Study Of Dual PH/Redox Sensitive Marine Laminarin-based Nanomedicine Carrier

Marine polysaccharides,which are used to resist tumor and virus,have a wide applocation prospect in the medical field.In addition,marine polysaccharides can significantly improve the body's immune function and enhance the effect of tumor targeting.Therefore,the modification of marine polysaccharides with specific functions has become an important part in current research.The modified marine polysaccharides can increase the hydrophilicity of drugs,improve the tumor targeting and reduce the toxicity on the body,which are widely used in drug delivery.It is of great significance to the resaerch of new drug formulations.According to the specificity of tumor microenvironment,functional groups such as ketal and disulfide bonds were modified on the surface of polymer polysaccharides to synthesize multifunctional nanocarrier materials with pH/Redox stimuli-triggering,named as Hematin-laminatin-S-S-MGK(HLDM).The nanocarrier was used to deliver photosensitiveness drugs to tumor site,so that the drug could accumulate in tumor site to induce antitumor effect.To put it simply,this study had proposed that photosensitizer(Pp IX)was encapsulated in dual pH/Redox stimuli-triggered marine polysaccharides-based nanocarrier(HLDM)to form micelles,and the properties in vitro and in vivo were also investigated.The experiments were as follows:1.Preparation and characterization of HLDM carriersFirstly,the synthesis and characterization of HLDM materials were completed.The laminarin was chosen as the polymer material,and the dithiodipropionic acid was used as an linker to make the connection with the Menthone 1,2-glycerol ketal(MGK).In order to enhance the hydrophobicity of the material,the hematin was linked to the structure of laminarin.Finally,the Hematin-laminatin-S-S-MGK(HLDM)nanocarriers were synthesized.The structure was characterized by nuclear magnetic resonance(1H-NMR)and confirmed by infrared spectroscopy(IR)and ultraviolet-visible spectroscopy(UV-Vis).Then the properties of the carrier material were investigated,which laid the foundation for the further study2.Preparation and characterization of Pp IX/HLDM micellesProtoporphyrin IX(Pp IX)as model drugs was encapsulated in the HLDM materials to obtain the Pp IX/Hematin-laminatin-S-S-MGK micelles,named as Pp IX/HLDM micelles.The properties of drug-loaded micelles,including the particle size,P.I,zeta potential,morphology,drug loading capacity(DL)and encapsulation efficiency(EE),were investigated.The results demonstrated that the particle sizes of Pp IX/HLDM micelles were 149.±35 nm,the zeta potential was 7.24±4.9 mV,the DL was 5.89±3.87%,the EE was 40.06±4%,and the P.I was 0.167±0.06.The results of the drug release study of Pp IX/HL DM micelles in vitro indicated that the cumulative release of HLDM/Pp IX micelle at low pH and high glutathione was 66.98%,which had the dual pH/Redox stimuli-triggered Pp IX on-demand release to achieve the desired purpose in the tumor microenvironment.3.The cell assay of Pp IX/HLDM micellersIn cell-level studies,the absorption of breast cancer,liver cancer,colon cancer and lung cancer cells was evaluated.The MCF-7 cells were selected according to the factors of proliferation,uptake and source.Cell uptake assay showed that HLDM materials had properties to response the tumor microenvironment to increased drug uptake,and the uptake was time-and concentration-dependent.After Pp IX was stimulated at a specific wavelength,the cells showed obvious green fluorescence in the reactive oxygen species(ROS)assay kit,indicating that photosensitizer could kill the tumor by producing ROS at a specific wavelength after uptake by the cells.In the cytotoxicity study,MTT method was used to evaluate the phototoxicities of Pp IX/HLDM micelles,the dark toxicity of Pp IX/HLDM micelles and the HLDM materials.The results showed that there was no significant difference in the dark toxicity of Pp IX/HLDM micelles with different concentrations.After given a certain wavelength,the phototoxicity enhanced with the increase of micelle concentration and intensified the inhibitory effect on tumor.In addition,there was no statistical difference in the cytotoxicity of HLDM carrier material,indicating that the toxic side effect of the material was small.The results of apoptosis experiment showed that Pp IX/HLDM micelles had a significant antitumor effect and could cause nuclear damage.4.The animal experiments of HLDM/Pp IX micellesIn animal level,the nude mice tumor models with MCF-7 cells were constructed to research the distribution of carrier materials in vivo.The results indicated that the HLDM materials could reach the tumor site and realize the effective release of the encapsulated drug in the tumor site.In pharmacodynamics experiment,MCF-7 cells were still used to construct tumor model of nude mice.The tumor-bearing nude mice were irradiated with specific wavelength after administration,and the growth inhibition of axillary tumors and pathological sections of tumor tissues in nude mice were observed.At the same time,the weight changes of tumor-bearing nude mice were recorded during the experiment cycle The results of tumor growth showed that the antitumor effect of HLDM micelles significantly enhanced under the condition of specific wavelength irradiation.The pathological sections of tumor tissue also proved that Pp IX/HLDM micelles could significantly increase the degree of tumor necrosis and decrease the number of tumor cells under the condition of specific wavelength.The results of body weight change of nude mice showed that Pp IX-loaded HLDM micelles had no significant effect on the body weight of nude mice.