ABSTRACT
Snake venom has special pharmacological activities and contains a array of small polypeptides that can antagonize integrins, therefore called disintegrins. Disintegrins can block integrin-dependent platelet aggregation, tumor growth, and tumor metastasis. A disintegrin fraction was isolated and purified from the venom of snake Gloydius brevicaudus (GBV). Its physical and chemical properties were characterized, and its biological activities were investigated. The crude venom of GBV were isolated by Superdex 75 gel filtration chromatography. The anti-platelet aggregation activity of the fractions was screened by the Born method. The fraction that shown anti-platelet activity was further purified with Sephadex G-25 gel filtration, DEAE Sepharose Fast Flow ion exchange chromatography, and Lichrospher C18 reversed-phase chromatography respectively. The purity of the active component was analyzed with SDS-PAGE (Tris-Tricine system) and high-performance liquid-phase chromatography (HPLC), with protein concentration determined by the Bradford method. The molecular weight was evaluated by the gel imaging method and mass spectrometry, and the isoelectric point was measured by disc isoelectric focusing electrophoresis. The protease activity was measured with the Rick method. The phospholipase A activity was determined by the automatic potentiometric titration method. Amino acid sequencing results were subjected to homology comparison using the BLAST program. Seven fractions (Ⅰ-Ⅶ) were isolated from GBV by gel filtration chromatography on Superdex 75 column. The fraction Ⅳ inhibited the platelet aggregation induced by ADP with molecular weight lower than 10 000 Da, suggesting a disintegrin component. A disintegrin named GBV-Ⅳ4 was purified from the fraction by Sephadex G-25 gel filtration, DEAE Sepharose Fast Flow ion-exchange and Lichrospher C18 reverse chromatography. It was homogeneous shown as a single band on SDS-polyacrylmide gel electrophoresis (SDS-PAGE, Tris-Tricine system) with molecular weight 8 746 Da as calculated by Image Master VDS system. The isoelectric point of GBV-Ⅳ4 was 6.3 by disc polyacrylamide gel electrophoresis. GBV-Ⅳ4 exhibited no detectable phospholipase A2 (PLA2) activity with the pH-stat technique or proteinase activity according to the method of Rick. GBV-Ⅳ4 is composed of 70 amino acids with RGD (Arg-Gly-Asp) active region and a molecular weight of 7 442 Dalton as assayed by Mass Spectrography. Characterization of GBV-Ⅳ4 is consistent with meta-chain disintegrin (70 amino acid sequence, six pairs of disulfide bond). Retrieved by Genbank, GBV-Ⅳ4 has high homology with other disintegrins. We concluded that GBV-Ⅳ4 is a novel disintegrin contained RGD. GBV-Ⅳ4 showed dose-dependent inhibition of ADP- or thrombin-induced platelet aggregation with IC50 0.339 or 0.577 μg·mL-1 respectively. In conclusion, a new disintegrin derived from the GBV snake venom and named GBV-Ⅳ4 containing RGD tripeptide sequence could inhibit platelet aggregation.
ABSTRACT
Objective To investigate the characteristics of 18F-Alfatide II PET/CT imaging in normal breasts and breast cancer lesions.Methods From March 2016 to August 2017,22 female patients(age:(52±10)years)with suspected breast malignant nodules or masses were prospectively enrolled.All patients underwent 18F-Alfatide II PET/CT imaging prior to biopsy or surgery.The imaging characteristics of normal breasts were assessed visually and the difference of maximum standardized uptake value(SUVmax)in normal breasts and uterus between patients with and without menopause was compared,SUVmax of cancer lesions and normal breasts was also compared.Breast cancer lesions were classified according to the distribution characteristics of radioactive uptake,and molecular subtypes ware determined by immunohistochemistry and fluorescence in situ hybridization.The SUVmax of different morphological and molecular subtypes were analyzed.Two-sample t test and Pearson or Spearman correlation analysis were used to analyze the data.Results There were 23 breast cancer lesions(one patient had bilateral breast cancer lesions and one had a history of one-side breast resection),20 normal breasts and 21 normal uteruses.Those normal breasts and uteruses didn't show