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Ping Shao,1 Danyang Dong,1 Yong Huang,2 Yan Wang,1 Lian Pan,3 Qunshan Zhu,2 Zheng Wu,2 Fengjuan Jiang,4 Wei Wei2 1Department of Spleen and Stomach Diseases, Yixing Traditional Chinese Medicine Hospital, Wuxi, 214200, People’s Republic of China; 2Department of General Surgery, Jiangdu People’s Hospital Affiliated to Yangzhou University, Yangzhou, 225200, People’s Republic of China; 3Pathology Department, Yixing Traditional Chinese Medicine Hospital, Wuxi, 214200, People’s Republic of China; 4Department of Gastroenterology, Jiangdu People’s Hospital Affiliated to Yangzhou University, Yangzhou, 225200, People’s Republic of ChinaCorrespondence: Wei Wei, Email 18952531159@yzu.edu.cnBackground: Gastric cancer (GC) as is the second deadliest malignancy still lacks rapid, simple and economical detection and early clinical screening techniques. Surface-enhanced Raman spectroscopy (SERS) is a spectroscopic technique based on the surface plasmon resonance of precious metal nanoparticles, which can effectively detect low-abundance tumor markers. Combining SERS technology with sensors has high potential in the diagnosis and screening of GC.Methods: A novel Au/Si nano-umbrella array (Au/SiNUA) was prepared as a SERS substrate and the substrate was functionalized using the corresponding tumor marker aptamers for the detection of clinical biological samples using a one-step recognition release mechanism. Optimization of aptamer and complementary chain concentrations and detection time for optimal sensor preparation.Results: Au/SiNUA were tested to have good SERS enhancement activity. The proposed aptamer biosensor has good specificity and stability, with a low detection time of 18 min and a limit of detection (LOD) at the fM level, which is superior to most of the methods reported so far; and the accuracy of the clinical assay is comparable to that of the ELISA method. The expression levels of PDGF-B and thrombin in the serum of GC patients and healthy individuals can be effectively detected and differentiated.Conclusion: The ultrasensitive and specific aptamer biosensor is highly feasible for the diagnosis and screening of GC and has good application prospects.Keywords: surface-enhanced Raman scattering, gastric cancer, aptamer, biosensor, Au/SiNUA

Dong Dong,1,2,* Cheng-Hui Hsiao,3,* Beppino C Giovanella,4,† Yifei Wang,2 Diana SL Chow,3 Zhijie Li11International Ocular Surface Research Center and Institute of Ophthalmology, Jinan University Medical School, Guangzhou, China; 2GuangZhou (Jinan) Biomedical Research and Development Center Co. Ltd, Guangzhou, China; 3Department of Pharmacological and Pharmaceutical Sciences, College of Pharmacy, University of Houston, Houston, TX, USA; 4CHRISTUS Stehlin Foundation for Cancer Research, Houston, TX, USA*These authors contributed equally to this work†Dr Beppino C Giovanella passed away in 2017.Background and aim: We have synthesized a novel lactone-stabilized camptothecin (CPT) analog named CZ48 and demonstrated its potent anticancer effects via bioconversion to the active CPT in earlier studies. Herein, we aimed to develop, optimize and characterize CZ48 nanosuspensions, for a sustained delivery of this drug in humans with an intravenous (i.v.) administration.Methods and materials: A three-factor, five-level central composite design (CCD) was employed to establish the impacts of the critical influencing factors (concentrations (wt%) of CZ48, polysorbate 80 (Tween-80), and Pluronic®, F-108 (F-108)) on the responses (particle size and zeta potential). Based on the quantitative influencing factor–response relationships, two optimized CZ48 nanosuspensions of 197.22 ± 7.12 nm (NS-S) and 589.35 ± 23.27 nm (NS-L) were developed with the zeta potential values of –26.5 mV and –27.9 mV, respectively.Results: CZ48 released from the nanosuspensions in a sustained manner in contrast to the rapid release from cosolvent in both PBS and human plasma. Moreover, NS-S exhibited more favored pharmacokinetic properties than NS-L, with a 31-fold prolonged elimination half-life of CPT, and a 2.4-fold enhanced CPT exposure over cosolvent. In efficacy study, NS-S exhibited significant tumor suppression and an improved survival rate with a higher tolerable dose, compared to CZ48 cosolvent.Conclusion: We have successfully developed CZ48 nanosuspensions with significantly favorable pharmacokinetics and improved efficacy using CCD approach. The formulation offers potential merits as a preferred candidate for clinical trials with the prolonged CPT exposure, which is known to correlate with the clinical efficacy.Keywords: sustained drug delivery, CZ48, camptothecin, nanosuspension, anticancer

Xue Yuan,1,2 Qian Xie,1 Keyu Su,1 Zhijie Li,3 Dong Dong,3 Baojian Wu1,2 1Research Center for Biopharmaceutics and Pharmacokinetics, College of Pharmacy, 2Guangdong Province Key Laboratory of Pharmacodynamic Constituents of TCM and New Drugs Research, 3International Ocular Surface Research Centre and Institute of Ophthalmology, School of Medicine, Jinan University, Guangzhou, China Abstract: Arenobufagin (ABG) is a major active component of toad venom, a traditional Chinese medicine used for cancer therapy. However, poor aqueous solubility limits its pharmacological studies in vivo due to administration difficulties. In this study, we aimed to develop a polymeric nanomicelle (PN) system to enhance the solubility of ABG for effective intravenous delivery. ABG-loaded PNs (ABG-PNs) were prepared with methoxy poly (ethylene glycol)-block-poly (D,L-lactic-co-glycolic acid) (mPEG-PLGA) using the solvent-diffusion technique. The obtained ABG-PNs were 105 nm in size with a small polydispersity index of 0.08. The entrapment efficiency and drug loading were 71.9% and 4.58%, respectively. Cellular uptake of ABG-PNs was controlled by specific clathrin-mediated endocytosis. In addition, ABG-PNs showed improved drug pharmacokinetics with an increased area under the curve value (a 1.73-fold increase) and a decreased elimination clearance (37.8% decrease). The nanomicelles showed increased drug concentrations in the liver and lung. In contrast, drug concentrations in both heart and brain were decreased. Moreover, the nanomicelles enhanced the anticancer effect of the pure drug probably via increased cellular uptake of drug molecules. In conclusion, the mPEG-PLGA-based nanomicelle system is a satisfactory carrier for the systemic delivery of ABG. Keywords: bufanolide steroid, arenobufagin, nanomicelles, mPEG-PLGA

Dong Dong,1* Xiao Wang,1* Huailing Wang,1 Xingwang Zhang,2 Yifei Wang,1 Baojian Wu2 1Guangzhou Jinan Biomedicine Research and Development Center, 2Division of Pharmaceutics, College of Pharmacy, Jinan University, Guangzhou, People’s Republic of China *These authors contributed equally to this work Introduction: SNX-2112 is a promising anticancer agent but has poor solubility in both water and oil. In the study reported here, we aimed to develop a nanocrystal formulation for SNX-2112 and to determine the pharmacokinetic behaviors of the prepared nanocrystals. Methods: Nanocrystals of SNX-2112 were prepared using the wet-media milling technique and characterized by particle size, differential scanning calorimetry, drug release, etc. Physiologically based pharmacokinetic (PBPK) modeling was undertaken to evaluate the drug’s disposition in rats following administration of drug cosolvent or nanocrystals. Results: The optimized SNX-2112 nanocrystals (with poloxamer 188 as the stabilizer) were 203 nm in size with a zeta potential of -11.6 mV. In addition, the nanocrystals showed a comparable release profile to the control (drug cosolvent). Further, the rat PBPK model incorporating the parameters of particulate uptake (into the liver and spleen) and of in vivo drug release was well fitted to the experimental data following administration of the drug nanocrystals. The results reveal that the nanocrystals rapidly released drug molecules in vivo, accounting for their cosolvent-like pharmacokinetic behaviors. Due to particulate uptake, drug accumulation in the liver and spleen was significant at the initial time points (within 1 hour). Conclusion: The nanocrystals should be a good choice for the systemic delivery of the poorly soluble drug SNX-2112. Also, our study contributes to an improved understanding of the in vivo fate of nanocrystals. Keywords: intravenous delivery, PBPK, tissue distribution, poloxamer 188

Background: The global prevalence of Dengue virus (DENV) infection poses a significant health risk, urging the need for effective vaccinations. Peptide vaccines, known for their capacity to induce comprehensive immunity against multiple virus serotypes, offer promise due to their stability, safety, and design flexibility. Spherical nucleic acid (SNA), particularly those with gold nanoparticle cores, present an attractive avenue for enhancing peptide vaccine efficacy due to their modularity and immunomodulatory properties., Methods: The spherical nucleic acid-TBB (SNA-TBB), a novel nanovaccine construct, was fabricated through the co-functionalization process of SNA with epitope peptide, targeting all four serotypes of the DENV. This innovative approach aims to enhance immunogenicity and provide broad-spectrum protection against DENV infections. The physicochemical properties of SNA-TBB were characterized using dynamic light scattering, zeta potential measurement, and transmission electron microscopy. In vitro assessments included endocytosis studies, cytotoxicity evaluation, bone marrow-dendritic cells (BMDCs) maturation and activation analysis, cytokine detection, RNA sequencing, and transcript level analysis in BMDCs. In vivo immunization studies in mice involved evaluating IgG antibody titers, serum protection against DENV infection and safety assessment of nanovaccines., Results: SNA-TBB demonstrated successful synthesis, enhanced endocytosis, and favorable physicochemical properties. In vitro assessments revealed no cytotoxicity and promoted BMDCs maturation. Cytokine analyses exhibited heightened IL-12p70, TNF-α, and IL-1β levels. Transcriptomic analysis highlighted genes linked to BMDCs maturation and immune responses. In vivo studies immunization with SNA-TBB resulted in elevated antigen-specific IgG antibody levels and conferred protection against DENV infection in neonatal mice. Evaluation of in vivo safety showed no signs of adverse effects in vital organs., Conclusion: The study demonstrates the successful development of SNA-TBB as a promising nanovaccine platform against DENV infection and highlights the potential of SNA-based peptide vaccines as a strategy for developing safe and effective antiviral immunotherapy., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Zhao et al.)

Background: Gastric lesions pose significant clinical challenges due to their varying degrees of malignancy and difficulty in early diagnosis. Early and accurate detection of these lesions is crucial for effective treatment and improved patient outcomes., Methods: This paper proposed a label-free and highly sensitive classification method for serum of patients with different degrees of gastric lesions by combining surface-enhanced Raman scattering (SERS) and machine learning analysis. Specifically, we prepared Au lotus-shaped (AuLS) nanoarrays substrates using seed-mediated and liquid-liquid interface self-assembly method for measuring SERS spectra of serum, and then the collected spectra were processed by principal component analysis (PCA) - multi-local means based nearest neighbor (MLMNN) model to achieve differentiation., Results: By employing this pattern analysis, AuLS nanoarray substrates can achieve fast, sensitive, and label-free serum spectral detection. The classification accuracy can reach 97.5%, the sensitivity is higher than 96.7%, and the specificity is higher than 95.0%. Moreover, by analyzing the PCs loading plots, the most critical spectral features distinguishing different degrees of gastric lesions were successfully captured., Conclusion: This discovery lays the foundation for combining SERS with machine learning for real-time diagnosis and recognition of gastric lesions., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Chen et al.)

