RESUMO
Objective To explore molecular fluorescence imaging features of the growth and metastasis of DsRed-marked mouse bladder carcinoma. Methods The study used lipofectamine 2000 transfection method,transferred on the BTT739 cells with plasmid chickenβ-actin-DsRed-Neo vector.The stably expressing BTT739-DsRed monoelonal cells were got with G418 selection.It randomly divided the 615 mouse of 24 into three groups,injected cell suspension on the hindlimb,the first and second group with BTT739-DsRed cell and the third group with BTT739 cell to found xenograft roodel.MAESTRO imager recorded fluorescence images of the growth and metastasis of the tumors in vivo and the fluorescence intensity was measured.The excitation wavelength was 560-580 nm,emission wavelength was 590-610 nm and exposure time was 5000 ms.After continuous observation of 4 weeks,every week killed the mouse of the second group and cut into image,made records of the red fluorescent mouse bladder cancer xenograft model,measured the tumor size and fluorescence sighal values; analyzed the relations between the tumor size and fluorescence signal values as well as between the whole image and cut image. Results DsRed tumor could be observed at the first week. Central local fluorescence loss could be detected at the second week, pathologically confirmed necrotic tumor tissue and a little connective tissue. At the fourth week, a local lymph node metastasis could be observed with no distant metastasis. The measured values of fluorescent signal were as follows: (89±18), (122±55), (133±69), (715±343)counts; the tumor size were as follows: (13±4), (45±22), (83±29), (253±67)mm2. The whole body image of tumor size were as follows: (12± 3),(50±23), (90±29), (290±74)mm2. The cut image of tumor size were as follows: (12±5), (72±30), (141±43), (524±237)mm2. The tumor size and fluorescent signal values reflect positive linear correlation with 0. 74 coefficient (t= 3. 97, P<0.05), whole body imaging and cut image reflect positive linear correlation with 0. 97 coefficient (t=10, P<0.05). The whole body image of tumor size was (70. 85±17.13) % of cut image. Conclusions Red fluorescent mouse bladder cancer xenograft model could observe the growth and metastasis of the tumor intuitively, continuously, and sensitively.As the tumor increased, the fluorescence range also increased, the fluorescence disappeared after tumor necrosis, the expression of the red fluorescent transferred after the metastasis of the tumor.
RESUMO
Objective To investigate the effects of different dosages of bone marrow mesenchymal stromal cells (BMSC) on lung fibrosis. Methods BMSCs with red fluorescence protein (RFP) from male FVB mice were cultured in vitro. Twenty-four female wild type FVB mice were randomly divided into four groups: normal group, model group, BMSC 1 group and BMSC 2 group (n = 6). Mouse pulmonary fibrosis models were induced by bleomycin via single intratracheal perfusion. Twenty-four h after model establishment, mice in BMSC 1 group and BMSC 2 group were injected with 1 × 10~6 BMSCs and 2 × 10~6 BMSCs, respectively through vena caudalis for each mouse. All the animals were sacrificed 21 d after model estalishment, and mouse lung tissue samples were obtained. The pathological changes were observed by light microscopy, the hydroxyproline ( Hyp) contents were measured by alkaline hydrolysis assay, the distribution of RFP( + ) BMSCs and quantitation of RFP were analysed by laser scanning confocal microscopy and immunohistochemistry, the expression of surfactant protein A (SP-A) was detected by immunohistochemistry, and the expression of transforming growth factor-β (TGF-β) and platelet-derived growth factor (PDGF) mRNA was detected by Real-time PCR. Results Compared with model group, the pulmonary fibrosis in BMSC 1 group was significantly alleviated, and that of BMSC 2 group became much more severe.A large number of RFP( +) BMSCs were found in fibrosis area of BMSC 2 group,which exhibited morphology similar to fibroblasts. As far as the expression of SP-A was concerned, normal group was higher than BMSC 1 group, BMSC 1 group was higher than BMSC 2 group and model group (P < 0. 05), while there was no significant difference between BMSC 2 group and model group (P >0. 05). Normal group, BMSC I group, model group and BMSC 2 group fell in the increase order by Hyp contents (P <0.01, P <0.05), and BMSC 2 group, BMSC 1 group, model group and normal group fell in the decrease order by expression of TCF-|$ and PDGF mRNA (P < 0.05). Conclusion Proper dose of BMSC has a favourable effect on bleomycin-induced lung fibrosis, while excessive dose of BMSC can aggravate the fibrosis.