Elastase-targeting biomimic nanoplatform for neurovascular remodeling by inhibiting NETosis mediated AlM2 inflammasome activation in ischemic stroke.

Neutrophil elastase (NE) is a protease released by activated neutrophils in the brain parenchyma after cerebral ischemia, which plays a pivotal role in the regulation of neutrophil extracellular traps (NETs) formation. The excess NETs could lead to blood-brain barrier (BBB) breakdown, overwhelming neuroinflammation, and neuronal injury. While the potential of targeting neutrophils and inhibiting NE activity to mitigate ischemic stroke (IS) pathology has been recognized, effective strategies that inhibit NETs formation remain under-explored. Herein, a biomimic multifunctional nanoplatform (HM@ST/TeTeLipos) was developed for active NE targeting and IS treatment. The core of the HM@ST/TeTeLipos consisted of sivelestat-loaded ditelluride-containing liposomes with ROS-responsive and NE-inhibiting properties. The outer shell was composed of platelet-neutrophil hybrid membrane vesicles (HMVs), which acted to hijack neutrophils and neutralize proinflammatory cytokines. Our studies revealed that HM@ST/TeTeLipos could effectively inhibit NE activity, thereby suppressing the release of NETs, impeding the activation of the AIM2 inflammasome, and consequently redirecting the immune response away from a pro-inflammatory M1 microglia phenotype. This resulted in enhanced neurovascular remodeling, reduced BBB disruption, and diminished neuroinflammation, ultimately promoting neuron survival. We believe that this innovative approach holds significant potential for improving the treatment of IS and various NE-mediated inflammatory diseases.

A biomimic anti-neuroinflammatory nanoplatform for active neutrophil extracellular traps targeting and spinal cord injury therapy.

Traumatic spinal cord injury (SCI) always leads to severe neurological deficits and permanent damage. Neuroinflammation is a vital process of SCI and have become a promising target for SCI treatment. However, the neuroinflammation-targeted therapy would hinder the functional recovery of spinal cord and lead to the treatment failure. Herein, a biomimic anti-neuroinflammatory nanoplatform (DHCNPs) was developed for active neutrophil extracellular traps (NETs) targeting and SCI treatment. The curcumin-loaded liposome with the anti-inflammatory property acted as the core of the DHCNPs. Platelet membrane and neutrophil membrane were fused to form the biomimic hybrid membrane of the DHCNPs for hijacking neutrophils and neutralizing the elevated neutrophil-related proinflammatory cytokines, respectively. DNAse I modification on the hybrid membrane could achieve NETs degradation, blood spinal cord barrier, and neuron repair. Further studies proved that the DHCNPs could reprogram the multifaceted neuroinflammation and reverse the SCI process via nuclear factor kappa-B (NF-κB) pathway. We believe that the current study provides a new perspective for neuroinflammation inhibition and may shed new light on the treatment of SCI.

Angiopep-2 conjugated biomimetic nano-delivery system loaded with resveratrol for the treatment of methamphetamine addiction.

Methamphetamine (METH) addiction can damage the central nervous system, resulting in cognitive impairment and memory deficits. Low target effects have limited the utility of anti-addiction drugs because the presence of the blood-brain barrier hinders the effective delivery of drugs to the brain. Angiopep-2 can recognize and target low-density lipoprotein receptor-associated protein 1 (LRP-1) on the surface of cerebral capillary endothelial cells, causing cross-cell phagocytosis, and thus has high blood-brain barrier transport capacity. Resveratrol (RSV) has been found to be a neuroprotective agent in many nervous system diseases. In our study, we modified Angiopep-2 on the surface of the erythrocyte membrane to obtain a modified erythrocyte membrane (Ang-RBCm) and coated RSV-loaded poly(ε-caprolactone)-poly(ethylene glycol) (PCL-PEG) nanoparticles with Ang-RBCm (Ang-RBCm@RSVNPs) to treat METH addiction. Our results showed that Ang-RBCm@RSVNPs can penetrate the blood-brain barrier and accumulate in the brain better than free RSV. Besides, mice treatetd with Ang-RBCm@RSVNPs showed less preference to METH-paired chamber and no noticeable tissue toxicity or abnormality was found in H&E staining images. Electrophysiological experiments demonstrated Ang-RBCm@RSVNPs could elevate synaptic plasticity impaired by METH. These indicated that Ang-RBCm@RSVNPs has better anti-addiction and neuroprotective effects. Therefore, Ang-RBCm@RSVNPs has great potential in the treatment of METH addiction.