any malignant change after being followed up for more than 1 year(one patient had uterine fibroids resection).There was a slight increase of radioactivity uptake in the cord-like connective tissue region at the margin of the gland in 11 mammary glands,and the SUVmax was higher than that of glandular tissue in the central region(1_81±0.67 vs 0.79±0.37;t = 6.771,P<0.00l).Of the 11 cases,except for one patient whose uterus was removed,the other 10 patients were accompanied by increased diffuse radioactivity of the uterus.SUVmax of 19 normal breast connective tissues(1.31±0.80)and uterus(3.80+1.79)were positively correlated(r = 0.785,P<0.05).For patients with/without menopause(n= 11 each group),the SUVmax of normal breast connective tissues(0.72±0.39 vs 1.81±0.67)and uterus(2.04±0.39 vs 5.11 + 1.06)were significantly different(t values:4.42 and 8.66,both P<0.01).Different levels of radioactive uptake were observed in all 23 breast cancer lesions,with SUVmax of 6.93±3.97,which was significantly higher than the nipple,connective tissue and glandular tissue of normal breasts(t values:6.784-7.559,all P<0.05).According to the characteristics of the radioactivity uptake distribution of the lesion,among the 23 breast cancer lesions,5 were mass type,3 were nodular type,4 were diffuse type,and 11 were multi-focal/multi-center type,and the SUVmax of multi-focal/multi-center type was the highest(F=3.55,P<0.05).The SUVmax of basal-like breast cancer lesions(2.49±1.67)was lower than the other three molecular subtypes.Lesions with high level human epidermal growth factor receptor 2(HER2)positive expression had higher SUVmax.Conclusions 18F-Alfatide II PET/CT imaging shows that normal breasts have a slight radioactive distribution,mainly concentrate in the nipple and connective tissues around the glandular,and the uptake have a positive correlation with the radioactive uptake of the uterus.The degree of radioactive uptake of breast cancer lesions is significantly higher than that of normal breasts.Breast cancer lesions with different moqjhological features all have obvious radioactive uptake,especially the multi-focal/multi-center type.Different molecular subtypes have different radioactive uptake levels.SUVmax is lower in basal-like breast cancer lesions,and higher in HER2 positive expression lesions.
ABSTRACT
Objective To investigate the biodistribution of 18F-Alfatide II in patients with breast diseases and to compare its uptake with 18F-fluorodeoxyglucose(FDG)uptake.Methods A total of 44 female patients(age:(50.7±8.0)years)with clinically suspected breast cancer from December 2015 to May 2017 were prospectively enrolled and underwent 18 F-Alfatide II and 18F-FDG PET/CT prior to treatment.By drawing regions of interest in normal organs and breast lesions,differences between 18F-Alfatide II uptake and l8F-FDG uptake were evaluated in all patients.Paired t test,two-sample t test and Wilcoxon rank sum test were used for data analysis.Results There were 53 breast lesions confirmed by histopathology in 44 patients.Among them,42 lesions were malignant and the others were benign.The uptake of 18F-Alfatide II was very low in the brain,vocal cords,lungs,blood pool and muscle.But the renal cortex and bladder had high 18F-Alfatide II accumulation.Different levels of 18F-Alfatide II uptake were found in other normal organs including normal breast tissue.There were differences(t values:2.04-41.65,all P<0.05)between 18F-Alfatide II and 18F-FDG maximum standardized uptake value(SUVmax)and mean standardized uptake value(SUVmean)in many normal organs except for the choroid plexus,salivary glands,liver,colon and normal breast tissue.The uptake of 18F-Alfatide II was significantly lower than 18F-FDG in breast cancer lesions(SUVmax:3.77±1.78 vs 7.37±4.48,SUVmean:2.25±0.98 vs 4.54±2,82;t values:4.89,4.82,both P< 0.05),but it was still higher in benign breast lesions(SUVmax:2.37±1.62,SUVmean:1.50±0.92;t val-ues:2.35,2.29,both P<0.05).Also,target/non-target(T/NT)of 18F-Alfatide II in breast cancer lesions was higher than that in benign breast lesions(5.32±3.08 vs 2.60±2.37;t = 2.72,P<0.05).Condusion The biodistribution of 18F-Alfatide II in patients is favorable and 18F-Alfatide II can be clinically used for breast cancer imaging.