Diabetic wounds pose a significant challenge in modern healthcare due to their chronic and complex nature, often resulting in delayed healing, infections, and, in severe cases, amputations. In recent years, nanotherapeutic approaches have emerged as promising strategies to address the unique pathophysiological characteristics of diabetic wounds. This review paper provides a comprehensive overview of the latest advancements in nanotherapeutics for diabetic wound treatment. We discuss various nanomaterials and delivery systems employed in these emerging therapies. Furthermore, we explore the integration of biomaterials to enhance the efficacy of nanotherapeutic interventions. By examining the current state-of-the-art research, challenges, and prospects, this review aims to offer valuable insights for researchers, clinicians, and healthcare professionals working in the field of diabetic wound care., Competing Interests: The authors declare that they have no competing interests in this work., (© 2024 Shi et al.)

Background: The inadequate perfusion, frequently resulting from abnormal vascular configuration, gives rise to tumor hypoxia. The presence of this condition hinders the effective delivery of therapeutic drugs and the infiltration of immune cells into the tumor, thereby compromising the efficacy of treatments against tumors. The objective of this study is to exploit the thermal effect of ultrasound (US) in order to induce localized temperature elevation within the tumor, thereby facilitating vasodilation, augmenting drug delivery, and enhancing immune cell infiltration., Methods: The selection of US parameters was based on intratumor temperature elevation and their impact on cell viability. Vasodilation and hypoxia improvement were investigated using enzyme-linked immunosorbent assay (ELISA) and immunofluorescence examination. The distribution and accumulation of commercial pegylated liposomal doxorubicin (PLD) and PD-L1 antibody (anti-PD-L1) in the tumor were analyzed through frozen section analysis, ELISA, and in vivo fluorescence imaging. The evaluation of tumor immune microenvironment was conducted using flow cytometry (FCM). The efficacy of US-enhanced chemotherapy in combination with immunotherapy was investigated by monitoring tumor growth and survival rate after various treatments., Results: The US irradiation condition of 0.8 W/cm 2 for 10 min effectively elevated the tumor temperature to approximately 40 °C without causing any cellular or tissue damage, and sufficiently induced vasodilation, thereby enhancing the distribution and delivery of PLD and anti-PD-L1 in US-treated tumors. Moreover, it effectively mitigated tumor hypoxia while significantly increasing M1-phenotype tumor-associated macrophages (TAMs) and CD8+ T cells, as well as decreasing M2-phenotype TAMs. By incorporating US irradiation, the therapeutic efficacy of PLD and anti-PD-L1 was substantially boosted, leading to effective suppression of tumor growth and prolonged survival in mice., Conclusion: The application of US (0.8 W/cm 2 for 10 min) can effectively induce vasodilation and enhance the delivery of PLD and anti-PD-L1 into tumors, thereby reshaping the immunosuppressive tumor microenvironment and optimizing therapeutic outcomes., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Xiang et al.)

Purpose: Incorporation of luvangetin in nanoemulsions for antimicrobial and therapeutic use in infected wound healing., Patients and Methods: Luvangetin nanoemulsions were prepared by high-speed shear method and characterized based on their appearance structure, average droplet size, polydispersity index (PDI), electric potential, storage stability. Optimized formulation of luvangetin nanoemulsion by Box-Behnken design (BBD). The antimicrobial activity and antimicrobial mechanism of luvangetin nanoemulsions against common hospital pathogens, ie, Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) , were investigated using luvangetin nanoemulsions. The biosafety of luvangetin nanoemulsion was evaluated through cytotoxicity, apoptosis, and reactive oxygen species (ROS) assay experiments using human normal epidermal cells and endothelial cells. Finally, the effect of luvangetin nanoemulsion on healing of infected wounds was investigated in B6 mice., Results: Luvangetin nanoemulsion formulation consists of 2.5% sunflower seed oil, 10% emulsifier Span-20 and 7 minutes of shear time, and with good stability. Luvangetin nanoemulsion produces antibacterial activity against S. aureus and E. coli by disrupting the structure of bacterial cell membranes. Luvangetin nanoemulsion are biologically safe for HaCat and HUVEC. Luvangetin nanoemulsion showed good therapeutic effect on MRSA infected wounds in mice., Conclusion: For the first time, developed a new formulation called luvangetin nanoemulsion, which exhibited superior antibacterial effects against Gram-positive bacteria. Luvangetin nanoemulsion has a favorable effect in promoting infected wound healing. We have combined luvangetin, which has multiple activities, with nanoemulsions to provide a new topical fungicidal formulation, and have comprehensively evaluated its effectiveness and safety, opening up new possibilities for further applications of luvangetin., Competing Interests: The authors have no relevant financial or non-financial interests to disclose., (© 2024 Chong et al.)

Purpose: This study aims to explore a novel scaffold for osteotendinous junction regeneration and to preliminarily verify its osteogenic and tenogenic abilities in vitro., Methods: A polycaprolactone (PCL) scaffold with aligned and orthogonal fibers was created using melt electrowriting (MEW) and fused deposition modeling (FDM). The scaffold was coated with Type I collagen, and hydroxyapatite was carefully added to separate the regions intended for bone and tendon regeneration, before being rolled into a cylindrical shape. Human adipose-derived stem cells (hADSCs) were seeded to evaluate viability and differentiation. Scaffold characterization was performed with Scanning Electron Microscope (SEM). Osteogenesis was assessed by alkaline phosphatase (ALP) and Alizarin red staining, while immunostaining and transcription-quantitative polymerase chain reaction (RT-qPCR) evaluated osteogenic and tendogenic markers., Results: Scaffolds were developed in four variations: aligned (A), collagen-coated aligned (A+C), orthogonal (O), and mineral-coated orthogonal (O+M). SEM analysis confirmed surface morphology and energy-dispersive X-ray spectroscopy (EDS) verified mineral coating on O+M types. Hydrophilicity and mechanical properties were optimized in modified scaffolds, with A+C showing increased tensile strength and O+M improved in compression. hADSCs demonstrated good viability and morphology across scaffolds, withO+M scaffolds showing higher cell proliferation and osteogenic potential, and A and A+C scaffolds supporting tenogenic differentiation., Conclusion: This study confirms the potential of a novel PCL scaffold with distinct regions for osteogenic and tenogenic differentiation, supporting the regeneration of osteotendinous junctions in vitro., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Ma et al.)

Adoptive T cells immunotherapy, specifically chimeric antigen receptor T cells (CAR-T), has shown promising therapeutic efficacy in the treatment of hematologic malignancies. As extensive research on CAR-T therapies has been conducted, various challenges have emerged that significantly hampered their clinical application, including tumor recurrence, CAR-T cell exhaustion, and cytokine release syndrome (CRS). To overcome the hurdles of CAR-T therapy in clinical treatment, cell-free emerging therapies based on exosomes derived from CAR-T cells have been developed as an effective and promising alternative approach. In this review, we present CAR-T cell-based therapies for the treatment of tumors, including the features and benefits of CAR-T therapies, the limitations that exist in this field, and the measures taken to overcome them. Furthermore, we discuss the notable benefits of utilizing exosomes released from CAR-T cells in tumor treatment and anticipate potential issues in clinical trials. Lastly, drawing from previous research on exosomes from CAR-T cells and the characteristics of exosomes, we propose strategies to overcome these restrictions. Additionally, the review discusses the plight in large-scale preparation of exosome and provides potential solutions for future clinical applications., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Hu et al.)

Acute liver injury (AIL), a fatal clinical disease featured with a swift deterioration of hepatocyte functions in the short term, has emerged as a serious public health issues that warrants attention. However, the effectiveness of existing small molecular antioxidants and anti-inflammatory medications in alleviating AIL remains uncertain. The unique inherent structural characteristics of liver confer it a natural propensity for nanoparticle capture, which present an opportunity to exploit in the formulation of nanoscale therapeutic agents, enabling their selective accumulation in the liver and thereby facilitating targeted therapeutic interventions. Significantly increased reactive oxygen species (ROS) accumulation and inflammation response have been evidenced to play crucial roles in occurrence and development of AIL. Nanozymes with ROS-scavenging capacities have demonstrated considerable promise in ROS elimination and inflammation regulation, thereby offering an appealing therapeutic instrument for the management of acute liver injury. In this review, the mechanisms of different type of ALI were summarized. In addition, we provide a comprehensive summary and review of the available ROS-scavenging nanozymes, including transition metal-based nanozymes, noble metal nanozymes, carbon-based nanozymes, and some other nanozymes. Furthermore, the challenges still need to be solved in the field of ROS-scavenging nanozymes for ALI alleviation are also discussed., Competing Interests: The authors report no conflicts of interest in this work., (© 2023 Sun et al.)

Introduction: Pathogenic respiratory RNA viruses, including influenza A virus (IAV), respiratory syncytial virus (RSV), and SARS-CoV-2, are major causes of causes of acute respiratory infection globally. Plant-derived exosome-like nanoparticles containing miRNAs have shown substantial cross-kingdom regulatory effects on both viral and human transcripts. Houttuynia cordata ( H. cordata ), a traditional Chinese medicine frequently used to treat respiratory diseases. However, the role of H. cordata- derived exosome-like nanoparticles (HELNs) and the miRNA they encapsulated are unclear., Methods: HELNs were isolated from fresh underground roots (uHELNs) and above ground stems and leaves (aHELNs) using differential centrifugation. The HELNs were identified using transmission electron microscopy, nanoparticle tracking analysis, and zeta potential. Small RNA sequencing and RT-PCR were employed to determine the miRNA expression in uHELNs and aHELNs. All genomes were sourced from the NCBI database. Target prediction of viral genomes was performed using RNAhybrid, while human target prediction was conducted using both RNAhybrid and Miranda. Functional enrichment analysis was applied to the predicted human targets to explore the hub targets and their roles in antiviral effects. The accessibility of miRNA target sites was determined through the MFOLD web server, and customized dual-luciferase reporter assays were administered to validate the computational findings., Results: A total of 12 highly enriched miRNAs were identified in both uHELNs and aHELNs. Upon prediction and verification, miR858a and miR858b were shown to target the NP gene in H1N1, while miR166a-3p targeted the ORF1ab in SARS-CoV-2. However, no valid miRNA targets were found for RSV. Regarding human transcripts, miR168a-3p, miR168b-3p, and miR8175 were found to inhibit MAPK3 expression, and novel_mir2 could suppress both AKT1 and MAPK3 expression., Discussion: This study sheds light on the collaborative antiviral mechanism of miRNAs in HELNs across two species and explores the potential antiviral scopes of both H. cordata miRNAs and HELNs., Competing Interests: The authors declare no competing interest in this work., (© 2023 Zhu et al.)

