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Introduction: Development of multi-, extensively-, and pandrug-resistant (MDR, XDR, and PDR) strains of Pseudomonas aeruginosa remains a major problem in medical care. The present study evaluated the effect of antimicrobial photodynamic therapy (aPDT) as a monotherapy and in combination with colistin against P. aeruginosa isolates., Methods: Two P. aeruginosa isolates recovered from patients with respiratory tract infections were examined in this study. Minimum inhibitory concentration (MIC) of colistin was determined by the colistin broth disk elution (CBDE) and the reference broth microdilution (rBMD) methods. aPDT was performed using the photosensitizer (Ps) riboflavin at several concentrations and a light-emitting diode (LED) emitting blue light for different irradiation times with or without colistin at 1/2 × MIC concentration., Results: Both PA1 and PA2 isolates were identified as colistin-resistant P. aeruginosa with a MIC ≥4 μg/mL by the CBDE and MICs of 512 μg/mL and 256 μg/mL, respectively, by the rBMD. In aPDT, neither riboflavin nor LED light alone had antibacterial effects. The values of colony forming units per milliliter (CFU/mL) in both isolates were significantly reduced by LED + Ps treatments in a time-dependent manner (LED irradiation time) and dose-dependent manner (Ps concentration). In comparison with control, treatment with Ps (50 μM) + LED (120 s) and Ps (100 μM) + LED (120 s) resulted in 0.27 log 10  CFU/mL and 0.43 log 10  CFU/mL reductions in PA1, and 0.28 log 10  CFU/mL and 0.34 log 10  CFU/mL reductions in PA2, respectively, (P < 0.01). The best results were obtained after the combination of aPDT followed by colistin, which increased bacterial reduction, resulting in a 0.41-0.7 log 10  CFU/mL reduction for PA1 and 0.35-0.83 log 10  CFU/mL reduction for PA2 (P = 0.001)., Conclusions: This study suggests the potential implications of aPDT in combination with antibiotics, such as colistin for treatment of difficult-to-treat P. aeruginosa infections., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest., (Copyright © 2024 Japanese Society of Chemotherapy, Japanese Association for Infectious Diseases, and Japanese Society for Infection Prevention and Control. Published by Elsevier Ltd. All rights reserved.)

Background: Streptococcus mutans has been implicated as a primary causative agent of dental caries and one of its important virulence properties is an ability to form biofilm on tooth surfaces. Thus, strategies to prevent and control S. mutans biofilms are requested. The present study aimed to examine the eradication of S. mutans planktonic and biofilm cells using riboflavin (Rib)-mediated antimicrobial photodynamic therapy (aPDT) enhanced by postbiotic mediators derived from Lactobacillus species., Materials and Methods: Minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of Rib and postbiotic mediators were determined. The antimicrobial and anti-biofilm effects of Rib-mediated aPDT (Rib plus blue light), Rib-mediated aPDT in combination with postbiotic mediators derived from Lactobacillus casei (LC) (aPDT + LC ), and Rib-mediated aPDT in combination with postbiotic mediators derived from Lactobacillus plantarum (LP) (aPDT + LP ) were evaluated. The anti-virulence potential of Rib-mediated aPDT, aPDT + LC , and aPDT + LP were assessed by measuring the expression of the gtfB gene using quantitative real-time polymerase chain reaction (qRT-PCR) at the highest concentrations of Rib, LC, and LP, at which the S. mutans had proliferation as the same as in the control (non-treated) group., Results: According to the results, the MIC doses of LC, LP, and Rib were 64 µg/mL, 128 µg/mL, and 128 µg/mL, respectively, while the MBC values of LC, LP, and Rib were 128 µg/mL, 256 µg/mL, and 256 µg/mL, respectively. Rib-mediated aPDT, aPDT + LP , and aPDT + LC showed a significant reduction in Log 10 CFU/mL of S. mutans compared to the control group (4.2, 4.9, and 5.2 Log 10 CFU/mL, respectively; all P < 0.05). The most destruction of S. mutans biofilms was observed after treatment with aPDT + LC followed by aPDT + LP and Rib-mediated aPDT (77.5%, 73.3%, and 67.6%, respectively; all P < 0.05). The concentrations of 31.2 µg/mL, 62.5 µg/mL, and 62.5 µg/mL were considered as the highest concentrations of LC, LP, and Rib, respectively, at which S. mutans replicates as same as the control group and were used for gtfB gene expression assay using qRT-PCR during Rib-mediated aPDT, aPDT + LP , and aPDT + LC treatments. Gene expression results revealed that aPDT + LP and aPDT + LC could decrease the gene expression level of gtfB by 6.3- and 5.7-fold, respectively (P < 0.05), while only 5.1-fold reduction was observed after Rib-mediated aPDT (P < 0.05)., Conclusion: Our findings indicate that aPDT + LP and aPDT + LC hold promise for use as a treatment to combat S. mutans planktonic and biofilms growth as well as anti-virulence as a preventive strategy to inhibit biofilms development via reduction of gtfB gene expression., (© 2024. The Author(s).)

Background: Peri-implantitis is a destructive inflammatory disease affecting both hard and soft tissues of the osseointegrated implant and causing bone loss and envelope surrounding the implant. The study aimed at evaluating the effect of Photodynamic therapy with Curcumin and Riboflavin on the level of decontamination of implant surface impregnated with Aggregatibacter actinomycetemcomitans (A.a) biofilm., Materials and Methods: In this experimental and laboratory study, 42 implants (4.3 mm in diameter and 8 mm in length) were infected with A.a. bacterial suspension. Then, the implants carrying A.a biofilm were randomly divided into seven groups (n = 6). The groups included: 1- a negative control group (without treatment), 2- a positive control group of Chlorhexidine 0.12 %, 3- a Curcumin (5 mg/ ml) group, 4- a Riboflavin (0.5 %) group, 5- an LED irradiation group (390-480 nm), 6- a photodynamic therapy with Curcumin group, and 7- a photodynamic therapy with Riboflavin group. Then, the implants were sonicated and the amount of CFU/mL of each sample was calculated. One-way ANOVA and Tamhane tests were used to analyze the data., Results: The lowest mean number of colonies of A.a (CFU/ mL) were seen in the following groups, respectively: the positive control group of Chlorhexidine 0.12 %, the photodynamic therapy with Curcumin group, the photodynamic therapy with Riboflavin group, the Curcumin (5 mg/ ml) group, the Riboflavin (0.5 %) group, the LED radiation group, and the negative control group. The use of photodynamic therapy with Curcumin significantly reduced the number of colonies of A.a (CFU/ mL) in comparison with the photodynamic therapy with Riboflavin group (p = 0.004), the Riboflavin group (p = 0.045), the LED radiation group (p = 0.012), and the negative control group (p = 0.007)., Conclusion: aPDT with Curcumin and LED can reduce A.a biofilm on implant surfaces and can be used as a safe and non-invasive disinfection method to reduce A.a biofilm on implant surfaces., (Copyright © 2023. Published by Elsevier B.V.)

Wounds infected by Pseudomonas aeruginosa (P. aeruginosa) biofilms are characterized by poor healing and by being long lasting. Pyocyanin and pyoverdine are exotoxins that contribute to P. aeruginosa pathogenicity in wound infections and are known as virulence factors. Despite the usefulness of antimicrobial photodynamic therapy (PDT) in the management of wound infections, biofilms are hurdle for microbial photoinactivation. Quorum sensing (QS) is a cell density-dependent chemical signaling system P. aeruginosa uses to regulate biofilm formation and virulence factors production. In the current study, QS attenuation was used in combination with PDT against P. aeruginosa biofilm cultured on skin explant. Iberin is a QS inhibitor that attenuates P. aeruginosa virulence and affects biofilm integrity. The antibiofilm and QS inhibitory activities of iberin in combination with either riboflavin or 5,10,15,20-Tetrakis(1-methyl-4-pyridinio) porphyrin tetra p-toluenesulfonate (TMP) mediated PDT were investigated using viable count method and pyocyanin and pyoverdine assays, respectively. No bactericidal activity was reported when iberin was added to a mature biofilm (24 h) followed by PDT. When added to a growing biofilm at multiple time points (0 h, 24 h and 48 h), iberin inhibited P. aeruginosa biofilm QS signaling system. This inhibitory effect resulted in an observable decrease in the levels of the QS-regulated virulence factors, pyocyanin and pyoverdine, without any effect on the growth of the biofilm cultures. These changes in biofilm virulence were associated with a decrease in biofilm resistance to PDT and caused bactericidal effect upon photosensitizers treatment and irradiation. Iberin-treated-riboflavin-mediated PDT resulted in a significant 1.3 log reduction in biofilm population. Similarly, iberin-treated-TMP-mediated PDT caused a significant 1.8 log reduction in biofilm population. The combination of QS inhibitor with PDT is a promising alternative antimicrobial therapy for the management of biofilms., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Riboflavin-5'-phosphate (FMN), an innocuous product of riboflavin (RF) phosphorylation, is vital for humans. FMN is sensitive to light illumination, as indicated by reactive oxygen species (ROS) formation. This investigation was undertaken to evaluate the influence of blue light illumination (BLI) and violet light illumination (VLI) upon FMN to develop a method to inhibit WiDr colon cancer cells by FMN photolysis. When FMN is subjected to BLI and VLI, it inhibits WiDr colon cancer cells by generating superoxide radical anions (O 2 •- ). The respective reduction rates are 42.6 and 81.9 % in WiDr colon cancer cells for FMN treated with BLI and VLI at 20 W/m 2 for 0.5 h. FMN treated with VLI inhibits WiDr colon cancer cells more effectively than BLI. Propidium iodide (PI) is a fluorescent dye that is used to detect abnormal DNA due to cell death by apoptosis or necrosis. The PI-positive count for nuclei increased significantly for the WiDr colon cancer cells that were treated with FMN under VLI at 20 W/m 2 for 0.5 h. FMN photolysis achieved using VLI allows efficient photodynamic therapy (PDT) by triggering the cytotoxicity of FMN on WiDr colon cancer cells., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Riboflavin, a water-soluble vitamin B2, possesses unique biological and physicochemical properties. Its photosensitizing properties make it suitable for various biological applications, such as pathogen inactivation and photodynamic therapy. However, the effectiveness of riboflavin as a photosensitizer is hindered by its degradation upon exposure to light. The review aims to highlight the significance of riboflavin and its derivatives as potential photosensitizers for use in photodynamic therapy. Additionally, a concise overview of photodynamic therapy and utilization of blue light in dermatology is provided, as well as the photochemistry and photobiophysics of riboflavin and its derivatives. Particular emphasis is given to the latest findings on the use of acetylated 3-methyltetraacetyl-riboflavin derivative (3MeTARF) in photodynamic therapy.