ABSTRACT
Objective To assess the imaging characteristics of 18F-Alfalide II in different tumorbearing mice and pharmacokinetics in Beagle dogs.Methods BALB/c nude mice(n-24)were used for subcutaneous tumor models(A549 and U87MG),orthotopic lung cancer models(A549)and orthotopic breast cancer models(MDA-MB-231)(n=6 in each group).18F-Alfatide II and 18F-fluorodeoxyglucose(FDG)microPET/CT images were compared in the 4 types of tumor-bearing nude mice models.18F-Alfatide II blocking experiment,biodistribution experiment and imaging studies in tumors of different growth cycles were performed in A549 subcutaneous tumor-bearing nude mice models.Pharmacokinetic experiments were carried out in Beagle dogs(n = 6)and CD-1 mice(n = 9).Two-sample t test was used to analyze the data.Results Compared with 18F-FDG,18F-Alfatide II microPET/CT images showed better imaging quality and contrast in subcutaneous A549,U87MG tumors and orthotopic A549(tumor/heart:4.50±1.17 vs 0.95±0.31;t = 4.125,P<0.01),orthotopic MDA-MB-231(tumor/muscle:6.60±1.53 vs 0.92±0.43;t = 3.984,P<0.01)transplantation nude mice models.18F-Alfatide II could specifically target A549 tumors,and the tumor uptake of 18F-Alfatide II was reduced by about 75% after pre-injection with cyclo(Arg-Gly-Asp-D-Tyr-Lys)(c(RGDyk)).18F-Alfatide II was rapidly cleared from the blood of Beagle dogs(T1/2 was(57.34±11.69)min).It was cleared in the form of prototype drug and(69.24±6.82)% of cumulative dose was excreted through the urine within 4 h after administration.Conclusions 18F-Alfatide II shows a higher target/non-target ratio than,18F-FDG in the imaging of A549,MDA-MB-231 and U87MG tumor-bearing nude mice models,which is more conducive to the diagnosis of tumor.18F-Alfatide II has excellent pharmacokinetic properties.
ABSTRACT
Objective To develop the automated preparation of 18F-Alfatide II using newly-designed 18F-minireactor and perform 18F-Alfatide D microPET/CT imaging in tumor.Methods The automated preparation of 18F-Alfatide H was developed by using 18F-microreactor and water phase Al18F-chelating method,and the radiochemical yield and quality analysis were measured.The nude mice bearing breast tumor ZR-75-1 and nasopharyngeal tumor CNE1 were established(n = 3 respectively).MicroPET/CT imaging was performed at 0.5,1.0 and 2.0 h after the injection of 18F-Alfatide II.The region of interest(ROI)was depicted and the tumor/muscle(T/M)ratio was calculated.Results 18F-Alfatide II was automatically prepared with the total synthesis time of 40 min,the radiochemical yield of(28±6)%(no decay corrected,n=11),and the radiochemical purity >97%.All quality analysis indexes accorded with the radiopharmaceutical requirements.18F-Alfatide II microPET/CT images of ZR-75-1 and CNE1 tumors were clear due to the high radioactivity uptake of tumor lesions(T/M ratio was greater than 4.0 at 1.0 h after injection).Conclusion Based on the 18F-minireactor,the,8F-Alfatide II can be prepared successfully with short synthesis time and high radiochemical yield,which can help the application studies in 18F-Alfatide II microPET/CT imaging.
ABSTRACT
The integrin is a family of membrane receptor proteins composed of heterologous dimer , it can identify the sequence RGD .Integrin excessive expressed in tumor cells and angiogenesis .Using their special relationship a-mong them, the RGD as the carrier, combined with the radioactive tracer , antineoplastic drugs and other substances,which has tumoraffin imaging ,targeted anticancer therapy ,anti-inflammatory,anti depression effect and bone and nerve repair , and so on .
ABSTRACT
Objective@#To observe the uptake of 99Tcm-3 polyethylene glycol Arg-Gly-Asp dimer (3PRGD2) in rat models of rheumatoid arthritis (RA), in order to provide theoretical foundation for early diagnosis of RA.@*Methods@#The healthy female SD rats were divided into collagen induced arthritis (CIA) group (n=100), osteoarthritis (OA) group (n=20) and control group (n=20). Bovine collagen type Ⅱ emulsion was used for arthritis induction to establish CIA models. OA models were established by injection of L-cysteine and papain. Gamma imaging was performed before and 15 d, 30 d after the model establishment. The mediastinum was selected as non-target (NT) area and the target (T)/NT ratios were calculated. The serum levels of vascular endothelial growth factor (VEGF), tumor necrosis factor-α (TNF-α), αVβ3 before and after the model establishment were measured. Pathological and immunohistochemical detection were performed. One-way analysis of variance, Pearson and Spearman correlation analyses were used to analyze the data.@*Results@#There were 58 CIA models successfully established. Before the model establishment, T/NT ratios of CIA group, OA group and control group were 0.215±0.049, 0.210±0.050, 0.209±0.051, respectively (F=0.093, P>0.05). The T/NT ratios of the above three groups were 0.405±0.230, 0.223±0.045, 0.211±0.049 (F=12.601, P<0.05) 15 d post-model establishment, and those were 0.572±0.182, 0.238±0.045, 0.212±0.055 (F=65.147, P<0.05) 30 d post-model establishment. T/NT ratios of CIA group were positively correlated with the levels of serum VEGF, TNF-α, αVβ3, pathological score and the levels of immunohistochemical markers (VEGF, TNF-α, αVβ3, CD31, CD34; r values: 0.391-0.721, rs values: 0.365-0.669, all P<0.05).@*Conclusion@#99Tcm-3PRGD2 can be specifically up-taken by RA lesions in rat models, thus has the potential in early synovial neovascularization imaging of RA.
