br the reactive oxygen species ROS contents in HeLa
the reactive oxygen species (ROS) contents in HeLa BYL-719 treated with diﬀerent samples were measured using 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) as an indicator. As shown in Fig. 6A, HeLa cells treated with Se and ION presented a typical green fluorescence be-longing to ROS, but those treated with MCDION-1 had a brighter fluorescence, which might be attributed to the synergistic eﬀect of Mn2+ ions and ION. In addition, the green fluorescence generated by MCDION-Se dramatically strengthened, possibly because the nano-Se coated onto MCDION-Se activated SOD and promoted the generation of SOARs in the cells, enhancing the ROS content in the cells. Subse-quently, the SOAR and SOD contents in the HeLa cells were measured using flow cytometry and Western blotting analysis, respectively. HeLa cells treated with Se displayed a strong fluorescence intensity belonging to SOARs (Fig. 6C), while the expression of SOD in cells incubated with MCDION-Se and nano-Se significantly increased in comparison with those incubated with MCDION-1 and the control group (Fig. 6D). These results confirmed that nano-Se coated on MCDION-1 could eﬀectively activate SOD and promote the generation of SOARs. Based on the above interesting results, the content of ROS in the HeLa cells was further quantitatively analyzed and was found to be in the order of Se < ION < MCDION-1 < MCDION-1+Se < MCDION-Se (Fig. 6B). This phenomenon occurred because an abundance of SOARs generated by MCDION-Se was catalyzed by active SOD and formed H2O2, and MCDION-Se could further decompose H2O2 into ·OH via a Fenton-like reaction, significantly increasing the content of ·OH in the cells and accelerating cell apoptosis. In addition, the ATP content in the cells dramatically decreased after incubation with MCDION-Se (Fig. 6E), implying that MCDION-Se could inhibit the production of energy and starve cancer cells.
The viability of HeLa and HK-2 cells treated with ION did not de-crease significantly, suggesting that the ION core possessed good bio-compatibility (Fig. 7A). Nevertheless, HeLa cells treated with MCDION-1 for 24 h showed a degree of apoptosis (Fig. 7B), which might be
because Mn2+ ions released from MCDION-1 limited the generation of ATP and accelerated the Fenton-like reaction, thus killing the cancer cells. In addition, MCDION-Se showed a strong cell inhibition ability in comparison with other groups, and this was because the synergistic
Fig. 4. (A) TEM image of MCDION-Se. (B) XPS full spectra and (C) Se 3p spectra of MCDION-1 and MCDION-Se. The color changes of the mixture of MCDION-Se, TMB, and H2O2 in (D) diﬀerent pH solutions and (E) diﬀerent concentrations of MCDION-Se at pH 5.5 solution. (F, G) The corresponding UV absorption spectra of D and E. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)
eﬀect of nano-Se and MCDION-1 increased the content of intracellular ·OH and broke the nutrition chain in the cells. HeLa cells treated with MCDION-Se for 48 h showed the same degree of apoptosis as those r> In the in vivo experiment, tumor-bearing BALB/C nude mice were treated with diﬀerent samples at a dose of 2 mg/kg via tail vein
Fig. 5. (A) T1 images of HeLa cells incubated with free Mn2+, MCDION-1, and MCDION-Se at diﬀerent concentrations of Mn2+ ions. (B) T1-weighted MR images of tumor-bearing mice acquired at pre-injection and post-injection of Mn2+, MCDION-1, and MCDION-Se. (C) The corresponding signal changes originating from the T1 images.
Fig. 6. (A) CLSM observation of intracellular ROS, all images shared the same scale bar. Flow cytometry analysis of (B) intracellular ROS and (C) SOAR in HeLa cells treated with diﬀerent samples. (D) Western blot analysis of HeLa cells treated with diﬀerent samples. (E) ATP content in HeLa cells treated with diﬀerent samples.