Antibody-Oriented Strategy and Mechanism for the Preparation of Fluorescent Nanoprobes for Fast and Sensitive Immunodetection.

Nanoprobes have been widely used in biomedical engineering. However, antibodies are generally conjugated onto nanoparticles disorderly, which reduces their antigen recognition ability. The existing antibody orientation approaches are usually complex. Here, we developed and demonstrated a simple antibody-oriented strategy for the lateral flow immunoassay of cardiac troponin I by conjugating antibodies onto polystyrene nanospheres at the optimal pH. The binding amount and orientation of antibodies as well as the detection sensitivity were significantly improved. Although pH regulation is commonly used to optimize antibody conjugation, this paper illustrates the mechanism of its antibody orientation enhancement ability for the first time and reveals the important influences of the density, the charge distribution and hydrophilicity of the antibody, the control of the velocities of physical adsorption and chemical coupling, and other factors on antibody orientation. It is of great significance to understand and regulate antibody conjugation on the surface of micro- or nanospheres to construct high-performance probes for in vitro diagnosis applications.

Preparation and In Vitro Cellular Uptake Assessment of Multifunctional Rubik-Like Magnetic Nano-Assemblies.

Through self-assembly of nanoparticles into high-order and stable structures of cubic clusters, high drug-loading rubik-like magnetic nano-assemblies (MNAs), possessing folic acid targeting and strong magnetism-enhanced cellular uptake capabilities, were built. In this study, the core of the cubic drug assemblies consisted of four monodisperse superparamagnetic iron oxide nanoparticles coated with layers of oleic acid (Fe3O4@OA), simultaneously encapsulating fluorescein, and Paclitaxol (Flu-MNAs and PTX-MNAs) for imaging and therapeutic applications. To enable preferential tumor cellular uptake by the nanocarriers, the outermost layer of Fe3O4 was functionalized with the new dual-oleic acid-polyethylene glycol-folic acid polymer (FA-PEG-Lys-OA2) as a "shell." The drug carriers exhibited excellent stability and biocompatibility, and showed high drug loading and excellent magnetic response In Vitro. Furthermore, preliminary evaluations of the drug carriers with Hela cells showed effective cellular targeting capability. In addition, the cubic assemblies enhanced anticancer efficiency for Hela cells compared to bare drugs. Especially, the applied external magnetic field further improved the uptake of the vectors, and thereby enhanced the inhibitory effect. In brief, all these results suggested that cubic assemblies could serve as potential strategies for targeted anticancer therapies.

Iron oxide nanoparticles in liquid or powder form enhanced osteogenesis via stem cells on injectable calcium phosphate scaffold.

The objectives of this study were to incorporate iron oxide nanoparticles (IONPs) into calcium phosphate cement (CPC) to enhance bone engineering, and to investigate the effects of IONPs as a liquid or powder on stem cells using IONP-CPC scaffold for the first time. IONP-CPCs were prepared by adding 1% IONPs as liquid or powder. Human dental pulp stem cells (hDPSCs) were seeded. Subcutaneous implantation in mice was investigated. IONP-CPCs had better cell spreading, and greater ALP activity and bone mineral synthesis, than CPC control. Subcutaneous implantation for 6 weeks showed good biocompatibility for all groups. In conclusion, incorporating IONPs in liquid or powder form both substantially enhanced hDPSCs on IONP-CPC scaffold and exhibited excellent biocompatibility. IONP incorporation as a liquid was better than IONP powder in promoting osteogenic differentiation of hDPSCs. Incorporating IONPs and chitosan lactate together in CPC enhanced osteogenesis of hDPSCs more than using either alone.

Fluorescent Nanoprobes with Oriented Modified Antibodies to Improve Lateral Flow Immunoassay of Cardiac Troponin I.

Performance of nanoprobes can often determine the detection level of Lateral immunochromatography. Traditional probes were limited by the quantity and orientation of antibodies, immune activity of the Fab region or binding strength between protein and substrate. This study developed a new efficient and robust technology to construct fluorescent nanoprobes with oriented modified antibodies, based on specific binding of the Fc region of antibody with streptococcal protein G (SPG) on the surface of polystyrene microspheres (MS) and subsequent covalent cross-linking at binding sites to firm them. Lateral flow immunoassay using these probes was applied for the detection of cardiac troponin I (cTnI). The significantly improved detection sensitivity demonstrated that antibody orientation on MS surfaces effectively enhanced immunological activities of probes compared with random immobilizing methods. Furthermore, performance evaluation results of lateral flow test strips met clinical requirements perfectly, including limit of detection (0.032 ng/mL), linearity ( R > 0.99), repeatability (CV < 10%), correlation ( R > 0.99), and heat aging stability. This research also employed heterophilic blocking reagent (HBR) to actively block redundant binding sites of SPG for the first time in order to eliminate false positive interferences, improving the sensitivity and precision of test results further.