ABSTRACT
Objective To evaluate 18F-fluorodeoxyglucose (FDG) PET/CT in the rabbit model of vulnerable plaques by correlation with 99Tcm-Arg-Gly-Asp (RGD) SPECT/CT imaging,lipid levels,pathological and immunohistochemical results.Methods Sixteen male New Zealand white rabbits were randomly divided into normal diet group (group A,n =4),stable plaque group (group B,n =4) and vulnerable plaque group (group C,n =8) using completely random grouping method.The animals were given abdominal aorta sham operation (groups A and B) or balloon injury of the abdominal aorta (group C) 2 weeks after feeding.Animals were injected with 18F-FDG and 99Tcm-RGD respectively at the end of 4,8 and 12 weeks.PET/CT was performed at 1,2 and 3 h post-injection.SPECT/CT was performed at 30 min post-injection.One rabbit was sacrificed at the end of 4 and 8 weeks after imaging studies,respectively.The others were sacrificed at the end of 12 weeks after imaging studies.All abdominal aortas were harvested.Pathology and immunohistochemistry analysis were performed.The data were analyzed by one-way analysis of variance and Pearson correlation analysis.Results There was no uptake in any group at 4th week and no uptake in group A or group B at 8th week.There was mild uptake in group B at 12th week and group C at 8th week.There was intense uptake in group C at 12th week,whereas both mean standardized uptake value (SUVmean) and maximum standardized uptake value (SUVmax) were significantly higher than the other two groups (F values:7.952,14.279,both P＜0.05).In group C,SUVmax(0.43±0.08,0.68±0.06,1.74±0.63) and SUV (0.37±0.03,0.56±0.03,1.26+0.23) had significant difference at 3 h post-injection for imaging at 4th,8th and 12th week (F values:10.939,39.747,both P＜0.05).At 12th week,there was a strong correlation between the uptake of 18 F-FDG and target/non-target (T/NT) ratio of 99Tcm-RGD in all groups(r values:0.748,0.709,both P＜0.05).Histopathology results showed that the plaques had rich macrophages and a small amount of smooth muscle cells in group C,little macrophages in group B,while no macrophages in group A.Conclusion 18F-FDG PET/CT might be an effective noninvasive method for early assessment of aortic vulnerability to atherosclerotic plaque.
ABSTRACT
Objective To investigate the value of 99Tcm-3PRGD2 SPECT/CT imaging in the diagnosis of benign and malignant lung lesions.Methods A retrospective analysis of 99Tcm-3PRGD2 planar imaging and chest SPECT/CT imaging with lung lesions before treatment was performed.Totally 37 cases including 27 malignant and 10 benign lesions confirmed pathologiclly were enrolled.Compared with hepar in planar imaging,the intake gray scale of 99Tcm-3PRGD2 in lung lesions was divided into low,moderate,and high elevations.The ratio of the highest intake value of the lesion (L) to the contralateral normal lung (N),the hepar (H),the mediastinum (Me) and the contralateral deltoid muscle (Mu) was calculated,and statistical analysis was performed to evaluate the diagnostic efficacy.The expression of immunohistochemistry integrin αvβ3 was performed through selecting 1 case of benign and malignant lung lesions,respectively.Results The intake of 99Tcm-3PRGD2 was higher in 37 patients with lung lesions.High elevation of malignant lesion was 66.67％ (18/27),low and moderate elevation of benign lesion was 90.00％ (9/10).The intake ratio of malignant lung lesions was higher than that of benign lesions (all P＜0.05).When L/N was greater than 5.45 or L/Mu was greater than 4.65,the sensitivity,specificity and accuracy of diagnosis of lung malignant lesions was 77.80％,80.00％ and 78.40％,respectively.Immunohistochemical results showed that malignant lung lesions highly expressed integrin αvβ3,and only a small number of benign lesions expressed it.Conclusion 99 Tcm-3PRGD2 SPECT/CT is valuable in the differential diagnosis of benign and malignant lung lesions.
