Polymeric micelles coated with hybrid nanovesicles enhance the therapeutic potential of the reversible topoisomerase inhibitor camptothecin in a mouse model.

Erythrocyte-based carriers can serve as theranostic platforms for delivery of imaging and therapeutic payloads. Engineering these carriers at micro- or nanoscales makes them potentially useful for broad clinical applications ranging from vascular diseases to tumor theranostics. Longevity of these carriers in circulation is important in delivering a sufficient amount of their payloads to the target. We have investigated the circulation dynamics of micro (∼4.95 μm diameter) and nano (∼91 nm diameter) erythrocyte-derived carriers in real time using near-infrared fluorescence imaging, and evaluated the effectiveness of such carrier systems in mediating photothermolysis of cutaneous vasculature in mice. Fluorescence emission half-lives of micro- and nanosized carriers in response to a single intravenous injection were ∼49 and ∼15 min, respectively. A single injection of microsized carriers resulted in a 3-fold increase in signal-to-noise ratio that remained nearly persistent over 1 h of imaging time. Our results also suggest that a second injection of the carriers 7 days later can induce a transient inflammatory response, as manifested by the apparent leakage of the carriers into the perivascular tissue. The administration of the carriers into the mice vasculature reduced the threshold laser fluence to induce photothermolysis of blood vessels from >65 to 20 J/cm2. We discuss the importance of membrane physicochemical and mechanical characteristics in engineering erythrocyte-derived carriers and considerations for their clinical translation.

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Intravital Vascular Phototheranostics and Real-Time Circulation Dynamics of Micro- and Nanosized Erythrocyte-Derived Carriers

Stimuli to excitable cells and various cellular processes can cause cell surface deformations; for example, when excitable cell membrane potentials are altered during action potentials. However, these cellular changes may be at or below the diffraction limit (in dendrites the structures measured are as small as 1 µm), and imaging by traditional methods is challenging. Using dual lenses incoherent holography lattice light-sheet (IHLLS-2L) detection with holographic phase imaging of selective fluorescent markers, we can extract the full-field cellular morphology or structural changes of the object’s phase in response to external stimulus. This approach will open many new possibilities in imaging neuronal activity and, overall, in light sheet imaging. In this paper, we present IHLLS-2L as a well-suited technique for quantifying cell membrane deformation in neurons without the actuation of a sample stage or detection microscope objective.

Deformation Measurements of Neuronal Excitability Using Incoherent Holography Lattice Light-Sheet Microscopy (IHLLS)

Optical tweezers across scales in cell biology.

Particles fabricated from red blood cells (RBCs) can serve as vehicles for delivery of various biomedical cargos. Flipping of phosphatidylserine (PS) from the inner to the outer membrane leaflet normally occurs during the fabrication of such particles. PS externalization is a signal for phagocytic removal of the particles from circulation. Herein, we demonstrate that membrane cholesterol enrichment can mitigate the outward display of PS on microparticles engineered from RBCs. Our in-vitro results show that the phagocytic uptake of cholesterol-enriched particles by murine macrophages takes place at a lowered rate, resulting in reduced uptake as compared to RBC-derived particles without cholesterol enrichment. When administered via tail-vein injection into healthy mice, the percent of injected dose (ID) per gram of extracted blood for cholesterol-enriched particles was ∼1.5 and 1.8 times higher than the particles without cholesterol enrichment at 4 and 24 h, respectively. At 24 h, ∼43% ID/g of the particles without cholesterol enrichment was eliminated or metabolized while ∼94% ID/g of the cholesterol-enriched particles were still retained in the body. These results indicate that membrane cholesterol enrichment is an effective method to reduce PS externalization on the surface of RBC-derived particles and increase their longevity in circulation.

Membrane Cholesterol Enrichment of Red Blood Cell-Derived Microparticles Results in Prolonged Circulation.

