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Objective::To investigate the effect of ancient recipe Yanghetang and its drug-contained serum on apoptosis of triple negative breast cancer cell line MDA-MB-231 based on the signal pathway of early growth response gene 1 (Egr1)/cyclin dependent inhibitor (p21). Method::The drug-containing serum of Yanghetang was prepared from rats, and MDA-MB-231 cells were cultured in vitro. The blank control group was set, and MDA-MB-231 cells in Yanghetang high, middle, and low dose groups (23.4, 11.7, 5.85 g·kg-1) were intervened for 24, 48, 72 h, respectively. After that, the cell counting kit-8(CCK-8) method was used to detect the cell proliferation of each group. The blank control group was set, while MDA-MB-231 cells in Yanghetang high, middle, and low dose groups (23.4, 11.7, 5.85 g·kg-1) were treated for 48 h, and then flow cytometry was used to detect the apoptosis of each group and the distribution of cell cycle. The expression of Egr1 and p21 mRNA in each group was detected by quantificational Real-time polymerase chain reaction (Real-time PCR), while the expression of Egr1 and p21 protein in each group was detected by Western blot. Result::After MDA-MB-231 cells were intervened by Yanghetang for 24, 48, 72 h, MDA-MB-231 cell proliferation was significantly inhibited in Yanghetang high and middle dose groups as compared with the blank control group (P<0.01). After MDA-MB-231 cells were intervened by Yanghetang for 48 h, the apoptosis was significantly increased in Yanghetang high and middle dose groups as compared with the blank control group (P<0.05, P<0.01). In the Yanghetang high, middle dose groups, the proportion of cell cycle G0/G1 phase decreased, and the proportion of G2/M phase increased (P<0.05, P<0.01). The mRNA and protein expressions of Egr1, p21 were increased in Yanghetang high and middle groups (P<0.05, P<0.01). Conclusion::Yanghetang can activate Egr1/p21 signaling pathway in MDA-MB-231 cells, increase the expression of Egr1 and p21, and cause G2/M cell cycle arrest, thereby inducing apoptosis of MDA-MB-231 cells.
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Objective: To investigate the association between early growth response gene 1 (EGR1) and Alzheimer's disease (AD) in Han Chinese people. Methods: A total of 715 AD patients and 760 health controls were recruited in two independent samples from Eastern China (382 AD patients and 426 normal individuals) and Southwest China (333 AD patients and 334 normal individuals). SNaPshot technique was utilized to analyse the single nucleotide polymorphism (SNP) of rs11743810. A public database was used to explore whether EGR1 gene was differentially expressed in the brain of AD patients and health controls. Then the protein-protein interaction (PPI) assessment was conducted using the STRING database, and the brain eQTL (expression quantitative trait loci) analysis was used to explore the difference in rs11743810 expression between different genotypes in different brain regions. Results: Cross-platform normalized data showed that there was significant difference of EGR1 expression in temporal cortex between AD patients and control subjects (|log2FC|=0.780, P=0.000 before FDR corrected; P=0.001 after FDR corrected). PPI analysis revealed that EGR1 was physically connected with amyloid precursor protein (APP) and clusterin (CLU) protein in the network. However, different genotypes of rs11743810 showed no significant difference in expression in 10 brain regions, and no significant difference in the genotype and allele frequency of rs11743810 between AD patients and controls were found in our two independent samples. Conclusion: The rs11743810 in EGR1 may not be major susceptibility gene site for AD in Han Chinese people.
