The effects of the inter-stimulus interval in magnetic stimulation on astrocyte migration and its mechanism / 中华物理医学与康复杂志

Objective To examine the effect of the inter-stimulus interval (ISI) in magnetic stimulation (MS) on astrocyte migration and its related mechanism.Methods Cultured astrocytes were treated with intermittent MS with intervals of 1,5 and 10 seconds.The PEA-15 inhibitor BisI (10 μmol/ml) and the ERK1/2 inhibitor U0126 (10 μmol/ml) were administered and cell migration assays evaluated the astrocytes' migration.The expression of phosphorylated ERK1/2 and PEA-15 was detected using Western blotting.Results The 1 second interval significantly facilitated astrocyte migration,the phosphorylation of PEA-15 and ERK1/2,and the expression of MMP-9 (browse matrix metalloproteinase-9).The addition of Bis I significantly reduced the production of phosphorylated ERK1/2 and MMP-9,as well as astrocyte migration induced by MS.In addition,pretreatment with U0126 also significantly decreased the astrocyte migration induced by MS.Conclusion 1s-ISI MS can induce PEA-15 activation and subsequently lead to ERK1/2 phosphorylation and upregulation of MMP-9,which may contribute to the migration of astrocytes.

Estradiol attenuates down-regulation of PEA-15 and its two phosphorylated forms in ischemic brain injury / 한국실험동물학회지

Estradiol exerts a neuroprotective effect against focal cerebral ischemic injury through the inhibition of apoptotic signals. Phosphoprotein enriched in astrocytes 15 (PEA-15) is mainly expressed in brain that perform anti-apoptotic functions. This study investigated whether estradiol modulates the expression of PEA-15 and two phosphorylated forms of PEA-15 (Ser 104 and Ser 116) in middle cerebral artery occlusion (MCAO)-induced injury and glutamate exposure-induced neuronal cell death. Adult female rats were ovariectomized to remove endogenous estradiol and treated with vehicle or estradiol prior to MCAO. Focal cerebral ischemia was induced by MCAO and cerebral cortices were collected 24 h after MCAO. Western blot analysis indicated that estradiol prevents the MCAO-induced decrease in PEA-15, phospho-PEA-15 (Ser 104), phospho-PEA-15 (Ser 116). Glutamate exposure induced a reduction in PEA-15, phospho-PEA-15 (Ser 104), phospho-PEA-15 (Ser 116) in cultured neurons, whereas estradiol treatment attenuated the glutamate toxicity-induced decrease in the expression of these proteins. It has been known that phosphorylation of PEA-15 is an important step in carrying out its anti-apoptotic function. Thus, these findings suggest that the regulation of PEA-15 phosphorylation by estradiol contributes to the neuroprotective function of estradiol in ischemic brain injury.

Construction and expression of eukaryotic expression vector of PEA-15 / 肿瘤研究与临床

Objective To construct the eukaryotic expression vector of pcDNA3.1-PEA-15 and express it in the human esophageal cancer (EC-109) cells,and to explore the effect of PEA-15 on EC-109 cells.Methods The PEA-15 gene was amplified from EC-109 by reverse transcriptase polymerase chain reaction (RT-PCR) and ligated to eukaryotic expression vector pcDNA3.1.After confirmation of recombinant plasmid was correctly by endonuc]eases digestion and DNA sequencing,the construct was transfected it into EC-109 through liposome inducing.The expression of PEA-15 in transfected EC-109 was detected by RT-PCR and Western blot.The cell growth inhibition ratio was evaluated by MTT assay.Results RT-PCR indicated that PEA-15 was highly expressed in EC-109 cells.The amplified fragment by RT-PCR was coincident with hypothesis enzyme digestion analysis and DNA sequencing confirmed that the pcDNA3.1-PEA-15 was constructed successfully.The expression of PEA-15 gene was increased obviously in the transfected EC-109 detected by RT-PCR and Western blot respectively (t =4.078,5.269,P ＜ 0.05).The cell growth inhibition ratio in the group which transfected pcDNA3.1-PEA-15 was significantly lower compared with the pcDNA3.1 transfect group after 72 hours (t =6.163,P ＜ 0.05).Conclusions The recombinant eukaryotic expression vector pcDNA3.1-PEA-15 is constructed successfully,and it can be expressed in EC-109.It also shows that PEA-15 has the function on cell growth which suggests that PEA-15 can inhibit the apoptosis of EC-109 cells and its expression involved in esophageal cancer development.