Thermo-Sensitive PLGA-PEG-PLGA Tri-Block Copolymer Hydrogel as Three-Dimensional Cell Culture Matrix for Ovarian Cancer Cells.

Thermo-sensitive hydrogels which could encapsulate cells and provide a three dimensional (3D) microenvironment have great potential in building new cell culture models in vitro. In this study, a thermal responsive hydrogel based on PLGA-PEG-PLGA tri-block copolymers was developed as matrix for 3D ovarian cancer cell culturing. The gelation of PLGA-PEG-PLGA tri-block copolymer was concentration-dependent. SEM images showed the pores were suitable for the formation of 3D cell structures. Cell morphological results showed that large aggregates of ovarian cancer cells (HO8910) were formed after cultured for 10 days. Therefore, hydrogel based on PLGA-PEG-PLGA tri-block copolymers hold potential as in vitro cell culture matrix for ovarian cancer cells.

Gold nanoparticles in injectable calcium phosphate cement enhance osteogenic differentiation of human dental pulp stem cells.

In this study, a novel calcium phosphate cement containing gold nanoparticles (GNP-CPC) was developed. Its osteogenic induction ability on human dental pulp stem cells (hDPSCs) was investigated for the first time. The incorporation of GNPs improved hDPSCs behavior on CPC, including better cell adhesion (about 2-fold increase in cell spreading) and proliferation, and enhanced osteogenic differentiation (about 2-3-fold increase at 14 days). GNPs endow CPC with micro-nano-structure, thus improving surface properties for cell adhesion and subsequent behaviors. In addition, GNPs released from GNP-CPC were internalized by hDPSCs, as verified by transmission electron microscopy (TEM), thus enhancing cell functions. The culture media containing GNPs enhanced the cellular activities of hDPSCs. This result was consistent with and supported the osteogenic induction results of GNP-CPC. In conclusion, GNP-CPC significantly enhanced the osteogenic functions of hDPSCs. GNPs are promising to modify CPC with nanotopography and work as bioactive additives thus enhance bone regeneration.

A Functional Iron Oxide Nanoparticles Modified with PLA-PEG-DG as Tumor-Targeted MRI Contrast Agent.

PURPOSE: Tumor targeting could greatly promote the performance of magnetic nanomaterials as MRI (Magnetic Resonance Imaging) agent for tumor diagnosis. Herein, we reported a novel magnetic nanoparticle modified with PLA (poly lactic acid)-PEG (polyethylene glycol)-DG (D-glucosamine) as Tumor-targeted MRI Contrast Agent. METHODS: In this work, we took use of the D-glucose passive targeting on tumor cells, combining it on PLA-PEG through amide reaction, and then wrapped the PLA-PEG-DG up to the Fe3O4@OA NPs. The stability and anti phagocytosis of Fe3O4@OA@PLA-PEG-DG was tested in vitro; the MRI efficiency and toxicity was also detected in vivo. RESULTS: These functional magnetic nanoparticles demonstrated good biocompatibility and stability both in vitro and in vivo. Cell experiments showed that Fe3O4@OA@PLA-PEG-DG nanoparticles exist good anti phagocytosis and high targetability. In vivo MRI images showed that the contrast effect of Fe3O4@OA@PLA-PEG-DG nanoparticles prevailed over the commercial non tumor-targeting magnetic nanomaterials MRI agent at a relatively low dose. CONCLUSIONS: The DG can validly enhance the tumor-targetting effect of Fe3O4@OA@PLA-PEG nanoparticle. Maybe MRI agents with DG can hold promise as tumor-targetting development in the future.

Preparation and in vivo safety evaluations of antileukemic homoharringtonine-loaded PEGylated liposomes.

