The advancement of safe nanomaterials for use as optical coherence tomography (OCT) imaging and stem cell-labeling agents to longitudinally visually track therapeutic derived retinal stem cells to study their migration, survival rate, and efficacy is challenged by instability, intracellular aggregation, low uptake, and cytotoxicity. Here, we describe a series of hybrid lipid-coated gold nanorods (AuNRs) that could solve these issues. These nanomaterials were made via a layer-by-layer assembly approach, and their stability in biological media, mechanism, efficiency of uptake, and toxicity were compared with a commercially available set of AuNRs with a 5 nm mesoporous silica (mSiO 2 )-polymer coating. These nanomaterials can serve as stem cell labeling and OCT imaging agents because they absorb in the near-infrared (NIR) region away from biological tissues. Although both subtypes of AuNRs were taken up by retinal pigment epithelial, neural progenitor, and baby hamster kidney cells, slightly negatively charged hybrid lipid-coated AuNRs had minimal aggregation in biological media and within the cytoplasm of cells (∼3000 AuNRs/cell) as well as minimal impact on cell health. Hybrid lipid-coated AuNRs modified with cell-penetrating peptides had the least toxicological impact, with >92% cell viability. In contrast, the more "sticky" AuNRs with a 5 nm mSiO 2 -polymer coating showed significant aggregation in biological media and within the cytoplasm with lower-than-expected uptake of AuNRs (∼5400 of AuNRs/cell) given their highly positive surface charge (35+ mV). Collectively, we have demonstrated that hybrid lipid-coated AuNRs, which absorb in the NIR-II region away from biological tissues, with tuned surface chemistry can label therapeutic derived stem cells with minimal aggregation and impact on cell health as well as enhance uptake for OCT imaging applications.
Gold nanorods (AuNRs) hold tremendous potential to improve the diagnosis and therapeutic options across the blood-retinal barrier to treat retinal diseases. For clinical ophthalmological translation, a fundamental understanding of how their physicochemical properties such as size, shape, charge, surface chemistry, and concentration, impact their stability biological environments, mechanism and efficiency of uptake, and toxicity is a necessity. Here we interrogated the uptake efficiency, biocompatibility, and stability of two subtypes of AuNRs with different types of surface coatings and varying charges, including a commercially available set of AuNRs with a 5 nm mSiO2-polymer coating and hybrid lipid-coated AuNRs developed in-house.Confocal and bright field microscopy images showed uptake of both subtypes of AuNRs in retinal pigment epithelium (RPE), neural progenitor (NP), and baby hamster kidney (BHK) cells.Transmission electron microscopy (TEM) confirms both types of AuNRs are taken up into the cytoplasm of the cells; however, larger aggregates of AuNRs are observed with the more positive and "sticky" AuNRs with a 5 nm mSiO2-polymer coating than the slightly negative hybrid lipidcoated AuNRs. Inductively Coupled Mass Spectroscopy (ICP-MS) confirm that ~3,000 of the slightly negative hybrid lipid-coated AuNRs cells and ~5,400 of the positively charged AuNRs with a 5 nm mSiO2-polymer coating (+35 mV) are taken up into RPE and BHK cell lines. Stability studies in a variety of cellular media showed that hybrid lipid-coated AuNRs are stable and disaggregated in water, 10 mM PBS buffer pH 7, and BHK media except for NP media. In contrast, the positively charged AuNRs with a 5 nm mSiO2-zeta polymer coating aggregated in all media, indicating more interactions with each other and components of the media. Bright-field and TEM confirm the presence of large aggregates of AuNRs on the surface and within the cytoplasm.Cytotoxicity studies both subtypes of AuNRs have an 80 ± 8 % cell viability, indicating mild toxicity. The hybrid lipid-coated AuNR with the cell-penetrating peptide had the least toxicological impact with a > 92 ± 7 % cell viability. Our study highlights the importance of evaluating the impact of the physicochemical features of each new nanoparticle design on their stability in biologically relevant environments and their impact on cellular uptake and toxicity in stem cell-derived therapeutic cells. Here we also provide a simple design strategy for tuning the surface chemistry of robust hybrid lipid-coated AuNRs to enhance cellular uptake to label stem cells with minimal aggregation and toxicity.
Gold nanorods (AuNRs) hold tremendous potential to improve the diagnosis and therapeutic options across the blood-retinal barrier to treat retinal diseases. For clinical ophthalmological translation, a fundamental understanding of how their physicochemical properties such as size, shape, charge, surface chemistry, and concentration, impact their stability biological environments, mechanism and efficiency of uptake, and toxicity is a necessity. Here we interrogated the uptake efficiency, biocompatibility, and stability of two subtypes of AuNRs with different types of surface coatings and varying charges, including a commercially available set of AuNRs with a 5 nm mSiO2-polymer coating and hybrid lipid-coated AuNRs developed in-house. Confocal and bright field microscopy images showed uptake of both subtypes of AuNRs in retinal pigment epithelium (RPE), neural progenitor (NP), and baby hamster kidney (BHK) cells. Transmission electron microscopy (TEM) confirms both types of AuNRs are taken up into the cytoplasm of the cells; however, larger aggregates of AuNRs are observed with the more positive and “sticky” AuNRs with a 5 nm mSiO2-polymer coating than the slightly negative hybrid lipid-coated AuNRs. Inductively Coupled Mass Spectroscopy (ICP-MS) confirm that ~3,000 of the slightly negative hybrid lipid-coated AuNRs cells and ~5,400 of the positively charged AuNRs with a 5 nm mSiO2-polymer coating (+35 mV) are taken up into RPE and BHK cell lines. Stability studies in a variety of cellular media showed that hybrid lipid-coated AuNRs are stable and disaggregated in water, 10 mM PBS buffer pH 7, and BHK media except for NP media. In contrast, the positively charged AuNRs with a 5 nm mSiO2-zeta polymer coating aggregated in all media, indicating more interactions with each other and components of the media. Bright-field and TEM confirm the presence of large aggregates of AuNRs on the surface and within the cytoplasm. Cytotoxicity studies both subtypes of AuNRs have an 80 ± 8 % cell viability, indicating mild toxicity. The hybrid lipid-coated AuNR with the cell-penetrating peptide had the least toxicological impact with a > 92 ± 7 % cell viability. Our study highlights the importance of evaluating the impact of the physicochemical features of each new nanoparticle design on their stability in biologically relevant environments and their impact on cellular uptake and toxicity in stem cell-derived therapeutic cells. Here we also provide a simple design strategy for tuning the surface chemistry of robust hybrid lipid-coated AuNRs to enhance cellular uptake to label stem cells with minimal aggregation and toxicity.
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