Motao Zhu

Physicochemical properties determine nanomaterial cellular uptake, transport and fate

Although a growing number of innovations have emerged in the fields of nanobiotechnology and nanomedicine, new engineered nanomaterials (ENMs) with novel physicochemical properties are posing novel challenges to understand the full spectrum of interactions at the nano-bio interface. Because these could include potentially hazardous interactions, researchers need a comprehensive understanding of toxicological properties of nanomaterials and their safer design. In depth research is needed to understand how nanomaterial properties influence bioavailability, transport, fate, cellular uptake, and catalysis of injurious biological responses. Toxicity of ENMs differ with their size and surface properties, and those connections hold true across a spectrum of in vitro to in vivo nano-bio interfaces. In addition, the in vitro results provide a basis for modeling the biokinetics and in vivo behavior of ENMs. Nonetheless, we must use caution in interpreting in vitro toxicity results too literally because of dosimetry differences between in vitro and in vivo systems as well the increased complexity of an in vivo environment. In this Account, we describe the impact of ENM physicochemical properties on cellular bioprocessing based on the research performed in our groups. Organic, inorganic, and hybrid ENMs can be produced in various sizes, shapes and surface modifications and a range of tunable compositions that can be dynamically modified under different biological and environmental conditions. Accordingly, we cover how ENM chemical properties such as hydrophobicity and hydrophilicity, material composition, surface functionalization and charge, dispersal state, and adsorption of proteins on the surface determine ENM cellular uptake, intracellular biotransformation, and bioelimination versus bioaccumulation. We review how physical properties such as size, aspect ratio, and surface area of ENMs influence the interactions of these materials with biological systems, thereby affecting their hazard potential. We discuss our actual experimental findings and show how these properties can be tuned to control the uptake, biotransformation, fate, and hazard of ENMs. This Account provides specific information about ENM biological behavior and safety issues. This research also assists the development of safer nanotherapeutics and guides the design of new materials that can execute novel functions at the nano-bio interface.

A doxorubicin delivery platform using engineered natural membrane vesicle exosomes for targeted tumor therapy.

Targeted drug delivery vehicles with low immunogenicity and toxicity are needed for cancer therapy. Here we show that exosomes, endogenous nano-sized membrane vesicles secreted by most cell types, can deliver chemotherapeutics such as doxorubicin (Dox) to tumor tissue in BALB/c nude mice. To reduce immunogenicity and toxicity, mouse immature dendritic cells (imDCs) were used for exosome production. Tumor targeting was facilitated by engineering the imDCs to express a well-characterized exosomal membrane protein (Lamp2b) fused to αv integrin-specific iRGD peptide (CRGDKGPDC). Purified exosomes from imDCs were loaded with Dox via electroporation, with an encapsulation efficiency of up to 20%. iRGD exosomes showed highly efficient targeting and Dox delivery to αv integrin-positive breast cancer cells in vitro as demonstrated by confocal imaging and flow cytometry. Intravenously injected targeted exosomes delivered Dox specifically to tumor tissues, leading to inhibition of tumor growth without overt toxicity. Our results suggest that exosomes modified by targeting ligands can be used therapeutically for the delivery of Dox to tumors, thus having great potential value for clinical applications.

Development of a mild mercaptoethanol extraction method for determination of mercury species in biological samples by HPLC-ICP-MS.

A mild, efficient and convenient extraction method of using 2-mercaptoethanol contained extractant solution combined with an incubator shaker for determination of mercury species in biological samples by HPLC-ICP-MS has been developed. The effects of the concentration of 2-mercaptoethanol, the composition of the extractant solution and the shaking time on the efficiency of mercury extraction were evaluated. The optimization experiments indicated that the quantitative extraction of mercury species from biological samples could be achieved by using 0.1% (v/v) HCl, 0.1% (v/v) 2-mercapoethanol and 0.15% (m/v) KCl extractant solution in an incubator shaker for shaking overnight (about 12h) at room temperature. The established method was validated by analysis of various biological certified reference materials, including NRCC DOLT-3 (dogfish liver), IAEA 436 (tuna fish), IAEA MA-B-3/TM (garfish filet), IAEA MA-M-2/TM (mussel tissue), GBW 08193 (bovine liver) and GBW 08572 (prawn). The analytical results of the reference materials were in good agreement with the certified or reference values of both methyl and total mercury, indicating that no distinguishable transformation between mercury species had occurred during the extraction and determination procedures. The limit of detection (LOD) for methyl (CH(3)Hg(+)) and inorganic mercury (Hg(2+)) by the method are both as 0.2microg L(-1). The relative standard deviation (R.S.D.s) for CH(3)Hg(+) and Hg(2+) are 3.0% and 5.8%, respectively. The advantages of the developed extraction method are that (1) it is easy to operate in HPLC-ICP-MS for mercury species determination since the extracted solution can be directly injected into the HPLC column without pH adjustment and (2) the memory effect of mercury in the ICP-MS measurement system can be reduced.