ABSTRACT
Objective To investigate the value of 99Tcm-3PRGD2 SPECT/CT imaging in the diagnosis of benign and malignant lung lesions.Methods A retrospective analysis of 99Tcm-3PRGD2 planar imaging and chest SPECT/CT imaging with lung lesions before treatment was performed.Totally 37 cases including 27 malignant and 10 benign lesions confirmed pathologiclly were enrolled.Compared with hepar in planar imaging,the intake gray scale of 99Tcm-3PRGD2 in lung lesions was divided into low,moderate,and high elevations.The ratio of the highest intake value of the lesion (L) to the contralateral normal lung (N),the hepar (H),the mediastinum (Me) and the contralateral deltoid muscle (Mu) was calculated,and statistical analysis was performed to evaluate the diagnostic efficacy.The expression of immunohistochemistry integrin αvβ3 was performed through selecting 1 case of benign and malignant lung lesions,respectively.Results The intake of 99Tcm-3PRGD2 was higher in 37 patients with lung lesions.High elevation of malignant lesion was 66.67％ (18/27),low and moderate elevation of benign lesion was 90.00％ (9/10).The intake ratio of malignant lung lesions was higher than that of benign lesions (all P＜0.05).When L/N was greater than 5.45 or L/Mu was greater than 4.65,the sensitivity,specificity and accuracy of diagnosis of lung malignant lesions was 77.80％,80.00％ and 78.40％,respectively.Immunohistochemical results showed that malignant lung lesions highly expressed integrin αvβ3,and only a small number of benign lesions expressed it.Conclusion 99 Tcm-3PRGD2 SPECT/CT is valuable in the differential diagnosis of benign and malignant lung lesions.
ABSTRACT
Objective To evaluate the effect and the anti-tumor mechanism of nucleoside combination on human hepatoma cell line Bel-7402 with Arg-Gly-Asp sequence labeled by ultrasmall superparamagnetic iron oxide (RGD-USPIO).Methods The tumor cells Bel-7402 of logarithmic phlyhase were divided into experimental group and control group,treated with 1 mmol/L nucleoside combination and 1640 medium respectively.The two group were co-cultured for 48 h,and were added RGD-USPIO and co-cultured for 6 h.Then the two groups were proceeded with MR scanning,and the signal intensity of T2WI were measured.After extraction of the total RNA and protein of experiment group and control group,the expression of integrin avβ3 was detected using real-time PCR and Western blot.Results The T2WI signal intensity of experimental group (997.35±42.83) was higher than that of control group (241.05±15.36,t 28.79,P＜0.01).Compared with control group,the expression of integrin αvβ3 mRNA in experimental group was (0.22±0.02) times (t=4.50,P＜0.01).According to Western blot,the protein bands of experimental group were relatively lighter than that of control group,the expression of integrin αvβ3 in experimental group was lower (t =11.88,P＜0.01).Conclusion Nucleoside combination has anti-tumor effect by inhibiting integrin ligand-receptor binding,and the anti tumor mechanism may be related to the induction of tumor cell apoptosis.MR molecular probes can conveniently and accurately evaluate the anti-tumor effect of nucleoside combination on Bel-7402 cells.
ABSTRACT
Objective To investigate the feasibility of a novel molecular probe 99Tcm-3P4-RGD2 in evaluating arterial plaque stability after atorvastatin intervention in rabbits with SPECT/CT. Methods Eighteen male New Zealand rabbits were randomly divided into group A (stable plaque), group B (vulnerable plaque), and group C (vulnerable plaque with statin intervention). All rabbits were fed with high-fat food for 12 weeks. After high-fat feeding for two weeks, sham surgery was performed on group A. In the meantime, abdominal aorta injury was performed on group B and group C. After that, rabbits of group C were given oral atorvastatin (2.5 mg·kg-1·d-1). 99Tcm-3P4-RGD2 SPECT/CT imaging was performed on each group at the end of 4, 8 and 12 weeks. T/NT ratios were calculated. Animals were sacrificed at the end of 12 week after imaging studies. The abdominal aortas were collected, imaged with SPECT/CT, and evaluated by pathological HE staining and immunohistochemical analysis. MVD was calculated. Differences among 3 groups were analyzed using one-way analysis of variance. Results There was no significant radioactive uptake in the abdominal aortas of three groups on the 4th week′s imaging. The radioactive uptake in abdominal aortas increased slightly on the 8th week, with the highest radioactive uptake in group B. The radioactivity in abdominal aortas of the 3 groups maintained increasing on the 12th week, with T/NT ratios of 1.579±0.217, 1.873±0.226 and 1.524±0.237, respectively (F=8.984, P<0.05). In ex vivo abdominal aorta images, especially images of group B, radioactivity in lesion sites was higher than that in normal tissue. Accordingly, results of HE staining showed that artery plaques of group A and group C were grade Ⅱ and group B was grade Ⅳ. The MVD of group A, B and C was 8.17±1.17, 15.86±1.07 and 7.17±1.60, respectively (F=9036, P<0.05). Conclusion 99Tcm-3P4-RGD2 SPECT/CT imaging has a high sensitivity in the evaluation of arterial plaque stability after statin intervention in rabbits.