In order to improve the in vivo safety and specific delivery efficiency of the antileukemic homoharringtonine (HHT) at the targets, the long-circulating PEGylated liposomes loaded with HHT (LCLipo-HHT) were prepared. Their physical characteristics, in vitro drug release, in vivo pharmacokinetic properties and elementary toxicity were evaluated. The mean diameter of the prepared LCLipo-HHT is 75.6 ± 3.2 nm and the zeta potential is -16.9 ± 2.5 mV. The entrapment efficiency of HHT in the liposomes is 69.5 ± 1.7%. In pharmacokinetic experiments, an increased plasma concentration as well as blood circulation time was obtained when distearoyl phosphoethanolamine-PEG 2000 lipid was added in the formulation, which results in enhancing drug delivery efficiency. Hemolysis test, vascular irritation test and acute toxicity test were used to demonstrate toxicity of LCLipo-HHT. Compared with clinical HHT injection dosage, LCLipo-HHT indicated no vascular irritation, good hemocompatibility, as well as much better safety. Therefore, the prepared LCLipo-HHT can be used as a promising anticancer formulation for antileukemic therapy in the future.

Antigen-specific killer polylactic-co-glycolic acid (PLGA) microspheres can prolong alloskin graft survival in a murine model.

BACKGROUND: The strategy of specifically depleting antigen-specific T cells can potentially be used for the treatment of allograft rejection and autoimmunity because it does not suppress the overall immune systems. METHODS: In this study, we generated killer polylactic-co-glycolic acid (PLGA) microspheres by covalently coupling major histocompatibility complex (MHC) class I antigens and apoptosis-inducing anti-Fas monoclonal antibody (mAb) onto PLGA microspheres. A modified double-emulsion method was used for the preparation of cell-sized PLGA microspheres. H-2K(b)/peptide monomers were generated in-house and analyzed through flow cytometry. The killer PLGA microspheres were administered intravenously into BALB/c mice (H-2K(d)) that had previously been grafted with skin squares from C57BL/6 mice (H-2K(b)). Tumor cell challenge and third-party mixed lymphocyte culture were used to assess the general immune functions of host. RESULTS: The alloskin graft survival was prolonged by 4 days. The killer PLGA microspheres could specifically deplete the H-2K(b) alloantigen-reactive CD8(+) T cells that infiltrated into the alloskin graft but not CD4(+) T cells, without impairment of host overall immune function. CONCLUSIONS: Here, we initially report that PLGA microspheres, which have been widely used as medicine-delivering carriers, were used to prepare antigen-specific killer complexes and treat allograft rejection. Our data highlight the therapeutic potential of this biocompatible and biodegradable antigen-specific killer effector for the treatment of allograft rejection and autoimmune disease.

Effective PEGylation of Fe3O4 Nanomicelles for In Vivo MR Imaging.

A practical and effective strategy for synthesizing PEGylated Fe3O4 nanomicelles is established. In this strategy, a magnetic fluid of the Fe3O4 nanomicelles was synthesized with amphiphilic PEGylated phospholipid as surfactant and soybean oil as stabilizer under simple mechanical stirring and subsequent ultrasonication. Transmission electron microscope (TEM) measurement indicated that the sample is monodisperse spherical Fe3O4 nanoparticles with internal core size of 9 nm and external nanomicelle shell thickness of 1.5 nm. The final hydrodynamic size of the sample is 19.5 nm and its zeta potential is - 38.5 mV, suggesting good stability of the magnetic nanomicelles in water. To assess the ability of magnetic nanomicelles to escape reticuloendothelial system (RES) uptake, in vitro cell phagocytosis experiments were conducted using murine macrophages (RAW264.7). The results indicated that the PEGylation can effectively prevent the uptake of the nanomicelles by the macrophages. Using a mouse model of 4T1 breast cancer, the nanomicelles provided a good magnetic resonance imaging (MRI) capability to sensitively detect tumor by enhanced permeability and retention (EPR) effect. The PEGylated monodisperse magnetic nanomicelles would become a potential contrast agent for passive targeting diagnosis of tumor by MR imaging.

Laser-assisted manipulation of Volta potential pattern on the TC4 surface for improved hBMSCs osteogenesis.

