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Objective:To evaluate the clinical efficacy and safety of modified chitosan eye drops on rabbit Candida albicans keratitis model.Methods:Ten healthy female New Zealand rabbits were used to establish the superficial Candida albicans keratitis model by the corneal surface lens method in the right eye.Slit lamp microscopy and corneal scraping and microscopic examination were performed to preliminarily determine whether the keratitis model has been successfully established, the rabbits were then randomly divided into a model group and a modified chitosan group by the random number table method.The successfully established rabbit models which were determined by fungal culture results were retained.Five normal rabbits receiving no intervention served as a normal control group.The experimental eyes in the normal control and modified chitosan groups were treated with modified chitosan eye drops, Six times a day for one week, and subsequently four times a day for one week.No treatment was administered to the model group.The changes of corneal lesions and ocular surfaces were examined by slit lamp microscopy every day.At 1, 7, 14, 21 and 28 days after modeling, the eye condition and corneal clinical scores was assessed by slit lamp microscopy.The corneal conditions in each group was observed for two weeks after drug withdrawal.Results:The corneal scraping and microscopic examination results of eight rabbits models showed that the fungal hyphae and spores were positive.The fungal culture results showed that the separated pathogen was Candida albicans.The success rate of modeling was 80%(8/10). The clinical scores in the model group at 7, 14 and 21 days after modeling were 14.50±0.58, 6.25±0.50 and 2.50±0.58, respectively, and were significantly higher than 7.25±1.26, 2.75±0.50 and 1.25±0.50 in the modified chitosan group (all at P<0.05). In the model group, corneal edema was significantly aggravated, and the central white ulcer area was enlarged within seven days after modeling.Between 7 and 28 days after modeling, the corneal ulcer was gradually healed, while the central corneal scar and neovascularization were remained.The average healing time was (24.5±2.6)days.In the modified chitosan group, the corneal infiltration was significantly alleviated within seven days after modeling, and the fungal hyphae and spores of corneal scraping were negative on the 14th day after modeling.The average healing time in the modified chitosan group was (13.5±1.3)days, which was significantly shorter than that in the the model group (P<0.01). No recurrence of keratitis was observed in the modified chitosan group after two weeks of drug withdrawal.The cure rate was 100%.In the normal control group, the conjunctival hyperemia, corneal edema, and lesions were not observed during topical administration.Conclusion:The treatment with modified chitosan eye drop is effective in a rabbit superficial Candida albicans keratitis model, and have no obvious toxic effects on ocular tissues.
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To investigate the effects of ultrasound combined with microbubbles on intracellular Ca2+ homeostasis in carboplatin ( CBP )‐treated A549 cell and its possible mechanisms of inhibiting A549 cell line activity . Methods According to whether SonoVue was used or not ,and the different dose of CBP ,the groups A‐F were arranged as the ultrasound( US) group( group A ) ,the ultrasound combined with microbubbles ( USMB) group( group B) ,the low dose CBP ( 100 μg/ml) + US group( group C) ,the low dose CBP+USMB group( group D) ,the high dose CBP ( 200 μg/ml)+ US group ( group E) and the high dose CBP+USMB group( group F) .A549 cells were bathed and washed by a calcium‐free buffer , loaded with Ca2+ indicator fluo‐4 AM . Real‐time images were acquired using laser confocal microscopy . T he fluorescence intensity of intracellular calcium ion concentration ( [ Ca 2+ ] i ) in individual living cell was observed and the calcium overload was analyzed . Results After ultrasound irradiation ,the normalized fluorescence intensity of [ Ca2+ ] i increased rapidly ,then returned to a new homeostasis ( selected cells in groups A ,B ,E ,F ) or experienced a second calcium oscillation ( some cells in group C and D ) . All the selected cells in group B and some cells in group C and D exhibited superimposed oscillations . T he calcium overloading time in group D was longer than those of any other groups . Four cells in group A experienced delayed calcium oscillations . Compared with group A ,the selected cells in other groups exhibited a larger amplitude of calcium oscillation ( all P < 0 .05 ) and the selected cells in group B and D exhibited calcium oscillation for a longer period of time( all P <0 .05) . Conclusions In the calcium‐free buffer ,US ,USMB , CBP+ US ,CBP + USMB are direct stimuli of calcium overload in A 549 cells . SonoVue ,CBP ,CBP +SonoVue are all synergistic stimuli of calcium overload in A 549 cells irradiated by ultrasound .US ,USMB and CBP may synergistically induce calcium release from intracellular store sites in A 549 cells . Calcium overload is a possible mechanism of ultrasound combined with microbubbles in assisting CBP chemotherapy .
