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Rhodamines have been continuously optimized in brightness, biocompatibility, and color to fulfill the demands of modern bioimaging. However, the problem of phototoxicity caused by the excited fluorophore under long-term illumination has been largely neglected, hampering their use in time-lapse imaging. Here we introduce cyclooctatetraene (COT) conjugated rhodamines that span the visible spectrum and exhibit significantly reduced phototoxicity. We identified a general strategy for the generation of Gentle Rhodamines, which preserved their outstanding spectroscopic properties and cell permeability while showing an efficient reduction of singlet-oxygen formation and diminished cellular photodamage. Paradoxically, their photobleaching kinetics do not go hand in hand with reduced phototoxicity. By combining COT-conjugated spirocyclization motifs with targeting moieties, these Gentle Rhodamines compose a toolkit for time-lapse imaging of mitochondria, DNA, and actin, and synergize with covalent and exchangeable HaloTag labeling of cellular proteins with less photodamage than their commonly used precursors. Taken together, the Gentle Rhodamines generally offer alleviated phototoxicity and allow advanced video recording applications, including voltage imaging.

Inspired by the design principle of pearl structure, a bottom-up flake powder self-assembly arrangement strategy, flake powder metallurgy, is used to prepare graphene films (GFs) reinforced CoCrFeNiMn high-entropy alloy (HEA) matrix composites with a pearl laminated structure. Flaky HEA powder was prepared by ball milling method and homogeneously mixed with Ni plated GFs. Vacuum hot-press sintering (VHPS) technique was carried out to solidify the mixed powders to obtain composites with uniform distribution of GFs(Ni) and flaky HEA. The results show that the bottom-up preparation strategy can effectively fabricate bionic laminated HEA matrix composites, and the composites have a distinct pearly laminated structure. The tensile strength of the composites with 5 vol% GFs(Ni) content reached 834.04 MPa, and the elongation reached 26.58 %. The compressive strength in parallel and perpendicular laminar directions reached 2069.66 MPa and 2418.45 MPa at 50 % strain, respectively. The laminated GFs(Ni)/HEA matrix composites possessed excellent strength and maintained good plasticity. In this study, the strengthening and toughening mechanism of the laminated GFs(Ni)/HEA matrix composites is discussed in detail, and the results show that the laminated structure and GFs(Ni) are favorable for the hardening and strengthening of the HEA matrix.

The direct oncolytic effect of Newcastle disease virus (NDV) depends on the following two aspects: the susceptibility of cancer cells to virus infection and the ability of virus itself to lyse cancer cells. First, we investigate the susceptibility of cancer cells to NDV infection, HepG2, MDA-MB-231, and SH-SY5Y cells were susceptible, A549, MCF7, and LoVo cells were less susceptible. To investigate the molecular mechanism responsible for cancer cell susceptibility, transcriptome sequencing was carried out. We found that the levels of alpha-sialic acid acyltransferase were upregulated in MDA-MB-231 cells compared with MCF7 cells, and the interferon was downregulated. Second, to optimize the oncolytic capacity of the wild-type rClone30, a series of chimeric viruses rClone30-Anh(HN), rClone30-Anh(F), and rClone30-Anh(HN-F) were constructed by exchanging the HN gene, F gene or both of non-lytic rClone30 strain with lytic strain Anhinga. rClone30-Anh(F) and rClone30-Anh(HN-F) enhanced the oncolytic effect of the rClone30, and this enhancement is more obvious in the susceptible cells. The oncolytic mechanism of rClone30-Anh(F) was analyzed by transcriptome analyses, in comparison with rClone30, rClone30-Anh(F) upregulated the expression of ATG5, Beclin 1, and MAP1LC3B, thus activating autophagy and promoting the production of syncytia. In conclusion, our study provides a strategy to enhance the oncolytic effect of rClone30.

NDV as an attractive candidate for oncolytic immunotherapy selectively lyses tumor cells but shows limited anti-tumor immunity. Immune co-stimulator OX40 ligand (OX40L) boosts anti-tumor immunity response by delivering a potent costimulatory signal to CD4+and CD8+T cells. To improve the anti-tumor immunity of NDV, the recombinant NDV expressing the murine OX40L (rNDV-mOX40L) was engineered. The viral growth kinetics was examined in CT26 cell lines. The ability of rNDV-mOX40L to express mOX40L was detected in the infected tumor cells and tumor tissues. The anti-tumor activity of rNDV-mOX40L was studied in the CT26 animal model. Tumor-specific CD4+, CD8+and OX40+T cells were examined by immunohistochemistry staining. The virus growth curve showed that the insertion of the mOX40L gene did not affect the growth kinetics of NDV. rNDV-mOX40L expresses mOX40L and effectively inhibits the growth of CT26 colorectal cancer in vivo. The tumor inhibition rate of the rNDV-mOX40L-treated group was increased by 15.8% compared to that of  NDV-treated group in the CT26 model. Furthermore, immunohistochemistry staining of tumor tissues removed from the CT26 model revealed that intense infiltration of tumor-specific CD4+, CD8+T cells, especially OX40+T cells were found in the rNDV-mOX40L-treated group. FACS showed that rNDV-mOX40L significantly enhanced the number of CD4+and CD8+ T cells in spleen. Moreover, compared to the NDV-treated group, the level of mouse IFN-γ protein in the tumor site increased significantly in the rNDV-mOX40L-treated group. Taken together, rNDV-mOX40L exhibited superior anti-tumor immunity by stimulating tumor-specific T cells and may be a promising agent for cancer immunotherapy.

Neuronal nuclei (NeuN) protein is a key RNA-associated protein to guide the transcription of RNA and regulate mRNA splicing in neurons. However, the lack of effective labeling and tracking method has hindered the elucidation of the biological mechanism of NeuN operation in living neurons to understand correlated central nervous system disorders. Here, we report a molecular probe that can be inserted into a neighboring hydrophobic–hydrophilic region in NeuN, which upon binding becomes capable of emitting light in red region. The NeuN specificity enables the probe imaging neuronal cells in primary brain regions including hippocampus, cerebellum, midbrain, and cingulate gyrus. The probes’ optical properties are such to enable stimulated emission depletion imaging showing for the first time the 3D structure of RNA tangling into NeuN in a living neuron with tens of nanometer resolution.