The Ti6Al4V (TC4) alloy, a prevalent biomedical material in orthopedics, still faces limitation of the insufficient osseointegration. To improve the bioactivity of TC4, introducing the electric environment onto the TC4 surface may be an effective way in the view of the necessity of endogenous electric microenvironment in bone regeneration. Herein, a Volta potential pattern was engendered on the TC4 surface via parallel laser patterning, so as to promote the osteogenic differentiation of cells. A 15 W laser successfully transformed the original α + ß dual phase towards radially distributed lath-like martensite phase in the laser treated region. The atomic lattice distortion between the heterogeneous microstructures of the laser treated and untreated regions leads to a significant Volta potential fluctuation on the TC4 surface. The Volta potential pattern as well as the laser-engraved microgrooves respectively induced mutually orthogonal cell alignments. The hBMSCs osteogenic differentiation was significantly enhanced on the laser treated TC4 surfaces in comparison to the surface without the laser treatment. Moreover, a drastic Volta potential gradient on the TC4 surface (treated with 15 W power and 400 µm interval) resulted in the most pronounced osteogenic differentiation tendency compared to other groups. Modulating the electric environment on the TC4 surface by manipulating the phase transformation may provide an effective way in evoking favorable cell response of bone regeneration, thereby improving the bioactivity of TC4 implant.

Biodegradable ferroelectric molecular crystal with large piezoelectric response.

Transient implantable piezoelectric materials are desirable for biosensing, drug delivery, tissue regeneration, and antimicrobial and tumor therapy. For use in the human body, they must show flexibility, biocompatibility, and biodegradability. These requirements are challenging for conventional inorganic piezoelectric oxides and piezoelectric polymers. We discovered high piezoelectricity in a molecular crystal HOCH2(CF2)3CH2OH [2,2,3,3,4,4-hexafluoropentane-1,5-diol (HFPD)] with a large piezoelectric coefficient d33 of ~138 picocoulombs per newton and piezoelectric voltage constant g33 of ~2450 × 10-3 volt-meters per newton under no poling conditions, which also exhibits good biocompatibility toward biological cells and desirable biodegradation and biosafety in physiological environments. HFPD can be composite with polyvinyl alcohol to form flexible piezoelectric films with a d33 of 34.3 picocoulombs per newton. Our material demonstrates the ability for molecular crystals to have attractive piezoelectric properties and should be of interest for applications in transient implantable electromechanical devices.

Imaging Structural and Electrical Changes of Aging Cells Using Scanning Ion Conductance Microscopy.

The local charge density and distribution of extracellular membranes play a crucial role in the various cellular processes, such as regulation and localization of membrane proteins, electrophysiological signal transduction, transcriptional control, cell growth, and cell death. In this study, a novel scanning ion conductance microscopy-based method is employed to extracellular membrane mapping. This method allows to not only visualize the dynamic topography and surface charge distribution around individual cells, but also distinguish the charge difference. To validate the accuracy and effectiveness of this method, the charge density on model sample surfaces are initially manipulated and the charge sensing mechanism using finite element modeling (FEM) is explored subsequently. By applying this method, both the extracellular charge distributions and topography structures of normal and senescent human dental pulp stem cells (hDPSCs) are able to monitor. Interestingly, it is observed that the surface charge became significantly more negative after cellular senescence. This innovative approach enables us to gain valuable insights into surface charge changes during cellular senescence, which can contribute to a better understanding of the underlying mechanisms and potential therapeutic strategies for age-related diseases.

Magnetic Delivery of Antigen-Loaded Magnetic Liposomes for Active Lymph Node Targeting and Enhanced Anti-Tumor Immunity.

Therapeutic cancer vaccines offer the greatest advantage of enhancing antigen-specific immunity against tumors, particularly for immunogenic tumors, such as melanoma. However, clinical responses remain unsatisfactory, primarily due to inadequate T cell priming and the development of acquired immune tolerance. A major obstacle lies in the inefficient uptake of antigen by peripheral dendritic cells (DCs) and their migration to lymph nodes for antigen presentation. In this context, the magnetic delivery of antigen-loaded magnetic liposomes (Ag-MLs) to actively target lymph node, is proposed. These magnetic responsive liposomes contain soluble mouse melanoma lysate and iron oxide nanoparticles in the core, along with the immunostimulatory adjuvant CpG-1826 incorporated into the lipid bilayer. When applied through magnetic targeting in the mouse melanoma model, Ag-MLs accumulate significantly in the target lymph nodes. This accumulation results in increased population of active DCs in lymph nodes and cytotoxic T lymphocytes (CTLs) within tumors, correlating with effective tumor growth inhibition. Overall, this study demonstrates the potential of magnetic targeting as an effective strategy for delivering cancer vaccines and activating the immune response, offering a novel platform for cancer immunotherapies.

