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Objective To investigate the feasibility of granzyme B (GB) promoter-controlled ferritin heavy chain (FTH1) reporter gene expression for monitoring the activation status of chimeric antigen receptor T cells (CAR-T) by magnetic resonance imaging (MRI). Methods Cytotoxic T lymphocytes (CTLs) were screened by Ficoll density gradient centrifugation and flow sorting. The GB promoter and FTH1 gene were ligated together with disialoganglioside 2 (GD2) CAR, and lentiviral vectors were transfected into CTLs to construct GD2-CAR-T/pGB-FTH1 cells. GD2-CAR-T/pCMV-FTH1, GD2-CAR-T, and T cells served as control cells. CytoTox96@non-radioactive cytotoxicity was used to detect the killing effect of each group of cells after co-culture with human neuroblastoma cells (SK-N-SH). ELISA was employed to detect the coincubation factor as well as the amount of GB secretion. Western blotting, Prussian blue staining and cellular MRI were applied to detect the expression of the FTH1 gene after co-culture. Results CTLs were successfully obtained, and then GD2-CAR-T/pGB-FTH1, GD2-CAR-T/pCMV-FTH1 and GD2-CAR-T cells were constructed. The killing effect, co-incubation factor and GB secretion of the above 3 groups of cells were significantly higher than those of the T cells, and the level of GB expression was highest at day 1, and then decreased in order at day 3 and day 7 after co-culturing with SK-N-SH cells. The relative expression of FTH1 and iron content of the GD2-CAR-T/pGB-FTH1 cells showed the same trend as GB expression, and the MRI signals were gradually increased. There were no significant differences in the relative expression of FTH1, iron content and MRI signals in the GD2-CAR-T/pCMV-FTH1 cells at all time points. No FTH1 expression or iron aggregation was observed in the GD2-CAR-T and T cells groups. Conclusion MRI based on the FTH1 reporter gene driven by the granzyme B promoter can reflect the GB expression level and tumor killing effect of CAR-T cells, which provides a potential real-time visual means to monitor the cell activation status for CAR-T therapy.

Lithium nitrate is an attractive lithium additive in the construction of high‐performance lithium metal anodes with a Li3N‐rich solid electrolyte interphase (SEI) layer. However, the eight‐electron transfer process induces high energy barriers between LiNO3 and Li3N. Herein, the inner Helmholtz plane is tuned on a Li deposition host to attain sluggish/rapid LiNO3 decomposition kinetics, resulting in different intermediate content distributions of Li species in the SEI. Notably, lithium oxynitride (LiNO) is identified as the decomposition intermediate, and experimental and simulation results confirm its role in obstructing LiNO3 decomposition. Moreover, the results reveal that the dipole–dipole interaction between LiNO and the polar V≡N bond can change the ionic/covalent character of the N═O bonds, considerably facilitating the energy transfer process of the N═O cleavage, and promoting a LiNO3 reduction to achieve a Li3N‐rich SEI. Consequently, when the electrolyte contains 0.37 m LiNO3, dendrite, and dead Li formation are suppressed effectively with the VN system, and an average Coulombic efficiency of 99.7% over 1000 cycles (1 mA cm−2, 1 mAh cm−2) can be attained. These results can promote the nitride oxidation break process and pave the way for fabricating high‐performance Li3N‐rich lithium metal batteries. [ABSTRACT FROM AUTHOR]

