A selenium-based NIR-II photosensitizer for a highly effective and safe phototherapy plan.

High efficiency, stability, long emission wavelength (NIR-II), and good biocompatibility are crucial for photosensitizers in phototherapy. However, current Food and Drug Administration (FDA)-approved organic fluorophores exhibit poor chemical stability and photostability as well as short emission wavelength, limiting their clinical usage. To address this, we developed Se-IR1100, a novel organic photosensitizer with a photostable and thermostable benzobisthiadiazole (BBTD) backbone. By incorporating selenium as a heavy atom and constructing a D-A-D structure, Se-IR1100 exhibits a maximum fluorescence emission wavelength of 1100 nm. Compared with FDA-approved indocyanine green (ICG), DSPE-PEGylated Se-IR1100 nanoparticles exhibit prominent photostability and long-lasting photothermal effects. Upon 808 nm laser irradiation, Se-IR1100 NPs efficiently convert light energy into heat and reactive oxygen species (ROS), inducing cancer cell death in cellular studies and living organisms while maintaining biocompatibility. With salient photostability and a photothermal conversion rate of 55.37%, Se-IR1100 NPs hold promise as a superior photosensitizer for diagnostic and therapeutic agents in oncology. Overall, we have designed and optimized a multifunctional photosensitizer Se-IR1100 with good biocompatibility that performs NIR-II fluorescence imaging and phototherapy. This dual-strategy method may offer novel approaches for the development of multifunctional probes using dual-strategy or even multi-strategy methods in bioimaging, disease diagnosis, and therapy.

Bioresponsive Nanoarchitectonics-Integrated Microneedles for Amplified Chemo-Photodynamic Therapy against Acne Vulgaris.

The excessive colonization of Propionibacterium acnes (P. acnes) is responsible for the genesis of acne vulgaris, a common inflammatory disease of skin. However, the conventional anti-acne therapies are always limited by various side effects, drug resistance, and poor skin permeability. Microneedles (MNs) are emerging topical drug delivery systems capable of noninvasively breaking through the skin stratum corneum barrier to efficiently enhance the transdermal drug penetration. Herein, MNs loaded with intelligent pH-sensitive nanoplatforms were constructed for amplified chemo-photodynamic therapy against acne vulgaris, jointly exerting antimicrobial and anti-inflammatory effects. The photosensitizer indocyanine green (ICG) was loaded into the zeolitic imidazolate framework-8 (ZIF-8) to improve its photostability, which would be triggered by 808 nm laser irradiation to generate cytotoxic reactive oxygen species (ROS) to result in oxidative damage and disturbed metabolic activities of P. acnes. In addition to the efficient drug delivery, the ZIF-8 carrier could selectively degrade in response to the acidic microenvironment of acne lesions, and the released Zn2+ also exhibited a potent antimicrobial activity. The fabricated ZIF-8-ICG@MNs presented an outstanding synergistic anti-acne efficiency both in vitro and in vivo. This bioresponsive microneedle patch is expected to be readily adapted as a generalized, modular strategy for noninvasive therapeutics delivery against superficial skin diseases.

A mitochondrial-metabolism-regulatable carrier-free nanodrug to amplify the sensitivity of photothermal therapy.

The oxidative phosphorylation inhibitor atovaquone (ATO) and the photosensitizer new indocyanine green (IR820) were self-assembled into carrier-free nanodrugs (IR820/ATO NPs) to achieve superior photothermal therapy (PTT), offering an attractive mitochondrial metabolism-regulatable approach for breast cancer treatment, where adenosine triphosphate (ATP) was downregulated along with downregulating the expression of heat shock proteins (HSPs) to amplify the sensitivity of PTT.

Glucose-Targeted Hydroxyapatite/Indocyanine Green Hybrid Nanoparticles for Collaborative Tumor Therapy.