Effects of oral administration of polystyrene nanoplastics on plasma glucose metabolism in mice.

Microplastic (MP) and nanoplastic (NP) induce neurotoxicity, cytotoxicity, and reproductive system toxicity in mammals. However, the impacts of NPs on the endocrine system are obscure. Here, monodisperse polystyrene nanoplastics (PS-NPs) were prepared by emulsion polymerization and the accumulation of fluorescent PS-NPs in various organs, including the liver, kidney, spleen, and pancreas, was examined. The oral administration of PS-NPs induced visceral organ injury, and the main toxicities were damage to hepatic function and the abnormity of lipid metabolism. Global transcriptome sequencing (RNA-Seq) revealed the impact of PS-NPs on the genes involved in reactive oxygen species (ROS) generation and the PI3K/Akt signaling pathway, which is associated with glucose metabolism in mice. Chronic exposure to PS-NPs significantly increased plasma glucose levels and ROS levels, but did not affect plasma insulin secretion. The phosphorylation of insulin receptor substrate (IRS)-1 at Ser307 was raised, which decreased the phosphorylation of Akt (at Ser473) in the PI3K/Akt pathway. Collectively, these findings suggested that the oral administration of PS-NPs significantly increased ROS, hepatic triglycerides, and cholesterol accumulation. The high levels of ROS disturbed the PI3K/Akt pathway, causing insulin resistance and increased plasma glucose in the mouse liver.

Surfactant-induced formation of honeycomb pattern on micropipette with curvature gradient.

Breath figure (BF) process is a facile method to prepare honeycomb structures by dynamic movements of condensed micrometer-sized water droplets at the interface of volatile fluid. Here, we aim to find answers to understand how the BF process occurs on micropipettes with curvature gradient and to understand the role of the surfactant in obtaining honeycomb patterns. Poly (L-lactic acid) (PLLA) chloroform solution with dioleoylphosphatidylethanolamine (DOPE) as surfactant was utilized. It is found that the honeycomb structure formed on the micropipettes changes remarkably with the gradually increased surface curvature. The variation trends of the arrangement and diameter of pores on the micropipettes with the increasing curvature are similar to the different time stages of BF process: smaller and sparse pores formed at higher curvature are similar to those formed at early stage of BF; regular honeycomb patterns formed at lower curvature are similar to those formed at the late stage of BF. Especially, the "semi-coalescence" hemispherical pores strings are found at high curvatures on PLLA-DOPE films, indicating the surfactant-induced coalescence of water droplets in BF process. The differences of drying speed of polymer solvent on micropipette with gradually increased curvatures make the printing of the pores at different BF stages on polymer film possible. These findings not only strongly support the mechanism of BF array formation, but also elucidate the surfactant-induced coalescence.

Effect of ZrN coating by magnetron sputtering and sol-gel processed silica coating on titanium/porcelain interface bond strength.

In this study, a coating technique was applied to improve the bond strength of titanium (Ti) porcelain. ZrN coating was prepared by magnetron sputtering, and silica coating was processed by a sol-gel method. The treated surfaces of the specimens were analyzed by X-ray diffraction, and the Ti/porcelain interface was investigated by scanning electron microscopy (SEM) and energy dispersive spectroscopy. The coated specimens appeared fully coherent to the Ti substrate. The fractured bonding surface was also investigated by SEM. The residual porcelain on the metal surface could be observed in the ZrN group and silica group, but there was no obvious porcelain remaining in the control group. A three-point-bending test showed that the bonding strength of the ZrN group (45.99 ± 0.65 MPa) was higher than the silica group (37.77 ± 0.78 MPa) (P < 0.001) and control group (29.48 ± 1.01 MPa) (P < 0.001), while that of the silica group was significantly higher than the control group (P < 0.001). In conclusion, conditioning the ceramic surface with ZrN and silica coatings resulted in a stronger Ti/porcelain bond. ZrN coating by magnetron sputtering was a more effective way to improve the bond strength between Ti and porcelain compared with sol-gel processed silica coating in this study.

Effect of silica coating on fracture strength of glass-infiltrated alumina ceramic cemented to dentin.

