Dynamic photodamage of red blood cell induced by CisDiMPyP porphyrin.

It is well-known that oxidative damage in red blood cell (RBC) usually causes morphological changes and increased membrane rigidity. Although many studies have focused on investigating how RBC responds to a photodynamic stimulus, the intermediate steps between membrane damage and hemolysis are not reported. To give a comprehensive insight into changes of RBC membrane property under different oxidative damage levels, we employed the photoactivation of CisDiMPyP porphyrin that primarily generates singlet oxygen 1O2 as oxidant species. We found that there were distinguishable characteristic damages depending on the 1O2 flux over the membrane, in a way that each impact of photooxidative damage was categorized under three damage levels: mild (maintaining the membrane morphology and elasticity), moderate (membrane elongation and increased membrane elasticity) and severe (wrinkle-like deformation and hemolysis). When sodium azide (NaN3) was used as a singlet oxygen quencher, delayed cell membrane alterations and hemolysis were detected. The delay times showed that 1O2 indeed plays a key role that causes RBC photooxidation by CisDiMPyP. We suggest that the sequence of morphological changes (RBC discoid area expansion, wrinkle-like patterns, and hemolysis) under photooxidative damage occurs due to damage to the lipid membrane and cytoskeletal network proteins.

Upconversion rare Earths nanomaterials applied to photodynamic therapy and bioimaging.

Light-based therapies and diagnoses including photodynamic therapy (PDT) have been used in many fields of medicine, including the treatment of non-oncological diseases and many types of cancer. PDT require a light source and a light-sensitive compound, called photosensitizer (PS), to detect and destroy cancer cells. After absorption of the photon, PS molecule gets excited from its singlet ground state to a higher electronically excited state which, among several photophysical processes, can emit light (fluorescence) and/or generate reactive oxygen species (ROS). Moreover, the biological responses are activated only in specific areas of the tissue that have been submitted to exposure to light. The success of the PDT depends on many parameters, such as deep light penetration on tissue, higher PS uptake by undesired cells as well as its photophysical and photochemical characteristics. One of the challenges of PDT is the depth of penetration of light into biological tissues. Because photon absorption and scattering occur simultaneously, these processes depend directly on the light wavelength. Using PS that absorbs photons on "optical transparency windows" of biological tissues promises deeper penetration and less attenuation during the irradiation process. The traditional PS normally is excited by a higher energy photon (UV-Vis light) which has become the Achilles' heel in photodiagnosis and phototreatment of deep-seated tumors below the skin. Thus, the need to have an effective upconverter sensitizer agent is the property in which it absorbs light in the near-infrared (NIR) region and emits in the visible and NIR spectral regions. The red emission can contribute to the therapy and the green and NIR emission to obtain the image, for example. The absorption of NIR light by the material is very interesting because it allows greater penetration depth for in vivo bioimaging and can efficiently suppress autofluorescence and light scattering. Consequently, the penetration of NIR radiation is greater, activating the biophotoluminescent material within the cell. Thus, materials containing Rare Earth (RE) elements have a great advantage for these applications due to their attractive optical and physicochemical properties, such as several possibilities of excitation wavelengths - from UV to NIR, strong photoluminescence emissions, relatively long luminescence decay lifetimes (µs to ms), and high sensitivity and easy preparation. In resume, the relentless search for new systems continues. The contribution and understanding of the mechanisms of the various physicochemical properties presented by this system is critical to finding a suitable system for cancer treatment via PDT.

Autophagy Regulation and Photodynamic Therapy: Insights to Improve Outcomes of Cancer Treatment.