ABSTRACT
Objective To analyze the compatibility of bone marrow mesenchymal stem cells (BMSCs) with amphiphilic peptide three-dimensional gel.Methods Three healthy 3-week old SD rats were taken for separating femur and tibia to obtain BMSCs,the BMSCs surface antigen was detected by flow cytometry;the 10 mg/mL RGD-cyclic amphiphilic peptide solution was added into the same volume of DMEM/F12 culture medium,after a few seconds,which was self assemble into three-dimensional gel.Three dimensional gel structure was observed by transmission electron microscope (TEM).1 × 106 cells/mL BMSCs suspension and RGDcyclic amphiphilic peptide were mixed to form a 3D culture system,1 × 106 cells/mL BMSCs suspension was mixed with polylysine to form a 2D culture system,the serum-free culture was conducted;the CCK-8 method was used to observe the cell growth situation,calcein acetoxy methyl ester/propidium iodide (PI) double standard staining was performed.The effect of RGD-cyclic amphiphilic peptide on the proliferation of BMSCs was observed by fluorescence microscopy.Results The separated and cultured BMSCs highly expressed CD29 and CD90,but lowly expressed or did not express CD34 and CD45;TEM showed that the gel was composed of multiple empty nanofibers with the nanofiber diameter of 2-5 nm and length of 100-1 000 nm;the molecular weight of synthetic peptides detected by mass spectrometry (MS) was 1 256.37,which was consistent with the theoretical value;the HPLC analysis showed that RGD-amphiphilic peptide purity was 95.88 ％;the calcein acetoxyl methyl ester/PI double staining showed that in the 3D culture system,a few BMSCs died after 30 min and the cells began to proliferate after 12 h,the proliferation was more active than that of 2D culture,and the difference was statistically significant (P＜0.05);CCK-8 cell count showed that the proliferation activity of 3D culture system was higher than that of 2D culture system,and the difference was statistically significant (P＜0.05).Conclusion RGD amphiphilic peptide has a good biocompatibility with BMSCs,and may become the tissue engineering scaffold material.
ABSTRACT
Objective To investigate the potential of 99Tcm-Durarnycin and 99Tcm-RGD in detecting vulnerable plaque in rabbit models.Methods Fifteen healthy New Zealand male rabbits were randomly divided into group A (control group,n =5),group B (stable plaque group,n =5) and group C (vulnerable plaque group,n =5).Animals were injected with 99Tcm-Duramycin and 99Tcm-RGD at the end of 4,8 and 12 weeks.SPECT/CT scanning was performed at 0.5 h post injection.One rabbit was sacrificed at the end of 4 weeks and one at the end of 8 weeks after imaging.The others were sacrificed at the end of 12 weeks after imaging studies.All aortas were collected.Intravascular ultrasound (IVUS) was performed at the end of 8,12 weeks before SPECT/CT scanning.The data was analyzed with paired t test.Results In group A,the aortas had little uptake of the two probes.In group B,the aortas showed obvious radioactive uptake of 99Tcm-Duramycin and 99Tcm-RGD at the end of 8 weeks and 12 weeks,while 99Tcm-Durarnycin gave better display than 99Tcm-RGD.In group C,99Tcm-Duramycin uptake was higher than 99Tcm-RGD uptake in the aorta.The T/NT ratios of 99Tcm-Duramycin and 99Tcm-RGD in group C were 2.14±0.34 and 1.46±0.34 (t=4.072,P＜0.05) at the end of 4 weeks,2.93±0.41 and 1.66±0.22 (t=5.578,P＜0.05) at the end of 8 weeks,3.25±0.29 and 1.81±0.28 (t=19.692,P＜0.05) at the end of 12 weeks.In isolated specimen of group C,the yellow lipid plaque of the intima bulged on the lumen at the end of 12 weeks.IVUS indicated that,at the end of 8 weeks and 12 weeks,the endometrial thickness of group C was (450±104) mm and (767±52) mm (t=44.024,P ＜ 0.05) respectively,and the rates of luminal stenosis were (29.30± 2.81) ％ and (37.98 ±6.41)％ (t =9.226,P＜0.05).Conclusions Both 99Tcm-Duramycin and 99Tcm-RGD may be used to detect vulnerable plaque at early time.99Tcm-Duramycin may detect vulnerable atherosclerotic plaque earlier than 99Tcm-RGD and provide better diagnostic image.