PURPOSE: To examine the availability of sol-gel processed silica coating for alumina-based ceramic bonding, and determine which silica sol concentration was appropriate for silica coating. MATERIALS AND METHODS: Sixty disks of In-Ceram alumina ceramic were fabricated and randomly divided into 5 main groups. The disks received 5 different surface conditioning treatments: Group Al, sandblasted; Group AlC, sandblasted + silane coupling agent applied; Groups Al20C, Al30C, and Al40C, sandblasted, silica coating via sol-gel process prepared using 20 wt%, 30 wt%, and 40 wt% silica sols, and then silane coupling agent applied. Before bonding, one-step adhesives were applied on pre-prepared ceramic surfaces of all groups. Then, 60 dentin specimens were prepared and conditioned with phosphoric acid and one-step adhesive. Ceramic disks of all groups were cemented to dentin specimens with dual-curing resin cements. Fracture strength was determined at 24 h and after 20 days of storage in water. RESULTS: Groups Al20C, Al30C, and Al40C revealed significantly higher fracture strength than groups Al and AlC. No statistically significant difference in fracture strength was found between groups Al and AlC, or among groups Al20C, Al30C, and Al40C. Fracture strength values of all the groups did not change after 20 days of water storage. CONCLUSION: Sol-gel processed silica coating can enhance fracture strength of In-Ceram alumina ceramic after bonding to dentin, and different silica sol concentrations produced the same effects. Twenty days of water storage did not decrease the fracture strength.

Xenon Nanobubbles for the Image-Guided Preemptive Treatment of Acute Ischemic Stroke via Neuroprotection and Microcirculatory Restoration.

Early lesion site diagnosis and neuroprotection are crucial to the theranostics of acute ischemic stroke. Xenon (Xe), as a nontoxic gaseous neuroprotectant, holds great promise for ischemic stroke therapy. In this study, Xe-encapsulated lipid nanobubbles (Xe-NBs) have been prepared for the real-time ultrasound image-guided preemptive treatment of the early stroke. The lipids are self-assembled at the interface of free Xe bubbles, and the mean diameter of Xe-NBs is 225 ± 11 nm with a Xe content of 73 ± 2 µL/mL. The in vitro results show that Xe-NBs can protect oxygen/glucose-deprived PC12 cells against apoptosis and oxidative stress. Based on the ischemic stroke mice model, the biodistribution, timely ultrasound imaging, and the therapeutic effects of Xe-NBs for stroke lesions were investigated in vivo. The accumulation of Xe-NBs to the ischemic lesion endows ultrasound contrast imaging with the lesion area. The cerebral blood flow measurement indicates that the administration of Xe-NBs can improve microcirculatory restoration, resulting in reduced acute microvascular injury in the lesion area. Furthermore, local delivery of therapeutic Xe can significantly reduce the volume of cerebral infarction and restore the neurological function with reduced neuron injury against apoptosis. Therefore, Xe-NBs provide a novel nanosystem for the safe and rapid theranostics of acute ischemic stroke, which is promising to translate into the clinical management of stroke.

Sphingosine 1-Phosphate Liposomes for Targeted Nitric Oxide Delivery to Mediate Anticancer Effects against Brain Glioma Tumors.

Specifically targeting glioblastoma multiforme (GBM) blood vessels and actively enhancing the permeability of the brain-blood-tumor barrier (BBTB) are two extremely difficult challenges currently hindering the development of effective therapies against GBM. Herein, a liposome drug delivery system (S1P/JS-K/Lipo) is described, which delivers the nitric oxide (NO) prodrug JS-K, O2 -(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl) piperazin-1-yl] diazen-1-ium-1,2-diolate, to GBM tumors using sphingosine-1-phosphate (S1P)-signaling molecules as active targeting lipid ligands. It is revealed that S1P/JS-K/Lipo actively penetrates the BBTB, aided by caveolin-1-mediated transcytosis, and it is demonstrated that the system specifically interacts with S1P receptors (S1PRs), which are highly expressed on GBM cells. Nondestructive ultrasound imaging in GBM mouse models is also utilized to observe microsized NO bubble production from JS-K, as catalyzed by the glutathione S-transferases (GSTs) resident in GBM cells. Given that these NO bubbles strongly promote GBM cell death in vivo, the S1PR-targeted liposome delivery system-which successfully achieves BBTB penetration and tumor targeted delivery of a complex multicomponent drug regimen-represents a promising approach for targeted therapies against GBM and other carcinomas characterized by elevated S1PR expression.