ROS responsive hydrogel for inhibition of MUC5AC against allergic rhinitis: A new delivery strategy for Ipratropium Bromide.

Allergic rhinitis (AR) is a chronic inflammatory disease of the nasal mucosa mediated by immunoglobulin E (IgE) after exposure to allergens. The bothersome symptoms of AR, such as runny nose and nasal congestion, affect millions of people worldwide. Ipratropium Bromide (IB), commonly used in clinical practice for treating AR, requires frequent administration through nasal spray and may cause significant irritation to the nasal mucosa. The induction of ROS is closely related to the initiation and symptoms of AR, and ROS will continue to accumulate during the onset of AR. To address these challenges, we have designed a drug delivery system that can be administered in liquid form and rapidly crosslink into a ROS-responsive gel in the nasal cavity. This system enables sustained ROS responsive release of IB in a high-concentration ROS environment at AR lesions, thereby alleviating AR symptoms. The gel demonstrated prolonged release of IB for up to 24 hours in rats. In the treatment of AR rat models, it improved their symptoms, reduced the expression of various inflammatory factors, suppressed MUC5AC protein expression, and decreased mucus secretion through a ROS responsive IB release pattern. Overall, this system holds promise as a better option for AR treatment and may inspire the design of nanogel-based nasal drug delivery systems.

Cancer cell membrane-camouflaged biomimetic nanoparticles for enhancing chemo-radiation therapy efficacy in glioma.

Glioblastoma multiforme (GBM) presents significant challenges in treatment, with current standard-of-care approaches offering limited efficacy and survival benefits. This necessitates the development of innovative therapeutic strategies to enhance treatment outcomes. Nanotechnology has emerged as a promising avenue in cancer therapy, offering targeted drug delivery and enhanced therapeutic efficacy. Polymeric nanoparticles, particularly those based on Poly (lactic-co-glycolic acid) (PLGA), have gained traction as drug carriers due to their biocompatibility and controlled release properties. However, their interception by macrophages poses challenges to effective drug delivery. Superparamagnetic iron oxide (SPIO) nanoparticles have shown promise as radiosensitizers, enhancing the efficacy of radiotherapy through the generation of reactive oxygen species (ROS). Moreover, cell membrane biomimetic drug delivery systems have garnered attention for their ability to improve biocompatibility and targeting capabilities. Leveraging these concepts, our study introduces a novel multifunctional platform, GM@P (T/S), comprising polymeric nanoparticles coated with cancer cell membrane. By encapsulating temozolomide (TMZ) and SPIO nanoparticles within GM@P (T/S), we aim to synergistically enhance the cytotoxic effects of chemotherapy and radiotherapy against GBM while overcoming limitations associated with conventional treatments. This innovative approach holds promise for addressing the unmet clinical needs in GBM therapy and advancing towards more effective and personalized treatment strategies.

Dendritic Polylysine with Paclitaxel and Triptolide Codelivery for Enhanced Cancer Ferroptosis through the Accumulation of ROS.

Recently, paclitaxel (PTX) was reported to increase intracellular lipid reactive oxygen species (ROS) levels, triggering cancer cell ferroptosis. Based on this, some efforts had been made to improve PTX treatment for non-small-cell lung cancer (NSCLC). Our previous studies demonstrated that triptolide (TPL) could improve the antitumor effect of PTX. Nevertheless, the poor solubility and side effects often limit the application of chemotherapy drugs. In this paper, we constructed a novel nanodrug delivery system (NDDS) chemosynthesis by PEGylated generation 3 (G3) dendritic polylysine coloaded with PTX and TPL (PTX-TPL-PEG-PLL, PTPP), which was endowed with the ability of tumor targeting and favorable solubility. In addition, we demonstrated that TPL could induce ROS generation by regulating the NF-κB signaling pathway to enhance the ferroptosis-induced effect of PTX. Besides, ferroptosis induced by PTPP could improve chemoresistance through inhibiting the level of P-gp, GPX4, and SLC7A11. Furthermore, we determined that ferroptosis may strengthen the immune response by increasing the expression of CD8+ T cells and IFN-γ+ cells while decreasing Treg cells. In general, PTPP may be a potential system for NSCLC treatment.

Orthogonally Sequential Activation of Self-Powered DNAzymes Cascade for Reliable Monitoring of mRNA in Living Cells.