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Objective To explore the effects of early growth response gene-1 (Egr-1) on bone marrow mesenchymal stem cells (BMSC) proliferation and osteogenic differentiation,which is aimed at providing new molecular targets for the treatment of osteoporosis.Methods Bone marrow was collected from adult men and the BMSCs were cultured primarily and observed by microscope.Meanwhile,flow cytometry was used for BMSCs phenotypic identification;After transfection of pcDNA3.1/Egr-1 into BM SCs,the level of BMSCs proliferation was determined by MTT respectively on the 2 d,4 d and 6 d;On the 7 d after transfection,the ALP activity assay was used for testing the ALP activity in BMSCs.And then,alizarin red S-calcium kit was used for measuring the calcified knots respectively on the 7 d,14 d and 21 d;On the 21 d after transfection,real-time qPCR and Western blotting were used respectively for measuring the expression of mRNA and protein of Egr-1,Runx2 and NDRG1;Further,BMSCs were transfected with Egr-1 siRNA,and the content of calcium nodules,ALP activity,the expression of Egr-1,Runx2 and NDRG1 were detected as above methods.Results The cells cultured in vitro showed high level of CD90 and CD29 and very low level of CD34 and CD45,which is accorded with the characteristic of BMSCs.The pcDNA3.1/Egr-1 transfection for BMSCs had no effect on cells prolifera tion.However,the calcified knots,ALP activity and the expression of Egr 1,Runx2 and NDRG1 were increased after transfection of pcDNA3.1/Egr-1 for BMSCs.In addition,Egr-1 siRNA showed the opposite effect with pcDNA3.1/Egr-1 transfection for BMSCs.Conclusion Egr-1 induces osteogenic differentiation of BMSCs by promoting NDRG1 but has no effects on proliferation of BMSCs.
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Objective To investigate the influence of early growth response gene-1 (EGR1) on the autophagy of host cells following infection with human T cell leukemia virus type 1 (HTLV-1).MethodsA HTLV-1-positive cell line MT2 was co-cultured with HeLa cells for 24 h to construct the virus early infection model.Immunoblotting assay was used to detect the expression of HTLV-1 core protein p19 and EGR1.Luciferase reporter gene analysis was used to detect the transcriptional activity of 5′-regulatory sequence of EGR1 at different time points after co-culturing.An effective small interfering RNA (siRNA) targeting EGR1 was screened out and transfected into HeLa cells by Lipofectamine 2000.Then the transfected HeLa cells were co-cultured with the HTLV-1-positive cell line MT2 for 24 h.Immunoblotting assay was used to detect HTLV-1 core protein p19, EGR1 and autophagy-related protein LC3.Real-time PCR was performed to detect viral load.Autophagosome was analyzed by immunofluorescence after co-culturing.Results The expression of EGR1 and the transcriptional activity of pEGR1-luc gradually increased after co-culturing HeLa cells with MT2 cells for 8 h (P<0.01).The expression of EGR1 was positively correlated with host cell autophagy following HTLV-1 infection.The effective siRNA for silencing the expression of EGR1 was obtained and named as siE2.The viral load, the expression of HTLV-1 core protein p19 and the proportion of LC3B/LC3A in the co-culture model were markedly down-regulated by RNA interference with siE2, which was concomitant with a persistent decrease of intracellular autophagosome (P<0.01).Conclusion EGR1 is associated with host cell autophagy and viral replication in HTLV-1 infection.
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Objective To explore the expression of early growth response gene-1 (EGR-1) in T cells that were positive for Tax protein of human T-cell leukemia virus type 1 (HTLV-1) and its possible reg-ulatory mechanism .Methods A series of expression structures carrying the regulatory elements of EGR-1 in different length and luciferase reporter genes were constructed .TaxP cells were transfected with the con-structs containing reporter genes and cultured with 5μmol/L of NF-κB inhibitor BAY 11-7082 or equal vol-ume of DMSO.After cultured for 24 hours the cells were collected to test the luciferase activity .BAY 11-7082 or equal volume of DMSO was added into the supernatant of TaxP cell culture to test the expression of EGR-1 protein by Western blot after 24 hours of culture .Tax and its mutants M22 and M47 were transfected into 293 T cells respectively to test the expression of EGR-1 protein by Western blot after 24 hours of culture . Results The expression structures carrying the regulatory elements of EGR-1 in different length and their mutants followed by luciferase reporter genes were successfully constructed .The luciferase activity in the cells transfected with the constructs containing the elements E 1 and E2 were higher than that transfected with E3, DelE and MutE, but the reporter gene expressions were decreased with the interference of BAY 11-7082 (P<0.01).However, there were no significant changes with the luciferase activity in the cells transfected by elements E3, DelE and MutE.Western blot analysis indicated that the expression of EGR-1 protein was significantly decreased with the interference of BAY 10-7082 .The expression of EGR-1 protein in M22 mu-tants-transfected 293 T cells were decreased significantly in comparison with those by wild type tax-and M47-transfected cells .Conclusion NF-κB was the key nuclear factor in regulating the expression of EGR-1 pro-tein in Tax-positive T cells .
