The Effect Of Apelin-13 On The Growth Of MNNG/HOS Osteosarcoma And Glucose Metabolism Related Studies

Objective : Establishment and evaluation of MNNG/HOS human osteosarcoma cells that were transplanted into animal models of host BALB/c male nuke mice at age of 4 weeks,are helpful to understand the possible effect of the development of MNNG/HOS human osteosarcoma tumor on blood glucose of the host animal and related changes radiologically,and to demonstrate the role of the glucose metabolism related PI3K/AKT cellular signal transduction pathway played in the progress.Meanwhile,the Apelin-13 were measured under the conditions of cell cultures or co-cultures of MNNG/HOS cell line with hFOB human osteoblast cell line and MNNG/HOS cell line with osteoblast taken from the host BALB/c nuke mice,and the mutual effects of Apelin-13 and the tumor growth on the glucose metabolism associated the genes and proteins of the PI3K/AKT pathway.Method: Establish tumor growth animal model in vivo by transplanting MNNG/HOS cells into the host male nuke mice at age of 4 weeks subcutaneously.The 80 mice had been divided into eight groups,four of these groups are control and the others are experimental group,both of control and experimental groups are sorted by time points of 1 week,2 weeks,3 weeks,4 weeks for each.Then blood was harvested for blood glucose and serum Apelin-13 tests.MRI scan was used for measuring the volume.The weights of the host bodies and the tumors were measured at the same time.The MNNG/HOS osteosarcoma cells and human hFOB cells or the osteoblast from the host mice are co-cultured for testing the glucose metabolism related genes and proteins expressions on the pathway of PI3K/AKT during the development of the tumor.Results: The change of signal intensity in MRI showed that tumor volume is increasing in progress.Body weight of nude mice is heavier than control group at 1 week(p<0.05),no significant difference at 2 weeks(p>0.05),lighter than control at 3 weeks(p<0.05),significant lighter than control(p<0.01).MRI proved to be an indicator of effective tool for evaluating development of osteosarcoma in vitro.Change in tumor volume succeeded to show significant positive correlation with body weight of host and quality of tumor sample.And tumor quality to body weight ratio is also positive correlation with serum glucose of nude mice hosts and negative correlation with the blood glucose.Apelin concentrations were significantly higher in mice hosts implanted with osteosarcoma at 3 or 4 weeks than in the controls.The highest apelin levels were found in mice hosts at 4 weeks and in hosts with lowest blood glucose levels at the same time.Serum ALP and LDH significantly decreased in mice hosts with regional and distant metastasis at 3 or 4 weeks.Serum apelin was significantly higher in cachectic mice than in the controls.Apelin was negetively correlated with ALP,LDH and serum glucose level.Apelin levels increased significantly in tumor tissues.Weak positive correlation between ALP and LDH levels in serum were observed.Serum glucose is decreasing gradually.Morphologically,at the same time points quantities of osteoblasts in IGF-1 groups are between control and clindamycin while cellular shapes are better and proteins related to proliferation of osteoblasts in IGF-1 groups are expressed more than those of clindamycin groups.13nmol/mL of IGF-1 is indicated to be an effective concentration to maintain the survival of osteoblasts cultured in 500 ug/ml clindamycin by using MTT of cellular viability test,RT-PCR and Western blotting.The agonist is capable of inducing the increased expression of mRNA and related proteins in activation of PI3K/AKT signaling pathway.The mRNA expression of IGF-1、Pi3kca、Pi3kr1、Akt、Irs1、Bmp2、Rras2 and the regulating proteins showed the activation of PI3K/AKT pathway enhance the survival of osteoblasts in inhibited environment.Conclusions: MRI correctly predicts osteosarcoma development in an animal model of nude mice as an effective tool.And in this animal model,positive correlations among tumor volume,tumor quality to host body weight and serum glucose were proved to be significant and effective for evaluating the relationship between osteosarcoma development and energy metabolism.This study proves the superiority of contrast-enhanced study over volumetric study by MRI,advantages of using the negative correlation between the blood glucose and tumor weight,and the efficacy of BALB/c nude mice model for development of MNNG/HOS osteosarcoma over an energy metabolism study.Apelin is associated with cachexia and metastasis processes of osteosarcoma in BALB/c nude mice.Reduction of blood glucose and increasing of serum ALP and LDH reflects acceleration of adipose and glucose energy metabolism consumption and continuous bone and/or bone tumor remodeling in relation to osteosarcoma progression.Correlation of apelin with ALP,LDH and blood glucose can reflect a presumable role of apelin in systemic energy metabolism response in development of osteosarcoma.MNNG/HOS human osteosarcoma cells transplanted into a BALB/c nuke mouse can be a reliable and stable animal model.Blood from the host mice...