Ratiometric imaging of tumor-related mRNA is significant, yet spatiotemporally resolved regulation on the ratiometric signals to avoid non-specific activation in the living cells remains challenging. Herein, orthogonally sequential activation of concatenated DNAzyme circuits is, first, developed for Spatio Temporally regulated Amplified and Ratiometric (STAR) imaging of TK1 mRNA inside living cells with enhanced reliability and accuracy. By virtue of the synthesized CuO/MnO2 nanosheets, orthogonally regulated self-powered DNAzyme circuits are operated precisely in living cells, sequentially activating two-layered DNAzyme cleavage reactions to achieve the two ratiometric signal readouts successively for reliable monitoring of low-abundance mRNA in living cells. It is found that the ratiometric signals can only be derived from mRNA over-expressed tumor cells, also irrespective of probes' delivery concentration. The presented approach could provide new insight into orthogonally regulated ratiometric systems for reliable imaging of specific biomarkers in living cells, benefiting disease precision diagnostics.

Enhanced Antitumor Efficacy of Novel Biomimetic Platelet Membrane-Coated Tetrandrine Nanoparticles in Nonsmall Cell Lung Cancer.

Nonsmall cell lung cancer (NSCLC) remains one of the leading causes of cancer-related death worldwide, posing a serious threat to global health. Tetrandrine (Tet) is a small molecule in traditional Chinese medicine with proven primary efficacy against multiple cancers. Although previous studies have demonstrated the potential anticancer effects of Tet on NSCLC, its poor water solubility has limited its further clinical application. Herein, a novel nanoparticle-based drug delivery system, platelet membrane (PLTM)-coated Tet-loaded polycaprolactone-b-poly(ethylene glycol)-b-polycaprolactone nanoparticles (PTeNPs), is proposed to increase the potency of Tet against NSCLC. First, tetrandrine nanoparticles (TeNPs) are created using an emulsion solvent evaporation method, and biomimetic nanoparticles (PTeNPs) are prepared by coating the nanoparticles with PLTMs. When coated with PLTMs, PTeNPs are considerably less phagocytized by macrophages than Tet and TeNPs. In addition, compared with Tet and TeNPs, PTeNPs can significantly inhibit the growth and invasion of NSCLC both in vitro and in vivo. With reliable biosafety, this drug delivery system provides a new method of sustained release and efficient anticancer effects against NSCLC, facilitating the incorporation of Tet in modern nanotechnology.

Acidity-responsive nanoplatforms aggravate tumor hypoxia via multiple pathways for amplified chemotherapy.

Since the hypoxia tumor microenvironment (TME) will not only limit the treatment effect but also cause tumor recurrence and metastasis, intratumoral aggravated hypoxia level induced by vascular embolization is one of the major challenges in tumor therapy. The chemotherapeutic effect of hypoxia-activated prodrugs (HAPs) could be enhanced by the intensified hypoxia, the combination of tumor embolization and HAP-based chemotherapy exhibits a promising strategy for cancer therapy. Herein, an acidity-responsive nanoplatform (TACC NP) with multiple pathways to benefit the hypoxia-activated chemotherapy is constructed by loading the photosensitizer Chlorin e6 (Ce6), thrombin (Thr), and AQ4N within the calcium phosphate nanocarrier via a simple one-pot method. In the acidic TME, TACC NPs could be degraded to release Thr and Ce6, resulting in the destruction of tumor vessels and consumption of intratumoral oxygen under laser irradiation. Therefore, the intratumoral hypoxia level could be significantly aggravated, further leading to the enhanced chemotherapeutic effect of AQ4N. With the guidance of in vivo fluorescence imaging, the TACC NPs exhibited excellent tumor embolization/photodynamic/prodrug synergistic therapeutic effects with good biosafety.

Potent nanoreactor-mediated ferroptosis-based strategy for the reversal of cancer chemoresistance to Sorafenib.