Nanoscale hydroxyapatite (nHA) is considered as a promising drug carrier or therapeutic agent against malignant tumors. But the strong agglomeration tendency and lack of active groups seriously hamper their usage in vivo. To address these issues, we fabricated an organic-inorganic hybrid nanosystem composed of poly(acrylic acid) (PAA), nHA, and indocyanine green (ICG), and further modified with glucose to give a targeting nanosystem (GA@HAP/ICG-NPs). These hybrid nanoparticles (∼90 nm) showed excellent storage and physiological stability assisted by PAA and had a sustained drug release in an acidic tumor environment. In vitro cell experiments confirmed that glucose-attached particles significantly promoted cellular uptake and increased intracellular ICG and Ca2+ concentrations by glucose transporter 1 (GLUT1)-mediated endocytosis. Subsequently, the excessive Ca2+ induced cell or organelle damage and ICG triggered photothermal and photodynamic effects (PTT/PDT) under laser irradiation, resulting in enhanced cell toxicity and apoptosis. In vivo tests revealed that the hybrid nanosystem possessed good hemocompatibility and biosafety, facilitating in vivo circulation and usage. NIR imaging further showed that tumor tissues had more drug accumulation, resulting in the highest tumor growth inhibition (87.89%). Overall, the glucose-targeted hybrid nanosystem was an effective platform for collaborative therapy and expected to be further used in clinical trials.

Validation of a Three-Dimensional Head and Neck Spheroid Model to Evaluate Cameras for NIR Fluorescence-Guided Cancer Surgery.

Near-infrared (NIR) fluorescence-guided surgery is an innovative technique for the real-time visualization of resection margins. The aim of this study was to develop a head and neck multicellular tumor spheroid model and to explore the possibilities offered by it for the evaluation of cameras for NIR fluorescence-guided surgery protocols. FaDu spheroids were incubated with indocyanine green (ICG) and then included in a tissue-like phantom. To assess the capability of Fluobeam® NIR camera to detect ICG in tissues, FaDu spheroids exposed to ICG were embedded in 2, 5 or 8 mm of tissue-like phantom. The fluorescence signal was significantly higher between 2, 5 and 8 mm of depth for spheroids treated with more than 5 µg/mL ICG (p < 0.05). The fluorescence intensity positively correlated with the size of spheroids (p < 0.01), while the correlation with depth in the tissue-like phantom was strongly negative (p < 0.001). This multicellular spheroid model embedded in a tissue-like phantom seems to be a simple and reproducible in vitro tumor model, allowing a comparison of NIR cameras. The ideal configuration seems to be 450 µm FaDu spheroids incubated for 24 hours with 0.05 mg/ml of ICG, ensuring the best stability, toxicity, incorporation and signal intensity.

Comparative effects of commonly used intraocular dyes on the viability of human retina Müller cells.

This study investigated the in vitro effect of various vital dyes in common clinical use on human Müller cell viability, and it compared the toxicity of these dyes using a cell culture model. Müller cells were exposed to a series of concentrations (1 %, 0.5 %, 0.25 %, and 0.125 % or 12.9 mM, 6.45 mM, 3.22 mM and 1.61 mM) of Indocyanine green (ICG) for 2, 24, 48, and 72 h. Similarly, groups of Müller cells were stained with "Heavy" brilliant blue G (HBBG), Trypan blue (TB) (0.15 %, or 1.56 mM), Membrane-blue-dual (MBD), and ICG (0.25 %, or 3.22 mM) or BBG (0.025 %, or 0.3 mM) with glucose (GS) (50 %, 66 % and 75 % or 2.78 M, 3.67 M and 4.17 M) for 30, 60, and 120 s. Cell viability was measured with the Cell Counting Kit-8 (CCK-8) and Lactate Dehydrogenase (LDH) release assays. We found that high stain concentration and long exposure time resulted in increased toxicity to Müller cells. Nevertheless, ICG seemed to be safe at the clinically relevant concentration of 0.25 % (3.22 mM) in the short time of exposure. TB was safer than both HBBG and MBD, especially HBBG. Hypertonic GS as a dilution was not safe for Müller cells, and the negative effect was more obvious in 0.025 % (0.3 mM) BBG than that in 0.25 % (3.22 mM) ICG. This is the first report to observe the cytotoxicity of commonly used stains in clinical on human Müller cells in vitro, and to provide some basis for further studies, including in vivo investigation.