Background: Olaparib, a poly (adenosine diphosphate-ribose) polymerase (PARP) inhibitor has demonstrated promising efficacy in patients with triple-negative breast cancer (TNBC) carrying breast cancer gene (BRCA) mutations. However, its impact on BRCA wild-type (BRCAwt) TNBC is limited. Hence, it is crucial to sensitize BRCAwt TNBC cells to olaparib for effective clinical practice. Novobiocin, a DNA polymerase theta (POLθ) inhibitor, exhibits sensitivity towards BRCA-mutated cancer cells that have acquired resistance to PARP inhibitors. Although both of these DNA repair inhibitors demonstrate therapeutic efficacy in BRCA-mutated cancers, their nanomedicine formulations' antitumor effects on wild-type cancer remain unclear. Furthermore, ensuring effective drug accumulation and release at the cancer site is essential for the clinical application of olaparib., Materials and Methods: Herein, we designed a progressively disassembled nanosystem of DNA repair inhibitors as a novel strategy to enhance the effectiveness of olaparib in BRCAwt TNBC. The nanosystem enabled synergistic delivery of two DNA repair inhibitors olaparib and novobiocin, within an ultrathin silica framework interconnected by disulfide bonds., Results: The designed nanosystem demonstrated remarkable capabilities, including long-term molecular storage and specific drug release triggered by the tumor microenvironment. Furthermore, the nanosystem exhibited potent inhibitory effects on cell viability, enhanced accumulation of DNA damage, and promotion of apoptosis in BRCAwt TNBC cells. Additionally, the nanosystem effectively accumulated within BRCAwt TNBC, leading to significant growth inhibition and displaying vascular regulatory abilities as assessed by magnetic resonance imaging (MRI)., Conclusion: Our results provided the inaugural evidence showcasing the potential of a progressively disassembled nanosystem of DNA repair inhibitors, as a promising strategy for the treatment of BRCA wild-type triple-negative breast cancer., Competing Interests: The authors report no conflicts of interest in this work., (© 2023 Fang et al.)

Purpose: This study aims to investigate the impact of enhancing subchondral bone repair on the efficacy of articular cartilage restoration, thereby achieving improved osteochondral regeneration outcomes., Methods: In this study, we modified the surface of nano-hydroxyapatite (n-HAp) through alkylation reactions to prepare n-HApMA. Characterization techniques, including X-ray diffraction, infrared spectroscopy scanning, thermogravimetric analysis, particle size analysis, and electron microscopy, were employed to analyze n-HApMA. Bioinks were prepared using n-HApMA, high porosity GelMA hydrogel, and adipose tissue derived stromal cells (ADSCs). The rheological properties of the bioinks during photocuring were investigated using a rheometer. Based on these bioinks, a biphasic scaffold was constructed. The viability of cells within the scaffold was observed using live-dead cell staining, while the internal morphology was examined using scanning electron microscopy. The stiffness of the scaffold was evaluated through compression testing. Scaffolds were implanted into the osteochondral defects of New Zealand rabbit knees, and microCT was utilized to observe the subchondral bone repair. Hematoxylin and eosin (H&E) staining, Masson's trichrome staining, and Safranin O/Fast Green staining were performed to assess the regeneration of subchondral bone and cartilage. Furthermore, immunohistochemical staining was employed to detect the expression of osteogenic and chondrogenic-related molecules., Results: Scaffold characterization revealed that surface modification enables the uniform distribution of n-HApMA within the GelMA matrix. The incorporation of 5% n-HApMA notably enhanced the elastic modulus and stiffness of the 6% high-porosity GelMA in comparison to n-HAp. Moreover, in-vivo study showed that the homogeneous dispersion of n-HApMA on the GelMA matrix facilitated the osteogenic differentiation of adipose-derived stem cells (ADSCs) and promoted osteochondral tissue regeneration., Conclusion: These findings suggest potential applications of the n-HApMA/GelMA composite in the field of tissue engineering and regenerative medicine., Competing Interests: The authors report no conflicts of interest in this work., (© 2023 Zheng et al.)

Purpose: Ultrasound molecular imaging (UMI) has proven promising to diagnose the onset and progression of diseases such as angiogenesis, inflammation, and thrombosis. However, microbubble-based acoustic probes are confined to intravascular targets due to their relatively large particle size, greatly reducing the application value of UMI, especially for extravascular targets. Extradomain B fibronectin (ED-B FN) is an important glycoprotein associated with tumor genesis and development and highly expressed in many types of tumors. Here, we developed a gas vesicles (GVs)-based nanoscale acoustic probe (ZD2-GVs) through conjugating ZD2 peptides which can specially target to ED-B FN to the biosynthetic GVs., Materials and Methods: ED-B FN expression was evaluated in normal liver and tumor tissues with immunofluorescence and Western blot. ZD2-GVs were prepared by conjugating ZD2 to the surface of GVs by amide reaction. The inverted microscope was used to analyze the targeted binding capacity of ZD2-GVs to MB49 cells (bladder cancer cell line). The contrast-enhanced imaging features of GVs, non-targeted control GVs (CTR-GVs), and targeted GVs (ZD2-GVs) were compared in three MB49 tumor mice. The penetration ability of ZD2-GVs in tumor tissues was assessed by fluorescence immunohistochemistry. The biosafety of GVs was evaluated by CCK8, blood biochemistry, and HE staining., Results: Strong ED-B FN expression was observed in tumor tissues while little expression in normal liver tissues. The resulting ZD2-GVs had only 267.73 ± 2.86 nm particle size and exhibited excellent binding capability to the MB49 tumor cells. The in vivo UMI experiments showed that ZD2-GVs produced stronger and longer retention in the BC tumors than that of the non-targeted CTR-GVs and GVs. Fluorescence immunohistochemistry confirmed that ZD2-GVs could penetrate the tumor vascular into the interstitial space of the tumors. Biosafety analysis revealed there was no significant cytotoxicity to these tested mice., Conclusion: Thus, ZD2-GVs can function as a potential UMI probe for the early diagnosis of bladder cancer., Competing Interests: Yanan Feng, Yongsheng Hao, Weijian Song and Litao Sun report a patent 202310218086.4 pending to Fei Yan. The authors report no other conflicts of interest in this work., (© 2023 Feng et al.)

Background: Breast cancer has become the most common cancer in women. Compare with other subtypes of breast cancer, triple-negative breast cancer (TNBC) is more likely to relapse and metastasize. Highly effective therapeutic strategies are desperately needed to be explored. In this study, a multifunctional nanoplatform is expected to mediate chemo-photothermal therapy, which can combine immunogenic cell death with checkpoint blockade to combat TNBC and distant metastasis., Methods: Poly (lactic acid-glycolic acid)-Poly (ethylene glycol) (PLGA-PEG) nanoparticles (NPs), a type of polymeric NPs, loaded with IR780, a near-infrared (NIR) dye, and doxorubicin (DOX) as the chemotherapeutic drug, were assembled by an improved double emulsification method (designated as IDNPs). The characterization, intracellular uptake, biosafety, photoacoustic (PA) imaging performance, and biodistribution of IDNPs were studied. Chemo-photothermal therapeutic effect and immunogenic cell death (ICD) were evaluated both in vitro and in vivo. The potency of chemo-photothermal therapy-triggered ICD in combination with anti-PD-1 immune checkpoint blockade (ICB) immunotherapy in eliciting immune response and treating distant tumors was further investigated., Results: IR780 and DOX were successfully loaded into PLGA-PEG to form the IDNPs, with size of 243.87nm and Zeta potential of -6.25mV. The encapsulation efficiency of IR780 and DOX was 83.44% and 5.98%, respectively. IDNPs demonstrated remarkable on-site accumulation and PA imaging capability toward 4T1 TNBC models. Chemo-photothermal therapy demonstrated satisfactory therapeutic effects both in vitro and in vivo, and triggered ICD efficiently. ICD, in combination with anti-PD-1, provoked a systemic antitumor immune response against distant tumors., Conclusion: Multifunctional IDNPs were successfully synthesized to mediate chemo-photothermal therapy, which combines immunogenic cell death with checkpoint blockade to combat TNBC and distant metastasis, showing great promise preclinically and clinically., Competing Interests: The authors report no conflicts of interest in this work., (© 2023 Zhu et al.)

Introduction: Triple-negative breast cancer (TNBC) is known to be the most aggressive form of breast cancer. Due to its high recurrence and mortality rates, the treatment of TNBC is a significant challenge for the medical community. Besides, ferroptosis is an emerging regulatory cell death that may provide new insights into the treatment of TNBC. As a central inhibitor of the ferroptosis process, the selenoenzyme glutathione peroxidase 4 (GPX4) is its classical therapeutic target. However, inhibition of GPX4 expression is quite detrimental to normal tissues. Ultrasound contrast agents, as an emerging visualization precision treatment, may provide a solution to the existing problem., Methods: In this study, nanodroplets (NDs) carrying simvastatin (SIM) were constructed using the homogeneous/emulsification method. Then, the characterization of SIM-NDs was systematically evaluated. Meanwhile, in this study, the ability of SIM-NDs combined with ultrasound-targeted microbubble disruption (UTMD) to initiate ferroptosis and its respective mechanisms of ferroptosis induction were verified. Finally, the antitumor activity of SIM-NDs was investigated in vitro and in vivo using MDA-MB-231 cells and TNBC animal models., Results: SIM-NDs exhibited excellent pH- and ultrasound-responsive drug release and noticeable ultrasonographic imaging ability, also showing good biocompatibility and biosafety. UTMD could promote increased intracellular reactive oxygen species and consume intracellular glutathione. However, SIM-NDs were efficiently internalized into cells under ultrasound irradiation, followed by the rapid release of SIM, which inhibited intracellular mevalonate production, and synergistically downregulated GPX4 expression, thereby promoting ferroptosis. Moreover, this combined treatment demonstrated strong antitumor ability in vitro and in vivo., Conclusion: The combination of UTMD and SIM-NDs presents a promising avenue for harnessing ferroptosis in the treatment of malignant tumors., Competing Interests: The authors declare that they have no competing interests., (© 2023 Liu et al.)

Purpose: To investigate the effect of macrophage exosomal long non-coding (lnc)RNAs on bone mesenchymal stem cell (BMSC) osteogenesis and the associated mechanism., Methods: Rat BMSCs and spleen macrophages were co-cultured with serum derived from the fracture microenvironment of rat tibia. BMSC osteogenesis was evaluated using Alizarin red staining and the expression of BMP-2, RUNX2, OPN , and OC mRNA. BMSC osteogenesis was evaluated after co-culture with macrophages stimulated using hypoxic conditions or colony-stimulating factor (CSF). The uptake of macrophage-derived exosomes by BMSCs was evaluated using the exosome uptake assay. High-throughput sequencing and bioinformatics analyses were performed to identify key lncRNAs in the macrophage exosomes. The effect of lncRNA expression levels on BMSC osteogenesis was also assessed using a lncRNA overexpression plasmid and siRNA technology. M1 and M2 macrophages were distinguished using flow cytometry and the key exosomal lncRNA was detected by in situ hybridization., Results: In the fracture microenvironment, macrophages (stimulated using either hypoxia or CSF) significantly increased the osteogenic ability of BMSCs. We showed that BMSCs assimilated macrophage-derived vesicles and that the inhibition of exosomal secretion significantly attenuated the macrophage-mediated induction of BMSC osteogenesis. The hypoxia condition led to the up-regulation of 310 lncRNAs and the down-regulation of 575 lncRNAs in macrophage exosomes, while CSF stimulation caused the up-regulation of 557 lncRNAs and the down-regulation of 407 lncRNAs. In total, 108 lncRNAs were co-up-regulated and 326 lncRNAs were co-down-regulated under both conditions. We eventually identified LOC103691165 as a key lncRNA that promoted BMSC osteogenesis and was expressed at similar levels in both M1 and M2 macrophages., Conclusion: In the fracture microenvironment, M1 and M2 macrophages promoted BMSC osteogenesis by secreting exosomes containing LOC103691165., Competing Interests: The authors declare no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (© 2023 Wang et al.)