The drug resistance of cancer cells is related to a variety of mechanisms, among which the destruction of redox homeostasis is one of the key factors. Ferroptosis, an intracellular iron-dependent form of cell death, is related to the production of oxidative stress. The accumulation of lipid peroxidation (LPO) during ferroptosis disrupts intracellular redox homeostasis, thereby affecting the sensitivity of tumor cells to drugs. In this work, we proposed a ferroptosis strategy based on LPO accumulation, reduced glutathione generation via inhibition of SLC3A2 protein and inactivated glutathione peroxidase 4 (GPX4) to reverse the chemoresistance of cancer cells. The Fenton reaction based on the ferroptosis-inducing nanoreactors (Au/Fe-GA/Sorafenib@PEG) not only generated hydroxyl radicals (·OH) under laser irradiation to realize the accumulation of LPO, but also depleted GSH to increase the accumulation of LPO. Meanwhile, the cystine uptake of cells was inhibited by Sorafenib, resulting in reduced GSH synthesis and inactivated GPX4. In vitro and in vivo experiments demonstrated AFG/SFB@PEG + Laser group could inactivate GPX4 and the enhanced ferroptosis can reverse chemo-resistance caused by continuous upregulation of GPX4 levels in cells through 'self-rescue'. The study proposed the mechanism and feasibility of ferroptosis to reverse drug resistance, providing a promising strategy for chemo-resistant cancer treatment. STATEMENT OF SIGNIFICANCE: Herein, we proposed a ferroptosis strategy based on LPO accumulation, reduced glutathione generation via inhibition of SLC3A2 protein, and inactivated glutathione peroxidase 4 (GPX4) to reverse chemoresistance of cancer cells. The Fenton reaction based on the ferroptosis-inducing nanoreactors (Au/Fe-GA/Sorafenib@PEG) not only generated hydroxyl radicals (·OH) under laser irradiation to realize the accumulation of LPO but also depleted GSH to increase the accumulation of LPO. Meanwhile, the cystine uptake of cells was inhibited by Sorafenib, resulting in reduced GSH synthesis and inactivated GPX4. In vitro and in vivo experiments demonstrated AFG/SFB@PEG + Laser group could inactivate GPX4 and the enhanced ferroptosis can reverse chemo-resistance caused by continuous upregulation of GPX4 levels in cells through 'self-rescue'.

Ferroptosis-induced anticancer effect of resveratrol with a biomimetic nano-delivery system in colorectal cancer treatment.

Ferroptosis is a novel form of programmed cell death impelled by iron-dependent lipid peroxidation, which may be a potential strategy for cancer therapy. Here we demonstrated for the first time that Resveratrol (RSV), a traditional Chinese medicine (TCM) chemical monomer, could effectually inhibit the growth of colon cancer cells through the ROS-dependent ferroptosis pathway. Mechanistically, RSV evoked the increase of reactive oxygen species and lipid peroxidation in colorectal cancer cells, and eventually lead to ferroptosis. Furthermore, RSV could promote ferroptosis by downregulating the expression of the channel protein solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase 4 (GPX4). To improve the delivery efficiency of RSV, a biomimetic nanocarrier was developed by coating RSV-loaded poly(ε-caprolactone)-poly(ethylene glycol) (PCL-PEG) nanoparticles with erythrocyte membrane (RSV-NPs@RBCm). The RSV-NPs@RBCm provide the possibility to escape macrophage phagocytosis and have a long circulation effect. In addition, when coupled with a tumor-penetrating peptide iRGD, which could trigger enhanced tissue penetration tumor-specifically, the delivery of RSV-NPs@RBCm into tumors would be significantly improved results from the in vivo study demonstrated an excellent treatment efficacy for CRC. Altogether, our study highlighted the therapeutic potential of RSV as a ferroptosis-inducing anticancer agent and when loaded into a biomimetic nanoplatform, it might pave the way for the application of RSV loaded nanosystems for colorectal cancer treatment.

Biomimetic Red Blood Cell Membrane-Mediated Nanodrugs Loading Ursolic Acid for Targeting NSCLC Therapy.

As one of the most common cancers worldwide, non-small-cell lung cancer (NSCLC) treatment always fails owing to the tumor microenvironment and resistance. UA, a traditional Chinese medicine, was reported to have antitumor potential in tumor models in vitro and in vivo, but showed impressive results in its potential application for poor water solubility. In this study, a novel biomimetic drug-delivery system based on UA-loaded nanoparticles (UaNPs) with a red blood cell membrane (RBCM) coating was developed. The RBCM-coated UANPs (UMNPs) exhibited improved water solubility, high stability, good biosafety, and efficient tumor accumulation. Importantly, the excellent antitumor efficiency of the UMNPs was confirmed both in vitro and in vivo in cancer models. In addition, we further investigated the antitumor mechanism of UMNPs. The results of Western blotting showed that UMNPs exerted an anticancer effect by inducing the apoptosis and autophagy of NSCLC cells, which makes it superior to free UA. In addition, body weight monitoring, hematoxylin and eosin (HE) analysis, and immunohistochemical (IHC) analysis showed no significant difference between UMNPs and the control group, indicating the safety of UMNPs. Altogether, the preparation of biomimetic UMNPs provides a promising strategy to improve outcomes in NSCLC.