Objective: The high incidence of endodontic failure is associated with the remnants of Enterococcus faecalis present within the intricate anatomies of the root canal system (RCS), often inaccessible by the current endodontic practices. This study was aimed at evaluating the effect of high-intensity focused ultrasound (HIFU) and photodynamic therapy (PDT) on E. faecalis biofilms in artificially infected root canals for the potential application in current endodontic practices., Methods: Forty-five single-rooted extracted teeth were instrumented using hand files, sterilized in an autoclave, infected with E. faecalis and incubated for 4 wk. The specimens were treated and identified as follows: Control, 4% sodium hypochlorite (NaOCl); riboflavin (1 mg/mL); light only; HIFU (250 kHz, 20 W, 60s); PDT; riboflavin/HIFU; light/HIFU; and riboflavin/HIFU/light. Bactericidal efficacy was determined by colony-forming units (CFU), (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) (MTT) assay, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM)., Results: Enterococcus faecalis biofilm exhibited significantly lower metabolic activity when treated with HIFU (250 kHz, 20 W, 60 s) compared with the control (4% NaOCl) and PDT groups. A similar phenomenon was observed with the CFU assay. HIFU remained the most effective treatment modality, with consistent results in CLSM and SEM., Conclusion: This study highlighted the potential application of HIFU as an adjunct drug-free, non-destructive root canal disinfection method for endodontic treatment, suggesting an alternative to the current gold standard of 4% NaOCl and PDT., Competing Interests: Conflict of interest The authors declare no competing interests., (Copyright © 2023 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.)

Objective: To evaluate the outcome of post space disinfection using Er,Cr:YSGG laser (ECYL), riboflavin photosensitizer (RFP), and photon-induced photoacoustic streaming (PIPS) along with ethylene diamine tetra acetic acid (EDTA) as final irrigant on the bond values of glass fiber post (GFP) to the canal wall. Materials and methods: Forty human mandibular premolars were subjected to root canal treatment after decoronation till the cementoenamel junction. Post space was prepared using a Gates-Glidden drill leaving 4 mm of gutta perch at the apical third of the root section. Random allocation of samples was performed into four groups based on the post space disinfection ( n  = 10 each): Group 1: 1% RFP and 17% EDTA, group 2: 5.25% sodium hypochlorite (NaOCl) and 17% EDTA (control), group 3: PIPS and 17% EDTA, and group 4: ECYL and 17% EDTA. GFP was luted using Rely X Unicem. A universal testing machine and stereomicroscope were used to assess the push-out bond strength (PBS) and failure mode, respectively. Analysis of variance determined the PBS amid different experimental groups at three root levels. Multiple group comparison using means of tested groups was identified using post hoc Tukey. Results: The coronal third of group 2 (5.25% NaOCl +17% EDTA) demonstrated the highest PBS (8.83 ± 0.11 MPa). However, the lowest bond integrity (4.15 ± 0.52 MPa) was displayed in an apical third of group 1 (RFP +17% EDTA). The intergroup comparison revealed that 5.25% NaOCl +17% EDTA at all three levels demonstrated comparable outcomes of PBS with PIPS +17% EDTA and ECYL +17% EDTA specimens ( p  > 0.05). However, riboflavin +17% EDTA demonstrated the lowest bond strength ( p  < 0.05). Conclusions: Er,Cr: YSGG and PIPS in combination with 17% EDTA demonstrated comparable outcomes of bond score with those of conventional gold standard irrigation regime 5.25% NaOCl.

Objectives: This study aimed to evaluate the inactivation potency of riboflavin-mediated blue diode laser photodynamic inactivation (PDI) against Enterococcus faecalis at planktonic and biofilm stages and also investigated its effect on the tooth color change., Materials and Methods: The antibacterial and antibiofilm activities of riboflavin mediated PDI against E. faecalis were investigated. The numbers of colony-forming units (CFUs)/mL were calculated. Teeth discoloration were evaluated using the CIE L*a*b* based color difference (ΔE)., Results: Antibacterial analysis indicated that the blue diode laser irradiation at 12, 18, 24, and 30 J/cm 2 alone and different concentrations of riboflavin solution (6.25 to 100 μM) reduced the number of CFU/mL of E. faecalis, but the reduction was not statistically significant (P > 0.05). Depending on the riboflavin concentration and the light dose, there can be as much as a 1-log effect on CFU/mL. In addition, E. faecalis biofilm was more affected with 30 J/cm 2 irradiation dosage and 100 μM riboflavin than other groups. Meanwhile, bacterial suspensions treated with 5.25% sodium hypochlorite (NaOCl) showed maximum biofilm inhibition and colony number reduction, compared with the control. The teeth exhibited clinically acceptable color change after riboflavin treatment at concentration ranging from 6.25 to 50 μM (ΔE < 3.7)., Conclusions: The riboflavin mediated PDI process is somewhat less effective than NaOCl but perhaps less toxic to tissues. It might be feasible to repeat the riboflavin + light treatment to further promote efficacy., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Photodynamic therapy (PDT) is a light triggered therapy by producing reactive oxygen species (ROS), but traditional PDT may suffer from the real-time illumination that reduces the compliance of treatment and cause phototoxicity. A supramolecular photoactive G-quartet based material is reported, which is self-assembled from guanosine (G) and 4-formylphenylboronic acid/1,8-diaminooctane, with incorporation of riboflavin as a photocatalyst to the G4 nanowire, for post-irradiation photodynamic antibacterial therapy. The G4-materials, which exhibit hydrogel-like properties, provide a scaffold for loading riboflavin, and the reductant guanosine for the riboflavin for phototriggered production of the therapeutic H 2 O 2 . The photocatalytic activity shows great tolerance against room temperature storage and heating/cooling treatments. The riboflavin-loaded G4 hydrogels, after photo-irradiation, are capable of killing gram-positive bacteria (e.g., Staphylococcus aureus), gram-negative bacteria (e.g., Escherichia coli), and multidrug resistant bacteria (methicillin-resistant Staphylococcus aureus) with sterilization ratio over 99.999%. The post-irradiated hydrogels also exhibit great antibacterial activity in the infected wound of the rats, revealing the potential of this novel concept in the light therapy., (© 2023 The Authors. Advanced Science published by Wiley-VCH GmbH.)

The photodynamic therapy (PDT) is considered as a noninvasive and photo-controlled treatment for various cancers. However, its potential is not fully developed as current clinically approved photosensitizers (PSs) mainly absorb the light in the UV-visible region (less than 700 nm), where the depth of penetration is inadequate for reaching tumor cells under deeper tissue layers. Furthermore, the lack of specific accumulation capability of the conventional PSs in the tumor cells may cause serious toxicity and low treatment efficiency. To address these problems, riboflavin (Rf) conjugated and amine-functionalized nitrogen-doped graphene quantum dots (am-N-GQD) are herein proposed. Rf functions as both photosensitizer and targeting ligand by indirect excitation through intra-particle fluorescence resonance energy transfer (FRET) via two-photon (TP) excited am-N-GQD, to enhance the treatment depth, and further am-N-GQD-Rf accumulation in cancer cells using Rf transporter family (RFVTs) and Rf carrier proteins (RCPs). The one-photon (OP) and two-photon(TP)-PDT effect and cellular internalization ability of the am-N-GQD-Rf were investigated in vitro in different cancel cell lines. Besides the excellent cellular uptake as well TP-PDT capability, the superior biocompatibility of am-N-GQD-Rf in vitro makes it promising candidate in PDT., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier B.V.)

Acinetobacter baumannii is a dangerous hospital pathogen primarily due to its ability to form biofilms on different abiotic and biotic surfaces. The present study investigated the effect of riboflavin- and chlorophyllin-based antimicrobial photodynamic therapy, performed with near-ultraviolet or blue light on the viability of bacterial cells in biofilms and their structural stability, also determining the extent of photoinduced generation of intracellular reactive oxygen species as well as the ability of A. baumannii to form biofilms after the treatment. The efficacy of antimicrobial photodynamic therapy was compared with that of light alone and the role of the photosensitizer type on the photosensitization mechanism was demonstrated. We found that the antibacterial effect of riboflavin-based antimicrobial photodynamic therapy depends on the ability of photoactivated riboflavin to generate intracellular reactive oxygen species but does not depend on the concentration of riboflavin and pre-incubation time before irradiation. Moreover, our results suggest a clear interconnection between the inactivation efficiency of chlorophyllin-based antimicrobial photodynamic therapy and the sensitivity of A. baumannii biofilms to used light. In summary, all the analyzed results suggest that riboflavin-based antimicrobial photodynamic therapy and chlorophyllin-based antimicrobial photodynamic therapy have the potential to be applied as an antibacterial treatment against A. baumannii biofilms or as a preventive measure against biofilm formation.