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Objective To investigate the clinical significance of restrictive foramen ovale ( RFO ) monitored by fetal echocardiography during the middle to late stage of pregnancy . Methods The detection rate ,echocardiographic features and outcome in fetuses with RFO without cardiac malformations from 7319 pregnant women received prenatal echocardiography were retrospectively reviewed and analyzed . Results RFO was found in 40 of 7319 (0 .55% ) fetuses . The inclusion criteria including a narrow right to left shunt of less than 2 .5 mm in diameter across atrial septum , enlarged right atrium , increased right-to-left ventricular size ratio ,and increased size ratio of main pulmonary artery to aorta were present in 40 fetuses . The direct ultrasound characters of RFO included limited opening of oval valve ( 70% ) and foramen ovale diameter less than 2 .5 mm (30% ) . And atrial septal aneurysm ( 62 .5% ) ,redundant primum atrial septum (57 .5% ) ,abnormal ductus arteriosus ( 57 .5% ) might also be present commonly in RFO . As the gestational weeks increased , the size ratio of right-to-left atrium , right-to-left ventricle and the main pulmonary artery to aorta also increased significantly( P =0 .004 , P <0 .001 , P <0 .001) . Among the 40 fetuses with RFO ,21 cases ( 52 .5% ) gave birth in full term ,8 cases ( 20% ) which were detected severe tricuspid regurgitation gave birth in early cesarean section ,5 cases ( 12 .5% ) had induced labor and 6 cases (15% ) were lost in the follow-up . Of the 29 newborns ,only 1 case died of heart failure ,and the other 28 subjects recovered both from heart structure or cardiac function within four months . Conclusions RFOwithout cardiac malformations presents echocardiographic features characterized by a narrow right to left shunt of less than 2 .5 mm in diameter across atrial septum . Fetal echocardiography can monitor the dynamic change of fetal heart structure and function based on the increase of right heart load and decrease of left heart volume ,which has important clinical significance for assessing fetal intrauterine condition and prognosis .
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Objective To investigate the effects of ultrasound combined with microbubbles on intracellular Ca2+ homeostasis in carboplatin ( CBP )-treated A549 cell and its possible mechanisms of inhibiting A549 cell line activity . Methods According to whether SonoVue was used or not ,and the different dose of CBP ,the groups A-F were arranged as the ultrasound(US) group(group A) ,the ultrasound combined with microbubbles ( USMB) group( group B) ,the low dose CBP ( 100 μg/ml) + US group( group C) ,the low dose CBP+USMB group( group D) ,the high dose CBP ( 200 μg/ml)+ US group ( group E) and the high dose CBP+USMB group( group F) .A549 cells were bathed and washed by a calcium-free buffer , loaded with Ca2+ indicator fluo-4 AM . Real-time images were acquired using laser confocal microscopy .The fluorescence intensity of intracellular calcium ion concentration ([Ca2+ ]i) in individual living cell was observed and the calcium overload was analyzed . Results After ultrasound irradiation ,the normalized fluorescence intensity of [Ca2+ ]i increased rapidly ,then returned to a new homeostasis (selected cells in groups A ,B ,E ,F) or experienced a second calcium oscillation ( some cells in group C and D) . All the selected cells in group B and some cells in group C and D exhibited superimposed oscillations . The calcium overloading time in group D was longer than those of any other groups . Four cells in group A experienced delayed calcium oscillations . Compared with group A ,the selected cells in other groups exhibited a larger amplitude of calcium oscillation( all P < 0 .05) and the selected cells in group B and D exhibited calcium oscillation for a longer period of time( all P <0 .05) . Conclusions In the calcium-free buffer ,US ,USMB , CBP+ US ,CBP + USMB are direct stimuli of calcium overload in A 549 cells . SonoVue ,CBP ,CBP +SonoVue are all synergistic stimuli of calcium overload in A 549 cells irradiated by ultrasound .US ,USMB and CBP may synergistically induce calcium release from intracellular store sites in A 549 cells . Calcium overload is a possible mechanism of ultrasound combined with microbubbles in assisting CBP chemotherapy .