Lactosylated IR820/DOX Co-Assembled Nanodrug for Synergetic Antitumour Therapy.

INTRODUCTION: Synergistic treatment integrating photothermal therapy (PTT) and chemotherapy is a promising strategy for hepatocellular carcinoma (HCC). However, the most commonly used photothermal agent, IR820, and chemotherapeutic drug, doxorubicin hydrochloride (DOX), are both hydrophilic molecules that suffer from the drawbacks of a short circulation time, rapid elimination and off-target effects. METHODS AND RESULTS: Herein, a novel nanodrug that combined HCC-targeted IR820 and DOX was developed based on excipient-free co-assembly. First, lactosylated IR820 (LA-IR820) was designed to target HCC. Then, the LA-IR820/DOX nanodrug (LA-IR820/DOX ND) was purely self-assembled without excipient assistance. The physicochemical properties and the chemo-photothermal antitumour activity of the excipient-free LA-IR820/DOX ND were evaluated. More importantly, the obtained LA-IR820/DOX ND exhibited 100% drug loading, remarkable HCC targeting and excellent antitumour efficacy. CONCLUSION: This excipient-free LA-IR820/DOX ND may be a promising candidate for the synchronous delivery and synergistic targeting of IR820 and DOX as a combined chemo-photothermal therapy.

Polylysine modified conjugated polymer nanoparticles loaded with the singlet oxygen probe 1,3-diphenylisobenzofuran and the photosensitizer indocyanine green for use in fluorometric sensing and in photodynamic therapy.

Conjugated polymer hybrid nanoparticles (NPs) loaded with both indocyanine green (ICG) and 1,3-diphenylisobenzofuran (DPBF) are described. The NPs are dually functional in that ICG acts as the photosensitizer, and DPBF as a probe for singlet oxygen (1O2 probe). The nanoparticle core consists of the energy donating host poly(9,9-dioctylfluorenyl-2,7-diyl)-co-(2,5-p-xylene) (PFP). The polymer is doped with the energy acceptor DPBF. Ratiometric fluorometric detection of singlet oxygen is accomplished by measurement of fluorescence at wavelengths of 415 and 458 nm. In addition, the shell of the positively charged polymeric nanoparticles was modified, via electrostatic interaction, with negatively charged PDT drugs ICG. The integrated nanoparticles of type ICG-DPBF-PFP display effective photodynamic performance under 808-nm laser irradiation. The 1O2 sensing behaviors of samples are evaluated based on the ratiometric fluorescent responses produced by DPBF and PFP. 1O2 can be fluorimetically sensed with a detection limit of 28 µM. The multifunctional nanoprobes exhibit effortless cellular uptake, superior photodynamic activity and a rapid ratiometric response to 1O2. Graphical abstractSchematic of a dual-functional nanoplatform for photodynamic therapy (PDT) and singlet oxygen (1O2) feedback. It offers a new strategy for self-monitoring photodynamic ablation. FRET: fluorescence resonance energy transfer. Indocyanine green is attached in the shell of nanoparticles, and 1,3-diphenylisobenzofuran is doped into the energy donating host conjugated polymer.

Tracing the molecular dynamics of living mitochondria under phototherapy via surface-enhanced Raman scattering spectroscopy.