Objective: To evaluate the bond efficacy and failure rates of rebonded metallic brackets after enamel reconditioning with chemical 37% phosphoric acid (PA) and natural and synthetic photosensitizers activated by PDT., Materials and Methods: A total of 50 non-cavitated, and cautiously extracted human premolars were congregated after sample size calculation. The enamel exterior was etched, washed, dried for adhesive application, and cured. Metallic brackets were then oriented and adapted to enamel surface using composite. Later, brackets were debonded from the surface via a Weingart plier. Enamel was finished for ensuing surface reconditioning. Ultimately, specimens were randomly distributed into five groups (n=10). Enamel surface before rebonding was reconditioned with curcumin photosensitizer (CP), riboflavin photosensitizer (RP), rose bengal photosensitizer (RBP), methylene blue photosensitizer (MBP), and 37% PA (control) respectively. After following reconditioning protocol, brackets were rebonded to the enamel exterior employing a composite adhesive system. Then, specimens were subjected to the universal testing machine for analyzing shear bond strength (SBS), and bond failures were predicted using an ARI index. One-way ANOVA and Tukey multiple comparison tests were used for statistical analysis at a variance value of p < 0.05., Results: Enamel reconditioned with 37% PA demonstrated the highest SBS for bracket rebonding, and the lowest SBS was presented by CP actuated by PDT. Enamel reconditioned with RP and RBP corroborated the analogous SBS outcome to 37% PA. Likewise, enamel surface treatment with MBP revealed a statistically significant result to CP for metallic bracket rebonding. The most prevalent failure scores anticipated among groups were 0 and 1 indicating an adhesive failure with the exemption of group 5 (control) that encountered more score 2 cohesive failure on debonding metallic brackets from enamel exterior., Conclusions: Rose bengal and riboflavin photosensitizers activated by photodynamic therapy with low ARI scores have the potential to be used as viable enamel reconditioning alternatives to 37% phosphoric acid for rebonding metallic brackets.

Background: Dental caries remain a significant global health challenge. Unfortunately, current dental materials lack sufficient antimicrobial power to address the pathogenic species involved in this disease. In this study the potential to load a dental resin blend (RB) with riboflavin (B2) for use in an antimicrobial photodynamic therapy (aPDT) approach was investigated., Methods: B2 was added to our experimental RB (0.1 - 10 wt%). Upon investigating the degree of conversion and specimen integrity of the RB as a function of B2 concentration, it was determined that loading should be restricted to 0.1, 1.0, and 1.5 wt%. Subsequent investigation included water sorption (WS) and solubility (SL), as well as shear bond strength (SBS) and flexural strength (FS) of the specimens after 24 h and 28-day water storage. Lastly, the antimicrobial response of Streptococcus mutans (S. mutans) biofilm following 6 h growth and 60 s of blue LED light (1.3 J/cm 2 ) in an aPDT-based approach was measured., Results and Conclusions: Adding up to 1.5 wt% B2 had minimal impact on the FS or SBS of the RB. However, aging for 28-days notably increased the FS by as much as 50% for the 1.5 wt% B2-loaded RB. In addition, adding 1.5 wt% B2 resulted in a significant reduction in WS/SL of the RB. Lastly, while adding B2 did not change the antimicrobial response, this was an initial study under these conditions and future investigation will seek to optimize light parameters to produce a more agonistic response. Overall, a riboflavin-loaded dental resin shows significant promise for utilization in restorative dentistry with aPDT., Competing Interests: Declarations of Competing Interest None., (Copyright © 2022. Published by Elsevier B.V.)

Aim: The present study aims to evaluate the bond integrity of composite restoration bonded to CAD after disinfection using pineapple peel extract (PPE) Diode laser (DL) and riboflavin (RF) activated by photodynamic therapy (PDT)., Material and Methods: Forty human permanent mandibular molars were collected, disinfected, and mounted in poly-vinyl pipes. Carious infected dentin was removed and CAD was disinfected. Samples were divided into 4 groups based on disinfection. Group 1: CAD disinfected with CHX; group 2: CAD disinfected with diode laser; CAD disinfected with PPE and CAD sterilized with riboflavin. Following disinfection, CAD surfaces were applied with the etch-and-rinse adhesive system and bonding agent and restored with a composite restoration. Samples were placed in a thermocycler to simulate oral aging and then mounted on the universal testing machine (UTM) for shear bond strength (SBS) testing. Failure mode was assessed using a steriomicroscoscope at 40X magnification. One-way analysis of variance (ANOVA) compared the mean SBS and failure type obtained to identify intergroup differences. Similarly, for multiple group comparison, the Tukey HSD test was used keeping the level of significance at 0.05., Results: CAD disinfected with chlorhexidine (CHX) established the highest bond integrity of the restoration. However, group 2 CAD disinfected with DL displayed the lowest bond values. It was also observed that dentin disinfected using CHX demonstrated comparable SBS values to CAD disinfected with PPE and CAD disinfected with RF., Conclusion: RF and PPE displayed comparable outcomes of SBS of composite restoration bonded to CAD to that of CHX. DL displayed lower bond integrity than the other investigated groups. Further studies are recommended to deduce the findings of the existing study., Competing Interests: Declaration of Competing Interest The author declares no conflict of interest., (Copyright © 2022. Published by Elsevier B.V.)

More than half of the global population lives in areas where the Aedes aegypti mosquito is present. Efforts have been made to deal with the population of this mosquito in the larval and adult stages to prevent outbreaks of diseases (Dengue, Zika, Chikungunya, and Yellow Fever). In this scenario, photodynamic inactivation may be an effective alternative method to control this vector population. To evaluate the efficacy of the riboflavin - B2 vitamin - as photosensitizer (PS) in the photodynamic inactivation of Ae. aegypti larvae, different concentrations (0; 0.005; 0.010; 0.025; 0.050; 0.075 and 0.100 mg mL -1 ) were evaluated under white light from RGB LEDs at a light dose of 495.2  J cm -2 . The results reveal that riboflavin can be successfully applied as a PS agent to photoinactivate Ae. aegypti larvae, showing its potential to deal with the larvae population., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Introduction: The present study aims to determine the effect of different post-space final irrigants ethylenediaminetetraacetic acid (EDTA), a mixture of Doxycycline citric acid and detergent (MTAD), Riboflavin (RF) and Pineapple peel extract (PPE) on the bond integrity of GFPs bonded to radicular dentin., Materials and Methods: Crowns of forty human mandibular premolars teeth were sectioned up to cementoenamel junction CEJ. Root canal treatment was performed. Gates Glidden drills were used to retrieve gutta-percha from the canal. All the specimens were randomly divided into four groups based on the final irrigant used. Group 1 (NaOCl with EDTA), group 2 NaOCl+ MTAD, group 3 NaOCl+RF and group 4 NaOCl+PPE. Cementation of glass fiber posts (GFPs) was performed using self-etch dual-cure resin cement. Sectioning was performed using a slow-speed saw from the coronal, middle, and apical third under constant water irrigation. A universal testing machine was used to assess push-out bond strength (PBS) and failure mode was evaluated using a Stereomicroscope at 40x magnification. One-way analysis of variance (ANOVA) and Tukey multiple comparison tests were used for statistical analysis (p = 0.05)., Results: Group 4 (2.5% NaOCl+ PPE) displayed the highest PBS at all three root levels (Cervical: 11.22 ± 0.74 MPa, Middle: 8.99 ± 0.97 MPa, and Apical: 6.00 ± 0.88 MPa). However, Group 1 (2.5% NaOCl+17% EDTA) demonstrated the lowest EBS (Cervical: 9.25 ± 0.17 MPa, Middle: 7.01 ± 1.84 MPa, and Apical: 4.41 ± 0.17 MPa). Inter-group comparison displayed that PBS to be comparable between group 1 and group 3 (2.5% NaOCl+ Riboflavin) (Cervical: 9.74 ± 0.41 MPa, Middle: 7.44 ± 1.18 MPa, and Apical: 4.21 ± 0.05 MPa) (p > 0.05). Whereas, group 2 (1.3% NaOCl+ MTAD) (Cervical: 9.32 ± 1.36 MPa, Middle: 7.17 ± 1.75 MPa, and Apical: 4.85 ± 0.19 MPa) and group 4 demonstrated comparable values of PBS (p > 0.05)., Conclusion: Pineapple peel extract and a mixture of Doxycycline citric acid and a detergent when used as a final irrigant demonstrated better bond integrity of GFPs to radicular dentin at all three levels coronal, middle and apical., (Copyright © 2022. Published by Elsevier B.V.)

The aim of this study was to assess the effect of antimicrobial photodynamic therapy (aPDT) with curcumin and riboflavin on three-week Enterococcus faecalis biofilm. At first the 15-mm root canals of 65 single rooted extracted human teeth (including maxillary incisors, mandibular and maxillary canines and mandibular premolars) were separated from the crown and were prepared with ProTaper instruments. After autoclave sterilisation, samples were inoculated with E. faecalis suspension, and incubated for three weeks. After ensuring biofilm formation by scanning electron microscopy (SEM) in two teeth, the remaining 63 teeth were randomly divided into seven groups (n = 9): aPDT + curcumin, aPDT + riboflavin, LED, curcumin, riboflavin, 5.25% NaOCl (positive control) and no intervention (negative control). For light source a LED unit with 390-480 nm wavelength (peak of 460 nm), power density of 1000 ± 100 mW cm -2 and mean energy density of 60 J cm -2 was used. The roots were horizontally sectioned into coronal, middle and apical thirds each with 5 mm thicknesses. Dentin chips with equal weight (1 ± 0.005 g) were collected from the root canal walls with Gates Glidden drills and were transferred into microtubes containing 1 mL of sterile saline and vortexed for 30 s. Next, 10 µL of the contents of each tube was serially diluted and eventually, 10 µL of each solution was cultured on BHI agar. The number of colony-forming units was determined. Data were analysed using the Kruskal-Wallis and Friedman tests. The colony reduction was not significantly different between NaOCl and either riboflavin + LED or Curcumin + LED. The 5.25% NaOCl group showed maximum reduction in colony count, compared with the negative control (P = 0.00). Groups with aPDT with Curcumin + LED (P = 0.005), and with riboflavin + LED (P = 0.011) showed significant reduction in colony count in all three canal thirds (P < 0.05) without any difference with one another. With significant reduction of E. faecalis colony count, aPDT with Curcumin and riboflavin can serve as an adjunct to routine root canal disinfection method., (© 2021 Australian Society of Endodontology Inc.)

