The effect of parasitic ions on riboflavin permeability and cross-linking effectiveness in iontophoresis-assisted scleral cross-linking / 生物医学工程学杂志

The effect of parasitic ions on the results of ultraviolet A (UVA) cross-linking in iontophoresis was still not clear. In this work, the porcine sclera was cross-linked by riboflavin lactate Ringer's solution (group A) and riboflavin normal saline (group B)

Blocking or enhancing effects of some basic emollients in UVA penetration

Abstract: Background: Topical agents used in combination with phototherapy or photochemotherapy may have both blocking or enhancing effects in ultraviolet rays. Objective: In this in vivo study, the effects of topical petrolatum, basis cream, glycerine, and olive oil on the transmission of ultraviolet A radiation were investigated. Methods: A test was performed to determine the minimal phototoxic dose on 29 volunteers with only psoralen plus ultraviolet A (PUVA) and then the same test was repeated with white petrolatum, basis cream, glycerine, olive oil, and sunscreen (0.3cc/25cm2). The effects of each agent on the minimal phototoxic dose were determined after 72 h. Results: When compared to pure PUVA, there was a statistically significant increase in the mean minimal phototoxic dose values by the application of white petrolatum (P = 0.011), but there was no significant increase or decrease in the mean minimal phototoxic dose values after the application of basis cream (P = 0.326), glycerine (P = 0.611) or olive oil (P = 0.799). Study limitations: Low number of patients Conclusion: The application of white petrolatum, which has a blocking effect, and also of basis cream immediately before PUVA therapy should not be recommended. Although we specify that glycerine and maybe olive oil can be used before photochemotherapy, there is a need for further research in larger series.

Photodynamic action of the red laser on Propionibacterium acnes

Abstract: Background: Photodynamic therapy is a therapeutic modality that has consolidated its activity in the photooxidation of organic matter, which arises from the activity of reactive oxygen species. Objective: To evaluate the effect of red laser 660nm with the photosensitizer methylene blue on Propionibacterium acnes in vitro. Method: The experimental design was distributed into four groups (1 - control group without the application of light and without photosensitizer, 2 - application of light, 3 - methylene blue without light, and 4 - methylene blue with light). Tests were subjected to red laser irradiation 660nm by four cycles of 5 minutes at 3-minute intervals. Results: It was evidenced the prominence of the fourth cycle (20 minutes) groups 2, 3 and 4. Study limitations: Despite the favorable results, the laser irradiation time photosensitizer associated with methylene blue were not sufficient to to completely inhibit the proliferation of bacteria. Conclusion: Further studies in vitro are recommended to enable the clinical application of this photosensitizer in photodynamic therapy.

The effects of riboflavin and ultraviolet light on keratocytes cultured in vitro / Avaliação do efeito da riboflavina e luz ultravioleta sobre os ceratócitos in vitro

ABSTRACT Purpose: To culture quiescent human keratocytes and evaluate the effects of ultraviolet light and riboflavin on human corneal keratocytes in vitro. Methods: Keratocytes were obtained from remaining corneoscleral ring donor corneas previously used in corneal transplant surgeries and cultured in DMEM/F12 with 2% FBS until confluence. Characterization of cultured cells was performed by immunofluorescence analysis for anti-cytokeratin-3, anti-Thy-1, anti-α-smooth muscle actin, and anti-lumican. Immunofluorescence was performed before and after treatment of cultured cells with either ultraviolet light or riboflavin. Corneal stromal cells were covered with collagen (200 µL or 500 µL) and 0.1% riboflavin, and then exposed to ultraviolet light at 370 nm for 30 minutes. After 24 hours, cytotoxicity was determined using MTT colorimetric assays, whereas cell viability was assessed using Hoechst 33342 and propidium iodide. Results: Cell cultures achieved confluence in approximately 20 days. Expression of the lumican was high, whereas no expression of CK3, Thy-1, and α-SMA was observed. After crosslinking, MTT colorimetric assays demonstrated a low toxicity rate, whereas Hoechst 33342/propidium iodide staining demonstrated a low rate of apoptosis and necrosis, respectively, in all collagen-treatment groups. Conclusion: Keratocytes can be successfully cultured in vitro and characterized by immunofluorescence using lumican. MTT colorimetric assays, and Hoechst 33342, and propidium iodide staining demonstrated a higher rate of cell death in cells cultured without collagen, indicating collagen protects keratocytes from the cytotoxic effects of ultraviolet light.

RESUMO Objetivo: Avaliar o efeito da aplicação da luz ultravioleta e riboflavina sobre ceratócitos da córnea humana in vitro. Métodos: Os ceratócitos foram obtidos a partir das rimas corneoesclerais remanescentes da trepanação de córneas previamente utilizadas em cirurgias de transplante de córnea e cultivadas em meio DMEM/F12 com 2% de FBS até atingir confluência. As culturas de células foram caracterizadas por imunofluorescência com os anticorpos K3 (marcador de células epiteliais), Thy-1 (marcador de fibroblasto) SMA (marcador de miofibroblasto) e Lumican (marcador de ceratócitos). Imunofluorescência também foi feita após o tratamento. As células do estroma da córnea foram cobertas com colágeno (200 µL e 500 µL) e 0,1% de riboflavina e exposta a luz UVA a 370 nm por 30 minutos. Após 24 horas, citotoxicidade foi determinada por ensaio de MTT e a viabilidade celular foi feita por Hoechst 33342/Iodeto de propideo. Resultados: As culturas de células atingiram confluência em aproximadamente 20 dias. Imunofluorescência apontou alta expressão para o marcador de ceratócitos (Lumican) e expressão negativa par os marcadores de células epiteliais (K3), fibroblasto (Thy-1) e miofibroblasto (α-SMA). Após o cross linking a análise de MTT mostrou baixa taxa de toxicidade e com a coloração de Hoechst 33342/Iodeto de propideo baixa taxa de apoptose e necrose respectivamente em todos os grupos que continham colágeno. Conclusão: As culturas de ceratócitos foram obtidas e caracterizadas por imunofluorescência através do marcador Lumican com sucesso. O ensaio de MTT e a coloração por Hoechst 33342 e iodeto de propídio, apresentaram maior índice de morte celular nos grupos que não continham colágeno, provando que protege as células contra os efeitos da luz UVA.