ABSTRACT
Objective To investigate the synthesis, in vivo biodistribution of 99Tcm-HYNIC-PEG4-E[PEG4-c (RGDfk)] 2 (99 Tcm-Galacto-RGD2), and its potential usage for targeted imaging of mice orthotopic glioma.Methods 99Tcm-Galacto-RGD2 was synthesized straightforward and its radiochemical purity and stability and distribution in mice were analyzed.MicroSPECT-CT imaging was done in a mice orthotopic glioma model, which had been set up with U87MG cells, after administration of 99Tcm-Galacto-RGD2.Region of interest (ROI) of glioma was drawn on SPECT-CT section images to quantify tumor uptake (％ ID/cm3).Glioma was harvested for pathological examination.Linear-regression was used to analyze the relationship between integrin αvβ3 and tumor uptake (％ID/cm3).Results The radiochemical purity of 99Tcm-Galacto-RGD2 was (97.7 ±0.8)％ and stable in vitro.Hynic-Galacto-RGD2 could specifically bind to integrin αv β3 of tumor cells with a IC50 of (18 ± 3) nmol/L.After tail vein injection, 99Tcm-Galacto-RGD2 was rapidly discharged from the blood, liver, kidneys and had a relative low concentration in normal brain tissue.MicroSPECT-CT imaging demonstrated that, after 60 min of injection, this drug was well uptaken by glioma tumor than that after 30 min (t =7.13 ,P ＜0.05), and the tumor to normal brain tissue (T/B) uptake ratio of 99Tcm-Galacto-RGD2 was 13.92± 3.43.Injection of HYNICGalacto-RGD2 2 min prior to 99Tcm-Galacto-RGD2 injection extensively reduced the uptake of radioactive drug in tumor tissue (t =11.36, P ＜ 0.05).Bland-Altman analysis showed that tumor volume based on SPECT-CT imaging measurement had almost same value with the tumor reference volume (95％ CI =-11.94％-11.92％).In addition, the tumor uptake of 99Tcm-Galacto-RGD2 and cellular integrin αvβ3 expression level had a linear relationship (R2 =0.896).Conclusions Stable 99Tcm-Galacto-RGD2 can be synthesized easily and is applicable for microSPECT-CT imaging analysis of orthotopic glioma in mice together with the evaluation of integrin αvβ3 level in tumor.
ABSTRACT
Objective To study the feasibility of a novel probe 99Tcm-HYNIC-2(poly-(ethylene glycol),PEG) 4-Dimer (Dimer:E-[c (RGDfK) 2]) as a potential imaging agent for integrin αv β3 positive tumors,and also to observe the influence of an angiogenesis inhibitor,endostar,on the biodistribution and tumor uptake of the tracer in tumor bearing nude mice.Methods The expression of integrin αv β3 in human glioma cells U87MG was determined with immunofluorescence staining before and after treatment with endostar.99Tcm-HYNIC-2PEG4-Dimer was prepared and administered in U87MG tumor bearing mice in 6 h after either administration of endostar (200 μl) or saline (control group) and then biodistribution study was performed.Other 16 mice were divided into endostar treated group (20 mg/kg) and control group (saline) and then gamma imaging was performed in the two groups.Statistical significance of differences between the two groups was assessed using two-sample t test.Results Radiochemical purity of 99Tcm-HYNIC-2PEG4-Dimer was exceeded 95％.The expression of integrin αvβ3 in U87MG cell was high and gradually decreased after treatment with endostar.There was a negative dose-effect relationship between the dose of endostar and the expression of integrin αvβ3 with the peak effect at the dose of 400 μg/ml.The distribution study in vivo showed that the tracer uptake of U87MG tumors was high,but it decreased after injection of endostar.At 90 min,the ％ID/g of endostar and control groups were 1.50±0.08 and 6.27±0.33,respectively (t =40.23,P＜0.05).The average T/NT ratios of 99Tcm-HYNIC-2PEG4-Dimer uptake in the endostar and control groups were 1.02±0.11 and 2.58±0.36,respectively (t =10.25,P＜0.05).The integrin αv β3 positive expression ratios of tumor in endostar and control groups were (33.1 ±2.7) ％ and (81.5±3.2) ％,respectively (t =32.60,P＜0.05).Conclusions The novel probe 99Tcm-HYNIC-2PEG4-Dimer may be a promising radiotracer for integrin αvβ3-positive tumor imaging.It may be used for monitoring the therapeutic effect of endostar and may be potentially used for screening the candidates of anti-angiogenesis therapy.
