Single Mesenchymal Stromal Cell Migration Tracking into Glioblastoma Using Photoconvertible Vesicles.

Reliable cell labeling and tracking techniques are imperative for elucidating the intricate and ambiguous interactions between mesenchymal stromal cells (MSCs) and tumors. Here, we explore fluorescent photoconvertible nanoengineered vesicles to study mMSC migration in brain tumors. These 3 µm sized vesicles made of carbon nanoparticles, Rhodamine B (RhB), and polyelectrolytes are readily internalized by cells. The dye undergoes photoconversion under 561 nm laser exposure with a fluorescence blue shift upon demand. The optimal laser irradiation duration for photoconversion was 0.4 ms, which provided a maximal blue shift of the fluorescent signal label without excessive laser exposure on cells. Vesicles modified with an extra polymer layer demonstrated enhanced intracellular uptake without remarkable effects on cell viability, motility, or proliferation. The optimal ratio of 20 vesicles per mMSC was determined. Moreover, the migration of individual mMSCs within 2D and 3D glioblastoma cell (EPNT-5) colonies over 2 days and in vivo tumor settings over 7 days were traced. Our study provides a robust nanocomposite platform for investigating MSC-tumor dynamics and offers insights into envisaged therapeutic strategies. Photoconvertible vesicles also present an indispensable tool for studying complex fundamental processes of cell-cell interactions for a wide range of problems in biomedicine.

Multiple dyes applications for fluorescent convertible polymer capsules as macrophages tracking labels.

Tracing individual cell pathways among the whole population is crucial for understanding their behavior, cell communication, migration dynamics, and fate. Optical labeling is one approach for tracing individual cells, but it typically requires genetic modification to induce the generation of photoconvertible proteins. Nevertheless, this approach has limitations and is not applicable to certain cell types. For instance, genetic modification often leads to the death of macrophages. This study aims to develop an alternative method for labeling macrophages by utilizing photoconvertible micron-sized capsules capable of easy internalization and prolonged retention within cells. Thermal treatment in a polyvinyl alcohol gel medium is employed for the scalable synthesis of capsules with a wide range of fluorescent dyes, including rhodamine 6G, pyronin B, fluorescein, acridine yellow, acridine orange, thiazine red, and previously reported rhodamine B. The fluorescence brightness, photostability, and photoconversion ability of the capsules are evaluated using confocal laser scanning microscopy. Viability, uptake, mobility, and photoconversion studies are conducted on RAW 264.7 and bone marrow-derived macrophages, serving as model cell lines. The production yield of the capsules is increased due to the use of polyvinyl alcohol gel, eliminating the need for conventional filtration steps. Capsules entrapping rhodamine B and rhodamine 6G meet all requirements for intracellular use in individual cell tracking. Mass spectrometry analysis reveals a sequence of deethylation steps that result in blue shifts in the dye spectra upon irradiation. Cellular studies on macrophages demonstrate robust uptake of the capsules. The capsules exhibit minimal cytotoxicity and have a negligible impact on cell motility. The successful photoconversion of RhB-containing capsules within cells highlights their potential as alternatives to photoconvertible proteins for individual cell labeling, with promising applications in personalized medicine.

Albumin microbubbles conjugated with zinc and aluminum phthalocyanine dyes for enhanced photodynamic activity.

Gas-liquid interfaces are reaching a particular interest in biomedicine. Microbubbles, ultrasound contrast agents of clinical routine, gained increasing attention as theranostic platforms due to the preserved acoustic response, drug conjugation capabilities, and applicability in biological barrier opening. A combination of microbubbles and photodynamic therapy agents can enhance the photodynamic effect, yet the evaluation of agent conjugation on microbubble stabilization and photodynamic effect is needed. Hence, two commercially available phthalocyanine photosensitizers - Holosens® (ZnPc) and Photosens® (AlPc) - were coupled with bovine serum albumin before microbubble synthesis. We demonstrated an albumin: phthalocyanine ratio of 1:1 and covalent attachment for ZnPc, a ratio of 1:3 with electrostatic binding for AlPc. Submicron-sized microbubbles (air- and SF6- filled) had a diameter of 0.8 µm. Albumin-phthalocyanine conjugates increased the microbubble concentration and shelf-life stability compared to plain ones. We hypothesized that phthalocyanine fluorescence lifetime values decreased after conjugation with microbubbles due to narrow distance between conjugates in the shell. Agents based on AlPc demonstrated higher photodynamic activity than agents based on ZnPc, and microbubbles preserved acoustic stability in human blood plasma. The biodistribution of AlPc-conjugated microbubbles was evaluated. We conclude that our microbubble platforms demonstrate greater photodynamic activity and prolonged stability for further applications in photodynamic therapy.