Introduction: Photodynamic therapy (PDT) has attracted increasing attention for tumor treatment because of its minimal invasiveness and specific spatiotemporal selectivity. However, insufficient tumor accumulation and low cellular uptake of photosensitizers limit its therapeutic efficacy., Methods: In this study, flexible hollow human serum albumin/catalase nanocapsules (HSA/CATs) were created using a core-assisted protein-coating method and combined with the photosensitizer chlorin e6 (HSA/CAT@Ce6) for PDT., Results and Discussion: Transmission electron microscopy (TEM) images demonstrate that HSA/CAT nanocapsules are flexible, with a uniform diameter (310 nm) and a well-defined hollow structure. Thanks to their flexibility, HSA/CAT@Ce6 nanocapsules show a higher cellular uptake than rigid nanoparticles. The nanocapsules effectively generate reactive oxygen species (ROS) in 4T1 cells because of their high cellular uptake and catalytic capacity, remarkably enhancing their in vitro PDT efficacy. In addition, the in vivo tumor accumulation of HSA/CAT@Ce6 nanocapsules is significantly larger than that of rigid nanoparticles and Ce6, meaning they are highly effective in tumor cell ablation. This demonstrates that our flexible nanoplatform holds great promise for enhancing PDT of tumor., Competing Interests: The authors report no conflicts of interest in this work., (© 2023 Shi et al.)

Background: Cancer cell-derived exosomal microRNAs (miRNAs) play critical role in orchestrating intercellular communication between tumor cells and tumor microenvironmental factors, including lymphatic endothelial cells (LECs). Nevertheless, the functions and underlying mechanisms of exosomal miRNAs in lymphatic metastasis and lymphangiogenesis in esophageal squamous cell carcinoma (ESCC) remain unclear., Methods: Small RNA sequencing, Gene Expression Omnibus (GEO) analysis and qRT‒PCR were performed to identify the candidate exosomal miRNAs involved in ESCC metastasis. Receiver operating characteristic curve analysis was conducted to evaluate the diagnostic potential of exosomal miR-10527-5p in predicting lymph node metastasis (LNM) status. An in vitro coculture system was used to investigate the effects of exosomal miR-10527-5p on ESCC cells and human LECs (HLECs), followed by a popliteal LNM assay in vivo. The relationship between miR-10527-5p and Rab10 was identified by dual-luciferase reporter, fluorescence in situ hybridization and qRT‒PCR assays. Then, a series of rescue assays were performed to further investigate whether Rab10 is involved in exosomal miR-10527-5p mediated ESCC metastasis., Results: MiR-10527-5p was found to be notably reduced in both the plasma exosomes and tumor tissues of ESCC patients with LNM, and plasma exosomal miR-10527-5p had a high sensitivity and specificity for discrimination of LNM status. Moreover, exosome-shuttled miR-10527-5p suppressed the migration, invasion and epithelial-to-mesenchymal transition (EMT) of ESCC cells as well as the migration and tube formation of HLECs via Wnt/β-catenin signaling in vitro and in vivo. Further investigation revealed that Rab10 was a direct target of miR-10527-5p, and re-expression of Rab10 neutralized the inhibitory effects of exosomal miR-10527-5p., Conclusion: Our study demonstrated that exosomal miR-10527-5p had a strong capability to predict preoperative LNM status and anti-lymphangiogenic effect. Exosomal miR-10527-5p inhibited lymphangiogenesis and lymphatic metastasis of ESCC in a vascular endothelial growth factor-C (VEGF-C)-independent manner, showing potential as a therapeutic target for ESCC patients., Competing Interests: The authors declare no conflicts of interest in this work., (© 2023 Xiao et al.)

Background: Nanomedicines have significant advantages in enhancing the oral bioavailability of drugs, but a deeper understanding of the underlying mechanisms remains to be interpreted. Hence, the present study aims to explain the uptake and trafficking mechanism for 6-MP nanomedicines we previously constructed., Methods: 6-MP loaded poly(lactide-co-glycolide) (PLGA) nanomedicines (6-MPNs) were prepared by the multiple emulsion method. The transcytosis mechanism of 6-MPNs was investigated in Caco-2 cells, Caco-2 monolayers, follicle associated epithelium (FAE) monolayers and rats, including transmembrane pathway, intracellular trafficking, paracellular transport and the involvement of transporter., Results: Pharmacokinetics in rats showed that the area under the curve (AUC) of 6-MP in the 6-MPNs group (147.3 ± 42.89 μg/L·h) was significantly higher than that in the 6-MP suspensions (6-MPCs) group (70.31 ± 18.24 μg/L·h). The uptake of 6-MPNs in Caco-2 cells was time-, concentration- and energy-dependent. The endocytosis of intact 6-MPNs was mediated mainly through caveolae/lipid raft, caveolin and micropinocytosis. The intracellular trafficking of 6-MPNs was affected by endoplasmic reticulum (ER)-Golgi complexes, late endosome-lysosome and microtubules. The multidrug resistance associated protein 4 (MRP4) transporter-mediated transport of free 6-MP played a vital role on the transmembrane of 6-MPNs. The trafficking of 6-MPNs from the apical (AP) side to the basolateral (BL) side in Caco-2 monolayers was obviously improved. Besides, 6-MPNs affected the distribution and expression of zona occludens-1 (ZO-1). The transport of 6-MPNs in FAE monolayers was concentration- and energy-dependent, while reaching saturation over time. 6-MPNs improved the absorption of the intestinal Peyer's patches (PPs) in rats., Conclusion: 6-MPNs improve the oral bioavailability through multiple pathways, including active transport, paracellular transport, lymphatic delivery and MRP4 transporter. The findings of current study may shed light on the cellular uptake and transcellular trafficking mechanism of oral nanomedicines., Competing Interests: The authors declare that they have no competing interests in this work., (© 2023 Zou et al.)

Purpose: The translation of nanocarrier-based theranostics into cancer treatment is limited by their poor cellular uptake, low drug-loading capacity, uncontrolled drug release, and insufficient imaging ability., Methods: In this study, novel hybrid nanogels were fabricated as theranostic nanocarriers by modifying chitosan (CTS)/tripolyphosphate (TPP) nanoparticles (NPs) with polyacrylic acid (PAA) and further conjugating cysteine-functionalized gold nanoparticles (AuNPs)., Results: The resultant nanogels, referred to as CTS/TPP/PAA@AuNPs (CTPA), exhibited excellent colloidal stability and a high encapsulation rate of 87% for the cationic drug doxorubicin (DOX). In the tumour microenvironment, the acidic pH and overexpression of lysozyme triggered CTPA@DOX to degrade and emit smaller nanoblocks (30-40 nm), which sequentially released the drug in a tumour-responsive manner. Cellular uptake experiments demonstrated that CTPA facilitates the entry of DOX into the cytoplasm. Furthermore, as visualised through AuNP-mediated computed tomography (CT) imaging, CTPA@DOX enabled favourable accumulation in the tumour. Our in vitro and in vivo data demonstrated that CTPA enabled advanced tumour cell-targeting delivery of DOX, which showed greater anti-tumour activity and biosafety than free DOX., Conclusion: The natural polymer CTS was developed for degradable nanogels, which can precisely track drugs with high antitumour activity. Additionally, the surface adjustment strategy can be assembled to achieve cationic drug loading and high drug-loading capacity, controlled drug release, and sufficient imaging ability. Therefore, multifunctional CTPA enables efficient drug delivery and CT imaging, which is expected to provide a valuable strategy for designing advanced theranostic systems., Competing Interests: The authors declare no conflicts of interest in this work., (© 2022 Liu et al.)

Cancer immunotherapy, a major breakthrough in cancer treatment, has been successfully applied to treat a number of tumors. However, given the presence of factors in the tumor microenvironment (TME) that impede immunotherapy, only a small proportion of patients achieve a good clinical response. With the ability to increase permeability and cross biological barriers, nanomaterials have been successfully applied to deliver immunotherapeutic agents, thus realizing the anti-cancer therapeutic potential of therapeutic agents. This has driven a wave of research into systems for the delivery of immunotherapeutic agents, which has resulted in widespread interest in nanomaterial-based drug delivery systems. Nanomaterial-based drug delivery systems are able to overcome the challenges from TME and thus achieve good results in cancer immunotherapy. If it can make a breakthrough in improving biocompatibility and reducing cytotoxicity, it will be more widely used in clinical practice. Different types of nanomaterials may also have some subtle differences in enhancing cancer immunotherapy. Moreover, delivery systems made of nanomaterials loaded with drugs, such as cytotoxic drugs, cytokines, and adjuvants, could be used for cancer immunotherapy because they avoid the toxicity and side effects associated with these drugs, thereby enabling their reuse. Therefore, further insights into nanomaterial-based drug delivery systems will provide more effective treatment options for cancer patients., Competing Interests: The authors declare that there is no conflict of interest., (© 2022 Jiang et al.)

Purpose: To enhance the osteoinductive effect of Hydroxyapatite (HA) in bone tissue engineering, this study manufactured polycaprolactone (PCL)/gelatin (GEL)/HA nanofibrous scaffolds incorporated with different ratios of attapulgite (ATP): HA (0:3, 0:0, 1:1, 2:1 and 3:0) by high-voltage electrospinning. The synergistic effect exerted by ATP and HA on bone formation was explored both in vivo and in vitro., Methods and Results: First, we determined the group composition and crystal structure of the nanosheets by Fourier transform infrared (FTIR) and X-ray diffraction (XRD) analyses. Then, the physical properties of the scaffolds, including the modulus of elasticity, porosity and water absorption were evaluated. Moreover, the surface microstructure of the nanofibrous scaffolds was captured by Scanning electron microscopy (SEM) and Transmission Electron Microscope (TEM). The biocompatibility of the fabricated scaffolds represented by cell counting kit 8 (CCK-8) and phalloidin staining was also assessed. Next, in vitro osteogenesis was evaluated. Real-time PCR, alkaline phosphatase (ALP) staining and Alizarin red S (ARS) staining results showed that the materials incorporated with HA and ATP at a ratio of 2:1 synergistically promoted more osteoblastic differentiation and extracellular mineralization than scaffolds doped with HA and ATP alone. Last, in vivo, Hematoxylin-Eosin staining (HE staining) and Masson staining showed that groups treated with HA and ATP acquired optimal patterns of bone regeneration., Conclusion: This study clarified for the first time that the combination of HA and ATP orchestrated biomaterial-induced osseointegration, and the synergistic effect was more significant when the ratio of ATP/HA was 2:1. This conclusion also provides new ideas and a scientific basis for the development of functionalized nanomaterials in bone tissue engineering., Competing Interests: The authors declare no conflicts of interest., (© 2022 Liu et al.)

