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Objective:To prepare pH-sensitive liposomes to avoid the degradation of monophosphoryl lipid A (MPLA) by lysosomes.Methods:Using DOPE and CHEMS as carrier materials, pH-sensitive liposomes were prepared by thin-film dispersion method. Particle sizes and Zeta potential of the liposomes were detected by dynamic light scattering. The morphological features of pH-sensitive liposomes under different pH conditions were observed by transmission electron microscopy. Flow cytometry was performed to detect the phagocytosis of liposomes by THP-1 and DC2.4 cells. Confocal laser microscopy was used to observed the colocalization of liposomes and lysosomes. BALB/c mice were immunized with hepatitis B surface antigen (HBsAg) using MPLA pH-sensitive liposome as an adjuvant. The levels of serum anti-HBs were quantitatively detected by ELISA. IFN-γ and IL-2 spot forming cells (SFCs) in mouse splenic lymphocytes were detected by ELISPOT.Results:The pH-sensitive liposomes were constructed with an average particle size of (90.90±1.13) nm, polydispersity index (PDI) of 0.076±0.013 and Zeta potential of (-27.900±0.666) mV. As the pH value of the solution decreased, the particle size increased significantly and the liposomes presented irregular shapes, indicating the pH-sensitive features. The phagocytosis rates by THP-1 cells and DC2.4 cells were 10.40% and 12.40% for pH-sensitive fluorescent liposomes, and 1.09% and 0.28% for fluorescent liposomes. Confocal laser microscopy revealed that pH-sensitive fluorescent liposomes were phagocytosed by THP-1 cells and existed in the cytoplasm, while fluorescent liposomes existed in lysosomes. Compared with MPLA liposomes, MPLA pH-sensitive liposomes could significantly improve the cellular immune response in mice. The levels of IFN-γ and IL-2 SFCs in the MPLA pH-sensitive liposome group were significantly higher than those in the MPLA liposome group ( P<0.01) and the non-adjuvant group ( P<0.001). Conclusions:The pH-sensitive liposome delivery system could improve the utilization of MPLA as an adjuvant.
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Objective:To express the recombinant varicella-zoster virus (VZV) gE Δ-Fc fusion protein using CHO cell expression system, and provide reference for screening candidate antigens of recombinant herpes zoster vaccines. Methods:A eukaryotic expression plasmid containing the gE Δ-Fc gene was transfected into CHO cells. Monoclonal cells were selected by methionine sulfoximine (MSX) pressure screening and limited dilution method to obtain the CHO cells secreting and expressing the gE Δ-Fc fusion protein. The expressed gE Δ-Fc fusion protein was purified by MabSelect SuRe affinity chromatography. The binding activity of gE Δ-Fc fusion protein to Fc receptors was identified by ELISA. Flow cytometry was used to detect the phagocytosis of antigens by DC2.4 cells. Antibody titers in serum samples of BALB/c mice immunized with the gE Δ-Fc fusion protein were detect by ELISA. Results:A CHO cell line stably expressing the gE Δ-Fc fusion protein was obtained. Flow cytometry suggested that the phagocytotic activity of DC2.4 cells against the gE Δ-Fc fusion protein was stronger than that against gE. Moreover, the gE Δ-Fc fusion protein could induce BALB/c mice to produce high titers of specific anti-VZV antibodies. Conclusions:The recombinant VZV gE Δ-Fc fusion protein expressed in CHO cells had a good immunogenicity. This study provided reference for screening candidate antigens of recombinant herpes zoster vaccines.
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Objective@#To explore and discuss the relationship between emotional intelligence and stress perception and adaptive ability of undergraduates majoring in nursing, and analyze whether stress perception plays an intermediary role in the relationship between emotional intelligence and adaptive ability.@*Methods@#A convenient sampling method was used to investigate 400 nursing undergraduates from 2 universities in Changchun. The survey tools were general information questionnaire, Emotional Intelligence Scale, Chinese version Perceived Stress Scale and College Student Adaptability Inventory.@*Results@#The emotional intelligence of nursing undergraduates was above average level (3.63±0.47) points, the perceived stress was below average level (2.84±0.42) points, and the adaptability was above average level (3.48±0.51) points. And the emotional intelligence was positively correlated with the adaptability (r=0.467, P<0.01), and the stress perception was negatively correlated with the adaptability (r=-0.519, P<0.01). The Perceived stress played partial intermediary role in the relationship between emotional intelligence and adaptability and the mediating effect accounts for the total effect was 22.65%.@*Conclusion@#The adaptability and nursing undergraduates′ emotional intelligence and perceived stress was closely related, and perceived stress could mediated the relationship between emotional intelligence and adaptability.
