Effect Of Hepatocarcinoma H22 Cell-derived Microparticles On The Growth Of Mice Transplanted Tumor

Background and ObjectiveIncreasing evidence indicates that tumor-derived microparticles(TMPs), as a medium of intercellular communication, in the microenvironment are involved in tumor progression. We isolated and extracted H22-derived microparticles(HMPs)by using differential centrifugation, established experimental animal model of liver cancer, and studied the influence of H22-derived microparticles on the growth of liver cancer. Further more, we used heparin and Dynasore to interpose the interaction between H22 cells and H22-derived microparticles, to further study the effect of H22-derived microparticles on the growth of liver cancer. Methods1. H22 cells were subcultured in the BALB/c mice abdominal cavity and centrifuged. H22 cells morphology was observed with hematoxylin and eosin(HE) staining.2. H22 cells were subcultured in the BALB/c mice abdominal cavity for three generations and induced with serum-free for 48 h. H22-derived microparticles were isolated by using differential centrifugation, observed by transmission electron microscopy(TEM). The number of HMPs(PS positive HMPs)in culture supernatants were measured by flow cytometry. Using Bradford method to quantify HMPs protein.3. H22 cells incubated with different doses of H22-derived microparticles were inoculated subcutaneously in BALB/c mice, and observing the effects of H22-derived microparticle on tumor growth.4. According to the results, choosing an effective dose of H22-derived microparticles, we treated them with heparin and Dynasore, further studied the effect of H22-derived microparticles on the growth of liver cancer. Results1. H22-derived microparticles, isolated by using differential centrifugation, were observed under the transmission electron microscopy, the result showed that their diameter was < 1μm, with different size, and also they were closed microvesicles. Using flow cytometry detected the count of HMPs and PS positive HMPs, which was indicated that the proportion of PS positive HMPs was 7.36% and 1×107 H22-derived microparticles were acduired totally, which was enough for all the experiment.2. Tumor volume in the high dose H22-derived microparticles group was larger than PBS control group, the difference was statistically significant(P<0.05), its tumor weight was also heavier compared with PBS control group and the low dose of H22-derived microparticles group, the difference was statistically significant(P<0.05). However, compared with the high dose group and low dose group respectively, the tumor volume and tumor weight in the middle dose group were no statistical difference(P> 0.05).3. For heparin intervention group, the results showed that tumor weight in the group of H22 cells incubated with H22-derived microparticles which treated with heparin(Heparin-HMPs-H22 group) was lighter than the untreated group(HMPs-H22 group), the difference was statistically significant(P<0.05). Similarly, the tumor volume was also smaller, which had a significantly statistical difference(P <0.01).4. For Dynasore intervention group, the results showed that tumor volume in the group of H22 cells incubated with H22-derived microparticles which treated with Dynasore(Dy-HMPs-H22 group) was smaller than the untreated group(HMPs-H22 group), the difference had statistical significance(P<0.05). Similarly, the tumor weight was also lighter, which had a statistical difference(P<0.05). Conclusions1. High dose of H22-derived microparticles may promote tumor growth in mice, thereby promoting the growth of tumor volume and weight.2. The role that H22-derived microparticles promote hepatoma cells growth can be inhibited by heparin and Dynasore.