Expression of PED/PEA-15 mRNA and protein in esophageal carcinoma / 肿瘤研究与临床

Objective To investigate the expression of PED/PEA-15 mRNA and protein in esophageal carcinoma tissue and their clinical significances.Methods The expression of PED/PEA-15 mRNA was detected by RT-PCR,and the expression of PED/PEA-15 protein was measured by immunohistochemistry in 50 cases of esophageal carcinoma,50 cases of corresponding paracarcinoma tissue,and 50 cases of corresponding normal esophageal tissue.Results The expression of PED/PEA-15 mRNA was 1.14±0.49 in esophageal carcinoma,which was significantly higher than that in para-carcinoma tissue (0.59±0.31) and normal esophageal carcinoma tissue (0.53±0.22) (F =44.085,P ＜ 0.001).The immunohistochemistry results showed that PED/PEA-15 protein expression was significantly higher than that in para-carcinoma tissue and normal esophageal tissue (x2 =36.967,P ＜ 0.001; x2 =26.272,P ＜ 0.001).The expression of PED/PEA-15 mRNA and protein were significantly associated with pathological grade,clinical stage of esophageal carcinoma (P ＜ 0.05),but were not significantly correlated to the age of onset,gender,pathological types (P ＞ 0.05).Conclusion The expression of PED/PEA-15 mRNA and protein are increased in esophageal carcinoma,which may play an important role in the occurrence and development of esophageal carcinoma.

Focal Cerebral Ischemia Induces Decrease of Astrocytic Phosphoprotein PEA-15 in Brain Tissue and HT22 Cells / 한국실험동물학회지

PEA-15 is a small phosphoprotein (15 kDa) that is enriched in brain astrocytes. PEA-15 acts as an important modulator of cellular function including apoptosis and signal integration. This study investigated the expression of PEA-15 in focal cerebral ischemic injury. Cerebral ischemia was surgically induced in adult male rats by middle cerebral artery occlusion (MCAO), and brains were collected 24 hr after MCAO. A proteomic approach demonstrated decreases of PEA-15 protein spots in MCAO-operated animals in comparison to sham-operated animals. Western blot analysis clearly demonstrated that MCAO induces decreases in PEA-15 levels. We previously showed that glutamate toxicity induces cell death in a hippocampus-derived cell line (HT22). Glutamate exposure induces decreases of PEA-15 levels in HT22 cells. The results of this study suggest that focal cerebral ischemia induces cell death through downregulation of PEA-15 protein.

Focal Cerebral Ischemia Induces Decrease of Astrocytic Phosphoprotein PEA-15 in Brain Tissue and HT22 Cells / 한국실험동물학회지

PEA-15 is a small phosphoprotein (15 kDa) that is enriched in brain astrocytes. PEA-15 acts as an important modulator of cellular function including apoptosis and signal integration. This study investigated the expression of PEA-15 in focal cerebral ischemic injury. Cerebral ischemia was surgically induced in adult male rats by middle cerebral artery occlusion (MCAO), and brains were collected 24 hr after MCAO. A proteomic approach demonstrated decreases of PEA-15 protein spots in MCAO-operated animals in comparison to sham-operated animals. Western blot analysis clearly demonstrated that MCAO induces decreases in PEA-15 levels. We previously showed that glutamate toxicity induces cell death in a hippocampus-derived cell line (HT22). Glutamate exposure induces decreases of PEA-15 levels in HT22 cells. The results of this study suggest that focal cerebral ischemia induces cell death through downregulation of PEA-15 protein.