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Despite improvements in medical and surgical therapies, a significant portion of patients with critical limb ischemia (CLI) are considered as "no option" for revascularization. In this work, a nitric oxide (NO)-boosted and activated nanovesicle regeneration kit (n-BANK) is constructed by decorating stem cell-derived nanoscale extracellular vesicles with NO nanocages. Our results demonstrate that n-BANKs could store NO in endothelial cells for subsequent release upon pericyte recruitment for CLI revascularization. Notably, n-BANKs enable endothelial cells to trigger eNOS activation and form tube-like structures. Subsequently, eNOS-derived NO robustly recruits pericytes to invest nascent endothelial cell tubes, giving rise to mature blood vessels. Consequently, n-BANKs confer complete revascularization in female mice following CLI, and thereby achieve limb preservation and restore the motor function. In light of n-BANK evoking pericyte-endothelial interactions to create functional vascular networks, it features promising therapeutic potential in revascularization to reduce CLI-related amputations, which potentially impact regeneration medicine.
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Células Endoteliales , Pericitos , Humanos , Femenino , Ratones , Animales , Células Endoteliales/fisiología , Óxido Nítrico , Isquemia/terapia , Células Madre , Neovascularización Fisiológica/fisiologíaRESUMEN
Hepatic macrophages represent a key cellular component of the liver and are essential for the progression of acute liver failure (ALF). We construct artificial apoptotic cells loaded with itaconic acid (AI-Cells), wherein the compositions of the synthetic plasma membrane and surface topology are rationally engineered. AI-Cells are predominantly localized to the liver and further transport to hepatic macrophages. Intravenous administration of AI-Cells modulates macrophage inflammation to protect the liver from acetaminophen-induced ALF. Mechanistically, AI-Cells act on caspase-1 to suppress NLRP3 inflammasome-mediated cleavage of pro-IL-1ß into its active form in macrophages. Notably, AI-Cells specifically induce anti-inflammatory memory-like hepatic macrophages in ALF mice, which prevent constitutive overproduction of IL-1ß when liver reinjury occurs. In light of AI-Cells' precise delivery and training of memory-like hepatic macrophages, they offer promising therapeutic potential in reversing ALF by finely controlling inflammatory responses and orchestrating liver homeostasis, which potentially affect the treatment of various types of liver failure.
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Células Artificiales , Fallo Hepático Agudo , Lesiones de Repetición , Animales , Ratones , Lesiones de Repetición/metabolismo , Macrófagos/metabolismo , Fallo Hepático Agudo/inducido químicamente , Fallo Hepático Agudo/tratamiento farmacológico , Fallo Hepático Agudo/prevención & control , Antiinflamatorios/efectos adversosRESUMEN
Acute liver failure (ALF) is a mortal and critical hepatic disease, in which oxidative stress, inflammation storm and hepatocyte death are crucial in the pathogenesis. Hence, in contrast to the control of a single link, a combination therapy targeting multiple pathogenic links of the disease will be a favorable means to control the progression of the disease. In this study, we constructed dimethyl itaconate-loaded liposomes modified with dodecyl gallate as a cocktail activator to investigate its functional role in acetaminophen (APAP)-induced ALF. Our results demonstrated that the cocktail activator acted on hepatocytes and triggered cocktail efficacy, thereby simultaneously attenuating APAP-induced hepatocyte damage and remodeling the damage microenvironment. The cocktail activator could effectively scavenge reactive oxygen species, inhibit excessive inflammatory responses and reduce cell death in impaired hepatocytes for detoxification. More importantly, the cocktail activator could remodel the damage microenvironment, thus further promoting hepatocyte expansion and specifically switching macrophages from the M1 to M2 phenotype for a favorable liver regeneration of ALF. Furthermore, in APAP-induced ALF mouse model, the cocktail activator improved liver function, alleviated histopathological damage and increased survival rate. In summary, these findings indicate that the cocktail activator may provide a promising therapeutic approach for ALF treatment as a nanomedicine.
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The majority (~80%) of patients with cancer do not derive clinical benefit from current immunotherapy, largely due to attenuation of immune responses imposed by robust immunosuppression at tumor sites. Here, a cell-based tumor antigen delivery strategy was developed to boost tumor-specific immunity. Notably, the platform constructing ferric oxide nanoparticle-trained macrophages loading tumor antigens (MFe-N) acquired an immunostimulatory program and functioned as the tumoritropic "cytokine-microfactories" to sustainably produce high levels of multiple therapeutic cytokines (GM-CSF, TNFα, and MIP-1α), which are important in activation of immune cells with antitumor potential. Indeed, MFe-N markedly enhanced recruitment of the professional antigen-presenting cells, dendritic cells (DCs), to the tumor sites of an established B16F10 mouse melanoma model. Subsequently, MFe-N effectively delivered tumor antigens to DCs by gap junction-mediated cell-to-cell transmission. And this trafficking was critical for DC maturation to augment antitumor T-cell responses. Simultaneously, the "cytokine-microfactories" elicited high production of the tumoricidal effectors, and in turn blunted the pro-angiogenic activity of tumor-associated macrophages, resulting in conversion of the tumor-supporting milieu to a tumoricidal function that favored infiltration of antitumor T-cells. The findings provided a novel "cytokine-microfactories" harnessing effective delivery of tumor antigens and production of therapeutic cytokines to robustly promote antigen presentation and reshape the tumor immune milieu for priming antitumor immunity. This can enhance existing T-cell mediated immunotherapeutic potency and extend the curative potential immunotherapy to a broader range of patients.
