In order to investigate the effect of vitamin A (VA) on the secretion of IFN-gamma and IL-4 in Mycoplasma Pneumoniae (MP)-induced A549 cells, A549 cells were co-cultured with MP for different time lengths and then the levels of IFN-gamma and IL-4 in the cell culture supernatants were detected before and after treatment with different concentrations of VA by using the enzyme-linked immunosorbent assay (ELISA). The results showed that the level of IFN-gamma and IL-4 in the supernatants of MP-induced A549 cells was much higher than that in non-induced cells (P<0.01). After application of VA, IL-4 level was not increased until the concentration of VA was up to 0.5x10(-5) mol/L (P<0.01). However, with concentration of VA increased up to 1x10(-4) mol/L, IL-4 was significantly suppressed (P<0.01). It was concluded that MP could induce the secretion of IFN-gamma and IL-4 in A549 cells. VA could inhibit the secretion of IFN-gamma and increase the IL-4 level in MP-induced A549 cells. However, high concentration of VA had an inhibitory effect on the secretion of IL-4 as well as on the IFN-gamma. These data provided a theoretical basis for the application of VA in MP pneumonia in the clinical practice.
Cell Line, Tumor ; Coculture Techniques ; Culture Techniques ; Interferon-gamma/*secretion ; Interleukin-4/*secretion ; Lung Neoplasms/pathology ; Mycoplasma pneumoniae/growth & development ; Mycoplasma pneumoniae/*physiology ; Vitamin A/*pharmacology

Objective To apply nitric oxide(NO) electrochemical microsensor in the real time detection of NO released from RAW 264 .3 cells infected by E .coli ,and to explore the application value of this NO microsensor in the research area of infection im‐munity against bacterium .Methods Taking NO microsensor to detect NO released from RAW 264 .3 cells respectively stimulated by E .coli of different densities and of 1 × 107 mL -1 for different time .Results The level of NO released from RAW 264 .3 cells was enhanced obviously when incubated with E .coli as compared with that of normal cells and the extent of incersase depended on the density of E .coli (P<0 .01) .The released level of NO increased gradually from the beginning and reached its peal at the time of 12 h then decreased slowly when incubated with E .coli of 1 × 107 mL -1 .Conclusion The electrochemical microsensor was applied in the real time detection of NO released from macrophages activated by E .coli successfully .

AIM: To construct an eukaryotic expression vector of human single-chain variable fragment ~against hepatitis B virus core protein (anti-HBc ScFv) and detect its expression in HepG2 cells. METHODS: Anti-HBc ScFv genes were amplified from the plasmids abstracted from positive clone and inserted into pEGFP-c1 vector that contained green fluorescent protein gene. The recombinant plasmids were transfected into HepG2 cells, and resistant clones were obtained by G418 selection. The expression of the gene of fusion protein was determined by fluorescent invert microscope and ELISA. RESULTS: Recombinant plasmids were successfully constructed. The plasmid transfected HepG2 cells were obtained by G418 selection. Specific fluorescence was observed in HepG2 cells 48 hours after transfection. ELISA analysis confirmed the expression of anti-HBc ScFv in the cells. CONCLUSION: The construction of human anti-HBc ScFv eukaryotic expression vector and its expression in HepG2 cells lay the foundation for advanced research of intracellular anti-HBc ScFv.

AIM: To Screen and identify human single-chain variable fragment (ScFv) specific to hepatitis B virus core protein and determine its gene sequence. METHODS: The recombinant phages were panned by HBcAg coated in a 96-pore plate and 48 clones were identified specific to HBc after three rounds of panning. The specificity of ScFv from the positive clone was determined by ELISA. Then, the soluble ScFv was expressed in E.coli. HB2151 and secreted in the supernatant. Subsequently, SDS-PAGE and dot blot were performed to identify the ScFv in the supernatant and cell lysate. The gene of ScFv specific to hepatitis B virus core protein was sequenced. RESULTS: The ScFv screened from phage antibodies has a specific combination character with hepatitis B virus core antigen. Soluble ScFv was confirmed to express in E.coli. HB2151 and secrete in the supernatant. The sequence of ScFv gene conformed to that of heavy chain and kappa chain of human immunoglubulin. CONCLUSION: Human ScFv specific to hepatitis B virus core protein has been identified by means of the phage display technology, and its gene sequence has been determined.

In order to investigate the effect of vitamin A (VA) on the secretion of IFN-γ and IL-4 in Mycoplasma Pneumoniae (MP)-induced A549 cells, A549 cells were co-cultured with MP for different time lengths and then the levels of IFN-γ and IL-4 in the cell culture supematants were detected before and after treatment with different concentrations of VA by using the enzyme-linked immunosorbent assay (ELISA). The results showed that the level of IFN-γ and IL-4 in the supernatants of MP-induced A549 cells was much higher than that in non-induced cells (P<0.01). After application of VA, IL-4 level was not increased until the concentration of VA was up to 0.5 × 10-5 mol/L (P<0.01).However, with concentration of VA increased up to 1 × 10-4 mol/L, IL-4 was significantly suppressed (P<0.01). It was concluded that MP could induce the secretion of IFN-y and IL-4 in A549 cells. VA could inhibit the secretion of IFN-γ, and increase the IL-4 level in MP-induced A549 cells. However,high concentration of VA had an inhibitory effect on the secretion of IL-4 as well as on the IFN-γ.These data provided a theoretical basis for the application of VA in MP pneumonia in the clinical practice.

Objective:To explore the risk factors of acute kidney injury (AKI) in acute respiratory distress syndrome (ARDS) patients with mechanical ventilation.Methods:A retrospective analysis was conducted. ARDS patients with mechanical ventilation admitted to ICU of Taizhou People's Hospital from January 2019 to December 2019 were enrolled. Patients were divided into the AKI group and non-AKI group according to whether the patients had AKI. Clinical characteristics and laboratory indicators of the two groups were compared. Risk factors of incidence of AKI in ARDS patients were analyzed. The Kaplan-Meier survival curve was drawn to evaluate the survival rates of the two groups.Results:A total of 120 ARDS patients with mechanical ventilation were included, and 57 patients (47.5%) developed AKI. Procalcitonin, increased basal creatinine, decreased pH and impaired consciousness were independent risk factors for AKI in ARDS patients with mechanical ventilation. Fifty-seven of the 120 patients died with a mortality of 38.3%. The Kaplan-Meier survival curve showed that the survival rate of the AKI group was significantly lower than that of the non-AKI group ( P<0.001). Conclusions:The incidence and mortality of AKI is high in ARDS patients with mechanical ventilation. Procalcitonin, increased basal creatinine, decreased pH and impaired consciousness are independent risk factors for AKI in ARDS patients with mechanical ventilation.