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Objective To investigate the impact of ultrasound-targeted microbubble destruction ( UTMD) combined with carboplatin sustained release microspheres on the temperature of the tumor microenvironment and the concentration of the drug release in target tumor . Methods Twenty tumor bearing rabbits were randomly assigned into 4 groups : the control group without any treatment ; the carboplatin-loaded PLGA microspheres (CPMs) group ,only intratumoral injection of carboplatin sustained-release microspheres ;the S2000 group ,diagnostic ultrasound irradiation using S2000 equipment on local chemotherapy tumor tissue combined with lipid microbubbles ;the AP-170 group ,the same treatment as the S2000 group except the application of AP-170 instead of S2000 . Far-infrared thermography was performed to measure the tumor temperature . The experimental equipment of extracorporeal circulation was set up . The experimental conditions were assigned into 4 groups :37℃ CPMs group ,37℃ S2000 group ,37℃ AP-170 group ,40℃ AP-170 group . After irradiation experiment ,the carboplatin absorbance was measured at 229 nm wavelength . The absorbance difference of each group was compared with the 37℃ CPMs group ,and each group was tested 10 times . Results The intratumorous temperature in AP-170 group increased to (40 .24 ± 0 .72) ℃ ,which was significantly different from all the other groups ( P < 0 .01).The in vitro drug release experiment showed that the absorbance in the 40 ℃ AP-170 group was 0 .1537 ± 0 .0094 ,which was significantly different from other groups ( P < 0 .01).Higher temperature promoted the release of chemotherapeutic drugs from carboplatin sustained-release microspheres , thus enhanced the chemotherapeutic effect .This might be the mechanism that UTMD affects the microenvironment in this experiment .Conclusions UTMD increases the permeability of tumor cells .The increase of temperature promotes more drugs into the microenvironment of tumor cells .Thus ,the local chemotherapeutic effect is enhanced in the tumor .
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Drugs for military special needs are important strategic materials for supporting military operational needs and maintaining national security,and are highly valued by Armed Forces in each country in the world.The U.S.military has accumulated much experience through the long-term management of drugs for military special needs.Here,The U.S.military and civilian integration management of drugs for military special needs between the U.S.DOD and FDA was analyzed in terms of research,purchase,use and storage,the secrets of success were summarized,and problems were identified.Finally,recommendations on military and civilian integration management of drugs for military special needs in China were proposed.
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OBJECTIVE:To systematically review the efficacy of Live combined bacillus subtilis and enterococcus faecium in the prevention and treatment of Antibiotic associated diarrhea(AAD),and provide evidence-based reference for clinical treatment. METHODS:Retrieved from CBM,CJFD and Wanfang database,randomized controlled trials (RCT)of Live combined bacillus subtilis and enterococcus faecium(test group)versus placebo(control group)in the prevention and treatment of AAD. After data ex-tract and quality evaluation,systematical evaluation was performed by using Rev Man 5.0 statistics software. RESULTS:A total of 21 RCT were included,involving 3 881 patients. According to the purpose of different purpose of druguse,the RCT were divided into prevention group and treatment group. Results of Meta-analysis showed the incidence of diarrhea in prevention test group was significantly lower than control group[OR=0.27 ,95% CI(0.22 ,0.32),P<0.001] ,the average hospitalization time in prevention test group was significantly less than control group [MD=-53.19,95%CI(-79.63,-26.75),P<0.001],and the total clinical ef-fective rate in treatment test group was significantly higher than control group[OR=6.76,95%CI(4.16,10.98),P<0.001]. CON-CLUSIONS:The efficacy of Live combined bacillus subtilis and enterococcus faecium are good in the prevention and treatment of AAD. However,due to the limit of methodological quality and sample size,it remains to be further verified with more rigorously designed and long-term follow-up of large-scale RCT.