Subcellular mitochondrion has become a target for improving the therapeutic efficiency and reducing side damage to normal cells via a combination of many therapeutic strategies. However, the underlying molecular mechanisms associated with cell death induced by subcellular dysfunction remain unknown or disputed. In this study, we investigated the dynamic molecular changes of living mitochondria upon phototherapy (photothermal therapy plus photodynamic therapy, PTT & PDT) by surface-enhanced Raman scattering spectroscopy (SERS) and intended to disclose the photo-induced cell death route in breast cancer cells (MCF-7) taking into account the mitochondrion. Indocyanine green (ICG), a Food and Drug Administration (FDA)-approved clinic blood-injection near-infrared angiographic contrast agent and a PTT & PDT drug, was used for the evaluation of the phototherapy effect. The results revealed that the content of phenylalanine (Phe) in mitochondria evidently increased during the phototherapy-induced cell death process. Moreover, the phototherapy-induced cell apoptosis was mainly regulated through the DNA structures. We expect that the understanding of mitochondrial molecular stress responses will be helpful for the diagnosis and therapy of cellular processes associated with mitochondria and provide valuable guidance for the further design and development of more effective therapeutic platforms and methods at the sub-cellular level.

In vivo irreversible albumin-binding near-infrared dye conjugate as a naked-eye and fluorescence dual-mode imaging agent for lymph node tumor metastasis diagnosis.

Tumor metastases account for about 90% of cancer-related death, among which lymphatic metastases play a pivotal role. Therefore, high-efficiency sentinel lymph node (SLN) identification is significant for lymph node (LN) metastasis diagnosis in clinic. Herein, a novel in vivo covalent albumin-binding near-infrared (NIR) fluorescent IR820-maleimide conjugate (IR-Mal) is firstly designed as a SLN dual-mode imaging agent. The IR-Mal conjugate exhibits bright blue appearance and its large Stokes shift (over 100 nm) increases the fluorescent imaging resolution effectively. The fluorescence intensity of covalent albumin-binding IR-Mal (BSA-IR-Mal) complex is considerably stronger than that of IR-Mal. In vivo, IR-Mal could rapidly covalently bind the tissue interstitial albumin following subcutaneous administration and BSA-IR-Mal complexes could specifically accumulate on LN, and detect both normal and metastatic SLN through naked-eye and fluorescence imaging with high resolution. Moreover, the light stability and enhanced fluorescence intensity of BSA-IR-Mal complex facilitates its diagnosis accuracy. These findings suggest that such in vivo irreversible albumin-binding fluorescence conjugates could serve as a new agent for dual-mode imaging and have a great potential to be applied in the SLNs imaging and diagnosis.

pH- and Enzyme-Sensitive IR820-Paclitaxel Conjugate Self-Assembled Nanovehicles for Near-Infrared Fluorescence Imaging-Guided Chemo-Photothermal Therapy.

The short lifetime and easy quick elimination of the near-infrared (NIR) dye new indocyanine green (IR820) in the body restrict its practical application as a photothermal agent in cancer therapy. Meanwhile, the drawback of poor water solubility of the chemotherapeutic drug paclitaxel (PTX) largely restricts its clinical applications. Herein, we, for the first time, combined IR820 and PTX in an "all-in-one" fluorescence imaging-guided chemo-photothermal therapy (PTT) platform by a rational design of a novel pH- and enzyme-sensitive IR820-PTX conjugate assembly. Specifically, the IR820-PTX conjugate nanoparticles exhibit an extremely high therapeutic agent content (IR820 and PTX, 95.7%). Besides the good stability in bloodstream, the IR820-PTX nanoparticles can target tumors for high accumulation via the enhanced permeation and retention effect. Particularly, our IR820-PTX nanoparticles simultaneously solve the obstacles of PTX poor solubility and the short lifetime of IR820 for cancer therapy. The simultaneous release of the free drug and dye can efficiently kill tumor cells by the combination of PTT and chemotherapy via NIR irradiation. Furthermore, the combined therapy can be imaging-guided by measuring the NIR fluorescence imaging resulting from the IR820 component. Therefore, our rationally designed pH- and enzyme-sensitive IR820-PTX conjugate nanoparticles provide an alternative "all-in-one" option for an efficient combinational dual-therapy and imaging.