Exosomes as Extrapulmonary Signaling Conveyors for Nanoparticle‐Induced Systemic Immune Activation

Evaluation of systemic biosafety of nanomaterials urgently demands a comprehensive understanding of the mechanisms of the undesirable interference and systemic signaling that arises between man-made nanomaterials and biological systems. It is shown that exosomes may act as signal conveyors for nanoparticle-induced systemic immune responses. Exosomes are extracellularly secreted membrane vesicles which act as Trojan horses for the dissemination and intercellular communication of natural nanosized particles (like viruses). Upon exposure to magnetic iron oxide nanoparticles (MIONs), it is possible to dose-dependently generate a significant number of exosomes in the alveolar region of BALB/c mice. These exosomes are quickly eliminated from alveoli into systemic circulation and largely transfer their signals to the immune system. Maturation of dendritic cells and activation of splenic T cells are significantly induced by these exosomes. Furthermore, exosome-induced T-cell activation is more efficient toward sensitized T cells and in ovalbumin (OVA)-sensitized mice than in the unsensitized counterparts. Activation of systemic T cells reveals a T helper 1 polarization and aggravated inflammation, which poses potential hazards to the deterioration of allergic diseases in OVA-sensitized mice. The studies suggest that exosomes may act as conveyors for extrapulmonary signal transduction in nanoparticle-induced immune systemic responses, which are the key in vivo processes of manufactured nanoparticles executing either biomedical functions or toxic responses.

Using ion-pair reversed-phase HPLC ICP-MS to simultaneously determine Cr(III) and Cr(VI) in urine of chromate workers

Urinary chromium speciation analysis can provide available information of the individual exposure levels of Cr(VI) compounds. An analytical method based on ion-pair reversed-phase HPLC combined with ICP-MS to simultaneously determine Cr(III) and Cr(VI) in human urine has been developed for assessing the occupational exposure to chromate. The separation conditions of the method, including the pH value, the concentrations of ion-pair reagent and methanol in the mobile phase were studied. Specially, a high-speed polyetheretherketone (PEEK) column and a typical sample introduction method were employed to avoid the exogenous chromium contamination during the analysis. The separation of Cr(III) and Cr(VI) could be finished within 4min with the detection limits as low as 0.03microgL(-1) at 100microL injections for both of them, providing a convenient method for routine analysis of chromium species. The chromium species in urine of chromate workers were monitored using the developed method. The statistical analysis showed a significant relationship (n=32, p<0.01) between the urinary Cr(VI) and the individual airborne exposure levels, indicating that the urinary Cr(VI) could be used as a convenient and suitable monitor for high level Cr(VI) occupational exposure.

Safety of Nanoparticles in Medicine

Nanomedicine involves the use of nanoparticles for therapeutic and diagnostic purposes. During the past two decades, a growing number of nanomedicines have received regulatory approval and many more show promise for future clinical translation. In this context, it is important to evaluate the safety of nanoparticles in order to achieve biocompatibility and desired activity. However, it is unwarranted to make generalized statements regarding the safety of nanoparticles, since the field of nanomedicine comprises a multitude of different manufactured nanoparticles made from various materials. Indeed, several nanotherapeutics that are currently approved, such as Doxil and Abraxane, exhibit fewer side effects than their small molecule counterparts, while other nanoparticles (e.g. metallic and carbon-based particles) tend to display toxicity. However, the hazardous nature of certain nanomedicines could be exploited for the ablation of diseased tissue, if selective targeting can be achieved. This review discusses the mechanisms for molecular, cellular, organ, and immune system toxicity, which can be observed with a subset of nanoparticles. Strategies for improving the safety of nanoparticles by surface modification and pretreatment with immunomodulators are also discussed. Additionally, important considerations for nanoparticle safety assessment are reviewed. In regards to clinical application, stricter regulations for the approval of nanomedicines might not be required. Rather, safety evaluation assays should be adjusted to be more appropriate for engineered nanoparticles.