Corneal collagen crosslinking (CXL) is a commonly used minimally invasive surgical technique to prevent the progression of corneal ectasias, such as keratoconus. Unfortunately, riboflavin/UV-A light-based CXL procedures have not been successfully applied to all patients, and result in frequent complications, such as corneal haze and endothelial damage. We propose a new method for corneal crosslinking by using a Ruthenium (Ru) based water-soluble photoinitiator and visible light (430 nm). Tris(bipyridine)ruthenium(II) ([Ru(bpy) 3 ] 2+ ) and sodium persulfate (SPS) mixture covalently crosslinks free tyrosine, histidine, and lysine groups under visible light (400-450 nm), which prevents UV-A light-induced cytotoxicity in an efficient and time saving collagen crosslinking procedure. In this study, we investigated the effects of the Ru/visible blue light procedure on the viability and toxicity of human corneal epithelium, limbal, and stromal cells. Then bovine corneas crosslinked with ruthenium mixture and visible light were characterized, and their biomechanical properties were compared with the customized riboflavin/UV-A crosslinking approach in the clinics. Crosslinked corneas with a ruthenium-based CXL approach showed significantly higher young's modulus compared to riboflavin/UV-A light-based method applied to corneas. In addition, crosslinked corneas with both methods were characterized to evaluate the hydrodynamic behavior, optical transparency, and enzymatic resistance. In all biomechanical, biochemical, and optical tests used here, corneas that were crosslinked with ruthenium-based approach demonstrated better results than that of corneas crosslinked with riboflavin/ UV-A. This study is promising to be translated into a non-surgical therapy for all ectatic corneal pathologies as a result of mild conditions introduced here with visible light exposure and a nontoxic ruthenium-based photoinitiator to the cornea. STATEMENT OF SIGNIFICANCE: Keratoconus, one of the most frequent corneal diseases, could be treated with riboflavin and ultraviolet light-based photo-crosslinking application to the cornea of the patients. Unfortunately, this method has irreversible side effects and cannot be applied to all keratoconus patients. In this study, we exploited the photoactivation behavior of an organoruthenium compound to achieve corneal crosslinking. Ruthenium-based organic complex under visible light demonstrated significantly better biocompatibility and superior biomechanical results than riboflavin and ultraviolet light application. This study promises to translate into a new fast, efficient non-surgical therapy option for all ectatic corneal pathologies., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have influenced the work reported in this paper., (Copyright © 2022 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Aim and Objectives: The aim of this study was to evaluate the effect of antimicrobial photodynamic therapy (aPDT) with riboflavin and curcumin on the shear bond strength (SBS) of the orthodontic brackets., Method and Material: A total of 45 human premolar teeth were used in this study. All teeth were examined under a stereomicroscope. The samples were divided into 3 groups including no intervention (control group), aPDT with riboflavin and 460 nm LED and aPDT with curcumin and 460 nm LED. After aPDT procedure, orthodontic brackets were bonded to enamel surfaces. Then, the samples were thermocycled for 3000 cycles. The brackets were then debonded using a universal testing machine. The adhesive remnant index (ARI) score was assessed. ANOVA with Post-hoc test was used to compare the SBS values between groups., Results: The highest SBS mean value was presented in curcumin group (25.95±3.68) whereas, the lowest SBS mean value was observed in riboflavin group (22.19±4.73). The mode of failure was mostly score 1 and score 2 in all groups. There was no significant difference in SBS values and ARI score between groups. Scanning electron microscope images of the curcumin and control groups showed the honey comb structure, while the microscopic view of the riboflavin group lacked this structure and had less porosity and irregularity after etching., Conclusion: Riboflavin and curcumin mediated antimicrobial photodynamic therapy both have acceptable bond strength of the orthodontic brackets and can be used before bonding to reduce inflammation and elimination of microbial biofilms., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Riboflavin-5-phosphate (RF) is the most commonly used photosensitizer in corneal cross-linking (CXL), but its hydrophilicity and negative charge limit its penetration through the corneal epithelium into the stroma. To enhance the corneal permeability of RF and promote its efficacy in the treatment of keratoconus, novel hibiscus-like RF@ZIF-8 microsphere composites [6RF@ZIF-8 NF (nanoflake)] are prepared using ZIF-8 nanomaterials as carriers, which are characterized by their hydrophobicity, positive potential, biocompatibility, high loading capacities, and large surface areas. Both hematoxylin and eosin endothelial staining and TUNEL assays demonstrate excellent biocompatibility of 6RF@ZIF-8 NF. In in vivo studies, the 6RF@ZIF-8 NF displayed excellent corneal permeation, and outstanding transepithelial CXL (TE-CXL) efficacy, slightly better than the conventional CXL protocol. Furthermore, the special hibiscus-like structures of 6RF@ZIF-8 NF meant that it has better TE-CXL efficacy than that of 6RF@ZIF-8 NP (nanoparticles) due to the larger contact area with the epithelium and the shorter RF release passage. These results suggest that the 6RF@ZIF-8 NF are promising for transepithelial corneal cross-linking, avoiding the need for epithelial debridement., (© 2022 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

The development of effective pathogen reduction strategies is required due to the rise in antibiotic-resistant bacteria and zoonotic viral pandemics. Photodynamic inactivation (PDI) of bacteria and viruses is a potent reduction strategy that bypasses typical resistance mechanisms. Naturally occurring riboflavin has been widely used in PDI applications due to efficient light-induced reactive oxygen species (ROS) release. By rational design of its core structure to alter (photo)physical properties, we obtained derivatives capable of outperforming riboflavin's visible light-induced PDI against E. coli and a SARS-CoV-2 surrogate, revealing functional group dependency for each pathogen. Bacterial PDI was influenced mainly by guanidino substitution, whereas viral PDI increased through bromination of the flavin. These observations were related to enhanced uptake and ROS-specific nucleic acid cleavage mechanisms. Trends in the derivatives' toxicity towards human fibroblast cells were also investigated to assess viable therapeutic derivatives and help guide further design of PDI agents to combat pathogenic organisms., (© 2022. The Author(s).)

Aim: To assess the influence of different photosensitizers activated by PDT as a disinfectant in comparison to conventional sodium hypochlorite (NaOCl) on the EBS (extrusion bond strength) of FRCP with radicular dentin., Methods: A total of fifty single-rooted human maxillary central incisors with fully developed apices were selected. Endodontic treatment of samples was performed using 10 K file to obtain patency than sequentially with a 25 K file followed by rotary pro tapers till F2 with constant irrigation. The canal was dried and obturated with corresponding gutta-percha and sealer. A Peso reamer was employed to prepare post space. Based on canal disinfection regimes, samples were divided into five groups. Group 1 Riboflavin (RF)+17%EDTA, group 2 Rose bengal (RB) +17%EDTA, group 3 Curcumin CP +17%EDTA, group 4 Porfimer sodium, Photofrin (PS) +17%EDTA and group 5 2.25% NaOCl +17% EDTA (control). Following disinfection, the canal space of all specimens was dried followed by FRCP cementation. Specimens were placed on a Universal testing machine (UTM) for EBS. The type of bond failure was evaluated using a stereomicroscope. ANOVA and Tukey multiple comparison tests were used to compare means., Results: The highest EBS was shown by group 1 canal disinfected with riboflavin (RF) and 17% EDTA at all three levels. The lowest EBS was displayed in group 5 canal cleaned with 2.25% NaOCl and 17% EDTA. Intragroup assessment disclosed a decrease in EBS from cervical one-third to apical one-third in all experimental groups. Intergroup comparison revealed group 4 using PS and 17% EDTA and group 5 canal disinfected with 2.25% NaOCl and 17% EDTA at all three levels of root structure coronal, middle, and apical exhibited comparable EBS (p>0.05)., Conclusion: Root canal dentin treated with different PS (RF, RB, CP) has the potential to be used as canal disinfection as it demonstrates better EBS than the conventional disinfecting regime (2.25% NaOCl +17% EDTA). PS and 17% EDTA as a canal disinfectant need further investigation., (Copyright © 2021. Published by Elsevier B.V.)

Aim: To assess the efficacy of photosensitizers (CP, riboflavin) and gaseous ozone in comparison to the conventional radicular dentin disinfectant (NaOCl) on push-out bond strength (PBS) of PFRC post cemented to radicular dentin., Material and Method: Human single-rooted teeth were collected, steriled implanted in polyvinyl pipes up to a cement-o-enamel junction and de coronated. Cleaning and shaping of the canal were performed using the crown down technique followed by obturation of the canal space. Canal space was prepared using peso reamers and samples were divided into four groups based on types of canal disinfectant protocols. Group 1: Riboflavin+ 17%EDTA; group 2: Curcumin Photosensitizer + 17% EDTA; group 3: Gaseous Ozone disinfection (O3) +17% EDTA and group 4 control 2.5% NaOCl +17% EDTA. Within the canal space, fiber post was cemented and cured, and thermocycled. PBS was evaluated using a Universal testing machine (UTM) and failure modes using a stereomicroscope at 40x magnification. One-way analysis of variance (ANOVA) was used to determine the mean and standard deviation of push-out bond strength (PBS). The Tukey multiple comparison tests (p = 0.05) was used to compare the means of PBS., Results: The highest PBS was displayed in group 2 CP+ 17% EDTA at all three root levels, coronal (8.81±0.61), middle (7.77±0.55), and apical (5.25±0.61). The lowest PBS was revealed in group 4, disinfected with 2.5% NaOCl +17% EDTA (control) at coronal (6.12±0.54), middle (5.46±0.84), and apical (3.00±1.88) levels. The most prevailed fracture mode was an adhesive failure (cement-dentin interface)., Conclusion: Radicular dentin disinfected with PDT using CP, riboflavin, and O3 displayed similar PBS at all root segments. NaOCl is a convenient, traditional, and commonly used disinfectant, and it's effects on PBS is still controversial., (Copyright © 2021. Published by Elsevier B.V.)