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Objective To explore and discuss the relationship between emotional intelligence and stress perception and adaptive ability of undergraduates majoring in nursing, and analyze whether stress perception plays an intermediary role in the relationship between emotional intelligence and adaptive ability. Methods A convenient sampling method was used to investigate 400 nursing undergraduates from 2 universities in Changchun. The survey tools were general information questionnaire, Emotional Intelligence Scale, Chinese version Perceived Stress Scale and College Student Adaptability Inventory. Results The emotional intelligence of nursing undergraduates was above average level (3.63 ± 0.47) points, the perceived stress was below average level (2.84 ± 0.42) points, and the adaptability was above average level (3.48 ± 0.51) points. And the emotional intelligence was positively correlated with the adaptability (r=0.467, P<0.01), and the stress perception was negatively correlated with the adaptability (r=-0.519, P<0.01). The Perceived stress played partial intermediary role in the relationship between emotional intelligence and adaptability and the mediating effect accounts for the total effect was 22.65% . Conclusion The adaptability and nursing undergraduates′emotional intelligence and perceived stress was closely related, and perceived stress could mediated the relationship between emotional intelligence and adaptability.
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Objective To investigate the risk factors and the influence of intravenous thrombolysis of acute cerebral infarction with cerebral microbleeds(CMBs)by SWI.Methods 1 64 patients with acute cerebral infarction were enrolled in this study.All pa-tients were scanned with routine MRI and SWI.According to the presence of CMBs on SWI,the patients were classified into two groups:CMBs group(73 cases)and non-CMBs group(91 cases).Past history was recorded and risk factors of CMBs were explored. 76 cases patients(including 35 cases of CMBs group and 41 cases of non-CMBs group)were treated by intravenous thrombolysis and rescanned with routine MRI and SWI to compare the changes in the number of CMBs and hemorrhage transformation 24 hours after thrombolysis.Results The difference age,hypertension,lacunar infarction and leukoaraiosis between the two groups were significant (P 0.05).Conclusion Acute cerebral infarction with CMBs are influenced by age,hypertension,lacunar infarction and leu-koaraiosis.Thrombolysis in acute cerebral infarction with CMBs can not augment the incidence of hemorrhagic transformation.
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objective To establish a molecular platform for screening thalidomide derivatives with potential high anti-tumor activities. Methods Human cereblon(CRBN) and Ikaros family zinc finger protein 1(IKZF1) cDNA were amplified and cloned into pXJ40-myc and pcDNA3-FLAG vectors respectively. Then the two constructs were transfected into 293T cells and the anti-tumor activities of thalidomide and its derivatives were reflected by their effects on the binding between CRBN and IKZF1 proteins. Results Both CRBN and IKZF1 were successfully expressed in 293T cells. Thalidomide and two of its derivatives were found to enhance the interaction between CRBN and IKZF1 significantly. Conclusion A molecular platform was successfully established for screening thalidomide derivatives with potential high anti-tumor activities and two thalidomide derivatives could potentiate the binding between CRBN and IKZF1, suggesting that they possess potential anti-tumor activities.
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Aim To provide practical method for screening human carbonic anhydrase Ⅱ(hCA Ⅱ) inhibitors in drug discovery.Methods hCA Ⅱ protein was obtained from induced BL21(DE3) E.coli containing plasmid pET-28b-hCA Ⅱ.The hCA Ⅱ activity was detected under pH 7.6 and 25℃ by its esterase activity which could decompose PNPA to increase the photoabsorption at 348 nm. After the assay conditions were finally selected, 35 new compounds were tested.Results A practical method for screening hCA Ⅱ inhibitors was successfully constituted by using recombinant hCA Ⅱ protein expressed in E.coli as the source of hCA Ⅱ enzyme.10 new compounds had better inhibitory effect and 9 new compounds had the same inhibitory effect on hCA Ⅱ compared with acetazolamide.Conclusions The hCA II inhibitor screening technique constituted in this work possesses advantages of being reliable, rapid, and practical. 19 new compounds are worth further research for developing high efficiency and low side effect drugs used for high-altitude illness.