Nanoparticle‐Induced Exosomes Target Antigen‐Presenting Cells to Initiate Th1‐Type Immune Activation

The mechanisms associated with the induction of systemic immune responses by nanoparticles are not fully understood, but their elucidation is critical to address safety issues associated with the broader medical application of nanotechnology. In this study, a key role of nanoparticle-induced exosomes (extracellularly secreted membrane vesicles) as signaling mediators in the induction of T helper cell type 1 (Th1) immune activation is demonstrated. In vivo exposure to magnetic iron oxide nanoparticles (MIONs) results in significant exosome generation in the alveolar region of Balb/c mice. These act as a source of nanoparticle-induced, membrane-bound antigen/signaling cargo, which transfer their components to antigen-presenting cells (APCs) in the reticuloendothelial system. Through exosome-initiated signals, immature dendritic cells (iDCs) undergo maturation and differentiation to the DC1 subtype, while macrophages go through classical activation and differentiation to the M1 subtype. Simultaneously, iDCs and macrophages release various Th1 cytokines (including interleukin-12 and tumor necrosis factor α) driving T-cell activation and differentiation. Activated APCs (especially DC1 and M1 subtypes) consequently prime T-cell differentiation towards a Th1 subtype, thereby resulting in an orchestrated Th1-type immune response. Th1-polarized immune activation is associated with delayed-type hypersensitivity, which might underlie the long-term inflammatory effects frequently associated with nanoparticle exposure. These studies suggest that nanoparticle-induced exosomes provoke the immune activation and inflammatory responses that can accompany nanoparticle exposure.

Understanding the Particokinetics of Engineered Nanomaterials for Safe and Effective Therapeutic Applications

Increasing numbers of engineered nanomaterials (ENMs) are being developed for therapeutic and diagnostic applications. However, the tunable and varied physicochemical properties of ENMs pose a new challenge for understanding their biological behavior, trafficking, and biodistribution. Herein the concept of particokinetics is introduced to address the dynamic biological behavior of ENMs at the molecular level (including gravitational sedimentation, dispersion, aggregation, and interaction with biomolecules in suspending media), cellular level (including cellular uptake, transport, biotransformation, and elimination), and whole-organism level (including absorption, distribution, metabolism, and excretion in vivo). Several mathematical modeling methods are introduced which guide a quantitative description of their biological behavior at different levels. Examples are also provided to delineate the impact of the physicochemical properties of ENMs on their particokinetics. A comprehensive understanding of the in vivo and in vitro particokinetics of ENMs will facilitate the design of tailor-made functional ENMs that act as highly effective and controllable drug-delivery systems with minimal side-effects.

A membrane vesicle-based dual vaccine against melanoma and Lewis lung carcinoma

In the past few years, cell-derived membrane vesicle-based tumor vaccines have been considered as valuable new tools for cancer immunotherapy. Despite promising results in cancer clinical trails, an improved method is urgently needed for high efficiency tumor vaccines for a broad spectrum of tumors. Here we developed a single membrane vesicle-based vaccine, which is active in repressing both melanoma (B16) and Lewis lung carcinoma (LLC) tumor growth. By using the intrinsic function of dendritic cells in the processing and presentation of antigens, we generated dendritic cell (DC)-derived membrane vesicles (DC-mv) bearing tumor antigens from both B16 and LLC cells. Vaccination with this DC-mv-based dual vaccine induced specific cytotoxic T lymphocytes (CTL)-dependent tumor rejection and suppressed the growth of both types of tumor xenografts in mice. In addition, induction of CTL by this vaccine resulted in cross-protection responses and consequently enabled significant enhanced anti-tumor effects, indicating the synergistic anti-tumor activity. Our study suggests that the DC-mv-based vaccine holds great potential as a highly effective, versatile, cell-free vaccine for inhibition of multiple types of tumor growth.

Applications of nanomaterials as vaccine adjuvants

Vaccine adjuvants are applied to amplify the recipients specific immune responses against pathogen infection or malignancy. A new generation of adjuvants is being developed to meet the demands for more potent antigen-specific responses, specific types of immune responses, and a high margin of safety. Nanotechnology provides a multifunctional stage for the integration of desired adjuvant activities performed by the building blocks of tailor-designed nanoparticles. Using nanomaterials for antigen delivery can provide high bioavailability, sustained and controlled release profiles, and targeting and imaging properties resulting from manipulation of the nanomaterials’ physicochemical properties. Moreover, the inherent immune-regulating activity of particular nanomaterials can further promote and shape the cellular and humoral immune responses toward desired types. The combination of both the delivery function and immunomodulatory effect of nanomaterials as adjuvants is thought to largely benefit the immune outcomes of vaccination. In this review, we will address the current achievements of nanotechnology in the development of novel adjuvants. The potential mechanisms by which nanomaterials impact the immune responses to a vaccine and how physicochemical properties, including size, surface charge and surface modification, impact their resulting immunological outcomes will be discussed. This review aims to provide concentrated information to promote new insights for the development of novel vaccine adjuvants.