Riboflavin under UVA radiation generates reactive oxygen species (ROS) that can induce various changes in biological systems. Under controlled conditions, these processes can be used in some treatments for ocular or dermal diseases. For instance, corneal cross-linking (CXL) treatment of keratoconus involves UVA irradiation combined with riboflavin aiming to induce the formation of new collagen fibrils in cornea. To reduce the damaging effect of ROS formed in the presence of riboflavin and UVA, the CXL treatment is performed with the addition of polysaccharides (dextran). Hyaluronic acid is a polysaccharide that can be found in the aqueous layer of the tear film. In many cases, keratoconus patients also present dry eye syndrome that can be reduced by the application of topical solutions containing hyaluronic acid. This study presents physico-chemical evidence on the effect of riboflavin on collagen fibril formation revealed by the following methods: differential scanning microcalorimetry, rheology, and STEM images. The collagen used was extracted from calf skin that contains type I collagen similar to that found in the eye. Spin trapping experiments on collagen/hyaluronic acid/riboflavin solutions evidenced the formation of ROS species by electron paramagnetic resonance measurements.

Purpose: This study assessed the potential of riboflavin-mediated photodynamic therapy (RF-PDT) against Candida albicans (C. albicans) and compared the effects of RF-PDT with other therapeutic modalities in terms of mechanical and surface characteristics of acrylic denture base material., Materials and Methods: Molds (10 × 10 × 2 mm) of acrylic resin specimens were prepared and underwent artificial ageing. C. albicans were grown aerobically over the specimens and divided into four groups (n = 10/group); Group-I: no decontamination; Group-II: nystatin suspension; Group-III: riboflavin 0.1% in darkness, Group-IV: blue LED light only, and Group-V: riboflavin 0.1% for 10 minutes (pre-irradiation time) and photoactivated by the blue LED light (light dose). Fungal viability was assessed using MTT assay and characterized using scanning electron microscopy and confocal laser microscopy (CLSM). Treated specimens were subjected to surface roughness (R a ), flexural strength (FS), and flexural modulus (FM)., Results: Group-I showed the highest C. albicans viability followed by Group-III. Nystatin group (Group-II) showed ∼50% of the viability while RF-PDT showed the least C. albicans viability among the four groups (p<0.05). On SEM, specimens treated with Group-IV and V showed almost clear and free from C. albicans that was evidenced on CLSM. Post-treated specimens and storage after 72 h revealed that FS was significantly higher for RF-PDT group as compared to any other group (>105.82 MPa) (p<0.05). The FM and Ra showed statistically no significant difference between the groups (p>0.05)., Conclusion: RF-PDT showed the highest anti-fungal capacity against C. albicans over acrylic denture surface without any surface deterioration., (Copyright © 2021. Published by Elsevier B.V.)

Combating triple-negative breast cancer (TNBC) is one of the greatest challenges in cancer therapy. This is primarily due to the difficulties in developing drug delivery systems that can effectively target cancer sites. In this study, we demonstrated a proof-of-principle concept using modified surfaces of poly(lactic-co-glycolic acid) nanoparticles linked with a riboflavin analogue (PLGA-CSRf) to obtain a dual-functional material. PLGA-CSRf nanoparticles were able to function as a drug delivery ligand and a photodynamic therapy agent for TNBC cells (MDA-MB-231). Biocompatibility of novel PLGA-CSRf nanoparticles was evaluated with both breast cancer and normal breast (MCF-10A) cells. In vitro studies revealed a six-fold increase in the cellular uptake of PLGA-CSRf nanoparticles in cancer cells compared with normal cells. The results demonstrate the ability of riboflavin (Rf) to enhance the delivery of PLGA nanoparticles to TNBC cells. The viability of TNBC cells was decreased following treatment with doxorubicin-encapsulated PLGA-CSRf nanoparticles in combination with UV irradiation, due to the photosensitizing property of Rf on the surface of the nanoparticles. This work demonstrated the ability of PLGA-CSRf to function both as an effective drug delivery carrier and as a therapeutic entity, with the potential to enhance photodynamic effects in the highly aggressive TNBC model., (© 2021 American Society for Photobiology.)

Purpose: To analyze the effect of riboflavin UV-A illumination on mRNA and protein expression of healthy (HCFs) and keratoconus human corneal fibroblasts (KC-HCFs), concerning the inflammatory markers NF-κB, iNOS, IL-6, and collagen 1 and 5 (Col 1/Col 5)., Methods: Keratocytes were isolated from healthy (n = 3) and keratoconus (KC) corneas (n = 3) and were cultivated in basal medium with 5% fetal calf serum, which resulted in their transformation into human corneal fibroblasts (HCFs/KC-HCFs). Cells underwent 0.1% riboflavin UV-A illumination for 250 s (CXL). NF-κB, iNOS, IL-6, Col 1, and Col 5 expression was investigated by qPCR and Western blot analysis. IL-6 concentration of the cell culture supernatant and cell lysate was determined by ELISA., Results: In untreated KC-HCFs, NF-κB (p = 0.0002), iNOS (p = 0.0019), Col 1 (p = 0.0286), and Col 5 (p = 0.0054) mRNA expression was higher and IL-6 expression was lower (p = 0.0057), than in healthy controls. In HCFs, CXL led to an increased NF-κB (p = 0.0286) and IL-6 (p = 0.0057) mRNA expression. The IL-6 concentration in the cell culture supernatant was increased in HCFs (p = 0.0485) and KC-HCFs (p = 0.0485) after CXL. CXL increased intracellular IL-6 concentration only in KC-HCFs (p = 0.0357). In the HCF group (p = 0.0286), an increased Col 1 mRNA expression after CXL could be observed., Conclusion: Our study confirmed altered gene expression in untreated KC-HCFs compared to untreated HCFs. Riboflavin UV-A illumination affected gene expression only in HCFs. Increased IL-6 concentration in the cell culture supernatant and cell lysate indicate a secondary inflammatory response of HCFs and KC-HCFs to riboflavin UV-A illumination.

Purpose: To assess the micro-tensile bond strength (μ-TBS), degree of conversion, and antimicrobial capacity of modified dentin adhesive using riboflavin photoinitiators., Materials and Methods: Three groups of adhesives (control; 0.1 % riboflavin-PDT and 0.5 % riboflavin PDT) were prepared and tested on 70 sound molar teeth. Fourier-transformed infrared spectroscopy (FTIR) was performed for riboflavin and riboflavin-modified adhesives to calculate the degree of conversion. Contact angle measurement was performed by dropping a 5 μL droplet of the adhesives onto polished dentin disk. For μ-TBS testing, the dentin beams were fixed on a micro-tensile tester and were loaded in tension at a cross-head speed of 1 mm per min until fracture. The viability of Streptococcus mutans biofilm was tested using MTT assay., Results: The spectrum at 1728 cm-1 indicates CO stretching frequency of riboflavin. The groups control and 0.1 % riboflavin PDT showed the highest degree of conversion followed by lowest degree of conversion of 0.5 % riboflavin adhesive after PDT. The control adhesive exhibited the highest contact angle over the dentin surface compared to the riboflavin-modified adhesive groups (p < 0.05). The lowest contact angle was observed for 0.1 % riboflavin that showed increased permeability of the adhesive onto the dentin surface (p < 0.05). The variables of immediate bonding (F = 12.328, p = 0.000) and ageing in artificial saliva (F = 41.559, p = 0.000) significantly affected the bond strength. For aged μ-TBS testing, the scores for 0.1 % riboflavin PDT was significantly higher as compared to the 0.5 % riboflavin PDT samples. The MTT resulted in a lower S. mutans viability for 0.5 % riboflavin PDT compared to 0.1 % riboflavin PDT and control adhesive (p < 0.05), irrespective of any time point, that is, after 24 h and 30 days ageing., Conclusions: The addition of riboflavin as photosensitizer in dentin adhesive demonstrated higher bond strength, excellent antimicrobial capability, and degree of conversion following PDT. The addition of riboflavin in dentin adhesive for PDT could be used as a potential restorative material in adhesive dentistry., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Over the past 20 years, corneal crosslinking (CXL) has been used by surgeons to halt progression in eyes with keratoconus. We reviewed the literature regarding the mechanism of action of CXL, the role of each of its components the strong biologic reaction, and their effects on cell interaction, proteins involved, wound healing, and cytotoxic reaction. CXL surgery involves a photochemical response in which ultraviolet light at a given wavelength and riboflavin participate. The combination of irradiation with UVA light and riboflavin leads to an intense process of apoptosis of keratocytes in the anterior stroma. Differences in light irradiation, as well as the importance of riboflavin and its vehicle, were also detailed. The surgery creates additional chemical bonds between the amino terminals of the collagen side chains and the proteoglycans of the extracellular matrix. A photosensitization reaction catalyzed by riboflavin classically involves the production of singlet oxygen. Microstructure studies show changes in the size of the fibril and potentially in the interfibrillar space, that the most significant changes related to the stiffening effect of CXL occur in the anterior third of the cornea and that short irradiation times, especially below 5 min, may not have the same biological effect. Changes in the riboflavin vehicle, with the incorporation of Hydroxypropyl methylcellulose as a carrier, can lead to faster diffusion and a more intense photochemical reaction. These are findings that can impact the optimal adjustment of irradiation time according to the riboflavin (and its carrier) used. Many studies have suggested that CXL is safe and effective in the standard and accelerated protocols that have been used by surgeons. After the initial depletion of anterior keratocytes, keratocyte density seems to return to average 6-12 months after surgery when corneas are examined with the confocal microscope., (Copyright © 2020. Published by Elsevier Ltd.)

