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Objective:To dynamically trace the migration and therapeutic effects of human bone marrow mesenchymal stem cells (MSCs) in mice with liver injury after cell transplantation through in vivo bioluminescent imaging (BLI).Methods:The MSCs were transfected with the lentivirus CMV-Luciferase2-mKate2 and mKate2 positive cells were purified and screened by fluorescence-activated cell sorting (FACS) after 96 h. The purified MSCs-R (MSCs-CMV-Luciferase2-mKate2) were used by in vitro and in vivo BLI. The mice (male BALB/c nude mice) were divided into 4 groups with 9 mice per group by random number table method, including (1) Liver injury experimental group: The liver injury model was established by intraperitoneal injection of CCl 4, and MSCS-R transplantation through spleen injection was performed 24 h later; (2) Control experimental group: The same volume of phosphate buffer (PBS) was injected intraperitoneally, and MSCS-R transplantation through spleen injection was performed 24 h later; (3) Liver injury group: Liver injury model was established and PBS was injected into the spleen;(4) Blank group: The mice were intraperitoneally injected of PBS.BLI was performed daily after cell transplantation until light signals disappeared in the liver region, and the pathological examination of liver tissue was obtained 14 d after MSCs-R transplantation. Linear regression analyses were performed to determine the correlation between the optical signal intensity and the number of cells, and statistical differences of the optical signal intensity between liver injury experimental group and control experimental group were evaluated using the Student′s t test. Results:The MSCs were readily transfected with lentivirus CMV-Luciferase2-mKate2 for 96 h. The transfected MSCs were purified by FACS and more than 95% of MSCs were mKate2 positive. The optical signal intensity of MSCs-R detected by BLI in vitro significantly correlated with cell numbers in vitro (R 2=0.980). In both of liver injury experimental group and control experimental group, cell migration to the liver was observed on the first day after intrasplenic transplantation of MSCs-R, and the optical signal intensity in the area of liver of liver injury experimental group was higher than that of control experimental group ( t=15.476, P<0.001). The optical signal intensity in the hepatic area was observed in 11 d after transplantation in liver injury experimental group, compared to control experimental group in 5 d. Optical signal was not detected in mice in the other two groups. Histopathology showed that the degree of liver injury after MSCs-R transplantation was significantly lower in liver injury experimental group than control experimental group. Conclusions:The dynamical migration of MSCs transplanted to the spleen and settled in the damaged liver could be tracked by BLI, and liver injury can prompt MSCs directionally migrate to the damaged tissues and play their role in repairing liver injury.
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Objective To obtain a thyroid cancer cell line integrated with luciferase and fluorescence,and to establish a metastatic nude mouse model of papillary thyroid cancer monitored with in vivo imaging system. Methods Lentivirus carrying recombinant plasmid containing luciferase gene and mCherry gene infected high malignant papillary thyroid cancer cell line (BHP10-3SCmice). The stable thyroid cancer cell line (BHP10-3mluc) labeled with luciferase and mCherry selected by hygromycin, analyzed for fluorescence with fluorescence microscopy and bioluminescence with in vitro analysis and in vivo imaging. 0.25 ml(5×106/ml)BHP10-3mluc cell suspension were injected into the left thigh intramuscular tissue of 5-week-old nude mice to establish BALB/C nude mice metastatic tumor model. The growth and metastasis of the tumors were monitored weekly with in vivo imaging system. The nude mice were sacrificed on the 42th day and the tissues with metastatic tumors were analyzed by fluorescent imaging ex vivo. The ectopic implanted tumors and metastatic lesions were verified by tissue sections with Hematoxylin and Eosin staining. Results BHP10-3mluc cells showed similar growth characteristics to the original BHP10-3SCmice cells and stably expressed luciferase and mCherry. At the end of the first week after ectopic implantation,the xenograft tumors were found and in the 6th week,the tumors were found to metastasized to adjacent lymph nodes and lungs,which was consistent with the results of pathology.The growth and metastasis of tumors can be accurately monitored with in vivo imaging system. Conclusions A PTC cell line stably expressing bioluciferase and fluorescence was successfully established.The nude mouse model of PTC metastatic tumor generated by intramuscular inoculation of those cells can be well monitored with in vivo imaging system, which provides ideal models for researches on tumor growth, metastasis,and drug treatment.
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Objective To observe the migration and inhibition mechanism of MicroRNA218-Robo1 pathway for breast cancer.Methods A total of 40 BALB/c-nu/nu female mice were randomly divided into four groups.Each group was transfected over-expression MicroRNA218 MDA-MB-231 breast cancer cells, co-over-expression MicroRNA218 and Robo1 MDA-MB-231 breast cancer cells, knock-down Robo1 MDA-MB-231 breast cancer cells and the control MDA-MB-231 breast cancer cells.The tumor volume was examined every two weeks.Results Tumor volume of MicroRNA218 group was obviously less than control group, tumor volume of Robo1 knock out group was obviously less than common MicroRNA218 high expression and Robo1 group, the difference was statistically significant;MicroRNA218 and Robol knockout group than the control group, the increase in breast cancer cells apoptosis, cell proliferation and angiogenesis is restrained.Conclusions MicroRNA218 inhibited the migration of breast cancer by down-regulating the expression of Robo1.
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MicroRNAs (miRNAs) are a novel class of small noncoding RNAs that regulate gene expression by targeting mRNAs for either cleavage or translational repression.Accumulated studies show that miRNAs aberrantly expressed in tumors are closely related with tumorogenesis and could be potential tumor biomarkers.It will be helpful in the diagnosis,prognosis and(or) predicting treatment response of tumor by imaging miRNAs expression.Several molecular imaging methods,including the optical imaging,radionuclide imaging and multimodality imaging,have been successfully used to visualize miRNAs,miRNAs,a new molecular imaging target,may explore a new field for tumor diagnosis and therapy.
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Molecular imaging is leading an important role in the era of molecular medicine. Optical imaging, a rising star in the filed of molecular imaging, largely consists of bioluminescent imaging and fluorescent imaging. It has been shown that well-aimed and creatively-built optical-imaging-reporters let researchers explore and answer a lot of biologically important questions in living subjects. Despite relatively short history, molecular optical imaging is rapidly being implemented not only in many clinical areas but also in various research fields from tracking gene expression, protein-protein interaction or migrating cells to molecular diagnosis and treatment.
Subject(s)
Diagnosis , Gene Expression , Linear Energy Transfer , Molecular Imaging , Molecular Medicine , Neurosciences , Optical ImagingABSTRACT
Molecular imaging is leading an important role in the era of molecular medicine. Optical imaging, a rising star in the filed of molecular imaging, largely consists of fluorescent imaging and bioluminescent imaging. In the fluorescence imaging, an illuminating light excites fluorescent reporters in the living subject, and a charged coupled device (CCD) camera collects an emission light of shifted wavelength. In the bioluminescent imaging, reporter genes code for the luciferase that is responsible for fireflies' glow. After the injection of the substrate iuciferin, animals carrying the luciferase gene are imaged with a supersensitive CCD camera to pick up the small number of photons transmitted through tissues. It has been shown that well aimed and creatively built reporters let researchers explore and answer a lot of biologically important questions in living subjects. Despite its relatively short history, optical imaging is rapidly being implemented in various clinical areas as well as research fields.