Acute graft-versus-host disease (aGVHD) remains a significant and severe complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT). Due to the occurrence of aGVHD, allo-HSCT significantly increases the mortality rate compared with autologous hematopoietic stem cell transplantation (auto-HSCT). In this study, auto-HSCT and allo-HSCT aGVHD mouse models were built to detect the difference in CD4+ lymphocyte in different tissues based on ribonucleic acid sequencing (RNA-Seq) analysis. Clustering analysis, functional annotation, and pathway enrichment analysis were performed on differentially expressed genes (DEGs). The protein-protein interaction (PPI) network was used to find hub genes. CD4+T cells were activated by MLR and cytokine stimulation. Cells were sorted out by a flow cell sorter. The selected genes were verified by qRT-PCR, histology, and immunofluorescence staining. The GSE126518 GEO dataset was used to verify the hub genes. Enrichment analysis revealed four immune-related pathways that play an important role in aGVHD, including immunoregulatory interactions between a lymphoid and a nonlymphoid cell, chemokine receptors binding chemokines, cytokine and cytokine receptor interaction, and the chemokine signaling pathway. At the same time, with the PPI network, 11 novel hub genes that were most likely to participate in immunoregulation in aGVHD were identified, which were further validated by qRT-PCR and the GSE126518 dataset. Besides, the protein expression level of Cxcl7 was consistent with the sequencing results. In summary, this study revealed that immunoregulation-related DEGs and pathways played a vital role in the onset of aGVHD. These findings may provide some new clues for probing the pathogenesis and treatment of aGVHD. [ABSTRACT FROM AUTHOR]

Magnetic resonance imaging (MRI) based on the ferritin heavy chain 1 (FTH1) reporter gene has been used to trace stem cells. However, whether FTH1 expression is affected by stem cell differentiation or whether cell differentiation is affected by reporter gene expression remains unclear. Here, we explore the relationship between FTH1 expression and neural differentiation in the differentiation of mesenchymal stem cells (MSCs) carrying FTH1 into neuron-like cells and investigate the feasibility of using FTH1 as an MRI reporter gene to detect neurally differentiated cells. By inducing cell differentiation with all-trans retinoic acid and a modified neuronal medium, MSCs and MSCs-FTH1 were successfully differentiated into neuron-like cells (Neurons and Neurons-FTH1), and the neural differentiation rates were (91.56±7.89)% and (92.23±7.64)%, respectively. Neuron-specific markers, including nestin, neuron-specific enolase, and microtubule-associated protein-2, were significantly expressed in Neurons-FTH1 and Neurons without noticeable differences. On the other hand, FTH1 was significantly expressed in MSCs-FTH1 and Neurons-FTH1 cells, and the expression levels were not significantly different. The R2 value was significantly increased in MSCs-FTH1 and Neurons-FTH1 cells, which was consistent with the findings of Prussian blue staining, transmission electron microscopy, and intracellular iron measurements. These results suggest that FTH1 gene expression did not affect MSC differentiation into neurons and was not affected by neural differentiation. Thus, MRI reporter gene imaging based on FTH1 can be used for the detection of neurally differentiated cells from MSCs. [ABSTRACT FROM AUTHOR]

Although mesenchymal stem cells (MSCs) have been demonstrated to possess a tumor-homing feature, their tropism to liver tumors has not been delineated in a visible manner. The aim of the present study was to evaluate the tumor-homing capacity of MSCs and to investigate the spatial and temporal distributions of MSCs in liver tumors using magnetic resonance imaging (MRI). MSCs were colabeled with superparamagnetic iron oxide (SPIO) particles and 4',6-diamidino-2-phenylindole (DAPI), and then transplanted into rabbits with VX2 liver tumors through intravenous injections. The rabbits were subjected to MRI before and at 3, 7 and 14 days after cell transplantation using a clinical 1.5-T MRI system. Immediately after the MRI examination, histological analyses were performed using fluorescence and Prussian blue staining. At 3 days after injection with labeled MSCs, heterogeneous hypointensity was detected on the MRI images of the tumor. At 7 days after transplantation, the tumor exhibited anisointense MRI signal, whereas a hypointense ring was detected at the border of the tumor. At 14 days after transplantation, the MRI signal recovered the hyperintensity. As demonstrated in the histological analyses, the distribution of the iron particles visualized with Prussian blue staining was consistent with the DAPI-stained bright fluorescent nuclei, and the particles corresponded to the hypointense region on the MR images. Thus, systemically administered MSCs could localize to liver tumors with high specificity and possessed a migration feature with active tumor growth. These results demonstrated that the targeting and distribution of the magnetically labeled stem cells in the tumor could be tracked for 7 days in vivo using a clinical 1.5-T MRI scanner. [ABSTRACT FROM AUTHOR]