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Objective To clarify the mechanism of immediate early response gene 5 (ler5)transcription induced by radiation. Methods Deletant construction, site-specific mutagenesis,electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) were used to forecast the promoter region,binding sites and transcription factors of Ier5 gene in HeLa cells.Results The promoter region of Ier5 gene might be in the region of Ier5 -8 deletant ( -408 - -238 bp).The Ier5 gene had two transcription factors of GCF and NFI,and GCF had two binding sites located in the region of - 388 - - 382 bp and - 274 - - 270 bp of Ier5 promoter.The binding site of NFI was located in - 362 --357 bp of Ier5 promoter. GCF could inhibit the expression of Ier5 gene and this inhibition was diminished when the radiation dose increased. In contrast, NFI increased the expression of Ier5.Conclusions The most possible region of Ier5 promoter is from -408 to - 238 bp which has two binding sites for the radiosensitivity transcription factors of GCF and NFI that could negatively and positively regulate the expression of Ier5 respectively.
RESUMO
Summary: The expression changes of early response genes due to ventilation with high volume in adult rats in vivo were observed. Forty SD male rats were randomly divided into control and 30, 60, 90 and 120 min ventilation groups, respectively (n=8 in each group). The animals were ventilated with tidal volume of 42 ml/kg and a PEEP level of 0 cmH2O at a rate of 40 breaths per minute in room air with a ventilator was given to the small animals. The expression of Egr-1, C-jun and IL-1β mRNA and proteins was detected by RT-PCR and immunohistochemical technique, respectively. The pathological changes in lung tissues were examined by HE staining. The results indicated that the expression of Egr-1, C-jun and IL-1β mRNA was detectable at 30th min after overventilation, but there was no significant difference in comparison with that in control group until overventilation for 60 min. However, at 90 and 120 min there was a significent increase as compared with 30 min or control group (P<0.05). The expression of Egr-1, C-jun and IL-1β deteced by immunohistochemical assay also showed a similar tendency of the gradual increase. In the 120 min ventilation group, the expression intensity of Egr-1, C-jun and IL-1β proteins in lung cells was the strongest and the nuclear translocation was increased markedly in comparison with any other groups (P<0.05). HE staining suggested that the degree of lung injury was aggravated gradually with the ventialtion going on and had a similar tendency to the expression of these early response genes and proteins. The current data suggested that overventilation activated and upregulated the expression of early response genes and the expression of these genes may be taken as the early signal to predict the onset and degree of lung injury. These results may demonstrated partially that the expression of early response genes induced by the mechanical stretch is associated with biochamic lung injury.