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Early growth response gene-1 is an immediate early response gene,a transcription factor which includ three zinc finger-like domains.Recentl studies demonstrate that early growth response gene-1 is involved in regulating cell cycle,proliferation and differentiation,and plays a critical role in injuring and repairing.Early growth response gene- 1 induces Target genes,and promotes extracellular environmental signal( such as oxygen deficit,oxidative stress,mechanical injury and cytokine)link with the complex biological responses.Intensive study to early growth respnse gene-1 will contribute to further illustrate the mechanism of the ischemia-reperfusion injury.This article summarizes from protein structure,signal transduction,biological functions and the relationship with ischemia- reperfusion injury.
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Objective To investigate the effect of nerve growth factor (NGF) on the neural cells and expression of early growth response gene-1 (Egr-1 gene) in the hippocampus discharge zone of childhood intractable epileptics. Methods Acutely dissociated cell suspensions of childhood hippocampus discharge zone with non specific pathological changes (n = 16) were exposed to NGF group and control group respectively. There were three subgroups exposed to NGF at the levels of 12.5, 50, 100 ng/ml in NGF group. After 4 hr culturing, the astrocytes and neurons were lablled by Bb immunostain with the specific markers such as GFAP and MAP2. The total number of neural cells was counted under inversion fluorescence microscope and the Egr-1mRNA expression was detected by semiquantitative RT-PCR analysis. Results There were significant differences of the numbers of neural cells survived in hippoeampus region between the NGF group and the non-added NGF group (P < 0.01). Among the different levels of NGF 12.5, 50, 100 ng/ml, the number of the total cultural neurons and GFAP(+) astrocytes, MAP_2(+) neurons were increased with ascending levels of NGF (P < 0.05). At the same time, the expression of Egr-1mRNA (A_(Egr-1)/A_(β-actin)) in the NGF groups was also increased (P < 0.01). There was positive correlation between the number of the survivable neural cells and Egr-1mRNA quantity (r = 0.780, P < 0.01) among the three NGF groups. Conclusions NGF can protect the neural cells of epileptic discharge zone by promoting the expression of Egr-1 gene.
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Objective To study the expression of early growth response gene-1(Egr-1)in esophageal carcinoma tissue,and to explore the relationship between Egr-1 and the survival time of the patients with esophageal carcinoma.Methods RT-PCR was performed to detect the expression of Egr-1 mRNA in68 cases of esophageal carcinoma.The relationship between survival time and prognosis was analyzed.Results The expression of Egr-1 mRNA was the lowest(0.567±0.404),(0.945±0.336)and(1.201±0.347)in esophageal carcinoma tissue,para-cancerous tissue and normal esophageal mucosa tissue,respectively(F=12.709,P＜0.05,P＜0.00).21 cases showed the positive expression of Egr-1 mRNA of both the esophageal carcinoma tissue and the para-cancerous tissue.21 cases showed the positive expression of Egr-1 mRNA in a single esophageal carcinoma tissue or the para-cancerous tissue.26 cases showed the negative expression of Egr-1 mRNA both in the esophageal carcinoma tissue and the paracancerous tissue.The positive rate of Egr-1mRNA expression was 65.71%and 30.00%in the groups of the survival time for three years and the groups of the survival time for one year(P＜0.05).The survival rates in the two groups with positive expression of Egr-1 mRNA were 94.44%and 86.96%,respectively(P＞0.05).Conclusion Decreased level of EGR1 expression may be related to esophageal carcinogenesis.The expression level of Egr-1 mRNA might be associated with the survival time of the patients with esophageal carcinoma.Egr-1 expression in esophageal carcinoma tissue may be of great value for determining prognosis of esophageal carcinoma.
