Cancer immunotherapy modulates immune cells to induce antitumor immune responses. Tumors employ immune checkpoints to evade immune cell attacks. Immune checkpoint inhibitors such as anti-PD-L1 antibody (aPD-L1), which is being used clinically for cancer treatments, can block immune checkpoints so that the immune system can attack tumors. However, immune checkpoint inhibitor therapy may be hampered by polarization of macrophages within the tumor microenvironment (TME) into M2 tumor-associated macrophages (TAMs), which suppress antitumor immune responses and promote tumor growth by releasing anti-inflammatory cytokines and angiogenic factors. In this study, we used exosome-mimetic nanovesicles derived from M1 macrophages (M1NVs) to repolarize M2 TAMs to M1 macrophages that release pro-inflammatory cytokines and induce antitumor immune responses and investigated whether the macrophage repolarization can potentiate the anticancer efficacy of aPD-L1. M1NV treatment induced successful polarization of M2 macrophages to M1 macrophages in vitro and in vivo. Intravenous injection of M1NVs into tumor-bearing mice suppressed tumor growth. Importantly, injection of a combination of M1NVs and aPD-L1 further reduced the tumor size, compared to the injection of either M1NVs or aPD-L1 alone. Thus, our study indicates that M1NV injection can repolarize M2 TAMs to M1 macrophages and potentiate antitumor efficacy of the checkpoint inhibitor therapy.

Development of localized inflammatory environments by M1 macrophages in the cardiac infarction region exacerbates heart failure after myocardial infarction (MI). Therefore, the regulation of inflammation by M1 macrophages and their timely polarization toward regenerative M2 macrophages suggest an immunotherapy. Particularly, controlling cellular generation of reactive oxygen species (ROS), which cause M1 differentiation, and developing M2 macrophage phenotypes in macrophages propose a therapeutic approach. Previously, stem or dendritic cells were used in MI for their anti-inflammatory and cardioprotective potentials and showed inflammation modulation and M2 macrophage progression for cardiac repair. However, cell-based therapeutics are limited due to invasive cell isolation, time-consuming cell expansion, labor-intensive and costly ex vivo cell manipulation, and low grafting efficiency. Here, we report that graphene oxide (GO) can serve as an antioxidant and attenuate inflammation and inflammatory polarization of macrophages via reduction in intracellular ROS. In addition, GO functions as a carrier for interleukin-4 plasmid DNA (IL-4 pDNA) that propagates M2 macrophages. We synthesized a macrophage-targeting/polarizing GO complex (MGC) and demonstrated that MGC decreased ROS in immune-stimulated macrophages. Furthermore, DNA-functionalized MGC (MGC/IL-4 pDNA) polarized M1 to M2 macrophages and enhanced the secretion of cardiac repair-favorable cytokines. Accordingly, injection of MGC/IL-4 pDNA into mouse MI models attenuated inflammation, elicited early polarization toward M2 macrophages, mitigated fibrosis, and improved heart function. Taken together, the present study highlights a biological application of GO in timely modulation of the immune environment in MI for cardiac repair. Current therapy using off-the-shelf material GO may overcome the shortcomings of cell therapies for MI.

Human mesenchymal stem cell (hMSC)-derived exosomes have been spotlighted as a promising therapeutic agent for cell-free regenerative medicine. However, poor organ-targeting ability and insufficient therapeutic efficacy of systemically injected hMSC-exosomes were identified as critical limitations for their further applications. Therefore, in this study we fabricated iron oxide nanoparticle (IONP)-incorporated exosome-mimetic nanovesicles (NV-IONP) from IONP-treated hMSCs and evaluated their therapeutic efficacy in a clinically relevant model for spinal cord injury. Compared to exosome-mimetic nanovesicles (NV) prepared from untreated hMSCs, NV-IONP not only contained IONPs which act as a magnet-guided navigation tool but also carried greater amounts of therapeutic growth factors that can be delivered to the target cells. The increased amounts of therapeutic growth factors inside NV-IONP were attributed to IONPs that are slowly ionized to iron ions which activate the JNK and c-Jun signaling cascades in hMSCs. In vivo systemic injection of NV-IONP with magnetic guidance significantly increased the amount of NV-IONP accumulating in the injured spinal cord. Accumulated NV-IONP enhanced blood vessel formation, attenuated inflammation and apoptosis in the injured spinal cord, and consequently improved spinal cord function. Taken together, these findings highlight the development of therapeutic efficacy-potentiated extracellular nanovesicles and demonstrate their feasibility for repairing injured spinal cord.

