[The supernatant of 4T1 breast cancer cell culture increases arginase 1 content in ANA-1 macrophages].

OBJECTIVE: To explore the effect of the supernatant of 4T1 murine breast cancer cell culture on arginase 1 (Arg-1) in ANA-1 macrophages in vitro by simulating the microenvironment of breast cancer. METHODS: The experimental ANA-1 macrophages were treated with the supernatant of 4T1 culture, and meanwhile, the control cells were cultured in the absence of the supernatant. Morphological changes of the ANA-1 macrophages were observed with a light microscope at 6, 8, 10, 24 hours, respectively. Real-time quantitative PCR (qRT-PCR) was performed to detect the levels of inducible nitric oxide synthase (iNOS) and Arg-1 mRNAs. Immunofluorescence and Western blotting were used to determine the levels of iNOS and Arg-1 proteins. RESULTS: The qRT-PCR indicated that the level of iNOS mRNA decreased in the experiment group compared with the control group, while Arg-1 mRNA level significantly increased compared with the control group and it reached a peak at the 8th hour. The immunofluorescence and Western blotting also demonstrated that Arg-1 protein expression was enhanced in the experimental group compared with the control group. However, iNOS protein expression was no significantly different between the experiment group and the control group. CONCLUSION: The supernatant of 4T1 cell culture increases Arg-1 production in ANA-1 macrophages.

[Autophagy and apoptosis of HeLa cells induced by recombinant human endostatin combined with hypoxia].

OBJECTIVE: To observe the impact of recombinant human endostatin (rhES) combined with hypoxia on the autophagy and apoptosis of human cervical cancer HeLa cells. METHODS: Under hypoxia, HeLa cells in logarithmic growth phase were treated with PBS, rhES, rhES combined with 3-methyladenine (3-MA, inhibitor of autophagy), respectively. Twenty-four hours later, cell apoptosis rate was detected using Hochest33528 staining and annexin V-FITC/PI staining and flow cytometry; the expressions of microtubule-associated protein light chain 3 (LC-3), Bcl-2, Bax proteins were determined using Western blotting. RESULTS: Hochest33528 staining suggested that cells of the hypoxia plus rhES group showed nuclear fragmentation; Annexin V-FITC/PI staining combined with flow cytometry revealed that the apoptosis rates were respectively (2.94±0.45)%, (21.38±0.92)% and (6.87±0.58)% in the three groups. It was significantly higher in the hypoxia plus rhES group than the other two groups. Western blotting showed that the expression levels of LC3 (LC-3II) and Bax increased and Bcl-2 decreased in the hypoxia plus rhES group as compared with the other two groups. CONCLUSION: Endostatin combined hypoxia can induce autophagy and apoptosis of HeLa cells.