Study On The Effect Of Doxorubicin-Loaded Gelatin Microspheres In The Treatment Of Bone Tumors

Objective:Doxorubicin is an important drugs for chemotherapy of osteosarcoma.However,it is an important challenge to realize the balance between effective concentration and side effects after systemic administration.It is necessary to improve the drug delivery method to improve the effective rate of drug delivery to specific sites and increase the drug concentration of doxorubicin at tumor sites,so as to achieve the purpose of target therapy.In this study,gelatin microspheres were used as drug delivery carrier of doxorubicin to explore the efficacy of doxorubicin in the treatment of bone tumors under the action of gelatin microsphere carrier.Methods:(1)Preparation and properties of doxorubicin gelatin microspheres:1.Doxorubicin bearing gelatin microspheres were prepared,glutaraldehyde was used as curing agent,and crosslinked and stored at low temperature;2.Scanning electron microscope was used to observe whether the shape and size of the prepared microspheres were normal;3.The loading rate and encapsulation rate were calculated by ultraviolet spectrophotometry;4.The release curve of doxorubicin gelatin microspheres in vitro was calculated.(2)In vitro and in vivo studies of doxorubicin gelatin microspheres in the treatment of osteosarcoma:1.The cell line used in this study was highly invasive osteosarcoma MNNG/HOS CL#5 cells.In vitro cytotoxicity test was performed by CCK8 method,and the semi-inhibitory concentration of the drug was calculated.2.A total of 40 nude mice(BALB/c nude mice)were used for subcutaneous tumor formation experiment.When the tumor in the mice grew to more than 80 mm~3,they were randomly divided into 4 groups with 10 mice in each group.Mice were injected with normal saline(control group),doxorubicin alone(DOX group),single gelatin microspheres(GM group)and doxorubicin-containing gelatin microspheres(GM+DOX group),respectively,and the changes of tumor tissue in each group were observed and compared.3.Hematoxylin-eosin staining was used to observe the morphological changes of tissues.4.The expressions of Ki67 and PCNA were observed by immunohistochemical staining.5.WB staining was used to observe the expression of related apoptotic proteins.6.m RNA expression levels of apoptotic protein-related genes were detected by RT-PCR.7.Statistical software SPSS 22.0 was used to collate and analyze the experimental data.The mean±standard deviation was expressed as the metrological data,and the Bonferroni multiple test method was used to compare the data in pairs.The difference between samples was considered statistically significant only when P<0.05 was measured.Results:(1)Preparation and properties of doxorubicin gelatin microspheres:1.The optimum material ratio for preparing doxorubicin gelatin microspheres DOX:GM was 1:10;2.The results of transmission electron microscopy showed that the gelatin microspheres had regular spherical shape,the size was about 40μm,the surface of the microsphere was smooth,the shape was good,no collapse,fold;3.Drug loading rate was(6.05±0.26)%,encapsulation rate was(84.27±3.10)%;4.Doxamycin gelatin microspheres accumulate release in vitro,continuously increasing release within 0-60 hours,with a gentle release curve and a continuous slow rise.(2)In vitro and in vivo studies of doxorubicin gelatin microspheres in the treatment of osteosarcoma:1.In vitro cytotoxicity tests,semi-inhibitory concentration of DOX nude drugs was higher than GM+DOX for 72 hours;2.Both DOX and GM+DOX groups had certain inhibitory effects on tumor in nude mice,and the tumor weight in GM+DOX group was reduced by 2.25 times compared with DOX group.And the tumor volume was significantly reduced over time.3.The positive rate of tissue cells in DOX and GM+DOX groups was significantly reduced,among which,the positive rate of Ki67 and PCNA in GM+DOX group was reduced by about 4 times.4.After the treatment of GM+DOX,the expression levels of AKT,ERK and key proteins in the m TOR pathway were significantly inhibited.5.The expression level of Bcl-2 in GM+DOX group was significantly lower than that of other apoptotic factors,and the expression levels of BAX,BID,p53 and Caspase family apoptosis proteins were up-regulated,and their corresponding m RNA expression levels were consistent,so as to promote the apoptosis of tumor cells.Conclusion:Doxorubicin gelatin microspheres can effectively inhibit the growth and invasion ability of tumor cells through sustained release drugs,and the expression levels of key proteins in tumor proliferation and anti-apoptosis were significantly inhibited,indicating that doxorubicin gelatin microspheres can effectively improve the efficiency of drug action.Compared with drug administration alone,it can significantly inhibit cell proliferation to prevent the progression of highly aggressive osteosarcoma.