Background: Satisfactory prognosis of breast cancer (BC) is limited by difficulty in early diagnosis and insufficient treatment. The combination of molecular imaging and photothermal therapy (PTT) may provide a solution., Methods: Fe 3 O 4 -Au shell nanoparticles (NPs) were prepared by thermal decomposition, seeded growth and galvanic replacement and were comprehensively characterized. After conjugated to PEG, NPs were used as MRI and PTT agents in vitro and in vivo., Results: Fe 3 O 4 -Au shell NPs which had uniform Janus-like morphology were successfully synthesized. The Fe 3 O 4 had a size of 18 ± 2.2 nm, and the Au shell had an outer diameter of 25 ± 3.3 nm and an inner diameter of 20 ± 2.9 nm. The NPs showed superparamagnetism, a saturation magnetization of 36 emu/g, and an optical absorption plateau from 700 to 808 nm. The Fe 3 O 4 -Au shell NPs were determined to possess good biocompatibility. After PEG coating, the zeta potential of NPs was changed from -23.75 ± 1.37 mV to -13.93 ± 0.55 mV, and the FTIR showed the characteristic C-O stretching vibration at 1113 cm -1 . The NPs' MR imaging implied that the T 2 can be shortened by Fe 3 O 4 -Au shell NPs in a concentration-dependent manner, and the Fe 3 O 4 -Au shell NPs coated with PEG at the molar ratio of 160 (PEG: NPs) showed the highest transverse relaxivity ( r 2 ) of 216 mM -1 s -1 . After irradiation at 0.65 W/cm 2 for 5 min, all cells incubated with the Fe 3 O 4 -Au shell -PEG160 NPs (Fe: 30 ppm, Au: 70 ppm) died. After administrated intratumorally, Fe 3 O 4 -Au shell -PEG160 notably decreased the signal intensity of tumor in T 2 WI images. Under the same irradiation, the temperature of tumors injected with Fe 3 O 4 -Au shell -PEG160 quickly rose to 54.6°C, and the tumors shrank rapidly and were ablated in 6 days., Conclusion: Fe 3 O 4 -Au shell -PEG NPs show good r 2 and PTT performance and are promising synergistic theranostic agents of MRI and PTT for BC., Competing Interests: The authors report no conflicts of interest in this work., (© 2021 Kang et al.)

Background: High levels of oxidants, such as reactive oxygen species (ROS) and reactive nitrogen species (RNS), are typical characteristics of an inflammatory microenvironment and are closely associated with a various inflammatory pathologies, eg, cancer, diabetes, atherosclerosis, and neurodegenerative diseases. Therefore, the delivery of anti-inflammatory drugs by oxidation-responsive smart systems would be an efficient anti-inflammatory strategy that benefits from the selective drug release in an inflammatory site, a lower treatment dose, and minimizes side effects., Purpose: In this study, we present the feasibility of an oxidation-sensitive PEGylated alternating polyester, methoxyl poly(ethylene glycol)- block -poly(phthalic anhydride- alter -glycidyl propargyl ether) (mPEG- b -P(PA- alt -GPBAe)), as novel nanocarrier for curcumin (CUR), and explore the application in anti-inflammatory therapy., Methods: The copolymers used were obtained by combining a click reaction and a ring-opening-polymerization method. CUR was loaded by self-assembly. The in vitro drug release, cytotoxicity toward RAW 264.7 cells and cellular uptake were investigated. Furthermore, the anti-inflammatory effects of CUR-loaded polymeric nanoparticles (NPs-CUR) were investigated in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages and tested in a murine model of ankle inflammation., Results: Fast drug release from NPs-CUR was observed in trigger of 1 mM H 2 O 2 in PBS. Compared with NPs and free drugs, the significant anti-inflammatory potential of NPs-CUR was proven in activated RAW 264.7 cells by inhibiting the production of TNF-α, IL-1β, and IL-6 and increasing the level of an anti-inflammatory cytokine IL-10. Finally, a local injection of NPs-CUR at a dose of 0.25 mg/kg suppressed the acute ankle inflammatory response in mice by histological observation and further reduced the expression of pro-inflammatory cytokines in the affected ankle joints compared to that of free CUR., Conclusion: Both the significant in vitro and in vivo anti-inflammatory results indicated that our oxidation responsive polymeric nanoparticles are promising drug delivery systems for anti-inflammatory therapy., Competing Interests: The authors report no conflicts of interest in this work., (© 2021 He et al.)

Introduction: Topical agents typically remain in the wound site for time duration that are too short to effectively eradicate MRSA tradition formation of BZK that can be maintained within the wound site for longer time periods, should be more effective., Methods: The novel chitosan and poly (D,L-lactide-co-glycoside) nanoparticles loaded with benzalkonium bromide (BZK) were designed, for the promotion wound healing after MRSA infection. The physical characterization of these nanoparticles, as well as their antibacterial activity in vitro, release profile in simulated wound fluid, cell toxicity, anti-biofilm activity, and their ability to improve the skin wound healing in a mouse model were also studied., Results: These novel nanoparticles were found to have a significant antibacterial activity ( p <0.01), both in vitro and in vivo test. The stronger anti-biofilm ability of the nanoparticles to inhibit the formation of bacterial biofilms, at a concentration of 3.33 μg/mL, and clear existing bacterial biofilms, at a concentration of 5 mg/mL, compared with its water solution. In addition, significant damage to bacterial cell walls also was found, providing insight into the mechanism of antibacterial activity., Conclusion: Taken together, these results demonstrated the ability of BZK-loaded nanoparticles in the promotion of skin wound healing with MRSA infection. The current findings open a new avenue for nanomedicine development and future clinical applications in the treatment of wounds., Competing Interests: The authors declared that there is no conflict of interest., (© 2021 Wu et al.)

Purpose: Malignant melanoma is one of the most devastating types of cancer with rapid relapse and low survival rate. Novel strategies for melanoma treatment are currently needed to enhance therapeutic efficiency for this disease. In this study, we fabricated a multifunctional drug delivery system that incorporates dacarbazine (DTIC) and indocyanine green (ICG) into manganese-doped mesoporous silica nanoparticles (MSN(Mn)) coupled with magnetic resonance imaging (MRI) and photothermal imaging (PI), for achieving the superior antitumor effect of combined chemo-photothermal therapy., Materials and Methods: MSN(Mn) were characterized in terms of size and structural properties, and drug loading and release efficiency MSN(Mn)-ICG/DTIC were analyzed by UV spectra. Photothermal imaging effect and MR imaging effect of MSN(Mn)-ICG/DTIC were detected by thermal imaging system and 3.0 T MRI scanner, respectively. Then, the combined chemo-phototherapy was verified in vitro and in vivo by morphological evaluation, ultrasonic and pathological evaluation., Results: The as-synthesized MSN(Mn) were characterized as mesoporous spherical nanoparticles with 125.57±5.96 nm. MSN(Mn)-ICG/DTIC have the function of drug loading-release which loading ratio of ICG and DTIC could reach to 34.25±2.20% and 50.00±3.24%, and 32.68±2.10% of DTIC was released, respectively. Manganese doping content could reach up to 65.09±2.55 wt%, providing excellent imaging capability in vivo which the corresponding relaxation efficiency was 14.33 mM -1 s -1 . And outstanding photothermal heating ability and stability highlighted the potential biomedical applicability of MSN(Mn)-ICG/DTIC to kill cancer cells. Experiments by A375 melanoma cells and tumor-bearing mice demonstrated that the compound MSN(Mn)-ICG/DTIC have excellent biocompatibility and our combined therapy platform delivered a superior antitumor effect compared to standalone treatment in vivo and in vitro., Conclusion: Our findings demonstrate that composite MSN(Mn)-ICG/DTIC could serve as a multifunctional platform to achieve a highly effective chemo-photothermal combined therapy for melanoma treatment., Competing Interests: The authors declare no competing financial interest in this work., (© 2021 Zhang et al.)

Purpose: Polyetheretherketone (PEEK) exhibits high mechanical strengths and outstanding biocompatibility but biological inertness that does not excite the cell responses and stimulate bone formation. The objective of this study was to construct submicro-nano structures on PEEK by femtosecond laser (FSL) for exciting the responses of MC3T3-E1 cells and gingival epithelial (GE) cells, which induce regeneration of bone/gingival tissues for long-term stability of dental implants., Materials and Methods: In this study, submicro-nano structures were created on PEEK surface by FSL with power of 80 mW (80FPK) and 160 mW (160FPK)., Results: Compared with PEEK, both 80FPK and 160FPK with submicro-nano structures exhibited elevated surface performances (hydrophilicity, surface energy, roughness and protein absorption). Furthermore, in comparison with 80FPK, 160FPK further enhanced the surface performances. In addition, compared with PEEK, both 80FPK and 160FPK significantly excited not only the responses (adhesion, proliferation, alkaline phosphatase [ALP] activity and osteogenic gene expression) of MC3T3-E1 cells but also responses (adhesion as well as proliferation) of GE cells of human in vitro. Moreover, in comparison with 80FPK, 160FPK further enhanced the responses of MC3T3-E1 cells/GE cells., Conclusion: FSL created submicro-nano structures on PEEK with elevated surface performances, which played crucial roles in exciting the responses of MC3T3-E1 cells/GE cells. Consequently, 160FPK with elevated surface performances and outstanding cytocompatibility would have enormous potential as an implant for dental replacement., Competing Interests: The authors report no conflicts of interest in this work., (© 2021 Xie et al.)

Background: Manganese Ferrite Nanoparticles (Mn-IONPs) are widely used in biomedical field and their cytotoxicity has been initially explored, but the mechanism remains obscure. The nano-bio interactions are believed to be crucial for cytotoxicity mechanism, while little data have been acquired., Methods: Mn-IONPs were synthesized by thermal decomposition of acetylacetonate precursor. After physicochemical characterization, we analyzed the metabolic conversion and removal of Mn-IONPs in RAW264.7 cells by Prussian blue staining, TEM, HRTEM and elemental quantitative analysis, followed by gene expression evaluation using quantitative RT-PCR., Results: Mn-IONPs were successfully synthesized. Both the uptake and cytotoxicity of Mn-IONPs on RAW264.7 cells were time- and dose-dependent. After internalized, Mn-IONPs were passed to daughter cells with passages on. Meanwhile, Mn-IONPs were exocytosed and digested to metal ions and further excreted out, resulted in the labeling rate and ions contents decreased gradually. As ion influx related genes, the expressions of ZIP14, IRP2, FtH and DMT1 were suppressed within 24 hours but overexpressed to a plateau at the 48th hour in a dose-dependent manner. At the 72nd hour, ZIP14 and DMT1 mRNA levels decreased toward normal, while IRP2 and FtH kept up-regulated. As efflux related genes, FPN, SLC30A10 and Hamp2 genes were up-regulated within 24-72 hours; SPCA1 was suppressed at the 24th and 72nd hour, while overexpressed at the 48th hour. All the efflux related genes' mRNA had a dose-dependent increasing manner at the corresponding time points., Conclusion: Mn-IONPs showed time- and dose-dependent cytotoxicity and cell labeling rate in RAW264.7 cells. Accompanying with the intracellular catabolic breakdown and exocytosis of Mn-IONPs, RAW264.7 cells also secreted and re-uptook manganese and iron ions to maintain intracellular homeostasis in the succeeding passages. And the metabolic conversion of Mn-IONPs in RAW264.7 cells can affect the expression of ZIP14, DMT1, FPN, SLC30A10, IRP2, FtH, Hamp2 and SPCA1 genes., Competing Interests: The authors report no conflicts of interest in this work., (© 2021 Zhang et al.)