Enhanced antitumor effect of icariin nanoparticles coated with iRGD functionalized erythrocyte membrane.

Lung cancer is the most common cause of incidence and mortality among tumor diseases. Icariin (ICA), a potential Chinese medicine monomer, has been reported to show outstanding antitumor effects. However, the hydrophobic nature and less tumor penetration limit its potential as a topical healing agent. There are few studies report the efficacy of ICA on lung cancer, moreover, there is no biomimetic targeted delivery system in the application of ICA. Herein, we firstly develop a novel ICA bionic targeted nano-preparation, camouflaged by the tumor penetrating peptide iRGD (cRGDKGPDC), functionalized red blood cell membrane (RBCM), has the increased solubility, utilized biocompatibility, and aggravated tumor penetration of ICA. In this study, we constructed the iRGD functionalized RBCM mimetic targeted ICA-loaded nanoparticles (iRINPs) and explored the anti-tumor effect of iRINPs against lung cancer with biochemical and behavioral analysis. The results suggested that iRINPs showed improved biocompatibility and stability, and reduced phagocytic uptakes by macrophages. Besides, the modification of iRGD significantly improved the targeting ability of iRINPs. In vitro and in vivo the treatment effects and safety assays showed that iRINPs attained better therapeutic effects than ICA by inhibiting A549 cell migration, proliferation and invasion, as well as reducing side effects of ICA. Overall, we expected that the new bionic nanocarriers would be a promising nano-platform for ICA in the precise therapy of lung cancer.

Targeted regulation of tumor microenvironment through the inhibition of MDSCs by curcumin loaded self-assembled nano-filaments.

One of the most important reasons underlying the resistance of tumors is the immune suppression induced by cancer cells. Myeloid-derived suppressor cells (MDSCs), which exerts pivotal functions in immunosuppression, is a key participator in tumor microenvironment and a novel target for cancer therapy. Here curcumin (Cur) was employed as a specific MDSCs repressor to inhibit the number and function of MDSCs. Moreover, a novel self-assembled nano-filament system was generated through the conjugation of Cur and a self-assembled peptide. In vivo study demonstrated the powerful antitumor effect of curcumin-loaded nano-filaments (Nano-Cur) with delayed tumor growth and longer survival. The immune status of tumor microenvironment (TME) was well improved by Nano-Cur treatment with increased T cell proliferation and activation as well as enhanced production of inflammatory mediators such as GM-CSF and IL-6, which revealed that Nano-Cur contributed to relieve the tumor burden by regulating and improving the TME. Furthermore, flow cytometry analysis implied the lower MDSCs levels under Nano-Cur treatment, which indicated that the anticancer effect of Nano-Cur may be associated with the inhibition of recruitment and accumulation of MDSCs in the TME. Therefore, Nano-Cur may be a novel therapeutic approach for lung cancer, and extensive studies of mechanisms are required to better understand how TME affects tumor progression and provide new insights into anticancer therapeutics.

Aggregation-Induced Emission Photosensitizer Synergizes Photodynamic Therapy and the Inhibition of the NF-κB Signaling Pathway to Overcome Hypoxia in Breast Cancer.

Triple-negative breast cancer (TNBC) is one of the most aggressive subtypes of breast cancer, and TNBC patients often develop resistance to endocrine or molecular targeted therapy. Thus, a search for effective treatments is urgently required. Photodynamic therapy (PDT) has been verified to be a successful therapy for cancer. However, this treatment is oxygen-consuming, thus considerably limiting the PDT outcomes. The present study introduced a multistage drug delivery system to alleviate hypoxia and enhance PDT efficiency. Specifically, aggregation-induced emission luminogen (AIEgen) TPE-Py was first introduced to achieve PDT properties, and natural naphthohydroquinone dimer Rubioncolin C (RC), a blocker of mitochondria-associated oxidative phosphorylation (OXPHOS) and an NF-κB inhibitor, was applied to suppress the O2 consumption of OXPHOS and mitigate hypoxia thereafter. Enhanced PDT efficiency was validated by in vitro and in vivo TNBC models. In terms of the mechanism, AIEgen-based PDT synergized with RC could induce a fatal burst of reactive oxygen species (ROS) and ROS-mediated apoptosis. Moreover, this combination promoted the effectiveness of PDT by inhibiting the NF-κB signaling pathway. All of these results demonstrated that the administration system not only achieved a synergistic anti-TNBC effect but also expanded the clinical application of AIEgen-based PDT by overcoming hypoxia and inhibiting the NF-κB signaling pathway.