Cancer is considered an age-related disease that, over the next 10 years, will become the most prevalent health problem worldwide. Although cancer therapy has remarkably improved in the last few decades, novel treatment concepts are needed to defeat this disease. Photodynamic Therapy (PDT) signalize a pathway to treat and manage several types of cancer. Over the past three decades, new light sources and photosensitizers (PS) have been developed to be applied in PDT. Nevertheless, there is a lack of knowledge to explain the main biochemical routes needed to trigger regulated cell death mechanisms, affecting, considerably, the scope of the PDT. Although autophagy modulation is being raised as an interesting strategy to be used in cancer therapy, the main aspects referring to the autophagy role over cell succumbing PDT-photoinduced damage remain elusive. Several reports emphasize cytoprotective autophagy, as an ultimate attempt of cells to cope with the photo-induced stress and to survive. Moreover, other underlying molecular mechanisms that evoke PDT-resistance of tumor cells were considered. We reviewed the paradigm about the PDT-regulated cell death mechanisms that involve autophagic impairment or boosted activation. To comprise the autophagy-targeted PDT-protocols to treat cancer, it was underlined those that alleviate or intensify PDT-resistance of tumor cells. Thereby, this review provides insights into the mechanisms by which PDT can be used to modulate autophagy and emphasizes how this field represents a promising therapeutic strategy for cancer treatment.

Liposome and polymeric micelle-based delivery systems for chlorophylls: Photodamage effects on Staphylococcus aureus.

Chlorophyll derivatives (Chls), loaded in F-127 polymeric micelles and DPPC liposomes as drug delivery systems (DDS), have been shown to be remarkable photosensitizers for photodynamic inactivation (PDI). Assays of photoinactivation of Staphylococcus aureus bacteria (as biological models) showed that the effectiveness of Chls in these nanocarriers is dependent on photobleaching processes, photosensitizer locations in DDS, singlet oxygen quantum yields, and Chl uptake to bacteria. These are factors related to changes in Chl structure, such as the presence of metals, charge, and the phytyl chain. The photodynamic activity was significantly greater for Chls without the phytyl chain, i.e., phorbides derivatives. Furthermore, the inactivation of S. aureus was increased by the use of liposomes compared to micelles. Therefore, this research details and shows the high significance of the Chl structure and delivery system to enhance the photodynamic activity. It also highlights the chlorophylls (particularly phorbides) in liposomes as promising photosensitizers for PDI.

Isomeric effect on the properties of tetraplatinated porphyrins showing optimized phototoxicity for photodynamic therapy.

The design of new photosensitizers (PS) with improved properties is essential for the development of photodynamic therapy as an alternative therapeutic method. The conjugation of porphyrins, well known PS, with platinum(ii) complexes, potent anticancer agents, may achieve new compounds with synergistic treatment effects and no side-effects. In this study, we synthesized para and meta isomers of free-base meso-tetra(pyridyl)porphyrins complexed to [PtCl(bipy)]+ units, and investigated their photophysics in solution and in lipid membrane vesicles, correlating with cell incorporation and viability results obtained from in vitro experiments using HeLa cells. Both porphyrins showed high singlet oxygen quantum yields and phototoxicity at the nanomolar scale, with green light irradiation (522 nm) and under very low light dose (1 J cm-2). The porphyrins showed LC50 values of 25 nM (meta) and 50 nM (para), which is remarkable for such mild conditions. Moreover, the phototoxicity difference between the isomers could be assigned to the higher amphiphilicity of the meta substituted porphyrin, which leads to improved lipid membrane interaction and cellular uptake compared to the para isomer.

Photodynamic Efficiency: From Molecular Photochemistry to Cell Death.

Photodynamic therapy (PDT) is a clinical modality used to treat cancer and infectious diseases. The main agent is the photosensitizer (PS), which is excited by light and converted to a triplet excited state. This latter species leads to the formation of singlet oxygen and radicals that oxidize biomolecules. The main motivation for this review is to suggest alternatives for achieving high-efficiency PDT protocols, by taking advantage of knowledge on the chemical and biological processes taking place during and after photosensitization. We defend that in order to obtain specific mechanisms of cell death and maximize PDT efficiency, PSes should oxidize specific molecular targets. We consider the role of subcellular localization, how PS photochemistry and photophysics can change according to its nanoenvironment, and how can all these trigger specific cell death mechanisms. We propose that in order to develop PSes that will cause a breakthrough enhancement in the efficiency of PDT, researchers should first consider tissue and intracellular localization, instead of trying to maximize singlet oxygen quantum yields in in vitro tests. In addition to this, we also indicate many open questions and challenges remaining in this field, hoping to encourage future research.