Foetal bovine serum can reduce toxicity of indocyanine green, brilliant blue G and trypan blue in ARPE-19 cellular model that suggests new surgical staining protocols for internal limiting membrane peeling procedure.

BACKGROUND: To investigate and compare the cytotoxicity of indocyanine green (ICG), brilliant blue G (BBG) and trypan blue (TB) using ARPE-19 cells that have been pre-treated/post-treated with balanced salt solution (BSS) or foetal bovine serum (FBS). METHODS: The cultured human retina pigment epithelium ARPE-19 cells were pre-treated/post-treated with BSS or FBS (represent the autologous serum in clinic) in parallel with cells being soaked with various concentrations of ICG, BBG and TB. The cells were then assessed for viability, growth rate, reactive oxygen species (ROS) level, mitochondrial membrane potential (Δψ) and mitochondrial mass as cytotoxic indices. For the FBS pre-treated cells, only ROS was examined. RESULTS: Using the MTT assay, cytotoxicity seemed to appear when the dye concentration was above 2.5 mg/mL for ICG but no cytotoxicity for BBG and TB at the concentrations used. Cell growth was arrested at a concentration 1 mg/mL when ICG or BBG were present but no arrest at any of the tested concentrations was found for TB with the cell-growth curve was slowest for ICG. Cellular ROS levels increased at all concentrations of all dyes, but the increasing slopes were decreased after FBS post-treatment washout. CONCLUSIONS: As a rinse buffer FBS performs much better than BSS in terms of cell rescue, which agrees with a clinical report when autologous whole blood was applied to macular hole surgery. However, FBS pre-treatment seems to be much better than FBS use as washout buffer in post-treatment.

Ultrahigh Sensitivity Multifunctional Nanoprobe for the Detection of Hydroxyl Radical and Evaluation of Heavy Metal Induced Oxidative Stress in Live Hepatocyte.

Hydroxyl radical (•OH) is an important marker of the progress of heavy metal induced oxidative stress. However, most reported probes and detection methods cannot meet the need of monitoring the •OH concentration within the whole progress because of the limited linear range. Besides, a low detection limit, high sensitivity, and good selectivity were also required. In this study, an ultrahigh sensitivity multifunctional nanoprobe (ICG-modified NaLuF4:Yb,Er) was developed to evaluate heavy metal induced oxidative stress by detecting •OH concentration, with a colorimetric, upconversion luminescence, and photothermal stepped method. This method has a broad linear detection range, from 16 pM to 2 µM, and a low detection limit of 4 pM. Besides, the nanoprobe showed less response to ions, amino acids, biomolecules, and other radical oxygen species (H2O2 and O2-) than •OH. This highly selective, highly sensitive probe with a broad linear detection range has great potential utility for monitoring •OH concentration in live hypatocyte within the progress of heavy metal induced oxidative stress, with probable in vivo applications in the future.

Nonclinical Profile of BLZ-100, a Tumor-Targeting Fluorescent Imaging Agent.

BLZ-100 is a single intravenous use, fluorescent imaging agent that labels tumor tissue to enable more complete and precise surgical resection. It is composed of a chlorotoxin peptide covalently bound to the near-infrared fluorophore indocyanine green. BLZ-100 is in clinical development for intraoperative visualization of human tumors. The nonclinical safety and pharmacokinetic (PK) profile of BLZ-100 was evaluated in mice, rats, canines, and nonhuman primates (NHP). Single bolus intravenous administration of BLZ-100 was well tolerated, and no adverse changes were observed in cardiovascular safety pharmacology, PK, and toxicology studies in rats and NHP. The single-dose no-observed-adverse-effect-levels (NOAELs) were 7 mg (28 mg/kg) in rats and 60 mg (20 mg/kg) in NHP, corresponding to peak concentration values of 89 400 and 436 000 ng/mL and area-under-the-curve exposure values of 130 000 and 1 240 000 h·ng/mL, respectively. Based on a human imaging dose of 3 mg, dose safety margins are >100 for rat and monkey. BLZ-100 produced hypersensitivity reactions in canine imaging studies (lethargy, pruritus, swollen muzzle, etc). The severity of the reactions was not dose related. In a follow-up study in dogs, plasma histamine concentrations were increased 5 to 60 minutes after BLZ-100 injection; this coincided with signs of hypersensitivity, supporting the conclusion that the reactions were histamine based. Hypersensitivity reactions were not observed in other species or in BLZ-100 human clinical studies conducted to date. The combined imaging, safety pharmacology, PK, and toxicology studies contributed to an extensive initial nonclinical profile for BLZ-100, supporting first-in-human clinical trials.

