| Part I Preparation and properties study of Fe3O4&IR-1061@PLGA self-assembled nanoprobesObjective To develop a self-assembled nanoprobe based on poly(lactic-co-glycol-ic acid)(PLGA)nanomicrospheres and evaluate its physicochemical properties,cytotoxicity and photothermal effects in vitro.Methods ? Single emulsion-solvent evaporation and magnetic separation purify-cation methods were used to prepare the Fe3O4&IR-1061@PLGA nanoprobes(FIP NPs),and whose stability and exosyndromes were investigated.? 4T1 murine breast cancer cells were incubated with different concentrations of FIP NPs for 24 h,whose cytotoxicity in vitro was evaluated by the methyl thiazolyl tetrazolium(MTT)assay.? The photothermal effect of FIP NPs at different concentrations in vitro was tested under the irradiation of 808 nm laser,and the MTT assay and Live-Dead assay were used to detect photothermal effect of FIP NPs to 4T1 murine breast cancer cells.Results ? The electronic particle and hydrodynamic size of FIP NPs was 98.4 nm and 182 nm,respectively,and whose hydrodynamic size and zeta potential in water and fetal bovine serum remained basically stable.The ultraviolet-visible-near infrared(UV-Vis-NIR)absorption spectra exhibited that FIP NPs possessed a strong NIR absorption band from 700 to 900 nm,with the maximum absorption intensity at 750 nm.Fluorescence emission spectroscopy under 808 nm laser excitation demonstrated an emission peak at 1095 nm.Which both showed blue-shifted absorption and emission spectrum by comparison with free IR-1061 molecules.? In vitro cytotoxicity assay showed no significant cytotoxic response of cells even at high nanoprobe concentrations up to 24 ?g/mL.? In vitro photothermal imaging evaluation showed that the FIP NPs presented excellent photothermal conversion effect and stability.At the same time,cell experiments showed that FIP NPs had significant photothermal killing and ablation effect on tumor cellsConclusion FIP NPs presented good colloidal stability,optical property and photothermal effect in vitro,and showed no obvious cytotoxicity.Part ? Fe3O4&IR-1061@PLGA self-assembled nanoprobes for multimodal imaging in lymph node metastasis of breast cancerObjective To investigate the feasibility of FIP NPs for magnetic resonance imaging(MRI)/optical imaging(OI)/photoacoustic imaging(PAI)/single-photon emission computed tomography(SPECT)in lymph node(LN)metastasis of breast cancer.Methods ? The LN metastasis model was established by injecting 4T1-Luc tumor cells into the foot pad of the mouse.? The FIP NPs at different concentrations were evaluated by in vitro performance of MRI/OI/PAI,meanwhile,FIP NPs were radiolabeled with radionuclide 99mTc and its radiolabel stability was then investigated.? In vivo MRI/OI/PAI/SPECT of LN metastasis was performed before injection of FIP NPs and at different time points within 8 hours after intratumoral injection of FIP NPs into the 4T1-Luc tumor,and then the changes of signal values in LN metastasis were observed and measured.Results? One week after foot pad injection of 4T1-Luc tumor cells,swollen LNs were palpated in the ipsilateral popliteal fossa of mice.Moreover,bioluminescence signals were observed both in the primary tumors of foot pad and swollen LNs of the ipsilateral popliteal fossa after intraperitoneal injection of D-fluorescein sodium salt.Furthermore,the LN metastasis was confirmed by in vitro anatomy and pathology.? The FIP NPs presented good contrast-enhanced effects of 01,PAI and MRI,and high transverse relaxivity of 172.73 mM-1 s-1.Not only that,FIP NPs could also easily achieve the radionuclide 99mTc labeling,whose radiochemical purity maintained above 95.0%within 24 hours.? In vivo imaging results demonstrated that the metastatic LN region presented darker effect in T2-weighted imaging and remarkably enhanced signals in NIR-?,PAI and SPECT after intratumorally injected with FIP NPs.In addition,the highest concentration and strongest signal of metastatic LN was observed 2 to 3 hours after intratumoral injection.Conclusion FIP NPs have the good multimodal imaging effect of MRI/OI/PAI/SPECT,and the precise identification and trace of LN metastasis of breast cancer in vivo were realized.Part ? Fe3O4&IR-1061@PLGA self-assembled nanoprobes for photothermal therapy in lymph node metastasis of breast cancerObjective To investigate the feasibility of FIP NPs for photothermal therapy(PTT)in LN metastasis of breast cancer.Methods ? Two 4T1-Luc tumor and LN metastasis bearing mice were intratum-oral injected with the same volume of saline and FIP NPs respectively via the primary tumor on the foot pad.2 hours later,the LNs metastasis in popliteal fossa were irradiated under 808 nm laser with the same power(1.25 W/cm2)for 10 minutes,and the temperature changes of LNs metastasis were real-time recorded by infrared thermal imager and then compared between the two mice.? Twelve 4T1-Luc tumor and LN metastasis bearing mice were randomly divided into three groups(n=4 per group):injection of FIP NPs without laser irradiation,injection of saline with laser irradiation,injection of FIP NPs with laser irradiation,and then the primary tumors of each group were surgical removal.The growth and bioluminescent signal changes of LNs metastasis,and whether there was distant metastasis were monitored.Results ? Study on the PTT warming effect of FIP NPs on LN metastases of breast cancer in vivo showed that the injection of FIP NPs mouse had a more significant warming effect than the injection of saline mouse.The temperature of the injection of FIP NPs mouse was increased up to 54? and increased by 19.3? while that of the injection of saline mouse was only increased by 8.7?.? It was found that 2 hours after intratumoral injection of FIP NPs combined with 808 nm laser irradiation and surgical removal of primary tumors could ablate the LN metastasis without recurrence and metastasis within 45 days,whereas,pulmonary metastasis and no trend of growth inhibition occurred in the other two groups.Conclusion FIP NPs have good PTT effect and are expected to be used in the PTT of LN metastasis of breast cancer patients in the future. |
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