Properties of chlorophyll and derivatives in homogeneous and microheterogeneous systems.

Chlorophyll (Mg-Chl) and its derivatives, zinc chlorophyll (Zn-Chl), copper chlorophyll (Cu-Chl), pheophytin (Pheo), pheophorbide (Pheid), and zinc chlorophyllide (Zn-Chld), were studied as to their acid-base equilibrium properties, hydrophobicity, stability, binding, and relative localization in neutral surfactant micellar systems. The stability order of metalochlorophyll (pH(M)) in acidic medium was found to be Cu-Chl > Zn-Chld > Zn-Chl > Mg-Chl. The apparent pK(a) for protonation of porphyrin ring nitrogens was around 1.0 for all derivatives. The pK(a) for protonation of carboxylate phorbide was 5.9 for Pheid and 2.4 for Zn-Chld. This difference was attributed to complexation of carboxylate with zinc. The hydrophobicity of chlorophyll in relation to the ability of partitioning the cell membrane lipid layer was estimated in the octanol/water biphasic system. Pheo, a more hydrophobic molecule, presented the highest partition coefficient (K(P)) in the organic phase, followed by Cu-Chl, Mg-Chl, Zn-Chl, Pheid, and Zn-Chld. The hydrophobic character was the key to relative drug location in the micellar systems. All studied derivatives interacted strongly with Tween 80 micellar systems, and particularly with P-123. For both surfactants, the order followed by binding constant (K(b)) was Zn-Chld > Pheo > Cu-Chl > Mg-Chl > Zn-Chl > Pheid, while binding constants estimated for the Chl containing the phytyl group correlated with K(P). Fluorescence quenching studies have shown that phorbides are located in a less hydrophobic region than the phytyl chain-containing derivatives, which are located preferentially in a deeper micellar microenvironment. Thus, the association of the chlorophylls with specific binding sites of micellar systems is strongly modulated by the presence of phytyl chains and metal coordinated to the porphyrinic ring.

Effects of metal and the phytyl chain on chlorophyll derivatives: physicochemical evaluation for photodynamic inactivation of microorganisms.

Chlorophyll compounds and their derivatives containing metal or phytyl chain can be used as photosensitizer in photodynamic inactivation of microorganisms (PDI). So, the physicochemical properties and antimicrobial effect of chlorophyll derivatives were investigated: Mg-chlorophyll (Mg-Chl), Zn-chlorophyll (Zn-Chl), Zn-chlorophyllide (Zn-Chlde), Cu-chlorophyll (Cu-Chl), pheophytin (Pheo) and pheophorbide (Pheid). The photobleaching experiments showed photostability according to Cu-Chl > Pheo ∼ Pheid ≫ Zn-Chl ∼ Zn-Chlde > Mg-Chl. This order was discussed in terms of metal and the phytyl chain presences. Pheid and Zn-Chl in aqueous Tween 80 solution exhibited highest singlet oxygen yield compared with the other derivatives. Chlorophyll derivatives (CD) with phytyl chain was limited by the self-aggregation phenomenon at high concentrations, even in micellar systems (Tween 80 and P-123). The antimicrobial effect of CD derivatives was investigated against Staphylococcus aureus, Escherichia coli, Candida albicans and Artemia salina. Pheid showed the best results against all organisms tested, Zn-Chlde was an excellent bactericide in the dark and Cu-Chl had no PDI effect. No correlation with CD uptake by microorganisms and darkness cytotoxicity was found. The physicochemical properties allied to bioassays results indicate that Mg-Chl, Pheo, Zn-Chl and Pheid are good candidates for PDI.