Background: Intracellular tension plays a crucial role in the destruction of the blood-brain barrier (BBB) in response to lesion stimuli. Tight junction structure could be primarily affected by tension activity. In this study, we aimed to determine the effects of extracellular BBB damage on intracellular tension activity, and elucidate the mechanism underlying the effects of intracellular protein nanoparticle-related osmotic pressure on BBB permeability., Methods: The intracellular tension for tight junction proteins occludin and ZO1 was evaluated using the fluorescence resonance energy transfer (FRET)-based tension probes and cpstFRET analysis. The changes in mobility ratios of occludin were evaluated via the fluorescence recovery after photobleaching (FRAP) test. The cytoplasmic osmotic pressure (OP) was measured using Osmometer. The count rate of cytoplasmic nanoparticles was detected by Nanosight NS300. The activation of cofilin and stathmin was examined by Western blot analysis. The BBB permeability in vivo was determined via the changes of Evans Blue (EB) injected into SD rats. The tight junction formation was assessed by the measurement of transendothelial electrical resistance (TEER). Intracellular calcium or chloride ions were measured using Fluo-4 AM or MQAE dyes., Results: BBB lesions were accompanied by changes in occludin/ZO1 tension. Increases in intracellular osmotic pressure were involved in alteration of BBB permeability, possibly through the depolymerization of microfilaments or microtubules and mass production of protein nanoparticles according to the Donnan effect. Recovery of protein nanoparticle-related osmotic pressure could effectively reverse the effects of changes in occludin/ZO1 tension under BBB lesions. Outward tension of intracellular osmotic potential also caused upregulation of membrane fluidity, which promoted nonselective drug influx., Conclusion: Our results suggest a crucial mechanical mechanism underlying BBB lesions, and protein nanoparticle-related osmotic pressure could be a novel therapeutic target for BBB lesion-related brain diseases., Competing Interests: The authors declare that they have no competing interests., (© 2021 Li et al.)

Background: Acute lymphoblastic leukemia (ALL) is the most common hematologic malignancy in children. It requires a long and rigorous course of chemotherapy treatments. 6-Mercaptopurine (6-MP) is one of the primary drugs used in chemotherapy. Unfortunately, its efficacy has been limited due to its insolubility, poor bioavailability and serious adverse effects. To overcome these drawbacks, we constructed 6-mercaptopurine (6-MP)-loaded nanomedicines (6-MPNs) with biodegradable poly(lactide-co-glycolide) (PLGA) to enhance the anticancer efficacy of 6-MP., Methods: We prepared the 6-MPNs using a double-emulsion solvent evaporation method, characterizing them for the physicochemical properties. We then investigated the plasma, intestinal region and other organs in Sprague Dawley (SD) rats for pharmacokinetics. Additionally, we evaluated its anticancer efficacy in vitro on the human T leukemia cell line Jurkat and in vivo on the ALL model mice., Results: The 6-MPNs were spherical in shape with uniform particle size and high encapsulation efficiency. The in vitro release profile showed that 6-MPNs exhibited a burst release that a sustained release phase then followed. The apoptosis assay demonstrated that 6-MPNs could improve the in vitro cytotoxicity in Jurkat cells. Pharmacokinetics profiles revealed that 6-MPNs had improved oral bioavailability. Tissue distribution experiments indicated that 6-MPNs increased the duodenum absorption of 6-MP, at the same time having a low accumulation of the toxic metabolites of 6-MP. The in vivo pharmacodynamics study revealed that 6-MPNs could prolong the survival time of the ALL model mice. The prepared 6-MPNs, therefore, have superior properties in terms of anticancer efficacy against ALL with reduced systemic toxicity., Conclusion: Our nanomedicines provide a promising delivery strategy for 6-MP; they offer a simple preparation method and high significance for clinical translation., Competing Interests: Yaru Zou and Dong Mei are co-first authors. The authors declare that they have no competing interests., (© 2021 Zou et al.)

Background: Intracerebral hemorrhage (ICH) is a common neurological crisis leading to high mortality and morbidity. Oxidative stress-induced secondary injury plays a critical role in neurological deterioration. Previously, we synthesized a porous Se@SiO 2 nanocomposite and identified their therapeutic role in osteonecrosis of the femoral head. Whether this nanocomposite is neuroprotective remains to be elucidated., Methods: A porous Se@SiO 2 nanocomposite was synthesized, and its biosafety was determined using a CCK-8 assay. The neuroprotective effect was evaluated by TUNEL staining, and intracellular ROS were detected with a DCFH-DA probe in SH-SY5Y cells exposed to hemin. Furthermore, the effect of the nanocomposite on cell apoptosis, brain edema and blood-brain barrier permeability were evaluated in a collagenase-induced ICH mouse model. The potential mechanism was also explored., Results: The results demonstrated that Se@SiO 2 treatment significantly improved neurological function, increased glutathione peroxidase activity and downregulated malonaldehyde levels. The proportion of apoptotic cells, brain edema and blood-brain barrier permeability were reduced significantly in ICH mice treated with Se@SiO 2 compared to vehicle-treated mice. In vitro, Se@SiO 2 protected SH-SY5Y cells from hemin-induced apoptosis by preventing intracellular reactive oxygen species accumulation., Conclusion: These results suggested that the porous Se@SiO 2 nanocomposite exerted neuroprotection by suppressing oxidative stress. Se@SiO 2 may be a potential candidate for the clinical treatment of ICH and oxidative stress-related brain injuries., Competing Interests: The authors declare no conflicts of interest for this work., (© 2021 Yang et al.)

Purpose: To synthesize echogenic chitosan/perfluorohexane nanodroplets (CNDs) for DKK-2 gene delivering in a spatiotemporally controlled manner in vitro., Methods: The characteristics, contrast-enhanced ultrasound imaging, DNA binding and DNase protection capacity, DKK-2 gene transfection and effects on LNCaP cells of these CNDs were investigated., Results: The obtained CNDs showed positive surface charges and could attract the genetic cargo with negative surface charges to form nanocomplexes. Agarose gel electrophoresis confirmed binding of the CNDs and pDNA. DKK-2 pDNA-loaded CNDs, in combination with ultrasound, ruptured and released DKK-2 pDNA, entering LNCaP cells through nano-scale pores in the cell membrane, which further reduced the proliferation of LNCaP cells., Conclusion: These stable and safe CNDs may be a promising choice to achieve efficient ultrasound-mediated gene delivery to specific tissues in a spatiotemporally controlled manner., Competing Interests: Xinxin Liu and Jie Li report a patent (Application of ultrasound-sensitive chitosan nanodroplets combined Dickkopf-2 Gene for the treatment of prostate cancer). The authors report no other potential conflicts of interest in this work., (© 2021 Liu et al.)

Background: Radiotherapy (RT), one of the main treatments for cervical cancer, has tremendous potential for improvement in the efficacy. Poly (ADP-ribose) polymerase (PARP) is a key enzyme in the repair of DNA strand breaks (DSB). Olaparib (Ola) is a PARP inhibitor that is involved in preventing the release of PARP from RT-induced damaged DNA to potentiate the effect of RT. Although the basic mechanism of Ola's radiosensitization is well known, the radiosensitization mechanism of its nanomedicine is still unclear. In addition, the lack of tumor tissue targeting is a major obstacle for the clinical success of Ola., Materials and Methods: In this study, we developed folate-conjugated active targeting olaparib nanoparticles (ATO) and investigated the anti-tumor effect of ATO combined with radiotherapy (RT) in nude mice using cervical cancer xenograft models. We used folate (FA)-conjugated poly (ε-caprolactone)-poly (ethyleneglycol)-poly (e-caprolactone) (PCEC) copolymer to prepare ATO via emulsification/solvent diffusion. Further, we evaluated ATO particle size, potential, encapsulation efficiency, and in vitro release characteristics, and evaluated the shape of ATO via transmission electron microscopy (TEM). We then performed MTT and cell uptake assays to detect cytotoxicity and targeting uptake in vitro. We investigated the anti-tumor properties of ATO in vivo by apoptosis test, 18 F-FDG PET/CT, and immunohistochemical analysis. Finally, the xenografted tumor in nude mice was subjected to RT and/or ATO treatment., Results: The results confirmed that ATO in combination with RT significantly inhibited tumor growth and prolonged survival time of tumor-bearing mice. This may be related to the inhibition of tumor proliferation and DNA damage repair and induction of cell apoptosis in vivo., Conclusion: The ATO developed in this study may represent a novel formulation for olaparib delivery and have promising potential for treating tumors with an over-expression of folate receptors., Competing Interests: The authors report no conflicts of interest in this work., (© 2020 Li et al.)

Objective: This study aimed to develop aptamer-anchored hyperbranched poly(amido amine) (HPAA) for the systemic delivery of miRNA-133a-3p and to evaluate its therapeutic potential against bone metastasis of prostate cancer in vivo and in vitro., Methods: A glutathione (GSH)-responsive cationic HPAA was prepared by the Michael addition reaction. Furthermore, HPAA-PEG was produced by PEGylation, and then the aptamer targeted to prostate-specific membrane antigen (PSMA) was conjugated to the HPAA-PEG. The obtained HPAA-PEG-APT could form nanocomplexes with miRNA-133a-3p through electrostatic adsorption., Results: The results of immunocytochemistry indicated that the complexes could target PSMA-expressing LNCaP cells. The ability of HPAA-PEG-APT to facilitate the delivery of miRNA-133a-3p into LNCaP cells was proven, and HPAA-PEG-APT/ miRNA-133a-3p demonstrated enhanced antitumor activity, lower cytotoxicity and better biocompatibility in vitro. Moreover, in a mouse tibial injection tumor model, the intravenous injection of the HPAA-PEG-APT/ miRNA-133a-3p complex significantly inhibited cancer growth and extended the survival time., Conclusion: This study provided an aptamer-anchored HPAA-loaded gene system to deliver miRNA-133a-3p for better therapeutic efficacy of bone metastasis of prostate cancer., Competing Interests: The authors report no conflicts of interest for this work., (© 2020 Ye et al.)

Background: Electrospinning is a widely used technology that can produce scaffolds with high porosity and surface area for bone regeneration. However, the small pore sizes in electrospun scaffolds constrain cell growth and tissue-ingrowth. In this study, novel drug-loading core-shell scaffolds were fabricated via electrospinning and freeze drying to facilitate the repair of tibia bone defects in rabbit models., Materials and Methods: The collagen core scaffolds were freeze-dried containing icariin (ICA)-loaded chitosan microspheres. The shell scaffolds were electrospun using collagen, polycaprolactone and hydroxyapatite materials to form CPH composite scaffolds with the ones containing ICA microspheres named CPHI. The core-shell scaffolds were then cross-linked by genipin. The morphology, microstructure, physical and mechanical properties of the scaffolds were assessed. Rat marrow mesenchymal stem cells from the wistar rat were cultured with the scaffolds. The cell adhesion and proliferation were analysed. Adult rabbit models with tibial plateau defects were used to evaluate the performance of these scaffolds in repairing the bone defects over 4 to 12 weeks., Results: The results reveal that the novel drug-loading core-shell scaffolds were successfully fabricated, which showed good physical and chemical properties and appropriate mechanical properties. Furthermore, excellent cells attachment was observed on the CPHI scaffolds. The results from radiography, micro-computed tomography, histological and immunohistochemical analysis demonstrated that abundant new bones were formed on the CPHI scaffolds., Conclusion: These new core-shell composite scaffolds have great potential for bone tissue engineering applications and may lead to effective bone regeneration and repair., Competing Interests: The authors report no conflicts of interest in this work., (© 2020 Zhao et al.)