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Antígenos de Neoplasias , Vacunas contra el Cáncer , Animales , Presentación de Antígeno , Citocinas , Células Dendríticas , Uniones Comunicantes , Humanos , Inmunoterapia , RatonesRESUMEN
Background: Exosome (Exo)-based chemotherapeutic drug delivery systems have been extensively investigated; however, the therapeutic potential of other subtypes of extracellular vesicles (EVs), in particular microvesicles (MiV), seem to be overlooked. Moreover, despite a general agreement on organ tropism of EVs, few studies have clearly demonstrated that EVs specifically target tumor tissue. Methods: Proinflammatory macrophage-derived EV subpopulations comprising apoptotic bodies (ApB), MiV and Exo were isolated under differential ultracentrifugation, and further analyzed using comparative proteomic and lipid approach. Results: On the basis of EV biogenesis pathways, our data demonstrated that MiV acquire the tumor-targeting capacity probably through inheritance of CCR2-enriched cell membrane which also drives the recruitment of donor cells to tumor sites. Further, our data validate MiV utilize SNARE-mediated membrane fusion to directly discharge doxorubicin to nucleus and bypass endocytic degradation. Conclusions: Compared with other EV subtypes, MiV loaded with doxorubicin gain significant benefits in chemotherapeutic outcomes including survival rate improvements in metastatic ovarian cancer. Therefore, MiV represent a potent alterative to Exo and synthetic liposomes (Lipo) for tumor-targeting drug delivery.
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Micropartículas Derivadas de Células/metabolismo , Quimiocina CCL2/metabolismo , Sistemas de Liberación de Medicamentos/métodos , Macrófagos/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Receptores CCR2/metabolismo , Proteínas SNARE/metabolismo , Animales , Antibióticos Antineoplásicos/farmacología , Línea Celular Tumoral , Quimiocina CCL2/genética , Doxorrubicina/farmacología , Femenino , Humanos , Macrófagos/inmunología , Fusión de Membrana , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Proteómica/métodos , Receptores CCR2/genética , Proteínas SNARE/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Surgical resection currently remains the mainstay of treatment for patients with gliomas of any grade. The maximum extent of surgical resection is associated with a long-term disease control; however, maximal resection of the brain tumor possibly results in additional neurological deficits. Therefore, improving the precision in brain tumor surgery by visual identification and screening of tumor cells can help to tackle this devastating disease. In the present study, BV2 microglial cells were engineered by iron oxide-nanoparticle stimulation as intraoperative optical imaging agent vehicles and loaded with near-infrared fluorescent dye DiD (DiDBV2-Fe) potentially for fluorescence-guided brain tumor surgery. Activation of BV2 microglial cells by citrate-stabilized iron oxide nanoparticles at a concentration of 62.5 µg mL-1 significantly inhibited M2 markers (arginase-1 and CD206), which is able to minimize risks of the immunosuppressive effects caused by the M2-like phenotype of microglial cells. Meanwhile, activated BV2 microglial cells showed up-regulation of arylsulfatase A, apolipoprotein E, transferrin, and ferritin heavy chain-1 gene expression that tends to promote microglia transport across the blood-brain barrier (BBB). Compared to DiDBV2 without iron oxide activation, DiDBV2-Fe indicated strong tumor tropism in response to monocyte chemoattractant protein-1 (CCL2) secreted by U87MG tumor cells. In vivo experiments proved that DiDBV2-Fe efficiently crossed the BBB and more than 90% fluorescence intensity generated by activated microglial cells was detected in the brain when administered through the carotid artery in an orthotopic glioblastoma mouse model. Notably, DiDBV2-Fe produced clear tumor border demarcation on near-infrared imaging and exhibited a superior tumor-to-brain fluorescence ratio to commercial 5-aminolevulinic acid. Accumulated DiDBV2-Fe induced a strong fluorescence signal in brain tumor tissue for a prolonged period (4-24 h), which is beneficial to perform complex and time-consuming brain operations. Overall, our study suggests that this newly engineered microglial cell has promise for enabling more accurate brain tumor imaging for fluorescence-guided resections.