Indocyanine Green: Historical Context, Current Applications, and Future Considerations.

BACKGROUND: Indocyanine green (ICG) is a dye used in medicine since the mid-1950s for a variety of applications in in cardiology, ophthalmology, and neurosurgery; however, its fluorescent properties have only recently been used in the intraoperative evaluation of tissue perfusion. METHOD: A literature review was conducted on the characterization and employment of ICG within the medical field. Historical and current context of ICG was examined while also considering implications for its future use. RESULTS: ICG is a relatively nontoxic, unstable compound bound by albumin in the intravascular space until rapid clearance by the liver. It has widespread uses in hepatic, cardiac, and ophthalmologic studies, and its use in analyzing tissue perfusion and identifying sentinel lymph nodes in cancer staging is gaining popularity. CONCLUSIONS: ICG has myriad applications and poses low risk to the patient. Its historical uses have contributed to medical knowledge, and it is now undergoing investigation for quantifying tissue perfusion, providing targeted therapies, and intraoperative identification of neurovascular anatomy, ophthalmic structures, and sentinel lymph nodes. New applications of ICG may lead to reduction in postoperative wound-related complications, more effective ophthalmologic procedures, and better detection and treatment of cancer cells.

Effect of Size-Dependent Photodestructive Efficacy by Gold Nanomaterials with Multiphoton Laser.

The photostability, photodestructive efficacy, two-photon excitation cross section, and two-photon fluorescence of gold nanoparticles conjugated with a hydrophilic photosensitizer, indocyanine green, via multiphoton laser exhibited an increased size effect in methicillin-resistant Staphylococcus aureus and A549 cancer cells that was dependent on the size of multifunctional gold nanomaterials, but the effect only occurred when nanomaterials within 100 nm in diameter were used. Besides, the enhanced effectiveness of photodestruction, photostability, and contrast probe indicated an additive effect in the therapeutic and imaging efficiency of multifunctional gold nanomaterials. Consequently, the preparation of the multifunctional gold nanomaterials and their use in biomedical applications via multiphoton laser is an alternative and potential therapeutic approach for killing bacteria and for ablating cancer cells.

PEDOT nanocomposites mediated dual-modal photodynamic and photothermal targeted sterilization in both NIR I and II window.

PEDOT nanoparticles with a suitable nanosize of 17.2 nm, broad adsorption from 700 to 1250 nm, and photothermal conversion efficiency (η) of 71.1%, were synthesized using an environmentally friendly hydrothermal method. Due to the electrostatic attraction between indocyanine green (ICG) and PEDOT, the stability of ICG in aqueous solution was effectively improved. The PEDOT nanoparticles modified with glutaraldehyde (GTA) targeted bacteria directly, and MTT experiments demonstrated the low toxicity of PEDOT:ICG@PEG-GTA in different bacteria and cells. Pathogenic bacteria were effectively killed by photodynamic therapy (PDT) and photothermal therapy (PTT) with PEDOT:ICG@PEG-GTA in the presence of near-infrared (NIR) irradiation (808 nm for PDT, and 1064 nm for PTT). The combination of the two different bacteriostatic methods was significantly more effective than PTT or PDT alone. The obtained PEDOT:ICG@PEG-GTA may be used as a novel synergistic agent in combination photodynamic and photothermal therapy to inactivate pathogenic bacteria in both the NIR I and II window.

