Dihydroethidium dhe

DHA was purchased from Aladdin Co. Ltd. (Shanghai, China). 1, 2-dioleoylsn-glycero-3-phosphoethanolamine (DOPE), cholesteryl hemisuccinate (CHEMS), BSO and FeSO₄•7H₂O were obtained from Sigma-Aldrich (St. Louis, MO, USA). Deferiprone (DEF) was purchased from Meyer Chemical Technology Co. Ltd (Shanghai, China). ROS Detection Kit, Glutathione Assay Kit, Annexin V-FITC/Propidium Iodide (PI) Cell Apoptosis Detection Kit, dihydroethidium (DHE), and Protein Extraction Kit were obtained from KeyGen Biotech. Co. Ltd. (Nanjing, China). BCA Protein Assay Kit was purchased from Beyo-time Institute of Biotechnology (Shanghai, China). The primary antibodies and secondary antibody against TfR and GAPDH were acquired from Affinity Biosciences (Changzhou, China). Fluorescein isothiocyanate (FITC), CellROX, LysoTracker Red, MitoTracker Red, Hoechst 33342, acridine orange (AO) and LIVE/DEAD™ Fixable Green Dead were obtained from Invitrogen (ThermoFisher Scientific, USA). Iron Colorimetric Assay Kit was purchased from BioVision (San Francisco, USA). 1, 2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-(polyethylene glycol) 2000-transferrin (DSPE-PEG₂₀₀₀-Tf) was synthesized by Xi-An Ruixi Biological Technology Co., Ltd. (Xi-An, China). Ultrapure water was prepared using a Millipore Simplicity System (Millipore, Bedford, USA).

PBMCs were isolated from the peripheral blood of patients and rat recipients by Ficoll density-gradient centrifugation according to the manufacturer’s instructions. Cell surface antigens (Ags) were characterized using a standard staining method and measured by flow cytometry. The surface markers of the cells were stained with mAbs against CD3, CD4, CD8. The proliferation of Jurkat T cells was measured by flow cytometric measurement of Ki-67 (BioLegend, USA). An apoptosis detection kit (Beyotime, China) was used for annexin V and PI staining. Dihydroethidium (DHE) (KeyGEN BioTECH, China) fluorescent probe for cytosolic reactive oxygen species (ROS) detection. Moreover, flow cytometry was used to assay for changes in LC3B and p-JNK levels in CD8⁺ T cell. Briefly, the collected cells were resuspended in PBS, then incubated with anti-LC3B and p-JNK primary antibody for 1 h, rinsed with PBS, incubated with fluorescent-conjugated secondary antibodies at room temperature for 30 min. The stained cells were detected by flow cytometry. The results were analyzed by flow cytometry.

CDDP, IR820, Hoechst 33342, methylthiazolyldiphenyl-tetrazolium bromide (MTT) and 1,3-diphenylisobenzofuran (DPBF) were purchased from Aladdin Company (Shanghai, China). PVPVA (trade name Kollidon VA64) was gifted by Basf Corporation (Shanghai, China). The polydimethylsiloxane (PDMS) MN molds were bought from Micropoint Technologies Pte Ltd (Singapore). The dihydroethidium (DHE) and the live/dead assay kit (Calcein AM/PI) were purchased from KeyGen Biotech (Nanjing, China). The total reactive oxygen species assay kit (DCFH-DA), the active caspase-3 assay kit (GreenNuc) and the TUNEL apoptosis assay kit were obtained from Beyotime Biotechnology (Shanghai, China). The anti-Ki67 and anti-cleaved caspase 3 primary antibodies were purchased from Servicebio Technology (Wuhan, China). Other regents were analytical grade.