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Neoplasias Encefálicas/diagnóstico por imagen , Ingeniería Celular/métodos , Colorantes Fluorescentes/administración & dosificación , Glioma/diagnóstico por imagen , Microglía/citología , Animales , Neoplasias Encefálicas/metabolismo , Línea Celular , Colorantes Fluorescentes/química , Glioma/metabolismo , Humanos , Nanopartículas de Magnetita , Ratones , Microglía/química , Microglía/metabolismo , Trasplante de Neoplasias , Imagen Óptica , RatasRESUMEN
Objective: Engineered immune cells (e.g., therapeutic T cells) provide a revolutionary approach to combat cancer. Certain activated immune cells can exquisitely sense and respond to the tumor microenvironment. Here, we propose a paradigm based on engineering macrophages to allow selective intercellular drug delivery and augmentation of antitumor activities by hijacking tumor microtube networks. Methods: Macrophages were engineered via anchoring lipopolysaccharides on the plasma membrane (LM). The tumor tropism of LM encapsulating doxorubicin (LM-Dox) was monitored by a real-time cell migration assay and small animal in vivo imaging. Monocyte chemoattractant protein-1 (CCL2) was measured by quantitative PCR and ELISA. Intercellular conduit formation was characterized by confocal laser scanning microscopy and scanning electron microscopy. LM-Dox activation of tumor-associated macrophages to release TNF-α was evaluated by western blot and immunofluorescence assays. The potential therapeutic effects of LM-Dox in a 3D tumor-immune model and a murine orthotopic lung cancer model were tested. Results: LM-Dox exhibited tumor tropism in response to CCL2 produced by A549 lung tumor cells and lung tumor tissues resulting in a remarkably higher amount of tumor accumulation than the case of Lipo-Dox (~ 4-fold). Intriguingly, LM-Dox accumulated at tumor sites hijacked the established tumor microtube networks and even stimulated microtube formation with tumor cells but not with normal cells to enable selective and rapid transport of the drug to tumor cells. Simultaneously, LM-Dox induced secretion of TNF-α in tumor-associated macrophages, which increased the antitumor activity of Dox. Thus, LM-Dox increased the inhibitory effects on tumor growth and metastasis in a mouse orthotopic lung cancer model and minimized the side effects of Dox-induced tumor invasion. Conclusion: Lipopolysaccharide-anchored macrophages that can hijack tumor microtube networks for selective drug transport may serve as versatile bioactive carriers of anticancer drugs. In the clinical context, these engineered microphages represent a personalized medicine approach that can be translated into potential use of patient-derived monocytes/macrophages for drug delivery by means of cell-to-cell communication.
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Antineoplásicos/administración & dosificación , Antineoplásicos/química , Sistemas de Liberación de Medicamentos/métodos , Lipopolisacáridos/química , Neoplasias Pulmonares/tratamiento farmacológico , Macrófagos/química , Células A549 , Animales , Membrana Celular/química , Movimiento Celular/efectos de los fármacos , Doxorrubicina/administración & dosificación , Doxorrubicina/química , Sistemas de Liberación de Medicamentos/instrumentación , Femenino , Humanos , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/fisiopatología , Macrófagos/inmunología , Ratones , Ratones Endogámicos BALB C , Microambiente Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
It is extremely difficult for cancer chemotherapy to control the peritoneal metastasis of advanced ovarian carcinoma given its inability to target disseminated tumors and the severe toxic side effects on healthy organs. Here, we report antitumor M1 macrophages developed as live-cell carriers that deliver anticancer drugs for the treatment of the metastatic ovarian carcinoma. Engineered doxorubicin-loaded M1 macrophages (M1-Dox) significantly enhanced tumor tropism by upregulation of CCR2 and CCR4 compared with their parent cells. Meanwhile, M1-Dox inhibited doxorubicin-induced tumor invasion, whereas commercial Lipo-Dox did not limit these side effects. Importantly, our data uncovered a drug delivery mechanism by which M1-Dox transferred drug cargoes into tumor cells via a tunneling nanotube pathway. The tunneling nanotube network acted as a transportation expressway for ultrafast drug delivery of M1-Dox, leading to efficient ovarian carcinoma cell death. Furthermore, genetic, pharmacological, and physical perturbations of these tunneling nanotubes obviously decreased drug transfer of M1-Dox, which further validated the evident correlation between drug delivery of M1-Dox and tunneling nanotubes. Finally, in peritoneal metastatic ovarian carcinoma-burdened mice, M1-Dox specifically penetrated into and accumulated deep within disseminated neoplastic lesions compared with commercial Lipo-Dox, resulting in reducing metastatic tumors to a nearly undetectable level and significantly increasing overall survival. Overall, the strategy of engineered macrophages for ultrafast and accurate drug delivery via the tunneling nanotubular expressway potentially revolutionizes the treatment of metastatic ovarian carcinoma.