The Study Of Melanin-manganese Nanoparticles For Tumor Imaging As A Novel Magnetic Resonance Contrast Agent

Objective:Melanin,a natural natural biopolymer,has marvellous biocompatibility and biodegradability.In this study,we designed melanin-manganese?MNP-Mn?nanoparticles as a novel magnetic resonance contrast agent.Combined with nanotechnology and magnetic resonance imaging?MRI?,which provides noninvasive anatomical and physiological information.It provides a new method to produce a novel MRI contrast agent for tumor diagnosis.Method:1.We synthesized MNP-PEG-Mn nanoparticles by one-step method.And the characteristics of MNP-PEG-Mn nanoparticles were measured by DLS,TEM,ICP-MS and so on.2.MNP-PEG-Mn was indicating the strong chelating stability by dialysis method in PBS solution?pH=7.4?and acidic buffer solution?pH=5.5?.3.In vitro MRI experiments of MNP-PEG-Mn and Gadodiamide were performed at 25°C in a 3.0T MR clinical scanner,and the r1 values were calculated from T1-weighted MR images.4.The cellular uptake behaviors and subsequent localization of MNP-PEG-Mn have been estimated by CLSM and bio-TEM after Hep-2 cells were incubated MNP-PEG-Mn.Hep-2 cells were employed to evaluate the cytotoxicity of MNP-PEG-Mn with various concentrations over 24 h using a standard MTT assay.5.To establish a laryngeal cancer model,then injected MNP-PEG-Mn into the mice,and the axial and coronal T1-weighted MR images were acquired pre-injection and at appropriate time intervals?0 h,0.5 h,24 h,48 h72 h and 120 h?after injection.The images and the data were collected.6.To study the biodistribution and the histopathological analysis of the mice which is injected MNP-PEG-Mn through tail intravenous,When the mice were sacrificed,then collected the tumor and the main apparatus at 24 h,5 days,and 30 days.Results:1.MNP-PEG-Mn was well dispersed with uniform size and shape,and the mean hydrodynamic diameter was about 5.6 nm.And the surface potential of the MNP-PEG-Mn was about-13.5±0.6 mV.The number of Mn²⁺chelated to the MNP-PEG was calculated to be about 55 per MNP.2.To check the leakage of Mn²⁺from MNP-PEG-Mn in both neutral and acidic solution.The results indicating the strong chelating stability of MNP-PEG-Mn,and only about 1%of the Mn²⁺were released from MNP-PEG-Mn in the first 2 h.3.Compared with MNP-PEG-Mn and Gadodiamide in a 3.0T MR clinical scanner,and the r1 value of MNP-PEG-Mn is 20.57 mM^-1s^-1,which was about 3.4 times higher than commercial Gadodiamide(6.00 mM^-1s^-1),revealed that MNP-PEG-Mn presented significantly brighter images than Gadodiamide at the same metalion concentration.4.MNP-PEG-Mn can be clearly seen in the cytoplasm after incubation for 24 h,as shown by the red fluorescence from rhodamine lighting up the nuclei,so as the thin sections in TEM.MNP-PEG-Mn could be located within endosomes or lysosomes. MTT assay also confirmed that MNP-PEG-Mn was safety even at Mn²⁺ concentrations up to 1600?g mL^-1.5.The axial T1-weighted MR images were acquired pre-injection and at different time intervals after injection,it can be seen that obvious enhanced signal and prominent positive contrast effects were achieved in the tumor region at 0.5 h after the injection,the MR signals of the tumor tissue became slightly weaker at the 24 h time interval,but could still be cleared by the kidney.Finally,the signal showed a sharp decrease at 48 h.6.The Mn²⁺levels were measured in various organs using ICP-MS.The results revealed that the nanoparticles were mainly cleared by the kidney and liver.The accumulation in various organs after 1 day is obviously higher than that after 5 days and 30 days.The level of the serum creatinine?SCr?and urea nitrogen?BUN? remained were still kept in normal.H&E staining indicated that MNP-PEG-Mn has no appreciable adverse effect on these main apparatus.Conclusion:A natural biopolymer system based on Mn²⁺-chelating melanin nanoparticles were developed as a T1 MRI contrast agent for tumor-targeted imaging.MNP-PEG-Mn had a much higher relaxivity than that of commercial Gadodiamide.Furthermore,the results of in vivo MRI displayed good tumor-targeting specificity,and the nanoparticles could be very efficiently eliminated by both the renal and hepatobiliary pathways,avoiding long-term accumulation.Therefore MNP-PEG-Mn showed great translation potential as an MRI contrast agent for tumor diagnosis.