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In a number of local wars and crisis in the region, the US military has shown the world its powerful military special drug development capabilities. To meet challenges including war wounds, chemical or biological weapons prevention , extreme environments as well as infectious diseases, the US military has developed a large number of military special needs medication. In this paper, in review of US medical product manuals and literature, we summarize the research progress in the US military drug prevention and treatment of chemical weapons injuries, drug prevention and treatment of infectious diseases, war injuries drugs, pharmaceutical and biological weapons and military operations medicament for the treatment of injury.
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Objective To analyse appearances of contrast-enhanced ultrasound(CEUS) of breast local adenosis,and to explore the value of CEUS in the diagnosis of the local adenosis of breast.Methods The data of conventional ultrasound and CEUS of local adenosis of breast in 110 cases confirmed by operation and pathology were analysed.The enhancement patterns,border of the lesion,time-intensity curve were evaluated respectively.Results Totally 151 lesions were detected by conventional ultrasound in 110 cases,61 lesions were diagnosed by conventional ultrasound for adenopathy or adenosis tumor,the diagnosis coincidence rate was 40.4% (61/151).By CEUS,106 lesions were diagnosed as adenopathy or adenosis tumor,the diagnosis coincidence rate was 70.2 % (106/151),higher than that of conventional ultrasound.The features of CEUS include:In contrast perfusion time to peak,103 lesions inhomogeneously enhanced in whole or in part (68.2 %).The enhancement degree of 121 lesions (80.1 %) were moderate enhancement or slightly high enhancement,a few lesions (11.9%,18/151) showed significantly enhanced,and very few lesions (7.9 %,12/151) for low enhancement.In contrast perfusion peak,107 (70.9 %) lesions had less clear boundary,no envelope,and had a certain continuity with the surrounding background breast tissue,in which 5 lesions were showed significantly increased in size by CEUS comparison with conventional ultrasound,and the boundary of 4 lesions was shaped like the burr,no regularity after enhancing.The boundary of 29.1% (44/151) of the lesion was clear.In the lesions with moderate enhancement or slightly high enhancement,the time-intensity curve mainly displayed same washin and same wash out (60/151,39.7 %) and fast washin and fast wash-out (65/151,43.0%).Conclusions CEUS image of focal breast adenosis has certain characteristics.CEUS can provide more evidences in diagnosis and differential diagnosis of breast adenosis,which can help to improve diagnostic confidence.
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The aims of this study were to investigate the utility of solid microneedle arrays (150 µm in length) in enhancing transdermal delivery of peptides and to examine the relationship between peptide permeation rates and D2O flux. Four model peptides were used (Gly-Gln-Pro-Arg [tetrapeptide-3, 456.6 Da], Val-Gly-Val-Ala-Pro-Gly [hexapeptide, 498.6 Da], AC-Glu-Glu-Met-Gln-Arg-Arg-NH2 [acetyl hexapeptide-3, 889 Da] and Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly-NH2 [oxytocin, 1007.2 Da]). The influence of microneedle pretreatment on skin permeation was evaluated using porcine ear skin with Franze diffusion cell. Peptide permeation across the skin was significantly enhanced by microneedle pretreatment, and permeation rates were dependent on peptide molecular weights. A positive correlation between D2O flux and acetyl hexapeptide-3 clearances suggests that convective solvent flow contributes to the enhanced transdermal peptide delivery. It is concluded that solid microneedle arrays are effective devices to enhance skin delivery of peptides.
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Objective To investigate the utilization of contrast-enhanced ultrasound (CEUS) for the detection of splenic artery steal syndrome (SASS) after orthotropic liver transplantation (OLT).Methods Color Doppler flow imaging (CDFI) were performed at various time points post-operatively.CEUS and celiac angiography were conducted in patients suspected of SASS.Results 9 patients were suspected of SASS by slim or undetectable hepatic arterial Doppler blood signals by CDFI at various time points postoperatively.CEUS in 9 patients showed a delayed and weak contrast-enhanced blood signal in the hepatic artery associated with a rapid and intense enhancement of portal venous blood.No narrowing of a hyperintense signal was observed in the hepatic artery by CEUS.The 9 diagnoses of SASS were proven by celiac angiography.Conclusions SASS is identified as a sluggish and weak hyperintense blood signal in the hepatic artery without the narrowing and interruption of hypeintense signal in CEUS.CEUS is an effective imaging modality for detection of SASS following OLT.