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Aim To investigate the effect on antagonism of benthiactzine and atropine against the function of muscarinic acetylcholine receptors by desensitization of nicotinic acetylcholine receptors. Methods The whole cell recording configuration of patch-clamp technique was used and the cell model was rat SCG neurons. To identify antagonists' antimuscarinic effect, mAchRs mediated IM-inhibition was measured and nicotine and oxotremorine were used. Results After de sensitization of nAChRs,the antimuscarinic effect ofboth benthiactzine and atropine decreased compared to the normal condition. The decreased antimuscarinic effect of benthiactzine was weaker than that of atro-pine. The antimuscarinic effect of both benthiactzine and atropine recovered gradually with the recovery of nAChRs from desensitization. Conclusion Desensiti-zation of nAChRs weakens the antagonists' antimuscarinic effect.
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AIMTo investigate the anti-atherosclerosis mechanisms of the novel compound PPVP. METHODSThirty-six male rats were a ssigned to control, model and PPVP 5 mg?kg -1 groups randomly. The rats in model and PPVP 5 mg?kg -1 groups were fed with hypercholesterol diet for inducing atherolcerosis model. The following variables were measured: HE stainin g of rat aorta; serum level of TC, LDL-CHO, HDL-CHO, IL-8, ET-1, PGI 2, TXA 2 and NO. RESULTSPPVP increased serum level of NO, decreased plasma level of TXA 2 and inhibited the over expression of IL-8. CONC LUSIONThe anti-atherogenic effect of PPVP seems to be closely involved in increasing serum NO level, decreasing plasma level of TXA 2, and inhibiting over expression of IL-8. The result also suggests that the anti-atherogenic ef fect of PPVP in high cholesterol-fed rat may not due to the regulation of plasm a lipid profile, plasma level of ET-1 and PGI 2.
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AIM To investigate the anti-atherosclerosis mechanisms of arecoline. METHODS Fifty-eight male rats were assigned to control, model, arecoline 1 mg?kg -1 and arecoline 5 mg? [FQ(12。46,X-WZ]-kg -1 groups randomly. The rats in model and arecoline-supplied groups were fed with hypercholesterol diet. The following variables were measured: HE staining of rat aorta; serum level of TC, LDL-CHO, HDL-CHO, IL-8, ET-1 and NO; expression of eNOS protein on rat aorta; expression of MCP-1, ICAM-1, CXCR-2 and eNOS mRNA on rat aorta. RESULTS Arecoline increased plasma level of NO and the expression of eNOS protein and mRNA, decreased plasma level of IL-8 and the expression of ICAM-1, MCP-1 and CXCR-2 mRNA on rat aorta. CONCLUSION The anti-atherogenic effects of arecoline seems to be closely involved in increasing plasma level of NO and eNOS protein and mRNA expression, decrease in plasma IL-8 level and down-regulation of the expression of ICAM-1, MCP-1 and CXCR-2 genes.-
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AIM To investigate the effects of iptakalim hydrochloride(Ipt) on potassium currents in cardiomyocytes derived from guinea pig and on the specific binding of glibenclamide (Gli) with sulfonylurea receptor(SUR_ 2A ) of ATP-sensitive potassium channel (K_ ATP ) in cardiac membranes derived from Wistar rats. The effects of Ipt on the association and dissociation kinetic processes of Gli binding with SUR_ 2A of K_ ATP in cardiac preparations were also determined. METHODS The effects of Ipt on potassium currents in cardiomyocytes were observed by using patch clamp technique(whole cell recording) after application of the drug in the bath. The experiments of the ass ociation and dissociation kinetic processes of K_ ATP blocker [ 3H]Gli binding with cardiac membranes were used. RESULTS (1)The potassium current-voltage curves (I-U curves) of cardiomyocytes derived from guinea pig were upward shifted by Ipt at the concentrations of 1 and 100 ?mol?L -1 . Within 5 minutes after application of the drug, the current amplitude increased to 124.9%?9.5%(n=5)and 151.6%?11.2%(n=7)of initial current amplitude respectively(P
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Aim To investigate the desensitization characteristics of neuronal ?4?2、?4?4 and ?7-nicotinic acetylcholine receptors heterologously expressed in the SH-EP1 cell line stimulated by prolonged of repetitive application of agonist.Methods The Whole cell recording configuration of patch-clamp technique was used to study the desensitization characteristics of ?4?2、?4?4 and ?7-nAChRs in cultured SH-EP1 cells three days after passage by rapid application of nicotine onto the soma of the cells.Results Inward currents were elicited by rapid application of agonist in all the three types of SH-EP1 cells in a concentration-dependent manner.?7-nAChRs desensitized the fastest and deepest in the three while ?4?4-nAChRs the slowest and lightest under prolonged stimulation.?4?2-nAChRs desensitized the fastest in the three under repetitive stimulation while ?4?4 and ?7-nAChRs didn't have obvious desensitization.Conclusion When stimulated by prolonged of repetitive application of agonist,neuronal ?4?2、?4?4 and ?7-nicotinic acetylcholine receptors had different desensitization characteristics,which is possibly associated with their roles in organophosphate poisoning inducing persistent stimulation of excessive accumulation of endogenous ACh and in nicotine addiction inducing repetitive stimulation of nicotine.