Purpose: To synthesize and evaluate a novel folate-conjugated ultrasonic nanobubble (HA-FOL-NB) loading low-molecular-weight hyaluronic acid (LMW-HA) for specific tumor-associated macrophages (TAMs) targeting and reeducation., Methods: The characteristics, cytotoxicity, contrast-enhanced ultrasound imaging (CEUS), and targeting ability to TAMs of HA-FOL-NBs were investigated. The TAMs reprogramming function of HA-FOL-NBs combining ultrasound targeted nanobubble destruction was assessed as well., Results: HA-FOL-NBs (about 342 nm) showed remarkable contrast enhancement images, and higher targeting ability due to the folate to folate receptor interactions. Combined with ultrasound targeted nanobubble destruction, HA-FOL-NBs could specifically deliver LMW-HA into TAMs, thus exhibited stronger reeducation effect compared with free LMW-HA., Conclusion: These folate-conjugated and LMW-HA-loaded nanobubbles, with targeted CEUS imaging and TAMs reeducation, are expected to be a potential approach for tumor therapy based on TAMs, especially folate receptor-positive ones., Competing Interests: The authors report no conflicts of interest in this work., (© 2020 Sun et al.)

Background: Synergistic chemoradiotherapy (CRT) has become a primary effective curative approach for many solid cancers. However, CRT is still associated with several obstacles, including the increases in side effects and systemic toxicity. Incorporating nanocarriers into CRT is a new and exciting approach to solve these obstacles. The purpose of the present study was to design a unique pH- and ultrasound-responsive perfluoropentane-encapsulated, paclitaxel (PTX)-loaded carboxymethyl chitosan nanodroplets (NDs) for combined imaging and synergistic CRT., Materials and Methods: The NDs were prepared by a homogenization/emulsion method. Their physicochemical properties, echogenicity and biocompatibility were evaluated. PTX-loaded NDs with a high loading efficiency and encapsulation efficiency were prepared and their pH-responsive drug release profile was determined by dialysis sack method. Then, PC3 cells were exposed to (1) PTX (4 μg/mL), (2) NDs (30 μg/mL), (3) PTX-loaded NDs (34 μg/mL), (4) RT (6 Gy), (5) RT (10 Gy), (6) combination of PTX (4 μg/mL), ultrasound (0.5 W/cm 2 , 30 s) and RT (6 Gy), (7) combination of NDs (30 μg/mL), ultrasound (0.5 W/cm 2 , 30 s) and RT (6Gy), (8) combination of PTX-loaded NDs (30 μg/mL), ultrasound (0.5 W/cm 2 , 30 s) and RT (6 Gy). 24 hrs later, CCK-8 assay, flow cytometry and migration assay were carried out to evaluate their therapeutic effects in CRT., Results: The desired NDs were successfully prepared, which were with round, spherical shapes, relatively smooth surfaces, core-shell structures and uniform in sizes (<300 nm with PDI<0.3 when at pH≧6.0). The NDs exhibited good abilities in pH-dependent charge conversion, biocompatibility and ultrasound contrast echogenicity. The in vitro drug release from PTX-loaded NDs (the highest loading efficiency and encapsulation efficiency were 20.35% and 91.58%) was pH dependent and exhibited an initial burst followed by a sustained drug release. The results of the CCK-8 assay, flow cytometry and migration assay all showed PTX-loaded NDs combined ultrasound and RT significantly enhanced cell responses in CRT., Conclusion: The pH- and ultrasound-responsive PTX-loaded NDs, which exhibited a high echogenicity, drug delivery ability and radiosensitization ability, could be a feasible option for combined imaging and novel enhancing approach in synergistic CRT., Competing Interests: The authors declare no conflicts of interest in this work., (© 2020 Shang et al.)

Introduction: Though widely studied for biomedical applications, the lack of current systemic studies on the in vivo fate of single-walled carbon nanohorns (SWCNHs) largely restricts their further applications, as real-time monitoring of their biodistribution remains a big challenge. Here, we aim to customize a label-free multispectral optoacoustic tomography (MSOT) method and systematically survey the fate of oxidized SWCNHs (SWCNHox) following different exposure routes by whole body imaging., Methods: Mice were given a suspension of SWCNHox with an average size of 136.4 nm via four different administration routes, and then imaged by MSOT., Results: After oral gavage, SWCNHox were mainly distributed in the gastrointestinal tract then excreted through the gut. Compared with the observation post first dosing, the accumulation of SWCNHox in the gastrointestinal tract was not obvious even after four-time oral gavage. Almost no SWCNHox were found at detectable levels in kidney, liver, blood and spleen. Following intravenous (iv) injection, SWCNHox were mainly presented and persisted in the spleen and liver, while very little in the kidney and almost none detectable in the intestine. SWCNHox accumulated significantly in the liver and spleen after four IV administrations. Following hypodermic and intramuscular injections, almost no SWCNHox could cross biological barriers and transport to the spleen, kidney or liver, likely due to their very low absorption rate. Almost all SWCNHox remained around the injection sites. For the first time, we have systematically investigated the in vivo fate of SWCNHs in a label-free and real-time manner., Conclusion: The findings of this study provide insights into the selection of appropriate exposure routes for potential biomedical applications of carbon nanomaterials., Competing Interests: The authors report no conflicts of interest in this work., (© 2019 Shi et al.)

Background: As an active ingredient of Chinese herbal medicine, quercetin (QU) can significantly induce apoptosis of tumor cells and give play to other effect such as decreasing both fibroblast population and collagen in cancer cell nest. However, the antitumor efficacy of quercetin was mostly evaluated at cellular level and rarely developed in vivo by intravenous injection, which may be ascribed to its inferior physicochemical properties including water insolubility, short plasma half-time, and insufficient enrichment in the tumor tissues., Methods: The DSPE-PEG was used to construct quercetin-loaded micelles, and the integrin ligand cRGDfK was grafted to modify the nanocarrier for enhancing its cancer-specific homing. The MALDI-TOF-MS, DLS, TEM, and UV were orderly operated to characterize guidance molecules and micelles by morphology, size distribution, Zeta potential, and drug encapsulation efficiency. In addition, the surface plasmon resonance study and real-time confocal analysis were employed to demonstrate α v β 3 integrin-overexpressing B16 cells-specific binding and uptake. After further pharmacodynamics studies in vitro and in vivo, we also evaluate systemic toxicity about cRGDfK-PM-QU., Results: The cRGDfK was successfully stitched with DSPE-PEG and modified on the surface of micelles. The ligand modification enhanced the negative charges of the micelles, but it did not induce significant changes in particle size. The quercetin micelles were about 15 nm in size and negatively charged, and had spherical morphology and high drug encapsulation efficiency. In vitro, the cRGDfK-modified micelles (cRGDfK-PM) showed α v β 3 integrin-overexpressing B16 cells-specific binding and uptake, and cRGDfK-PM-QU (QU loaded in cRGDfK-PM) induced more significant cell apoptosis and cytotoxic effects against B16 tumor cells than counterpart micelles (PM-QU). In vivo, the cRDGfK modification enhanced enrichment in B16 tumor tissue, improved the therapeutic efficacy of the quercetin-loaded micelles against B16 tumor, and exhibited lower systemic and pulmonary toxicity compared with counterpart micelles in the mouse mode., Conclusion: Quercetin as a natural product has triggered increasing interest in the antitumor field. In this study, cRGDfK-modified DSPE-PEG micelles significantly optimized quercetin therapeutic efficacy and pulmonary toxicity as well as lowered systemic toxicity., Competing Interests: The authors report no conflicts of interest in this work., (© 2019 Xu et al.)

Background: Beyond magnetic resonance imaging (MRI), which has been widely used clinically, molecular MRI (mMRI) can further provide qualitative and quantitative information at the cellular and molecular levels. However, the diagnostic accuracy may not be satisfactory via single-contrast mMRI due to some interferences in vivo. T 1 /T 2 dual-contrast MRI using the same contrast agent (CA) could significantly improve the detection accuracy. Therefore, in this study, we fabricated poly(ethylene glycol) (PEG)-coated, manganese-doped iron oxide nanocomposites (Mn-IONPs@PEG) as T 1 /T 2 dual-contrast CA, and evaluated its feasibility of T 1 /T 2 dual-contrast MRI in vitro and in vivo., Methods: Mn-IONPs were prepared by the thermal decomposition of iron-eruciate and manganese-oleate complexes and were coated with 1,2-Distearoyl-sn-glycero-3-phosphoethanolamine-N-(methoxy[polyethylene glycol]-2000) (DSPE-PEG 2000). The physicochemical properties and cytotoxicity of the Mn-IONPs were fully characterized, followed by MRI in vitro and in vivo., Results: Ultrasmall 3 nm-sized nanoparticles were successfully prepared and were identified using transmission electron microscopy (TEM), high-resolution TEM, and X-ray diffraction. After coating with DSPE-PEG, the Mn-IONPs@PEG displayed excellent hydrophilicity and good biocompatibility. Due to the manganese-doping and PEG coating, the Mn-IONPs@PEG showed good relaxivity in vitro. Especially, the Mn-IONPs@PEG coated with DSPE-PEG following a mass ratio to Mn-IONPs of 1:20 showed harmonious longitudinal relaxivity ( r 1 = 7.1 mM -1 s -1 ) and transversal relaxivity ( r 2 = 120.9 mM -1 s -1 ), making it a better candidate for T 1 /T 2 dual-contrast mMRI. After administrated via a caudal vein, the Mn-IONPs@PEG can induce significant enhancement in both T 1 -weighted and T 2 -weighted MR images and the time at 10 mins after injection was regarded as a suitable time for imaging because both the T 1 and T 2 enhancement were optimum at that time., Conclusion: The obtained Mn-IONPs@PEG exhibited good r 1 and r 2 and was a reasonable candidate for T 1 /T 2 dual-contrast mMRI., Competing Interests: The authors report no conflicts of interest in this work., (© 2019 Xiao et al.)

