To explore the regulation of eIF4E, we screened the protein interacting with eIF4E from human cDNA library by using yeast two-hybrid system. Several clones interacting with eIF4E were identified. One of them was homologous with HUWE1 (HECT, UBA and WWE domain containing 1, also named as ARF-BP1, HECTH9 or HUWE1). Cell co-immunoprecipitation showed that eIF4E could bind to HUWE1 in mammalian cells. We also found that HUWE1 bearing the HECT domain is necessary for its association with eIF4E.
Animals ; Eukaryotic Initiation Factor-4E ; metabolism ; Humans ; Ubiquitin-Protein Ligases ; metabolism

In this paper, the action of suet oil in the preparation of self-assembled micelles of the active flavonoids in Epimedium in the simulated human environment was researched. Twelve suet oil samples were collected from different growing areas and different positions of sheep or goat to simulate the formation of micelles. Then the effects of the fatty acids in suet oil on the preparation of self-assembled micelles were studied furthermore. The results showed that the micelles had a dispersed state and spherical smooth surface. To compare the diameter, potential, encapsulation efficiency and drug loading of the 12 batches micelles, the micelles prepared by the suet oil from Qinghai were more stable and had a higher encapsulation efficiency. The fatty acids in suet oil could promote the formation of self-assembled micelles, but the whole suet oil had a better effect. Above all the study, we confirmed that the suet oil promoted the formation of self-assembled micelles of the flavonoids in Epimedium, it laid foundation for further research about increasing the efficacy of Epimedium and improved the absorption of the active flavonoids in Epimedium.
Animals ; Chemistry, Pharmaceutical ; methods ; China ; Drug Carriers ; chemistry ; Electric Conductivity ; Epimedium ; chemistry ; growth & development ; Fatty Acids ; chemistry ; Flavonoids ; chemistry ; Geography ; Goats ; Humans ; Micelles ; Oils ; chemistry ; Sheep

Histocytochemistry ; methods ; Humans ; Pathology, Clinical ; methods ; Specimen Handling ; methods ; Staining and Labeling ; methods

Neurotensin receptor-1 (NTR1), which can stimulate the intracellular cascade signal pathway, belongs to the large superfamily of G-protein coupled receptors. NTR1 is related to the occurrence and development of several kinds of diseases. In order to screen the inhibitors for the cancers associated with NTR1 protein, we established a CHO (Chinese hamster ovary) cell line in which human neurotensin receptor-1 was highly expressed. The method is to construct the recombinant plasmid which was lysed with the hNTR1 gene and transfect it into CHO cells. After selected with G418, the cell line was evaluated by Western blotting analysis and calcium flux assays. Through the calcium flux assays on FlexStation 3, we got the EC50 value of neurotensin peptide which is the natural NTR1 agonist, and the IC 50 value of SR48692 which is the known NTR1 antagonist. The established human CHO/NTR1 cell line can be used to study the profile of NTR1 biological activity and further screen of NTR1 antagonists and agonists.
Animals ; CHO Cells ; Calcium Signaling ; Cricetinae ; Cricetulus ; Humans ; Pyrazoles ; pharmacology ; Quinolines ; pharmacology ; Receptors, Neurotensin ; antagonists & inhibitors ; genetics ; metabolism ; Transfection

BACKGROUND: After neural precursor cells (NPCs) induced from embryonic stem cells (ESCs) have been grafted into the brain, it would still keep some potency of proliferation and differentiation, strong plasticity and integration into the host neural tissues, which would help to observe the therapeutic effect of PD.OBJECTIVE: To observe the differentiation of mouse ESCs into NPCs and the therapeutic effect of NPCs after being transplanted on the behavior of Parkinson disease (PD) rats.DESIGN: Randomly and controlled animal experiment.SETTING: Staff Room of physiology and Staff Room of Neurobiology, Depayment of Basic Medical Sciences, the Third Military Medical University of Chinese PLA.MATERIALS: Totally 50 healthy adult Wistar rats were chosen and randomly divided into experimental group (n=45) and control group (n=5).METHODS: ① 5 μ L (2 g/L)6-hydroxydopamine (6-OHDA) was injected into substantia nigra pars compacta and ventral tegmental area two points in the experimental group to prepare PD rats, and normal saline with the dosage of 5 μL per point was injected into the rats in the control group.Behavioral test began at 1 week after operation to measure successful rate of model establishing, once a week for 7 consecutive weeks. ② Totally 20 successful PD rat models were chosen to perform corpus striatum NPCs with the dosage of 2 μL [the count of cell suspension was (5-8)×106/μL].The other 5 rats were given 2 μL normal saline at corpus striatum as normal saline control group.MAIN OUTCOME MEASURES: ① Successful rate of PD model. ②Effect of NPCs transplantation on the rotation times of PD models. ③ Distribution of transplanted NPCs in vivo, and survival and differentiation.RESULTS: ①6 weeks later, totally 33 of 45 rats in the experimental group achieve the standard of PD model . ② About 85% of mouse ESCs were differentiated into Nestin-positive NPCs 5 days after the embryoid bodies formed in the bacterial dishes and cultured in the N2 serum-free medium. ③The rotation times of the PD rats was significantly decreased after the intracerebral transplantation of NPCs as compared with normal control group. Most of the NPCs grafted into striatum of PD rats were survived, and some were differentiated into TH-positive neurons.CONCLUSION: The mouse ESCs-derived NPCs could be transplanted into striatum of PD rats, and then differentiated into TH-positive neurons,which leads to the obvious decrease of rotation times.