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Aim To establish a cell model heterologously expressing Kir6.x and SUR subunits of K_(ATP)channels and to study the selectivity of iptakalim on different subtypes of K_(ATP) channels.Methods Cell were transfected with the pcDNA vector containing the coding sequenced of Kir6.x and SUR using liposome and the pEGFP-N1 vector encoding for green fluorescent protein was added for easy identification of transfected cells.Using immunocytochemical technique,we detected the expression of Kir6.x and SUR protein in[FL(2K2]transfected cells.The electrophysiological experiment was performed after transfection 48-72 h.In whole cell configuration,the effects of K_(ATP) channel openers iptakalim,pinacidil and diazoxide on the clone currents in transfected HEK-293 cells and the antagonism of K_(ATP) channel inhibitor glibenclamide were evaluated.Results Immunocytochemical study identified the expression of Kir6.x and SUR protein in transfected cells.The electrophysiological experiment showed that in cells with recombinant expression of the Kir6.1/SUR2B channel complex,iptakalim(100 ?m),pinacidil(100 ?m)and diazoxide(200 ?m)significantly increased the clone current from(-88.0?30.8) pA to(-2042.6?127.3) pA,(-1431.9?142.4) pA and(-1104.7?228.9) pA,respectively,at-100 mV,and the actions were inhibited by glibenclamide(10 ?m).In cells expressed with Kir6.2/SUR2A channel,both iptakalim and pinacidil activated the currents,which was sensitive to glibenclamide.While diazoxide had no effects on the clone currents.In contrast,in cells with Kir6.2/SUR1channel,diazoxide increased the activity of recombinant K_(ATP) channels,while iptakalim and pinacidil had little effects.Conclusion From these observations,the effects of K_(ATP) channels openers with different chemical structures on the subtypes of K_(ATP) channels were diverse.Iptakalim showed selective action on Kir6.1/SUR2B and Kir6.2/SUR2A,but not Kir6.2/SUR1 K_(ATP) channels
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Aim To investigate the effects of newly synthesized 2, 3-dimethyl-2-butylamine derivatives on potassium channels in cortical neurons from rats.Method On cultured cortical neurons, the influences of those novel compounds on I A and I K currents were evaluated using whole-cell patch clamp technique. Result On cortical neurons, these compounds (5),(6),(7),(9) showed no effects on outward potassium currents, while the compounds (4),(8) exhibited inhibition of I K currents. But on a rterial smooth muscle cells(SMCs), all the six compounds had potassium channels opening activities. While the other three compounds had no effects on potassium channels of SMCs, the compound (3) decreased the I K currents evoked on neu rons.Conclusion The novel 2,3-dimethyl-2-butylamine derivati ves had the different modulation of potassium channels on cortical neurons as SM Cs.
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AIM To investigate the effects of iptakalim hydrochloride (Ipt) on cerebral neuronal sodium, calcium and potassium channels. METHODS The effects of Ipt on sodium, calcium and potassium channels in cultured hippocampal neurons was observed using whole-cell recording technique. RESULTS Ipt had no significant effect on Na +, Ca 2+ currents, but significantly increased the outward K + currents. This enhancement of Ipt was inhibited by synchronously application of glibenclamide. Upon the administration of Ipt 1, 10, 100 ?mol?L -1, the outward K + current was significantly increased to (121.9?8.1)%, (114.7?7.1)%, (109.1?10.0)% respectively (n=6～9, P
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AIM To investigate the effects of the novel antihypertensive drug iptakalim hydrochloride on potassium currents in artery smooth muscle cells. METHODS The effects of iptakalim hydrochloride on potassium currents in smooth muscle cells derived from rat pulmonary arteries were observed by using patch clamp technique(whole cell recording) after application of the drug in the bath. RESULTS The potassium current-voltage curves ( I-U curves) of smooth muscle cells derived from normotensive rat pulmonary arteries were up-ward shifted by iptakalim hydrochloride(0.1,1,10 and 100 ?mol?L -1 ). Within 5 minutes after application of the drug, the current amplitude could increase to[(118.6?15.9)%, P