Purpose: Multispecies biofilms play a significant role in persistent infections. Furthermore, by interspecies transfer of antibiotic resistance genes, multispecies biofilms spread antibiotic resistance. The purpose of this study was to investigate the effect of Photodynamic Antimicrobial Chemotherapy (PACT) using riboflavin on mono and multi species biofilms., Methods: For this we used two clinically relevant opportunistic pathogens species E. coli and S. aureus as mono-species and multispecies biofilms. We did broth dilution assay for antibacterial, crystal violet assay for biofilms and fluorometric study for reactive oxygen species (ROS) and extracellular polymeric substance (EPS) production by phenol-HCl method., Results: Antibacterial study revealed that photo-illuminated riboflavin shows bactericidal effect against each bacteria and their mix culture. E. coli was found to be little more resistant than S. aureus. Crystal violet assay revealed photo-illuminated riboflavin shows anti-biofilms activity against both mono and mix species biofilms. But mix species biofilms were more resistant to PACT than mono species biofilms. Further study revealed this may be due to the interaction between different EPS production, hence in mix species biofilms EPS production is less affected after PACT than mono species biofilms. We found photo-illuminated riboflavin increased the intracellular ROS production., Conclusion: Photo-illuminated riboflavin shows bactericidal and anti-biofilms effect against each bacteria and their mix culture. Photo-illuminated increased intracellular ROS production, which may induce the oxidative stress and destroy the respiratory system of bacteria., (Copyright © 2020 Elsevier B.V. All rights reserved.)

In this study, we established an experimental human corneal stroma model of simulated cornea tissue composed of thin anterior cornea strips layers obtained from small incision lenticular extraction (SMILE) surgery. We investigated the biomechanical effect of ultraviolet-A- riboflavin cross-linking at different depths of corneal stroma model and correlated it with stromal microstructural changes examined by transmission electron microscopy (TEM). Corneal strips were harvested from fresh human corneal lenticules obtained after SMILE surgery. Experimental models (n = 34) were established by superimposing the corneal lenticule strips until their thickness reached close to 500 μm. Corneal cross-linking (CXL) was performed subsequently using standard or accelerated protocol. Elasticity and viscosity were quantified using stress-strain extensometer. TEM was used to visualize the collagen fiber diameter and interfibrillar spacing. The relative change in Young's modulus (rel. ΔE) decreased nonlinearly with increasing stromal depth both in the standard and accelerated groups. Compared to the sham controls, the rel. ΔE in standard and accelerated CXL groups increased significantly in the anterior 400 μm and 275 μm depth, respectively. Also, the relative change in stress (rel. ΔS) was significantly lower after standard and accelerated CXL compared to sham controls. Depth analysis showed similar results for the elastic effect. TEM images showed a small, non-significant increase in fibril diameter. The interfibrillar spacing decreased significantly after standard and accelerated CXL in the anterior-mid stromal region. We noted that the increase of corneal stiffness correlated with decrease in interfibrillar spacing after CXL. The stiffening effect was depth dependent. The effect of accelerated CXL was less in the deep corneal stromal regions compared to standard CXL., (Copyright © 2020. Published by Elsevier Ltd.)

Purpose : Corneal collagen crosslinking (CXL) through an intact epithelium (epi-on) at high irradiance could potentially improve patient comfort, visual recovery, and clinical workflow compared to conventional epi-off CXL. However, intact epithelium limits stromal delivery of the oxygen, photosensitizer, and ultraviolet-A (UV-A) radiation needed to drive CXL. This ex vivo study evaluated three different epi-on CXL protocols compared to positive and negative controls, specifically focusing on the impact of supplemental oxygen. Endpoints included stromal oxygen levels, stiffness of crosslinked tissue, and acute flattening of whole eyes. Materials & Methods : Ex vivo porcine eyes were held in a custom environmental chamber. Intrastromal oxygen levels were continuously measured before, during, and after UV illumination by a fiberoptic probe inserted into a laser-cut flap. Accelerated, high irradiance, epi-on CXL protocols using riboflavin formulated with benzalkonium chloride (BAC) were studied, with and without supplemental oxygen. These were compared to an alternate, low irradiance, epi-on protocol using riboflavin formulated with sodium iodide. Both negative (no CXL) and positive (epi-off modified Dresden protocol) controls were performed. Post-CXL elastic modulus was measured using extensiometry and anterior tangential curvature was measured using a Scheimpflug tomographer. Results : Protocols including supplemental oxygen resulted in an approximately 5-fold increase in stromal oxygen levels prior to CXL. During epi-on, high-irradiance UV-A delivery under hyperoxic conditions, an aerobic state was maintained. Conversely, under normoxic conditions, stromal oxygen rapidly depleted to 0-5% for all other protocols. The combination of supplemental oxygen, BAC formulation, and high-irradiance UV-A resulted in the largest biomechanical changes and most pronounced flattening effects of the three epi-on protocols. Conclusions : Ex vivo analysis of stromal oxygen levels, corneal stiffness, and acute anterior curvature change indicates that simultaneous optimization of the oxygen environment, riboflavin formulation, and UV-A protocol can significantly increase the effects of corneal collagen crosslinking.

In this study, we evaluate the use of riboflavin-mediated collagen photocrosslinking as an experimental tool to modulate extracellular matrix (ECM) mechanical properties in 3D in vitro tumor models. Using this approach in conjunction with 3D pancreatic tumor spheroid transplants, we show that the extent of matrix photocrosslinking in reconstituted hydrogels with fixed protein concentration scales inversely with the extent of invasive progression achieved by cells infiltrating into the surrounding ECM from primary transplanted spheroids. Using cross-linking to manipulate the extent of invasion into ECM in conjunction with imaging-based treatment assessment, we further leverage this approach as a means for assaying differential therapeutic response in primary nodule and ECM-invading populations and compare response to verteporfin-based photodynamic therapy (PDT) and oxaliplatin chemotherapy. Treatment response data shows that invading cell populations (which also exhibit markers of increased EMT) are highly chemoresistant yet have significantly increased sensitivity to PDT relative to the primary nodule. In contrast, the oxaliplatin treatment achieves greater growth inhibition of the primary nodule. These findings may be significant in themselves, while the methodology developed here could have a broader range of applications in developing strategies to target invasive disease and/or mecahanobiological determinants of therapeutic response in solid tumors., (© 2019 American Society for Photobiology.)

Introduction: Enzyme-induced corneal crosslinking (CXL) may be a promising and effective method to stiffen the cornea. However, there have been no reports on the effect of corneal CXL with enzyme in vivo., Objective: To evaluate the biomechanical efficacy after in vivo enzyme-induced corneal CXL in rabbits using noninvasive imaging., Methods: The eyes of 10 white New Zealand rabbits were unilaterally treated with CXL enzyme (transglutaminases [Tgases]). The contralateral eyes served as negative controls. In every rabbit, a 1 U/mL solution of Tgases was applied to the corneal surface. Corneal dynamic deformation by an air puff was measured using high-speed Scheimpflug imaging on day 14 after the procedure. Biomechanical parameters, central corneal thickness, and intraocular pressure were recorded and assessed. Then, corneal buttons were excised from euthanized animals for hematoxylin and eosin staining to evaluate changes in corneal collagen distribution and cell damage., Results: Some biomechanical parameters showed statistically significant changes after in vivo CXL. The deformation amplitude and maximum corneal velocity during the second applanations were significantly lower than those of the untreated control group. The second applanations were significantly increased compared to the control group. No changes were found in other parameters. The morphology of the corneal stroma was similar in CXL-treated and untreated corneas. No obvious cell apoptosis was observed., Conclusions: Corneal stiffness increased after in vivo CXL induced by Tgases. This CXL method may be useful for corneal ectasia., (© 2019 S. Karger AG, Basel.)

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of infections in humans. Photodynamic therapy using blue light (450 nm) could possibly be used to reduce MRSA on different human tissue surfaces without killing the human cells. It could be less harmful than 300-400 nm light or common disinfectants. We applied blue light ± riboflavin (RF) to MRSA and keratinocytes, in an in vitro liquid layer model, and compared the effect to elimination using common disinfection fluids. MRSA dilutions (8 × 10 5 /mL) in wells were exposed to blue light (450 nm) ± RF at four separate doses (15, 30, 56, and 84 J/cm 2 ). Treated samples were cultivated on blood agar plates and the colony forming units (CFU) determined. Adherent human cells were cultivated (1 × 10 4 /mL) and treated in the same way. The cell activity was then measured by Cell Titer Blue assay after 24- and 48-h growth. The tested disinfectants were chlorhexidine and hydrogen peroxide. Blue light alone (84 J/cm 2 ) eliminated 70% of MRSA. This dose and riboflavin eradicated 99-100% of MRSA. Keratinocytes were not affected by blue light alone at any dose. A dose of 30 J/cm 2 in riboflavin solution inactivated keratinocytes completely. Disinfectants inactivated all cells. Blue light alone at 450 nm can eliminate MRSA without inactivation of human keratinocytes. Hence, a high dose of blue light could perhaps be used to treat bacterial infections without loss of human skin cells. Photodynamic therapy using riboflavin and blue light should be explored further as it may perhaps be possible to exploit in treatment of skin diseases associated with keratinocyte hyperproliferation.

Purpose: A follow-up experimental study on the exposure of animal tarsal plate to ultraviolet-A radiation aimed at establishing an optimum range for safe irradiation conditions., Methods: Sheep tarsus specimens were excised postmortem and then subjected to irradiation with ultraviolet-A rays (wavelength 365 nm) at higher irradiances than those reported in an initial study, using a laboratory radiation source. The mechanical properties (tensile strength and Young's modulus) of irradiated and nonirradiated samples were evaluated in a mechanical tester. The test and control specimens were examined histologically with an aim to assess the effects of radiation upon the meibomian glands and tarsal collagen networks, and to establish a safe range for the exposure irradiance level., Results: As expected, irradiation induced both stiffening and strengthening of the tarsal plate specimens. At an irradiance of 50 mW/cm for 3-minute exposure, these effects were at their maximum level, after which a decline in mechanical characteristics were observed. No destruction of the tarsal connective tissue or the meibomian glands were noticed up to an irradiance of 125 mW/cm for 3-minute exposure, corresponding to a fluence of 22.5 J/cm. Histology revealed that the collagen network surrounding the glands were packed more compactly following irradiation. At a fluence of 45 J/cm, massive destruction of periglandular collagen-rich network and meibocytes were demonstrated histologically., Conclusions: The study indicates that irradiation of tarsal collagen leading to tissue stiffening shall be carried out at levels of fluence between 10 and 15 J/cm, a region that is deemed safe. The exposure time can be adjusted according to the surgeon's decision.Safe irradiation conditions are established for the exposure of ex vivo ovine tarsus to ultraviolet-A radiation as a potentially effective treatment for eyelid laxity in human patients.