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ObjectiveTo make ultrasound anatomy teaching samples of odynopoeia fetus heart with complex abnormity consistent with echocardiographic view,and utilize them in echo teaching.MethodsTen odynopoeia foetus hearts were cutted comparing to different echocardiographic imaging respectively.Results Dissections of fetus heart were obtained including 5 cases of single cardiac ventricle(2 case with interruption of aortic arch),4 cases of double outlet of right ventricular,and 1 case of hypoplastic left heart syndrome with serious coarctation of the aorta.The apical long axis view,the three blood vessel view,as well as the long axis view of the aortic arch were typical views of ultrasound diagnosis and teaching.ConclusionsIt is profit to raise and accurate rate of fetus echocardiography diagnosis by studying dissection of fetus heart anatomic structure with complex abnormity,which would make ultrasound imaging visualization.
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Objective To investigate the effect of microbubble (MB) contrast agents impacted by different parameters such as acoustic pressure, frequency, duration of exposure, and MB concentration under flowing condition. Methods A capillary flow mimic model was set up for observation and analysis of MB displacement and aggregation under stereomicroscope. Results The displacement and aggregation of ultrasonic occurred significantly at the frequency of 2.0 MHz than 1.0 MHz and 0.5 MHz. Under low acoustic pressure, MBs were not visually disrupted but the flow slowed down. The aggregation and deflection applying to MB was stronger in the tube happened at the MB concentration of 7×10~7/ml than 7×10~5/ml, but did not when the concentration rose to 7×10~9/ml because of the high viscosity. The ultrasound exposure time could not affect significantly in displacement and MB aggregation. Conclusion MB contrast agents can be manipulated under some ultrasound parameters. It is expected to be physically modulated in blood vessels, in order to increase targeted adhesions for many therapeutic purposes.
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Objective To assess the left ventricular systolic asynchronicity in chronic ischemic model with real-time three-dimensional echocardiography (RT-3DE), and to explore the affection of low-dose dobutamine to it. Methods A chronic ischemic model was induced by placing an Ameroid constrictor in the left circumflex(LCX) in swines,then full volume RT-3DE was performed by Philips iE33 with X3-1 probe combining rest and stress(dobutamine stress echocardiography, DSE) every week after LCX constriction.Ten normal pigs before operation served as controls (group A). Examination of all the models post operation were grouped into group B (mild stenosis, LCX stenosis<50% ), group C (moderate stenosis, LCX stenosis 50%~75%) and group D (severe stenosis, LCX stenosis≥75%) according to the results of coronary angiography. Images were copied to QLAB 5.2 postprocess workstation,and 3DQA software was used to analyze the full volume data sets. The time to the point with minimal systolic volume (Tmsv) in each segment was taken to derive the following indexes of systolic synchrony: the maximum difference of Tmsv (Tmsv-dif) and standard deviation(Tmsv-SD) among various segments and standard index (Tmsv-dif% and Tmsv-SD%), to evaluate left ventricular dyssynchrony. Tmsv3-6 represented the maximum difference of Tmsv between lateral segment and posterior septum (Tmsv3-5: between lateral segment and inferior) in basal level. Results Tmsvl2-Dif%, Tmsv6-Dif%, Tmsv3-6% and Tmsv3-5% under stress condition in group C and D were significantly higher than those at rest;all the data in group D were significantly higher than in group A and B, and in group C higher than group A ( P <0.05,0.01 ). Compared with group A,Tmsv6-Dif,Tmsv3-6 and Tmsv3-5 in group B were significantly increased under stress condition,and so did their standardize data under both rest and stress conditions ( P < 0.05, 0. 01 ). Conclusions RT-3DEcombined with DSE could display sensitively the left ventricular asynchrony caused by chronic ischemia,and that will be more significant in lateral wall in LCX stenosis than in normal segments.