Background: Poly (ADP-ribose) polymerase (PARP) is a key enzyme in the repair process of DNA strand breaks (DSBs). Olaparib (Ola) is a PARP inhibitor that is involved in arresting PARP release from radiotherapy (RT)-induced damaged DNA to potentiate the effect of RT. Although the underlying mechanisms for the radiosensitization effects of Ola are well understood in vitro, the radiosensitization effects in vivo are still unclear. Moreover, poor water solubility and severe toxicity are two major impediments for the clinical success of Ola., Materials and Methods: Here, we developed olaparib nanoparticles (Ola-NPs) and investigated their radiosensitization mechanisms and toxicity using human non-small-cell lung cancer xenograft models in mice., Results: The prepared Ola-NPs showed a mean size of 31.96±1.54 nm and a lower polydispersity index of about 0.126±0.014. In addition, the sensitization enhancement ratio of Ola-NPs (3.81) was much higher than that of free Ola (1.66). The combination of Ola-NPs and RT (Ola-NPs + RT) significantly inhibited tumor growth and prolonged survival in mice. The mechanism of enhanced antitumor efficacy might be related to the inhibition of DSB repair and the promotion of cell apoptosis in vivo. No additional toxicity caused by Ola-NPs was observed., Conclusion: This study demonstrated the principle of using Ola-NPs as a potent radiosensitizer to improve the therapeutic effect of RT relative to free Ola ( P <0.05 in all cases)., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Background: Red blood cell membrane-coated nanoparticle (RBCm-NP) platform, which consist of natural RBCm and synthetic polymeric core, can extend circulation time in vivo with an improved biocompatibility and stability of this biomimetic nanocarrier. To achieve better bioavailability of antitumor drugs that were loaded in RBCm-NPs, the functionalization of coated RBCm with specific targeting ability is essential. Bispecific recombinant protein anti-EGFR-iRGD, containing both tumor penetrating peptide (internalizing RGD peptide) and EGFR single-domain antibody (sdAb), seems to be an optimal targeting ligand for RBCm-NPs in the treatment of multiple tumors, especially colorectal cancer with high EGFR expression., Materials and Methods: We modified the anti-EGFR-iRGD recombinant protein on the surface of RBCm-NPs by lipid insertion method to construct iE-RBCm-PLGA NPs and confirmed the presentation of active tumor-targeting ability in colorectal cancer models with high EGFR expression when compared with RBCm-PLGA NPs. In addition, potential anti-tumor drug gambogic acid (GA) was loaded into the NPs to endow the antitumor efficiency of iE-RBCm-GA/PLGA NPs. It was simultaneously evaluated whether GA can reach better biocompatibility benefiting from the improved antitumor efficiency of iE-RBCm-GA/PLGA NPs in colorectal cancer models., Results: We successfully modified anti-EGFR-iRGD proteins on the surface of biomimetic NPs with integrated and stable "shell-core" structure. iE-RBCm-PLGA NPs showed its improved targeting ability in vitro (multicellular spheroids [MCS]) and in vivo (nude mice bearing tumors). Besides, no matter on short-term cell apoptosis at tumor site (terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling [TUNEL]) and long-term tumor inhibition, iE-RBCm-GA/PLGA NPs achieved better antitumor efficacy than free GA in spite of the similar effects of cytotoxicity and apoptosis to GA in vitro., Conclusion: We expect that the bispecific biomimetic nanocarrier can extend the clinical application of many other potential antitumor drugs similar to GA and become a novel drug carrier in the colorectal cancer treatment., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Purpose: Ultrasound (US) molecular imaging provides a non-invasive way to visualize tumor tissues at molecular and cell levels and could improve diagnosis. One problem of using US molecular imaging is microbubbles challenges, including instability, short circulation time, and poor loading capacity and penetrability. It is urgent to design new acoustic contrast agents and new imaging methods to facilitate tumor-targeted imaging. In this study, phase-shift poly lactic-co-glycolic acid (PLGA) nanoparticles modified with folate as an efficient US molecular probe were designed and the long-term targeted imaging was achieved by low-intensity focused US (LIFU) irradiation., Methods: A new 5-step method and purification procedure was carried out to obtain uniform folic acid polyethylene glycol PLGA (PLGA-PEG-FA), the structure of which was confirmed by 1 H nuclear magnetic resonance spectroscopy and thin-layer chromatography. Perflenapent (PFP) was wrapped in PLGA-PEG-FA by a double emulsion solvent evaporation method to obtain PFP/PLGA-PEG-FA nanoparticles. The targeted ability of the resulting nanoparticles was tested in vivo and in vitro. LIFU irradiation can irritate nanoparticle phase-shift to enhance tumor imaging both in vivo and in vitro., Results: PLGA-PEG-FA was a light yellow powder with a final purity of at least 98%, the structure of which was confirmed by 1 H nuclear magnetic resonance spectroscopy and thin-layer chromatography. Highly dispersed PFP/PLGA-PEG-FA nanoparticles with spherical morphology have an average diameter of 280.9±33.5 nm, PFP load efficiency of 59.4%±7.1%, and shells, thickness of 28±8.63 nm. The nanoparticles can specifically bind to cells expressing high folate receptor both in vivo and in vitro. Ultrasonic imaging was significantly enhanced in vitro and in vivo by LIFU irradiation. The retention time was significantly prolonged in vivo., Conclusion: Phase-shift PFP/PLGA-PEG-FA nanoparticles induced by LIFU can significantly enhance ultrasonic imaging, specifically targeting tumors expressing folate receptor. As a potential targeting acoustic molecular probe, PFP/PLGA-PEG-FA nanoparticles can be used to achieve targeted localization imaging., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

In the current study, we synthesized prostate cancer-targeting poly(lactide- co -glycolic acid) (PLGA) nanobubbles (NBs) modified using A10-3.2 aptamers targeted to prostate-specific membrane antigen (PSMA) and encapsulated paclitaxel (PTX). We also investigated their impact on ultrasound (US) imaging and therapy of prostate cancer. PTX-A10-3.2-PLGA NBs were developed using water-in-oil-in-water (water/oil/water) double emulsion and carbodiimide chemistry approaches. Fluorescence imaging together with flow cytometry verified that the PTX-A10-3.2-PLGA NBs were successfully fabricated and could specifically bond to PSMA-positive LNCaP cells. We speculated that, in vivo, the PTX-A10-3.2-PLGA NBs would travel for a long time, efficiently aim at prostate cancer cells, and sustainably release the loaded PTX due to the improved permeability together with the retention impact and US-triggered drug delivery. The results demonstrated that the combination of PTX-A10-3.2-PLGA NBs with low-frequency US achieved high drug release, a low 50% inhibition concentration, and significant cell apoptosis in vitro. For mouse prostate tumor xenografts, the use of PTX-A10-3.2-PLGA NBs along with low-frequency US achieved the highest tumor inhibition rate, prolonging the survival of tumor-bearing nude mice without obvious systemic toxicity. Moreover, LNCaP xenografts in mice were utilized to observe modifications in the parameters of PTX-A10-3.2-PLGA and PTX-PLGA NBs in the contrast mode and the allocation of fluorescence-labeled PTX-A10-3.2-PLGA and PTX-PLGA NBs in live small animals and laser confocal scanning microscopy fluorescence imaging. These results demonstrated that PTX-A10-3.2-PLGA NBs showed high gray-scale intensity and aggregation ability and showed a notable signal intensity in contrast mode as well as aggregation ability in fluorescence imaging. In conclusion, we successfully developed an A10-3.2 aptamer and loaded PTX-PLGA multifunctional theranostic agent for the purpose of obtaining US images of prostate cancer and providing low-frequency US-triggered therapy of prostate cancer that was likely to constitute a strategy for both prostate cancer imaging and chemotherapy., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Single-wall carbon nanohorns (SWCNHs), which have multitudes of horn interstices, an extensive surface area, and a spherical aggregate structure, offer many advantages over other carbon nanomaterials being used as a drug nanovector. The previous studies on the interaction between SWCNHs and cells have mostly emphasized on cellular uptake and intracellular trafficking, but seldom on epithelial cells. Polar epithelium as a typical biological barrier constitutes the prime obstacle for the transport of therapeutic agents to target site. This work tried to explore the permeability of SWCNHs through polar epithelium and their abilities to modulate transcellular transport, and evaluate the potential of SWCNHs in drug delivery. Madin-Darby canine kidney (MDCK) cell monolayer was used as a polar epithelial cell model, and as-grown SWCNHs, together with oxidized and fluorescein isothiocyanate-conjugated bovine serum albumin-labeled forms, were constructed and comprehensively investigated in vitro and in vivo. Various methods such as transmission electron microscopy and confocal imaging were used to visualize their intracellular uptake and localization, as well as to investigate the potential transcytotic process. The related mechanism was explored by specific inhibitors. Additionally, fast multispectral optoacoustic tomography imaging was used for monitoring the distribution and transport process of SWCNHs in vivo after oral administration in nude mice, as an evidence for their interaction with the intestinal epithelium. The results showed that SWCNHs had a strong bioadhesion property, and parts of them could be uptaken and transcytosed across the MDCK monolayer. Multiple mechanisms were involved in the uptake and transcytosis of SWCNHs with varying degrees. After oral administration, oxidized SWCNHs were distributed in the gastrointestinal tract and retained in the intestine for up to 36 h probably due to their surface adhesion and endocytosis into the intestinal epithelium. Overall, this comprehensive investigation demonstrated that SWCNHs can serve as a promising nanovector that can cross the barrier of polar epithelial cells and deliver drugs effectively., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

The intraoperative precision cleaning of lymph nodes (LNs) is an essential component of treating neoplastic disease. To develop efficient probes for the targeted detection of LNs that could act as carriers for the specific diagnosis and treatment of metastatic LNs in the future, dextran-coated iron oxide nanoclusters (DIONs) were synthesized using a one-pot coprecipitation procedure. These modified DIONs have good water dispersibility, cytocompatibility, an optimum size, and a stable, dark brown color for LN imaging. In this study, cytotoxicity was evaluated using lymphatic endothelial cells (LECs) to predict biosafety and biocompatibility. Most importantly, the effectiveness of DIONs in mapping perigastric LNs in Sprague Dawley rats following injection into the gastric submucosal layer was demonstrated. In addition, a long-term tracing in vivo (from 4 days to 3 months) indicated that the DIONs had good biosafety and biocompatibility according to an evaluation of the behavior and blood biochemistry of the rat and a histopathological examination of the important organs., Competing Interests: Disclosure The authors report no conflicts of interest in this work.

Recently, photothermal therapy has become a promising strategy in tumor treatment. However, the therapeutic effect was seriously hampered by the low tissue penetration of laser. Therefore, in this study, radiofrequency (RF) with better tissue penetration was used for tumor hyperthermia. First, one type of gold nanorods (AuNRs) suitable for RF hyperthermia was selected. Then, poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) loaded with AuNRs and docetaxel (DTX) (PLGA/AuNR/DTX) NPs were constructed. Finally, manganese dioxide (MnO 2 ) ultrathin nanofilms were coated on the surfaces of PLGA/AuNR/DTX NPs by the reduction of KMnO 4 to construct the PLGA/AuNR/DTX@MnO 2 drug delivery system. This drug delivery system can not only be used for the combined therapy of chemotherapy and RF hyperthermia but can also produce Mn 2+ to enable magnetic resonance imaging. Furthermore, the RF hyperthermia and the degradation of MnO 2 can significantly promote the controlled drug release in a tumor region. The in vitro and in vivo results suggested that the PLGA/AuNR/DTX@MnO 2 multifunctional drug delivery system is a promising nanoplatform for effective cancer theranostic applications., Competing Interests: Disclosure The authors report no conflicts of interest in this work.