Luminescent ruthenium(II)-containing metallopolymers with different ligands: synthesis and application as oxygen nanosensor for hypoxia imaging.

A series of Ru(II)-containing metallopolymers with different polypyridyl complexes, namely [Ru(N^N)2(L)](PF6)2 (L = bipyridine-branched polymer; N^N = bpy: 2,2'-bipyridine (Ru 1); phen: 1,10-phenanthroline (Ru 2); dpp: 4,7-diphenyl-1,10-phenanthroline (Ru 3)), were synthesized with the motive that adjusting π-conjugation length of ligands might produce competent luminescent oxygen probes. The three hydrophobic metallopolymers were studied with 1H NMR, UV-Vis absorption, and emission spectroscopy, and then were utilized to prepare biocompatible nanoparticles (NPs) via a nanoprecipitation method. Luminescent properties of the NPs were investigated against dissolved oxygen by steady-state and time-resolved spectroscopy respectively. Luminescence quenching of the three NPs all followed a linear behavior in the range of 0-43 ppm (oxygen concentration), but Ru 3-NPs exhibited the highest oxygen sensitivity (82%) and longest emission wavelength (λex = 460 nm; λem = 617 nm). In addition, external interferons from cellular environments (e.g., pH, temperature, and proteins) had been studied on Ru 3-NPs. Finally, dissolved oxygen in monolayer cells under normoxic/hypoxic conditions was clearly differentiated by using Ru 3-NPs as the luminescent sensor, and, more importantly, hypoxia within multicellular tumor spheroids was vividly imaged. These results suggest that such Ru(II)-containing metallopolymers are strong candidates for luminescent nanosensors towards hypoxia. Graphical abstract.

Facile synthesis of fluorinated nanophotosensitizers with self-supplied oxygen for efficient photodynamic therapy.

Tumor hypoxia severely reduces the efficiency of photodynamic therapy (PDT) through the insufficient supply of oxygen. In this work, we reported on a design of fluorinated nanophotosensitizers (NPSs) prepared by a facile reprecipitation-encapsulation method, with the aim of addressing the issue of hypoxia. The fluorinated NPSs consisted of a hybrid particle core of perfluorosiloxane-polystyrene, doped with a fluorinated photosensitizer, and a biocompatible poly-l-lysine shell. Compared with non-fluorinated counterpart NPSs that are similarly prepared except for the replacement of perfluorosiloxane with alkoxysilane, the fluorinated NPSs saturated with O2 exhibit approximately 3.5 fold higher singlet oxygen production yield and higher in vitro PDT efficiency due to the O2-carrying capability of intra-particle 'F-C' bonds.

A fluorescent nanoprobe for real-time monitoring of intracellular singlet oxygen during photodynamic therapy.

Sensing of intracellular singlet oxygen (1O2) is required in order to optimize photodynamic therapy (PDT). An optical nanoprobe is reported here for the optical determination of intracellular 1O2. The probe consists of a porous particle core doped with the commercial 1O2 probe 1,3-diphenylisobenzofuran (DPBF) and a layer of poly-L-lysine. The nanoparticle probes have a particle size of ~80 nm in diameter, exhibit good biocompatibility, improved photostability and high sensitivity for 1O2 in both absorbance (peak at 420 nm) and fluorescence (with excitation/emission peaks at 405/458 nm). Nanoprobes doped with 20% of DPBF are best suited even though they suffer from concentration quenching of fluorescence. In comparison with the commercial fluorescent 1O2 probe SOSG, 20%-doped DPBF-NPs (aged) shows higher sensitivity for 1O2 generated at an early stage. The best nanoprobes were used to real-time monitor the PDT-triggered generation of 1O2 inside live cells, and the generation rate is found to depend on the supply of intracellular oxygen. Graphical abstract A fluorescent nanoprobe featured with refined selectivity and improved sensitivity towards 1O2 was prepared from the absorption-based probe DBPF and used to real-time monitoring of the generation of intracellular 1O2 produced during PDT.

Sensitive detection of PDT-induced cell damages with luminescent oxygen nanosensors.

In this work luminescent nanosensors specifically created for intracellular oxygen (ic-O2) were utilized to assess photodynamic therapy (PDT) -induced cell damages. Firstly, ic-O2 was demonstrated to be consumed much faster than extracellular O2 with respective O2 nanosensors. Using the ic-O2 nanosensors, PDT-treated cells with different degree of impairment were then resolved according to the oxygen consumption rate (OCR). The evolving trend of cytotoxicity derived from OCRs was in agreement with cell viability obtained from 3-(4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Moreover, the direct damage of PDT on cell mitochondria was successfully detected by monitoring respiration instantly after PDT treatment, which is actually beyond the scope of MTT assay. These results suggest that fluorescence sensing of ic-O2-associated cell respiration is promising and even may become a standardized method, complementary to MTT assay, to evaluate PDT-induced cytotoxicity.

Synthesis and optimization of ZnPc-loaded biocompatible nanoparticles for efficient photodynamic therapy.

Zinc(ii) phthalocyanine (ZnPc) is a promising photosensitizer for PDT but suffers from aggregation in a physiological aqueous environment. In this paper, a class of biocompatible polymeric nanoparticles (NPs) was prepared to encapsulate ZnPc molecules. Mostly because of the planar structure, ZnPc molecules were difficult to be encapsulated into the polymeric NPs unless further coated with a thick poly-l-lysine (PLL) layer. The PLL shell endowed the NPs with good biocompatibility, efficient cellular uptake, and potential bioconjugation. The degree of aggregation (DOA) of ZnPc molecules in PLL-NPs was thoroughly investigated based on self-defined relative DOA, and a loading capacity of 4 wt% was deduced as the turning point for aggravating aggregation. Similarly, the optimal loading capacity of ZnPc was determined to be 4% according to the 1O2 generation rate, demonstrating the feasibility of the DOA approach. Polymers with large rigid units (PVK and PFO) were also utilized to relieve the aggregation of ZnPc in NPs. Taking advantage of the optimized ZnPc-loaded NPs, high PDT efficacy was demonstrated in HepG2 cells and in tumor-bearing mice as well. Both high in vitro and in vivo PDT efficacy and biocompatibility are demonstrated. Aside from affording a class of efficient biocompatible nanophotosensitizers, this work is also instructive to design other types of ZnPc-based nanocarriers, in which aggregation should be well considered.

A Pyrene@Micelle Sensor for Fluorescent Oxygen Sensing.

For most fluorescent oxygen sensors developed today, their fabrication process is either time-consuming or needs specialized knowledge. In this work, a robust fluorescent oxygen sensor is facilely constructed by dissolving pyrene molecules into CTAB aqueous solution. The as-prepared pyrene@micelle sensors have submicron-sized diameter, and the concentration of utilized pyrene can be reduced as low as 0.8 mM but still can exhibit dominant excimer emission. The excimer fluorescence is sensitive to dissolved oxygen in both intensity and lifetime, and the respective Stern-Volmer plot follows a nonlinear behavior justified by a two-site model. Because of the merits of large Stokes shift (~140 nm), easy fabrication, and robustness, the pyrene@micelle sensors are very attractive for practical determination of oxygen.