation treatment, which was significantly lower after 12 hours of sedation than that before sedation, and HR in dexmedetomidine group was significantly lower than that in control group after 12 hours of sedation (bpm: 84.0±13.9 vs. 92.8±15.4 at 12 hours; 81.0±16.7 vs 92.6±12.7 at 24 hours, both P < 0.05). PaO 2 increased and RR decreased in all three groups after ventilation. PaO 2 in dexmedetomidine group and midazolam group were significantly higher than that in the control group after 12 hours of sedation [cmH 2O (1 cmH 2O = 0.098 kPa): 79.0±6.5, 79.0±8.9 vs. 70.0±7.8, both P < 0.05]; the decreases of RR in dexmedetomidine group and midazolam group were significant than that in control group after 1 hour of sedation (bpm: 34.0±3.9, 33.8±4.6 vs. 39.0±3.6, both P < 0.05). There were no differences of MAP, HR, PaO 2 and RR between dexmedetomidine group and midazolam group at different time points. The sleep duration in dexmedetomidine group and midazolam group were significantly longer than that in the control group (hours: 4.9±1.9, 5.8±2.4 vs. 3.0±1.8, both P < 0.05), but there was no difference between dexmedetomidine group and midazolam group ( P > 0.05). Adverse events occurred in all three groups. In midazolam group, there were 2 cases of excessive sedation with fall of tongue and abdominal distension, including 1 case of aspiration, 1 case receiving intubation due to refractory hypoxemia and 1 case due to unconsciousness. In dexmedetomidine group, there were 2 cases of bradycardia, 1 case of intubation due to refractory hypoxemia. In control group, 4 cases underwent intubation due to refractory hypoxemia. Conclusions:Non-invasive mechanical ventilation is an important respiratory support technology for patients with severe COVID-19. Appropriate sedation can increase the efficiency of non-invasive mechanical ventilation. Dexmedetomidine is more effective and safer than midazolam in these patients, but attention should be paid to HR and blood pressure monitoring.

To prepare polyclonal antibodies (PcAb) against UspA1 of Moraxella catarrhalis (Mc), we used bioinformatic analysis to determine the surface exposed region in this protein that holds the antigen epitopes. Then the corresponding coding sequences for this fragment was artificially synthesized according to the codon usage of Escherichia coli. The gene fragment was then subcloned into the prokaryotic expression vector pET-28a(+) and expressed in E. coli rosseta (DE3), and then the recombinant UspA1-His proteins were purified. Two New Zealand white rabbits were immunized with this protein to prepare antiserum. The resulting PcAb was then purified from the antiserum with Protein A affinity column. The results of fluorescence antibody assay, enzyme linked immunosorbent assay and Western blotting analysis showed that the PcAb could specifically recognize the surface exposed region of UspA1 on Mc. The preparation of the PcAb laid a foundation of further development of rapid detection technique for M. catarrhalis.