ABSTRACT
Objective To evaluate the biological potential of surface topography of biomimetic matrix combined PRP gels with RGD modification. Methods Surface topography of nβ-TCP/Cs/PCL matrix was made by Nd: YAG laser, tissue-engineered bone was constructed in the following ways: ADSCs were loaded to nβ-TCP/Cs/PCL matrix with PRP gels plus RGD modification (group A), ADSCs were cultured to nβ-TCP/Cs/PCL matrix with RGD modification (group B), ADSCs implantation with topography-surfaced treatment of matrix (group C), ADSCs cultivation with smooth-surfaced treatment of matrix (group D). SEM and CLM were used to observe cellular pattern, survival rate, cell activity, ALP and collagen type Ⅰ level were detected at 1, 4, 8, 12, 16, 20, 24,28 days. Runx2 and OPG expressions were assayed at different interval. Results Under observation of SEM and CLM, new tissue showed more remarkable cell proliferation with abundant ECM in group A. Compared with other groups, the survival rate in group A was significantly higher (88.16 ± 1.29, P ＜ 0.05),and the level of cell activity, ALP, collagen type Ⅰ were significantly higher (0.92± 0.13, 87.27 ± 3.08, 93.27 ± 3.91, P＜ 0.05), and remarkable expression of Runx2 and OPG was also seen. Conclusion Topography-surfaced treatment of matrix combined PRP gels with RGD modification enhances the cell proliferation and acts as a feasible osteopromotive method.
ABSTRACT
Objective:To study the effects of ~(125)I-(α_v)ASODN on the in vitro invasive ability of heptocellular carcino-ma cell line(HepG2) through PEI-RGD-mediated receptor process. Methods: Intergrin α_v-specific antisense oligonucle-otide was labeled with ~(125)I, and PEI-RGD/~(125)I-(α_v)ASODN complex was prepared by combining ~(125)I-(α_v)ASODN with polyethyleneimine derivative PEI-RGD. PEI-RGD/~(125)I-(α_v)ASODN complex was transferred into HepG2 cells through the receptor-mediated process. The effect of PEI-RGD/~(125)I-(α_v)ASODN complex on the invasive ability of HepG2 cells was examined by Boyden chamber invasive assay. Results: (1) The labeling yield and radiochemical purity of ~(125)I-(α_v) ASODN were(73.78±4.09)% and(96.68±1.38)%, respectively, and the labeled compound had a good stability in vitro after 48 h at 37℃; (2) The ability of HepG2 cells to uptake PEI-RGD/~(125)I-(α_v)ASODN reached its peek ([12.77±0.85] % ) when PEI-RGD/~(125)I-(α_v)ASODN was at 4 μl/2 μg ([12.77±0.85] %), and then gredually decreased thereafter. So the dosage of PEI-RGD/~(125)I-(α_v)ASODN for the following experiment was chosen as 2 μl/1 μg; (3) The invasive capacity of HepG2 cells was significantly reduced in PEI-RGD/~(125)I-(α_v)ASODN group compared with those in other experiment and control groups (P <0.01 ). Conclusion: ~(125)I-(α_v)ASODN mediated by PEI-RGD can effectively inhibit the invasive capacity of HepG2 cells.
ABSTRACT
Objective To introduce the current research status, value and development future of Arg-Gly-Asp (RGD) peptides in diagnosis and treatment of neoplasms. Methods The current literatures on advances about RGD peptides in diagnosis and treatment of neoplasms were reviewed. Results RGD peptides, specificly recognizing and combining with integrin receptors, exist in extracellular matrix (ECM) of many kinds of organisms. After combining with integrin receptors, extrinsic RGD peptides can prevent tumor cells from adhering to ECM and migrating as the competitive inhibitor of intrinsic RGD peptides, suppress agiogenesis and induce tumor cells apoptosis, showing potential value of tumor specific imaging by targetal labelling neoplasms and treating tumors combining with other methods.Conclusion RGD peptides may be a new drug for diagnosis and treatment of neoplasms.
ABSTRACT
Objective To explore the role of extracellular signal-regulated kinase(ERK)in the apoptosis of hepatic stellate cells(HSC) induced by Arg-Gly-Asp-Ser (RGDS) tetrapeptide. Methods The cultrued HSC cells were divided into six groups including untreated control, fibronertin (FN), FN+RGDS (25 mg/L), FN+RGDS (50 mg/L), FN+RGDS(100 mg/L) and FN+Arg-Gly-Glu-Ser(RGES, 100 mg/L) groups. 3H-thymidine incorporation, transmission electron microscopy and TUNEL were employed to estimate the influence of RGDS on proliferation and apoptosis of HSC. The HSC adhesion rates were observed by toluidine blue colorimetric assay. The expressions of ERK mRNA and ERK protein in HSC were detected by RT-PCR and Western blotting. Results Compared with control and fibronectin groups, RGDS tetrapeptide at concentrations of 25 mg/L, 50 mg/L and 100 mg/L inhibited the proliferation of HSC(P