Because of poor engraftment and safety concerns regarding mesenchymal stem cell (MSC) therapy, MSC-derived exosomes have emerged as an alternative cell-free therapy for myocardial infarction (MI). However, the diffusion of exosomes out of the infarcted heart following injection and the low productivity limit the potential of clinical applications. Here, we developed exosome-mimetic extracellular nanovesicles (NVs) derived from iron oxide nanoparticles (IONPs)–incorporated MSCs (IONP-MSCs). The retention of injected IONP-MSC–derived NVs (IONP-NVs) within the infarcted heart was markedly augmented by magnetic guidance. Furthermore, IONPs significantly increased the levels of therapeutic molecules in IONP-MSCs and IONP-NVs, which can reduce the concern of low exosome productivity. The injection of IONP-NVs into the infarcted heart and magnetic guidance induced an early shift from the inflammation phase to the reparative phase, reduced apoptosis and fibrosis, and enhanced angiogenesis and cardiac function recovery. This approach can enhance the therapeutic potency of an MSC-derived NV therapy.

Liposomes are clinically used as drug carriers for cancer therapy; however, unwanted leakage of the encapsulated anticancer drug and poor tumor-targeting efficiency of liposomes may generate toxic side effects on healthy cells and lead to failure of tumor eradication. To overcome these limitations, we functionalized liposomes with a photosensitizer (KillerRed, KR)-embedded cancer cell membrane (CCM). A lipid adjuvant was also embedded in the lipocomplex to promote the anticancer immune response. KR proteins were expressed on CCM and did not leak from the lipocomplex. Owing to the homotypic affinity of the CCM for the source cancer cells, the lipocomplex exhibited a 3.3-fold higher cancer-targeting efficiency in vivo than a control liposome. The liposome functionalized with KR-embedded CCM and lipid adjuvant generated cytotoxic reactive oxygen species in photodynamic therapy and effectively induced anticancer immune responses, inhibiting primary tumor growth and lung metastasis in homotypic tumor-bearing mice. Taken together, the lipocomplex technology may improve liposome-based cancer therapy.

Currently, most biological research relies on conventional experimental techniques that allow only static analyses at certain time points in vitro or ex vivo . However, if one could visualize cellular dynamics in living organisms, that would provide a unique opportunity to study key biological phenomena in vivo . Intravital microscopy (IVM) encompasses diverse optical systems for direct viewing of objects, including biological structures and individual cells in live animals. With the current development of devices and techniques, IVM addresses important questions in various fields of biological and biomedical sciences. In this mini-review, we provide a general introduction to IVM and examples of recent applications in the field of immunology, oncology, and vascular biology. We also introduce an advanced type of IVM, dubbed real-time IVM, equipped with video-rate resonant scanning. Since the real-time IVM can render cellular dynamics with high temporal resolution in vivo , it allows visualization and analysis of rapid biological processes.

In a native muscle microenvironment, electrical and mechanical stimuli exist in the form of action potentials and muscle contraction. Here, a cell culture system is developed that can mimic the in vivo microenvironment and provide these stimuli to cultured cells, and it is tested whether the stimulation can promote myogenic differentiation of human umbilical cord blood mesenchymal stem cells (hUCBMSCs). A thermosensitive, stretchable, and piezoelectric substrate (TSPS) is fabricated by polydimethylsiloxane spin-coating of aligned ZnO nanorods and subsequent poly(N-isopropylacrylamide) grafting on the polydimethylsiloxane surface. Pulsatile mechanoelectrical cues are provided to hUCBMSCs cultured on the TSPS by subjecting the TSPS to cyclic stretching and bending, resulting in significant promotion of myogenic differentiation of hUCBMSCs as well as intracellular signaling related to the differentiation. After differentiation ex vivo, the cells are detached from the TSPS in the form of cell sheet fragments. Injection of the cell sheet fragments of differentiated cells into injured mouse skeletal muscle shows improved cell retention and muscle regeneration as compared to injection of either undifferentiated cells or differentiated dissociated cells. This system may serve as a tool for research on the electrical and mechanical regulation of stem cells and may be used to potentiate stem cell therapies.

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