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Background To investigate the expression of the early growth response gene-1 ( Egr-1 ) mRNA after focal cerebral ischemia / reperfusion in rats.Methods Ten healthy male SD rats weighing 200 ～ 250 g were used to create model of focal cerebral ischemia. The expression of Egr-1 after focal cerebral ischemia/reperfusion in rats was determined using in situ hybridization and RT-PCR.Results (1) The result of the in situ hybridization: A trace amount of Egr-1 mRNA expressed in the neurons and the glial cells in the sham operated group. The expression of Egr-1 mRNA at the ischemic side increased dramatically following ischemia and reached peaks after 4 hours' reperfusion. Egr-1 expression started to subside following 22 hours' reperfusion and further decreased following 166 hours' reperfusion, which was still significantly higher than that in the sham operated group. (2) The result of RT-PCR: The expression of Egr-1 mRNA at the ischemic side was significantly higher than that in the sham operated group at all time points after ischemia/reperfusion in the rats(P ＜0. 01). Expression of Egr-1 increased 2 h after ischemia and reached the peak 4 h following reperfusion, and then decreased dramatically at 46 h after reperfusion which was still higher than that in the sham operated group (P ＜ 0. 01). As the ischemia/reperfusion period prolonged, the expression of Egr-1 mRNA increased gradually, but still detectable even 166 h following reperfusion. The expression of Egr-1 was significantly higher than that in the sham operated group at all time points (P ＜0. 01).Conclusions The expression of Egr-1 mRNA increase in the neurons and the glial cells after ischemia/reperfusion, which may have protective effects on ischemic brain tissues.
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Objective To evaluate the role of early growth response gene1 (Egr-1) in liver injury in taurocholate-induced acute pancreatitis rat model. Methods Twenty-four male rats were divided into 4 groups. Egr-1 immunohistochemistry staining and pancreatic pathologic scoring were assessed, and the serum levels of AST, LDH, TNF-? and IL-1 ? were measured. Egr-1 mRNA levels of primary hepatocyte culture stimulated with TNF-? and pretreated primary hepatocytes with PD98059 followed by TNF-? stimulation were also observed. Results Egr-1 expression of liver correlated with degree of liver injury in acute pancreatitis in rat, and high Egr-1 mRNA expression of primary hepatocyte was observed when hepatocyte injury occurred. After pretreated with PD98059, TNF-?-stimulated hepatocytes showed a lower level of Egr-1 mRNA compared with hepatocytes without pretreatment. Conclusions Egr-1 may play an important role in liver injury in acute pancreatitis, and this effect depended partly upon extracellular signal-regulated kinase 1/2 (ERK1/2) pathway.
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Objective To investigate the expression of early growth response gene-1 in autogenous vein graft in rats, and its role in intimal hyperplasia (IH).Methods Autogenous vein graft model was established in 90 Wistar rats. Grafts were harvested at 1、2、6、24 hours, and 3、7、14、28、42 days after surgery. Normal veins serve as control. Egr-1 mRNA was measured by RT-PCR and in situ hybridization. Western blotting and immunohistochemistry were used to detect the protein expression of Egr-1. Results The expression of Egr-1 mRNA and Egr-1 protein experienced biphasic changes. At 1 hour, the value of gene expression of Egr-1 mRNA was 2. 21?0. 85, declining at 6 and 24 hours, and upgrading at 7 days, culminating at 3.16?1.14 at 28 days. Egr-1 protein expression was detected at 2 hours, with the positive expression rate of 31%?6%. The level of Egr-1 protein was decreased from 24 hours to 3 days. A peak was reached at 41%?8% at 28 days. Immunoreative Egr-1 was located within vascular smooth muscle cells (VSMCs) and monocytes/macrophages in the media at the early phase of day 7 and 14, and in neointimal and medial VSMCs at later phase of day 28. Egr-1 was also present in the endolumial endothelial cells. Conclusion In autogenous vein graft, Egr-1 plays an important role in the proliferation of VSMCs in autogenous vein graft.