Purpose: To determine the relative impact of contact lens- assisted corneal cross-linking (CACXL) and standard protocol CXL (CXL) on regional corneal stiffness using Brillouin microscopy., Methods: CXL and CACXL were performed on 30 intact fresh porcine eyes (15 per group). Depth profile of stiffness variation and averaged elastic modulus of anterior, middle, and posterior stroma were determined by Brillouin maps. Corneas were cut into strips to conduct mechanical stress-strain tests after Brillouin microscopy to evaluate stiffness difference between CXL and CACXL. Each eye served as its own control., Results: CXL had a greater impact on corneal stiffness, with a maximum increase of 5.74% compared to 3.99% for CACXL (P < .001). CXL increased longitudinal modulus by 7.8% in the anterior, 1.7% in the middle, and -0.7% in the posterior regions compared to CACXL, which increased longitudinal modulus by 5.5% in the anterior (P < .001), 1.2% in the middle (P = .15), and -0.4% in the posterior regions (P = .60). Mechanical stress-strain tests showed that at 10% strain averaged Young's modulus was 5 MPa for CXL and 2.97 MPa for CACXL (P < .001)., Conclusions: Both CACXL and standard protocol CXL induced significant corneal stiffening primarily concentrated in the anterior cornea. CACXL leads to less stiffening compared with CXL. An attenuated but continuous stiffening effect can be observed through the whole cornea for both CACXL and CXL, although CACXL has a smaller stiffness gradient. [J Refract Surg. 2019;35(11):721-728.]., (Copyright 2019, SLACK Incorporated.)

Purpose: To investigate the safety of blue light scleral cross-linking (SXL) by evaluating changes in biological parameters in the retina and choroid in the eyes of rhesus macaques (Macaca mulatta)., Methods: Fifteen 3-year-old macaques (30 eyes) were randomly divided into three groups (n = 5). SXL was performed via riboflavin (0.5%) and blue light (460 nm) at the location of the equatorial sclera. Right eyes served as experimental eyes, and left eyes as control eyes. One quadrant of each right eye was irradiated in group A, two quadrants of each right eye and one quadrant of each left eye were irradiated in group B, and two quadrants of each right eye were irradiated in group C. Optical coherence tomography, optical coherence tomography angiography, and flash electroretinography (f-ERG) examinations were performed at baseline and 1 week, 1 month, 3 months, and 6 months after SXL. Additionally, retinal tissue alterations were detected via transmission electron microscopy at 1 week postoperatively., Results: There were no significant differences between experimental eyes and control eyes in retinal thickness, vessel density of retinal superficial capillary plexus, and choroid thickness in any of the groups at any of the time points investigated (p > 0.05). Significant reductions in f-ERG parameters were detected 1 week postoperatively in the experimental eyes of groups A and C (p < 0.05), but they gradually recovered, and there was no significant difference 1 month postoperatively (p > 0.05). Ultrastructural changes were evident in the retinal layers of SXL eyes. In group B, there were no significant differences between the right and left eyes at any of the follow-up time points investigated., Conclusions: Blue light SXL can cause transient retina damage. The f-ERG parameters reductions and retinal ultrastructural changes were found at early stage, even though there were not significant changes in retinal thickness, vessel density of retinal superficial capillary plexus, and choroid thickness after blue light SXL. The long-term intraocular safety of the blue light SXL technique should be investigated further.

Purpose: To evaluate the efficacy of corneal cross-linking (CXL) as adjuvant therapy for the treatment of fungal ulcerative keratitis., Methods: Forty-one patients with fungal ulcerative keratitis were recruited and assigned into two randomized controlled groups. These groups were treated with CXL combined with antifungal medications (CXL-M) or antifungal medications alone (M). The ulcers were assessed by slit-lamp biomicroscopy, slit-lamp images, in vivo confocal microscopy (IVCM), and anterior segment optical coherence tomography (AS-OCT). The patients were followed up before surgery/first visit (FV), 1 day after surgery, 1 and 2 weeks, and 1, 2, 3, 4, 5, and 6 months after surgery/FV., Results: In the cured patients, the area of corneal ulcers, the duration of ulcer healing, the time to non-observed fungal hyphae by IVCM, the number of antifungal medications, the frequency of administered medications, and the maximum ulcer depth decreased significantly after CXL (all P < 0.05) compared with the M group. There were no significant differences in either corneal thickness or epithelial thickness of ulcers after healing between 5 and 6 months after surgery in the CXL-M group, while these were increased significantly at 6 months compared with 5 months after FV in the M group (both P < 0.05)., Conclusions: In our study, CXL accelerated healing of the fungal ulcers, shortened the treatment duration, and minimized the need for medications and surgery. It appears that CXL is an effective procedure and adjuvant therapy for managing fungal keratitis.

Purpose: To assess corneal thickness changes with isotonic riboflavin (RF) solution with hydroxylpropyl methylcellulose in patients undergoing accelerated corneal collagen crosslinking (CXL) with and without an eyelid speculum., Methods: Fifty-two eyes of 48 patients with progressive keratoconus were enrolled in this study. The patients in this study were divided into 2 groups: in group 1 an eyelid speculum was removed during 20-minute RF (0.1%) + hydroxylpropyl methylcellulose (Mediocross M; Avedro Inc, Waltham, MA) instillation, and in group 2 the eyelid speculum was retained in place during the entire CXL procedure. All patients underwent accelerated CXL using continuous ultraviolet-A (UVA) light exposure at 9 mW/cm for 10 minutes; total energy dose was 5.4 J/cm. Intraoperative ultrasound pachymetry measurements were obtained before and after epithelial removal, after RF loading, and after UVA light exposure at 5 and 10 minutes., Results: The preoperative pachymetric measurements decreased in both groups after the removal of epithelium [group 1 (n = 26): -25 μm, group 2 (n = 26): -31 μm, P = 0.234]. Although the thinnest pachymetry significantly increased after soaking in both group 1 (52.26 μm) and group 2 (27.88 μm, P < 0.001), closure of the eyelids during RF instillation further increased the pachymetry readings (P < 0.0001). The corneal thickness remained stable in both groups during UVA irradiation at 5 and 10 minutes (P > 0.05)., Conclusions: Closure of the eyelids further induces corneal swelling that may offer an advantage to improve safety of the procedure particularly in thin corneas.

Background: Although antimicrobial photodynamic therapy (aPDT) has been shown to be efficient in killing planktonic periodontopathogenic bacteria, its activity on established biofilms is very limited. The aim of the present in-vitro study was to evaluate the potential effect of hydrogen peroxide as a pretreatment for aPDT., Methods: aPDT consisting of riboflavin as photosensitizer and illumination by a LED lamp emitting in the blue spectrum for 30 s and 60 s (aPDT60) was combined with a pretreatment with 0.25% and 3% hydrogen peroxide. The antimicrobial activity of these treatments was determined against eight oral species (incl. Porphyromonas gingivalis and Tannerella forsythia) and against eight-species biofilms. Treatment of biofilms in an artificial pocket model included a mechanical removal of the biofilm., Results: Against planktonic bacteria, pretreatment with hydrogen peroxide increased killing of planktonic bacteria, after aPDT60 no viable bacteria were detected in 7 of 8 strains. In biofilms formed on well-plates, aPDT60 reduced bacterial counts only by 0.53 log10 cfu, whereas reduction was closed to 4 log10 or higher when 3% hydrogen peroxide was used. When biofilms were treated in the periodontal-pocket model, reduction of cfu was less than 0.5 log10 after mechanical therapy or aPDT60 only, however no bacteria were detected after mechanical biofilm removal followed by the use 3% of hydrogen peroxide and aPDT60., Conclusions: aPDT using riboflavin and blue LED light applied after mechanical removal of biofilm and adjunctive 3% hydrogen peroxide solution appears to represent an alternative for antimicrobial periodontal therapy., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Purpose: There has been a recent explosion in the variety of techniques used to accomplish corneal cross-linking (CXL) for the treatment of ectatic corneal diseases. To understand the success or failure of various techniques, we review the physicochemical basis of corneal CXL and re-evaluate the current principles and long-standing conventional wisdom in the light of recent, compelling, and sometimes contradictory research., Methods: Two clinicians and a medicinal chemist developed a list of current key topics, controversies, and questions in the field of corneal CXL based on information from current literature, medical conferences, and discussions with international practitioners of CXL., Results: Standard corneal CXL with removal of the corneal epithelium is a safe and efficacious procedure for the treatment of corneal ectasias. However, the necessity of epithelium removal is painful for patients, involves risk and requires significant recovery time. Attempts to move to transepithelial corneal CXL have been hindered by the lack of a coherent understanding of the physicochemistry of corneal CXL. Misconceptions about the applicability of the Bunsen-Roscoe law of reciprocity and the Lambert-Beer law in CXL hamper the ability to predict the effect of ultraviolet A energy during CXL. Improved understanding of CXL may also expand the treatment group for corneal ectasia to those with thinner corneas. Finally, it is essential to understand the role of oxygen in successful CXL., Conclusions: Improved understanding of the complex interactions of riboflavin, ultraviolet A energy and oxygen in corneal CXL may provide a successful route to transepithelial corneal CXL.

