Effects Of Gold Nano-Clusters On Osteoclasts And Breast Cancer Bone Metastases

Osteoporosis is a common degenerative disease of bone structure,which is characterized by decreased bone mass and degradation of bone microstructure,resulting in bone fragility and increased risk of fracture.When osteoclasts’ bone absorption rate exceeds osteoblasts’ bone formation rate for various reasons,it will lead to the imbalance of bone loss and bone growth,bone loss and degradation of bone structure and strength,and eventually lead to osteoporosis.The disease is most common in postmenopausal women and the elderly.With the development of medical technology,people’s life expectancy is extended,the aging society is speeding up,and the prevalence of osteoporosis is rising rapidly,which has become a serious public health problem in the world.Therefore,more and more attention has been paid to the prevention and treatment of osteoporosis.At present,there is no effective pathological treatment for this disease.The incidence rate of breast cancer is the highest in the world amone females.Because the mammary vein flows back into the paravertebral vein,about 65% of breast cancer will have spinal metastasis,forming osteolytic destruction.In the late stage,it is easy to cause fracture and nerve compression.Most patients show obvious pain and paralysis symptoms,and the quality of life is seriously damaged.The survival time of the patients was short,and the 5-year survival rate was less than 20%.These two diseases are accompanied by over activation and differentiation of osteoclasts.Therefore,if a drug possesses ability to inhibit the activation and differentiation of osteoclasts,as well as the migration,proliferation and chemotaxis of tumor cells,it may have the potential to become a new type of anti osteoporosis and anti-tumor bone metastasis drug.Au nano-clusters are nano materials formed by the combination of gold elements,proteins and peptides.They have many biological characteristics.It has been proved that suitable Au nano-clusters can inhibit the physiological and pathological activities of monocyte macrophages.We speculated that osteoclasts,as mononuclear macrophages in bone microenvironment,could also be inhibited by appropriate gold nanoclusters.So,it may have the possibility of stabilizing bone microenvironment and alleviating osteoporosis.At the same time,if the Au nano-clusters can inhibit the biological activities of tumor cells at the same time,it will be a potential anti-tumor drug for bone metastasis as well.So,we designed and carried out the experiment according to our above conjecture.In this project,we synthesized two stable gold nanoclusters,bovine serum albumin Au nano-clusters(Au BSA)and glutathione Au nano-clusters(Au GSH),and characterized their physical and chemical properties by fluorescence detection,transmission electron microscopy,dynamic light scattering detection and mass spectrometry.Furthermore,we verified the effect of Au BSA and Au GSH on the differentiation of primary mouse bone marrow macrophages(BMMs)into osteoclasts.After evaluating the cytotoxicity,BMMs of mice was stimulated to differentiate into osteoclasts by RANKL,and the process was inhibited by adding safe concentration Au nano-clusters.The inhibition effect was evaluated by TRAP staining,F-Actin staining and bone slice test.Finally,the activities of oxidoreductase,nuclear factor kappa-B(NF-κB)signaling pathway related proteins and osteoclast activation related genes in BMMs were detected by enzyme-linked immunosorbent assay(ELISA),Western blot and q RT-PCR.At the cellular level,the effects of two Au nano-clusters on the proliferation and osteoclastic differentiation of BMMs were evaluated.We cultured human breast cancer cell line MDA-MB-231 and normal human breast epithelial cell line MCF-10 A,tested the cytotoxicity of the two materials and compared the gold element endocytosis;verified the influence of the material on the migration ability of breast cancer cells through wound healing experiment and cell migration experiment;evaluated the influence of the material on the proliferation ability of breast cancer cells through the cell clone formation experiment;and established the effect of the two Au nano-clusters on the regulation of osteoclast recruitment and differentiation by conditional culture and ELISA.The expression of NF-κB signal pathway related proteins in osteoclasts was also detected by western blot.Finally,the mouse model of allograft tumor bearing in tibial medullary cavity was designed,and the damage of tumor bone was observed and evaluated.Our results showed that Au BSA and Au GSH could inhibit the differentiation of BMMs into osteoclasts and the osteolytic function of osteoclasts.At the same time,our cell experiments and animal experiments verified that Au BSA and Au GSH can inhibit the proliferation and migration of breast cancer cell MDA-MB-231,and the ability of tumor cells stimulate and chemotactic osteoclast to cause osteolytic damage.Through our research,we have demonstrated that Au BSA and Au GSH,two kinds of Au nano-clusters,can inhibit the activation and differentiation of osteoclasts,and inhibit the migration,proliferation and chemotaxis of breast cancer cells without damaging the normal cells.It has the potential to become a new anti osteoporosis drug,anti-tumor bone metastasis and secondary bone destruction drug.