Covalent IR820-PEG-diamine nanoconjugates for theranostic applications in cancer.

Near-infrared dyes can be used as theranostic agents in cancer management, based on their optical imaging and localized hyperthermia capabilities. However, their clinical translatability is limited by issues such as photobleaching, short circulation times, and nonspecific biodistribution. Nanoconjugate formulations of cyanine dyes, such as IR820, may be able to overcome some of these limitations. We covalently conjugated IR820 with 6 kDa polyethylene glycol (PEG)-diamine to create a nanoconjugate (IRPDcov) with potential for in vivo applications. The conjugation process resulted in nearly spherical, uniformly distributed nanoparticles of approximately 150 nm diameter and zeta potential -0.4±0.3 mV. The IRPDcov formulation retained the ability to fluoresce and to cause hyperthermia-mediated cell-growth inhibition, with enhanced internalization and significantly enhanced cytotoxic hyperthermia effects in cancer cells compared with free dye. Additionally, IRPDcov demonstrated a significantly longer (P<0.05) plasma half-life, elimination half-life, and area under the curve (AUC) value compared with IR820, indicating larger overall exposure to the theranostic agent in mice. The IRPDcov conjugate had different organ localization than did free IR820, with potential reduced accumulation in the kidneys and significantly lower (P<0.05) accumulation in the lungs. Some potential advantages of IR820-PEG-diamine nanoconjugates may include passive targeting of tumor tissue through the enhanced permeability and retention effect, prolonged circulation times resulting in increased windows for combined diagnosis and therapy, and further opportunities for functionalization, targeting, and customization. The conjugation of PEG-diamine with a near-infrared dye provides a multifunctional delivery vector whose localization can be monitored with noninvasive techniques and that may also serve for guided hyperthermia cancer treatments.

The effects of vital dyes on retinal pigment epithelium cells in oxidative stress.

PURPOSE: To determine the effect of the most commonly used vital dyes in vitrectomy [trypan blue at 0.15% concentration and indocyanine green (ICG) at 0.5% concentration] on the viability of human retinal pigment epithelium (RPE) cell lines (ARPE-19) exposed to oxidative stress. METHODS: ARPE-19 cells unexposed or exposed to oxidative stress (hypoxic chamber) were treated for 1 min with one of the dyes. RPE proliferation was measured by (3)H-thymidine incorporation, adhesion by ability to adhere to fibronectin, and safety by annexin V staining. RESULTS: Proliferation: The dyes affected the proliferation of RPE cells differently under non-hypoxic and hypoxic conditions (p = 0.001). In non-hypoxic conditions, there was no statistically significant difference between the proliferation of the treated (both dyes) and untreated RPE cells (p = 0.279). Under hypoxia, both dyes significantly suppressed proliferation, more so with ICG (p = 0.001). Adhesion: The dyes affected adhesion differently under non-hypoxic and hypoxic conditions (p = 0.04). In non-hypoxic conditions, both increased the adhesive properties of RPE cells to fibronectin, ICG more than trypan blue (p = 0.001). Under hypoxia, both dyes suppressed adhesion, with no statistically significant difference between treated and non-treated RPE cells. Apoptosis: Both dyes increased early apoptosis of RPE cells compared with no treatment (p = 0.001), ICG more than trypan blue. Hypoxia increased the apoptosis of both dyes compared to non-hypoxic conditions (p = 0.02). CONCLUSIONS: In hypoxic conditions, both dyes showed an inhibition of RPE adhesion to fibronectin and proliferation capacity and an increase in early apoptosis compared with non-hypoxic conditions. Apoptosis was greater in ICG-treated RPE cells than in trypan blue-treated cells.