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Objective To compare two effective preparation methods for paclitaxel-loaded lipid microbubbles, and to evaluate the physiochemical properties as for acoustic activated drug delivery. Methods Paclitaxel-loaded lipid microbubbles were prepared with two methods: one was mixed with phospholipids directly (Ⅰ); the other was added in triacetin, then mixed with phospholipids (Ⅱ). Concent ration, size, pH, drug entrapment efficiency, drug-loading amounts of these two kinds of paclitaxel-loaded lipid microbubbles were studied, while drug release with ultrasound and tumor imaging enhanced on rabbit breast tumor were observed. Results There was no significant difference in tumor imaging between two kinds of microbubbles which could be ruptured by low energy ultrasound. Compared with Ⅰ, the mean diameter of Ⅱ decreased significantly ([ 1.07±0.38] μm vs [2.79± 0.41] μm, P<0.01), the surface potencial was higher ([19.10±0.32] mV vs [-5.90±0.21] mV, P<0.01), whereas entrapment efficiency and drug-loading amounts increased markedly ([ 95.00±1.22]% vs [36.10±4.74]%, P<0.01; [5.60±0.11]% vs [0.50±0.04]%, P<0.01). Conclusion The Ⅱ paclitaxel-loaded lipid microbubbles added triacetin have an important clinical value.
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An analytical method of fluorescent dye-labeled oligonucleotides was established by ion pair reversed phase high performance liquid chromatography(IP-RP-HPLC) which was improved by optimizing the effects of triethylamine-acetic acid(TEAA)(0-0.15 mol/L), pH(4.5-7.0) and gradient. Comparing the retention of 5, 10 and 15-mer unlabeled oligonucleotides with that of 5'-carboxyfluorescein(5'FAM) labeled oligonucleotides, the mechanism of fluorescent dye-labeled oligonucleotides retention was studied. In addition, TaqMan~(TM) probes as wellas other common fluorescent dye-labeled oligonucleotides were concerned. The results showed that the best resolution of different length fluorescent dye-labeled oligonucleotides was observed under the condition of 0.01 mol/L TEAA and pH 7.0. The retention behavior of fluorescent dye-labeled oligonucleotides was different from that of unlabeled oligonucleotides significantly, and therefore they can be separated completely. The results indicated that the retention of unlabeled oligonucleotides enhanced with the increase of the length of molecule. In contrast, the retention of fluorescent dye-labeled oligonucleotides was reduced with the increase of the length of molecule. For the hydrophobicity of fluorescent dyes made a great impact on the retention, a longer retention time the labeled oligonucleotides would take while the hydrophobicity of fluorescent dyes was higher. However, the effect of the hydrophobicity was limited as the length was increased to a certain level.
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Objective To explore the value of diagnostic ultrasound mediated microbubble destruction in improving the myocardial perfusion and left ventricular systolic function when cooperated with the mecsenchymal stem cells(MSCs) transplantation in rabbit myocardial ischemia. Methods One week after myocardial ischemia (MI) modeling,36 rabbits were divided into 3 groups,the control group(group Ⅰ) ,intravenous injection of MSCs group(group Ⅱ) and ultrasound + microbubble + MSCs group (group Ⅲ). Myocardial contrast enhancement (MCE) was performed and quantification analysis of anterior wall was assessed with Photoshop. Left ventrieular systolic function was assessed with M-mode echocardiography and bi-plane Simpson's method. CD34 expression in heart was detected with immunohistochemisty(IHC). Western blotting was applied to detect the level of VEGF in three groups. Results The differences of gray scale analyzed with histogram of Photoshop in anterior wall of ischemia myocardium between the group Ⅰ and group Ⅱ or group Ⅲ were significant,and P value was 0. 032 and 0. 000 , respectively. There were significant differences of FS between group Ⅲ (30. 43±4.09)% and group Ⅱ (26.29±2.93)%, P<0.01, and similar to group Ⅰ (19.28 ± 2.84)%. The difference of EF(%) between group Ⅲ and group Ⅱ was significant [(61.5±5.8 vs 53.6±4. 71), P<0. 05] ,or markedly significant between group Ⅲ and group Ⅰ [(61.5±5.8 vs 42.6± 5.0), P <0.01]. EF(%) assessed with bi-plane Simpson's method was significantly increased from (34.64 ± 4.59) in group Ⅰ to (41.78 ± 4.21) in group Ⅱ and (48.6±3.96) in group Ⅲ. The expression of CD34 assessed with immunohistochemistry was the highest in group Ⅲ. The level of VEGF with western blotting in group Ⅲ was significantly higher than other two groups. Conclusions It is an efficacious transplantation means of MSCs infusion under the ultrasound mediated microbubles destruction in improving the myocardial perfusion and cardiac systolic function.