Objective To study the increased radiosensitivity of tumor cells by triterpene acids of loquat leaf (TAL) and mechanism.Methods C6 and MG63 cells were pretreated by TAL,and then were exposed to X-rays at 1,2,3,5,7 Gy.Clonogenic assay was used to evaluate those tumor cells survival fraction (SF) to calculate the sensitization enhancement ratios (SER) of TAL.The cellular micronuclei ratios of tumor cells were analyzed by micronuclei assay.Additionally,the changes of Rad51 and XRCC4 levels were observed by RT-PCR and Western blot.Results D0 values were decreased to 1.31 and 2.85 Gy in C6 and MG63 cells by TAL pretreatment respectively.The SER value of the effect of TAL on C6 and MG63 cells was 1.73 and 2.04,respectively.There was a statistical difference in the cellular micronuclei ratios between tumor cells with TAL above 2 Gy and those without TAL (C6:t =-8.372--2.476,P ＜0.05 ; MG63:t =-4.03--2.557,P ＜ 0.05).TAL attenuated the expression of XRCC4 at transcriptional and translational level,but not for Rad51,a key gene in homologous recombination repair (HRR).Conclusions TAL pretreatment could increase the lethal effect of X-rays on tumor cells in vitro.The mechanism might be involved in the inhibition of non-homologous end joining (NHEJ).

Objective To investigate the inhibitory effects of overexpression of PTEN on renal epithelial-mesenchymal trans-differentiation induced by TGF-β1,and the signaling transduction mechanism.Methods HKC cells were transfected with GFP-PTEN via lipofectAMINE2000.The efficiency of transfection was detected by fluorescence microscope.The expression of PTEN protein and mRNA in the translected cells was detected by Western blot and RT-PCR respectively.The experiment was divided into four groups:normal group,TGF-β1 stimulation group,GFP-PTEN+TGF-β1 group and empty vector+TGF-β1 group.The expression of E-cadherin,a-SMA,Akt and p-Akt was detected by Western blot.Results Most ceils transfected with GFP-PTEN expressed GFP.The expression of PTEN protein and mRNA was strongly increased when HKC cells were transfected with GFP-PTEN(all P<0.05).In both TGF-β1 stimulation group and empty vector+TGF-β1 group,the expression level of E-cadherin was lower(all P<0.05),while that of p-Akt and a-SMA was higher than in normal group(both P<0.05).The expression level of p-Akt and a-SMA in GFP-PTEN+TGF-β1 group was Iower(both P<0.05),while that of E-cadherin was higher than in TGF-β1 stimulation group and empty vector+TGF-β1 group(both P<0.05).The expression of Akt was similar in the four groups.Conclusion Overexpression of PTEN can inhibit renal epithelial-mesenehymal trans-differentiation induced by TGF-β1 through suppressing the activation of PI3K/Akt signal pathway.

Adult ; Delivery of Health Care ; Female ; Health Education ; Health Knowledge, Attitudes, Practice ; Humans ; Male ; Occupational Health

Objective:To study the effect of N-acetyltransferase 10(Naa10p)expression on the sensitivity of tongue squamous cell carcinoma Tca8113 cells to pingyangmycin(PYM).Methods:The lentiviral vectors of LV-shNaa10p,LV-Naa10p and control LV-NC were transfected into Tca8113 cells,the infection efficiency were identified,then the sensitivity of the infected cells to PYM was determined by MTS assay.Results:The IC50 of Tca8113-LV-shNaa10p group,Tca8113-LV-Naa10p group and Tca8113-LV-NC group were (20.772 ±0.106)μg/ml,(2.157 ±0.123)μg/ml and (6.301 ±0.069)μg/ml respectively(between groups P <0.05).Con-clusion:Knock down of Naa10p may reduce and over expression of Naa10p can increase the sensitivity of Tca8113 cells to PYM.

Objective To investigate the level and value of serum IgE, anti-IgE, FcεRⅠα, anti-FcεRⅠin autoimmune liver disease ( AILD) .Methods In this case-control study, the serum samples and clinical data of 77 patients with hepatosis were collected between May and November 2014 from the department of gastroenterology of Shanghai First People′s Hospital.These patients had positive results about the liver-related autoimmune antibodies, including 33 cases of AILD, 44 cases of other chronic liver disease. 64 healthy persons were collected as control group.Serum mast cell-associated anti-IgE, FcεRⅠα, anti-FcεRⅠwere detected by Enzyme-linked Immuno sorbent Assay ( ELISA) .Serum IgE, IgM and IgG were detected by rate scatter nephelometry.Differences among AILD, other chronic liver disease and healthy control were assessed.Results were compared using Mann-Whitney U test.Results Mast cell-associated anti-IgE, FcεRⅠα, anti-FcεRⅠin liver-related autoimmune antibodies positive patients were significantly higher than healthy control [1.74(1.16 -2.88)mg/L, 14.86(4.39 -26.23)mg/L, 47.22(36.89 -55.29)mg/L and 1.23(0.95-1.58)mg/L, 1.87(1.52-2.33)mg/L, 35.40(24.74-44.89)mg/L, respectively;U=1614,556.5,1319.5, P<0.01].FcεRⅠαwas significantly higher in other chronic liver disease patients than AILD patients [18.40(7.35-30.64)mg/L and 6.25(2.49-22.29), respectively;U=445, P<0.01] .Conclusion Mast cell-associated anti-IgE, FcεRⅠα, anti-FcεRⅠwere increased in liver-related autoimmune antibodies positive hepatosis patients.However, FcεRⅠαwas lower in AILD than other chronic liver disease.Mast cell-associated anti-IgE、FcεRⅠαand anti-FcεRⅠ molecules involved in the inflammatory lesion of liver disease.