Author(s): Lu, T; Anvari, B | Abstract: Ovarian cancer is the deadliest gynecological cancer in women. It is a highly metastatic disease with pelvis, regional lymph nodes, and peritoneal cavity as major sites for tumor deposits. Mechanical properties of ovarian cancer cells can play a major role in metastasis as the cells detach from the ovaries and undergo deformation during the metastatic process. Herein, we have characterized the viscoelastic properties of the plasma membrane of normal epithelial (IOSE364) and cancerous (SKOV3) ovarian cells by optical tweezers and quantitative phase imaging. Using optical tweezers, we obtained time-resolved force profiles associated with membrane tethers pulled from the cells. We used quantitative phase imaging to measure the diameter of membrane tethers, and subsequently, estimated the membrane bending modulus and membrane tension in the tether. Our results indicate that the force (190 ± 76npN) (mean ± standard deviation) required to separate the membrane of SKOV3 cells from the cytoskeleton was significantly lower (p = 0.0004) than the force (350 ± 81npN) for IOSE364 cells. The mean stiffness (2.8 ± 0.8npN/μm) of membrane tethers pulled from SKOV3 cells was significantly lower (p = 0.032) than the value for IOSE 364 cells (3.7 ± 0.8npN/μm). Mean value of the force relaxation characteristic time associated with diffusive flow of lipids was also significantly lower (p = 0.018) for SKOV3 membranes (12.9 ± 6.9ns) as compared to the value for IOSE 364 membranes (20.4 ± 6.2ns). Similarly, the mean value of the membrane bending modulus for SKOV3 cells [(0.51 ± 0.23) × 10–18nJ] was significantly lower (p = 0.007) than the value for IOSE364 cells [(1.29 ± 0.32) × 10–18nJ]. Overall, our results suggest that the membranes of SKOV3 cells are less resistant to mechanical deformation. Increased membrane susceptibility to mechanical deformation may be a facilitating factor in the metastatic behavior of cancerous ovarian cells. Characterization of membrane biomechanics may provide a useful diagnostic biomarker for assessment of the metastatic potential of ovarian cancer, and a target for development of therapeutics.

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Characterization of the Viscoelastic Properties of Ovarian Cancer Cells Membranes by Optical Tweezers and Quantitative Phase Imaging

We have fabricated constructs from erythrocytes that contain the near-infrared (NIR) dye, indocyanine green (ICG). We refer to these constructs as NIR erythrocyte mimicking transducers (NETs). Mechanical properties of NETs can play an important role in the circulation kinetics and biodistribution of these particles. We characterize the mechanical properties of erythrocytes, hemoglobin-depleted erythrocytes ghosts (EGs), and micron-sized NETs (μNETs) through analysis of membrane fluctuations measured by quantitative phase imaging, and forces associated with membrane tethers pulled by optical tweezers. EGs were prepared from erythrocytes by hypotonic treatment. μNETs were prepared through hypotonic loading of 25 μM ICG into EGs. Quantitative phase images were obtained by a common-path interferometric phaseshifting system. Approximating the membrane as a sheet of springs, we estimated the stiffness of the membrane of erythrocytes, EGs, and µNETs as 3.0 ± 0.6 pN/μm, 6.5 ± 2.1 pN/μm, and 8.0 ± 2.1 pN/μm. Optical tweezers experiments yielded a similar trend. Differences in membrane stiffness suggest that the circulation dynamics of μNETs may be altered as compared to native erythrocytes.

Mechanical characterization of erythrocyte-derived optical microparticles by quantitative phase imaging and optical tweezers

We have combined optical tweezers platform with quantitative phase imaging to characterize the mechanical properties of cells. Our results indicate that the bending modulus of ovarian cancer cells is lower than that of non-cancerous cells.

Thompson Lu

Papers

Characterization of the Viscoelastic Properties of Ovarian Cancer Cells Membranes by Optical Tweezers and Quantitative Phase Imaging

Mechanical characterization of erythrocyte-derived optical microparticles by quantitative phase imaging and optical tweezers

Combined Optical Tweezers and Quantitative Phase Imaging for Mechanical Characterization of Ovarian Cells