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Objective To investigate the effect of microbubbles mediated ultrasound insonation on proliferation and apoptosis of vascular smooth muscle cells (VSMCs) in different phase of cell cycle. Methods Rat thoracic aortic VSMCs were cultured in vitro by the method of tissue adherence. The cells were synchronized by the methods of serum starvation and double thymidine block. The synchronization results were detected by flow eytometer. VSMCs in different phases of cell cycle were exposed to 1 MHz continuous waves ultrasound for 120 s at intensity 0.3 W/cm2 in the presence of lipid-coated microbubbles (1 ml/L). Apoptosis of VSMCs was analyzed by AnnexinV/PI staining using flow eytometry. The proliferation and the proliferating cell nuclear antigen(PCNA) protein expression of VSMCs were detected by MTT assay and immunoeytochemistry, respectively. Results The synchronized G0/G1 and S phase VSMCs were achieved, with synchronized rates to 89.53 % and 66.87 %, respectively. Ultrasound sonication for 120 s with microbubbles could significantly inhibit the proliferation and downregulate the PCNA expression of S phase VSMCs,but the proliferation and PCNA expression of G0/G1 phase VSMCs were not affected. After treatment of ultrasound with microbubbles, the apoptotic ratio were found to reach (7.05 ± 2.04)% in G0/G1 phase VSMCs and (27.01 ±3.87)% in S phase VSMCs. Conclusions Microbubbles mediated ultrasound insonation can significantly inhabit the proliferation and induce apoptosis in VSMCs at proliferation stage.
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Objective To investigate the effect of ultrasound irradiation combined with liposome membrane microbubbles on the reorgnization of cytoskeleton in vascular smooth muscle cells (VSMCs). Methods Rat thoracic aortic VSMCs were cultured in vitro. VSMCs were exposed to 1 MHz continuous waves ultrasound radiation for 120 s at intensity 0.3 W/cm2in the presence of liposome membrane microbubbles (1 μl/ml) after treated with platelet derived growth factor-BB (PDGF-BB). The reorganizations of microfilaments, microtubules and intermediate filaments were examined by using immunofluorescence and fluorocytochemistry techniques. Results There was a substantial increase in the expression of F-actin and assembly of long bundles of stress fibers in the transversed cell body when treated with PDGF-BB. Neither alterations of β-tubulin nor of vimentin cytoskeletal protein organization were observed in PDGF-BB treated cells as compared to those of the contol group. After ultrasound irradiation combined with liposome membrane microbubbles, the expression of F-actin, β-tubulin and vimentin were reduced along with the simultaneous changes in microfilaments, microtubles and intermediate filaments array. Conclusions Ultrasound irradiation combined with liposome membrane microbubbles can induce significant changes in cytoskeleton structure of VSMCs cultured in vitro.
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Objective To investigate characteristics and ultrasonic imaging of the nano-scale ultrasound-enhanced contrast in vivo. Methods Nano-scale microbubbles were prepared by machine vibration and low speed centrifugation. The surface morphology and average size distribution of microbuble were measured. The contrast-enhanced effect of the hahn-scale microbubbles in the normal rabbit liver was observed,and compared with micro-scale microbubbles. Results The nano-scale microbubbles had a good shape and uniform distribution by light microscope and transmission electron microscope, with average diameter of 623.4 nm and surface electric potential of 1.3 inV. The contrast imaging study in vivo showed the nano-scale microbubbles could significantly enhance ultrasonic imaging of rabbit livers, which had no obvious difference with micro-scale microbubbles. Conclusions The nano-scale ultrasound contrast agent is stable and effective for the enhancement of ultrasound imaging, which is based on development of miniaturizing targeting ultrasound contrast agent.