Azulene-Containing Squaraines for Photoacoustic Imaging and Photothermal Therapy.

Photoacoustic imaging (PAI) guided photothermal therapy (PTT) can realize real-time diagnosis and in situ treatment of cancer at the same time. Absorption in the near-infrared (NIR) region with large molar extinction coefficient (ε) and high value of photothermal conversion efficiency (PCE) are key prerequisites for photothermal agents (PTAs) to realize dual PAI and PTT treatments. Squaraines have stable quinoid structures with strong planarity and rigidity, in favor of the NIR absorption and high ε values. On the other hand, azulene derivatives mostly have very faint fluorescence emission, which is beneficial for photothermal transformation. Herein, two azulene-containing squaraines Az-SQ-1 and Az-SQ-2 are synthesized as high-performance PTAs. In comparison with Az-SQ-1, Az-SQ-2 possesses larger εmax of 3 × 105 M-1 cm-1 at 780 nm in organic solution and higher PCE of 53.2% in the form of nanoparticles under 808 nm laser irradiation. Accordingly, Az-SQ-2 NPs present stronger photoacoustic signals (about 15.1-times the background signal) and more efficient suppression of tumor growth. Our research indicates that the introduction of azulene unit to traditional NIR dyes is a simple but effective approach to obtain outstanding PTAs in the aspect of phototheranostics.

Superior in vitro anticancer effect of biomimetic paclitaxel and triptolide co-delivery system in gastric cancer.

As a well-known anticancer drug, paclitaxel (PTX), a first-line chemotherapeutic agent, remains unsatisfactory for gastric cancer therapy. It is reported that triptolide (TPL) could enhance the anti-gastric cancer effect of PTX. Considering the poor solubility of both drugs, we developed a red blood cell membrane-biomimetic nanosystem, an emerging tool in drug delivery, to co-load paclitaxel and triptolide (red blood cell membrane coated PTX and TPL co-loaded poly(lactic-co-glycolic acid) [PLGA] nanoparticles, RP(P/T)). The successful preparation was confirmed in terms of particle size, morphology, and surface markers assays. This biomimetic system could prolong circulation and escape immune surveillance. And these properties were verified by stability, in vitro drug release, and cellular uptake assays. Moreover, the MTT and colony formation assays demonstrated the superior anti-proliferation effect of the RP(P/T) to free drugs. The enhanced antitumor effects of RP(P/T) on migration and invasion were also evaluated by wound-healing and transwell assays. Overall, the bionic co-delivery nanoplatform with improved efficacy in vitro is a promising therapy for gastric cancer.

Using Fluorescence On/Off to Trace Tandem Nanofiber Assembly/Disassembly in Living Cells.

To track an intact biological process inside cells, continuous showing of the assembly/disassembly process is needed and fluorescence is advantageous in characterizing these processes. However, using fluorescence "on/off" to observe a sequential assembly/disassembly process in living cells has not been reported. Herein, we rationally designed a probe PEA-NBD-Yp and employed its fluorescence "on/off" to trace tandem assembly/disassembly of nanofibers in living HeLa cells. In vitro experiments validated that PEA-NBD-Yp could be efficiently dephosphorylated by ALP to yield PEA-NBD-Y, which self-assembled into nanofibers with the NBD fluorescence "on". Also, the PEA-NBD-Y nanofiber was disassembled by GSH, accompanied by fluorescence "off". Living cell imaging (together with ALP-inhibition or GSH-blocking) experiments sequentially showed the self-assembling nanofibers on the cell outer membrane with fluorescence "on" (On1), translocated inside cells (On2), and disassembled by GSH with fluorescence "off" (Off2). We anticipate that our strategy of one probe conferring temporal "on/off" fluorescence signals might provide people with a new tool to deeply understand a biological event in living cells in the near future.