Great success has been achieved in recent years in the development of synthetic or assembled nanobiomaterials. Among these, biomolecule-based nanoarchitectures with special optical property are of particular interest. Here, we demonstrate that vitamin B 2 nanocrystals assembled as nanorods can be obtained with precise control. Excitingly, such one-dimensional nanostructures not only exhibit intrinsic optical waveguiding properties but also the ability to sensitize oxygen to produce reactive oxygen species. With these properties, we applied the obtained vitamin B 2 nanorods under remotely localized light illumination into single tumour cells in vitro for anticancer photodynamic therapy. Further, vitamin B 2 nanorods were explored for in vivo photodynamic therapy by using a tumour model. With such bionanostructures, new features and functions of vitamin B 2 and its derivatives have been developed., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Purpose: To evaluate the ocular biological parameter difference between scleral corneal cross-linking (CXL) and control eyes in rhesus monkeys by using a rebound tonometer, A-scan ultrasonography, retinoscopy, optical coherence tomography, and electroretinography (ERG)., Methods: Six rhesus monkeys were used in this study, with ages ranging from 3 to 3.5 years. One eye of each rhesus monkey was randomly selected to receive riboflavin/ultraviolet-A CXL in the temporal quadrant of the equatorial sclera and the contralateral eye served as an intra-individual control. The ocular biological parameters were repeatedly measured in both eyes of the monkeys before scleral CXL and 1 week, 1 month, and 3 months postoperatively., Results: The intraocular pressure, refractive state, total axial length, and axial dimensions of the anterior chamber, crystalline lens, vitreous chamber, and central corneal thickness were not statistically significantly different between the control and cross-linked specimens at the different time periods (each P > .05). No obvious changes in the waveform of the standard full-field ERGs were observed in the control and cross-linked specimens. There were no statistically significant differences between the control and cross-linked specimens in the dark-adapted 0.01 ERG, the dark-adapted 3.0 ERG, the light-adapted 3.0 ERG, and the amplitudes of the a-wave and b-wave for the different time periods (each P >.05)., Conclusions: The scleral CXL laboratory technique might not significantly affect the ocular biological parameters of the rhesus monkey in the early postoperative period, but long-term effects and histological changes still need to be investigated further. [J Refract Surg. 2019;35(5):333-339.]., (Copyright 2019, SLACK Incorporated.)

Purpose: Contact lens-assisted corneal cross-linking (CACXL) has been proposed for the cross-linking treatment in thin corneas. The aim of this study was to assess the biomechanical efficacy of this treatment., Methods: Post-mortem porcine eyes were treated with standard cross-linking and with cross-linking placing a contact lens soaked with isoosmolar riboflavin solution on the debrided cornea with or without an adherent precorneal riboflavin film of up to 100 μm thickness. Three soft contact lenses (Air Optix Aqua, SofLens and Galifa) with different degrees of hydrophilic properties were tested. After cross-linking with a surface UVA irradiance of 3 mW/cm² for 30 min (fluence 5.4 J/cm²), a 400 μm deep anterior corneal flap was created using a lamellar rotating microkeratome. Biomechanical stress-strain measurements and thermal shrinkage tests were performed., Results: In the Air Optix Aqua group (30% hydration) without riboflavin film, Young's modulus and stress at 8% strain were increased significantly versus untreated controls and the effect was 92.4% respectively 86.35% of the standard CXL value. In the SofLens group (59% hydration) without riboflavin film, Young's modulus and stress at 8% strain were increased significantly versus untreated controls and the effect was 67.04% respectively 65.28% of the standard CXL value. In the Galifa group (72% hydration) without riboflavin film, Young's modulus and stress at 8% strain were increased significantly versus untreated controls and the effect was about 68.48% respectively 75.52% of the standard CXL value. In all samples with a precorneal riboflavin film under the contact lens, there was no significant biomechanical effect compared to the untreated controls. Similarly, in the hydrothermal experiments at 70°C, there was a typical mushroom pattern with increased resistance to thermal shrinkage in the anterior stroma after standard CXL, a markedly reduced mushroom effect using a riboflavin-soaked contact lens only and no effect with the use of a riboflavin-soaked contact lens plus a precorneal riboflavin film., Conclusion: The biomechanical effect of CACXL in porcine corneas is about one-third less than after standard CXL. The efficacy of CACXL might be improved by reducing or omitting the riboflavin film on the contact lens. Further risk assessment studies are necessary., (© 2018 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.)

Purpose: We retrospectively investigated the efficacy of corneal crosslinking (CXL) on progressive keratoconus in a Japanese population and compared the outcomes of conventional and accelerated CXL., Study Design: A retrospective cohort study METHODS: A total of 108 consecutive eyes in 95 patients (75 men; 21.9 ± 6.2 years) with progressive keratoconus were enrolled. The epithelium was ablated in all eyes. After presoaking the corneal stroma in riboflavin, UV-A was irradiated at 3.0 mW/cm 2 (conventional CXL) for 30 min on 23 eyes and 18.0 mW/cm 2 for 5 min (accelerated CXL) on 85 eyes. Best spectacle-corrected visual acuity (BSCVA), manifest refraction, keratometric value, corneal thickness, corneal endothelial cell density (ECD), intraocular pressure, and complications were evaluated at 1, 3, 6, and 12 months after the procedure., Results: BSCVA, manifest refraction, ECD, and corneal thickness did not change significantly after both procedures. The keratometric value was significantly decreased from the preoperative value at 12 months (p < 0.001). Progression to more than 1.0 D after CXL was observed in 10 eyes (9.3%). The ΔKmax was negatively associated with preoperative Kmax (p < 0.001) and positively associated with preoperative thinnest corneal thickness (p < 0.001). Both treatment modules showed no significant difference in all parameters., Conclusion: CXL was as effective in treating keratoconus in Japanese patients as in individuals of other ethnicities. Overall, CXL could be performed using either the conventional or accelerated approach to halt the progression of keratoconus in Japanese populations.

Purpose: The purpose of this study was to determine whether corneal collagen crosslinking (CXL) inhibits hemangiogenesis and lymphangiogenesis during acute corneal inflammation in an in vivo rat model., Methods: Inflammatory corneal neovascularization was induced by suture placement into a rat cornea. At day 3 after suture, a CXL protocol using riboflavin and UVA was administered after mechanical epithelial debridement. Hemangiogenesis and lymphangiogenesis were analyzed morphometrically. CD45 and CD68 immunostaining evaluated corneal leucocyte and macrophage immune cell infiltration, respectively. A TUNEL assay detected stromal cell apoptosis. Quantitative RT-PCR analysis identified angiogenic and lymphangiogenic genes as well as proinflammatory cytokine expression. Western blot analysis characterized vascular endothelial cell CD31 and lymphatic vessel endothelial hyaluronan receptor (LYVE-1) protein expression., Results: CXL treatment significantly reduced corneal pathologic suture-induced hemangiogenesis and lymphangiogenesis 7 days after suture emplacement, but this procedure failed to affect hemangiogenesis and lymphangiogenesis 14 days after suture. Increased cell apoptosis and reduced CD45+ and CD68+ cell infiltration were evident in CXL-treated rats on days 7 and 14 after suture emplacement. CXL treatment significantly decreased angiogenic and lymphangiogenic mRNA expression levels and both CD31 and LYVE-1 protein expression levels, whereas it increased proinflammatory cytokine levels on day 7 after suture emplacement. However, on day 14 after corneal neovascularization, angiogenic and lymphangiogenic mRNA gene expression levels were upregulated along with hematic CD31 and lymphatic LYVE-1 protein expression., Conclusions: CXL treatment only temporarily inhibits corneal inflammatory-associated hemangiogenesis and lymphangiogenesis in vivo. Such insight suggests that future studies are warranted to develop novel CXL strategies with longer-lasting effectiveness in attenuating hemantic- and lymphatic-related corneal diseases.

Purpose: To characterize the mechanical property improvement of riboflavin and ultraviolet light corneal crosslinking (CXL) procedure in artificially swollen human and porcine corneas., Setting: Computational Biomechanics Research Laboratory, Mechanical and Industrial Engineering Department, University of Illinois at Chicago, Illinois, USA., Design: Experimental study., Methods: Porcine and human donor corneas were crosslinked at different hydration levels using riboflavin-dextran solutions of different osmolality. Four porcine groups (H w [hydration in mg H 2 O/mg dry tissue] = 3.3 ± 0.2 [SD]; 4.0 ± 0.1; 5.1 ± 0.1; 5.6 ± 0.1) and 3 human groups (H w  = 3.2 ± 0.1; 3.9 ± 0.2; 5.3 ± 0.3) were considered. The mechanical properties were measured by uniaxial tensile experiments during which the hydration of samples was the same as the hydration at which they were crosslinked. Tensile properties of 2 porcine groups (H w  = 5.1 ± 0.1; 5.6 ± 0.1) were also measured when their average hydration was lowered to 4.0 mg H 2 O/mg dry tissue., Results: The CXL procedure significantly increased tensile properties of both human and porcine samples in each hydration group (P < .05). The improvement in tensile properties was hydration-dependent, that is, samples crosslinked at higher hydration levels showed lower maximum tensile stress. The behavior of samples crosslinked at different initial hydration but tested mechanically at similar hydration showed insignificant difference (P = .7)., Conclusion: Increasing the hydration of porcine and human corneal samples before the CXL treatment had insignificant influence on tensile property improvement, as measured by testing specimens at similar hydration., (Copyright © 2018 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.)

Riboflavin (vitamin B2) has been thought to be a promising antitumoral agent in photodynamic therapy, though the further application of the method was limited by the unclear molecular mechanism. Our work reveals that riboflavin was able to recognize G-T mismatch specifically and induce single-strand breaks in duplex DNA targets efficiently under irradiation. In the presence of riboflavin, the photo-irradiation could induce the death of tumor cells that are defective in mismatch repair system selectively, highlighting the G-T mismatch as potential drug target for tumor cells. Moreover, riboflavin is a promising leading compound for further drug design due to its inherent specific recognition of the G-T mismatch., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)