Six-month outcomes of corneal crosslinking with dextran-free isotonic riboflavin solution / Resultados após seis meses de crosslinking de córnea com solução isotônica de riboflavina sem dextrano

ABSTRACT Purpose: To analyze the short-term clinical and topographic outcomes in patients with keratoconus after corneal collagen cross-linking treatment (CXL) with dextran-free isotonic riboflavin solution. Methods: In this retrospective case series, 26 eyes from 26 patients with keratoconus were studied. The best corrected visual acuity (BCVA) and refractive and topographic findings were analyzed at a 6-month follow-up. Results: The mean BCVA (Snellen lines) values before and 1, 3, and 6 months after CXL were 0.51 ± 0.2, 0.48 ± 0.2, 0.57 ± 0.2, and 0.64 ± 0.2, respectively, and the difference between the preoperative and 6-month values was statistically significant (p=0.006). The mean spherical equivalent refraction decreased from -5.6 ± 2.4 diopters (D) preoperatively to -5.0 ± 2.1 D, and mean simulated keratometry decreased from 48.5 ± 2.5 D to 47.8 ± 2.6 D at 6 months. (p=0.145 and p=0.001, respectively). In addition, the maximum keratometry decreased progressively and significantly from the preoperative value during follow-up (p=0.003). The central and minimal corneal thicknesses, including those of the epithelium, also decreased from 442.8 ± 25.6 µm and 430.5 ± 23.9 µm preoperatively to 420.7 ± 31.8 µm and 409.3 ± 28.7 µm at the most recent follow-up (p<0.001), respectively. No intraoperative or postoperative complications were observed. Conclusions: CXL with dextran-free isotonic riboflavin solution appears to be a safe treatment alternative for keratoconus and yields sustained short-term improvements in visual acuity, keratometric readings, and corneal thickness. However, long-term results are needed to confirm these outcomes.

RESUMO Objetivo: Analisar os resultados clínicos e topográficos curto prazo após crosslinking (CXL) de córnea com solução isotônica de riboflavina sem dextrano, em pacientes com ceratocone. Método: Estudamos 26 olhos de 26 pacientes com ceratocone, nesta série retrospectiva de casos. Melhor acuidade visual corrigida (BCVA), refração e achados topográficos foram analisados aos 6 meses de acompanhamento. Resultados: BCVA pré-operatória (linhas de Snellen) foi de 0,51 ± 0,2. BCVA após CXL foram de 0,48 ± 0,2, 0,57 ± 0,2 e 0,64 ± 0,2 no 1º, 3º e 6º meses, respectivamente. A diferença entre a BCVA pré-operatória e mais recente foi estatisticamente significativa (p=0,006). O equivalente esférico médio diminuiu de -5,6 ± 2,4 dioptrias (D) no pré-operatório para -5.0 ± 2.1 D e a média da ceratometria simulada diminuiu de 48,5 ± 2,5 D para 47, 8± 2,6 D aos 6 meses. (p=0,145 e p=0,001, respectivamente). A ceratometria máxima diminuiu progressivamente durante o acompanhamento com as mudanças sendo significativamente diferentes do valor pré-operatório (p=0,003). As espessuras corneanas central e mínima, diminuiram de 442,8 ± 25,6 µm e 430,5 ± 23,9 µm para 420,7 ± 31,8 µm e 409,3 ± 28,7 µm, respectivamente, na visita mais recente (p<0,001). Não foram observadas complicações intraoperatórias e pós-operatórias. Conclusões: CXL com solução de riboflavina isotônica sem dextrano parece ser uma opção segura de tratamento para o ceratocone com melhora mantida na acuidade visual, ceratometria e espessura corneana, no curto prazo. Resultados a longo prazo são necessários para confirmar estes resultados.

Factors affecting visual acuity after accelerated crosslinking in patients with progressive keratoconus / Fatores que afetam a acuidade visual após crosslinking acelerado entre pacientes com ceratocone progressivo

ABSTRACT Purpose: The present study aimed to report the outcomes of patients with progressive keratoconus who were treated via accelerated crosslinking (CXL) 6 months earlier and to determine the factors that promoted improved visual acuity after treatment. Methods: This retrospective study included 35 eyes of 34 patients with progressive keratoconus who underwent CXL. Topographical measurements were obtained preoperatively and in the first, third, and sixth months postoperatively using a rotating Scheimpflug camera. The uncorrected visual acuity (UCVA), best-corrected visual acuity (BCVA), flat keratometry (K) value (K1), steep K value (K2), average K value (avgK), topographic cylindrical value (Cyl), apical keratoscopy front (AKf), apical keratoscopy back (AKb), symmetry index front (SIf), symmetry index back (SIb), and thinnest point of the cornea (ThkMin) were recorded. Results: At the 6-month follow-up, the mean UCVA and BCVA values were improved, and the K values remained stable. Statistically significant decreases in AKf (p=0.04) and the thinnest point of the cornea (p=0.001) and a statistically significant increase in AKb (p=0.01) were observed. A correlation analysis revealed that the preoperative BCVA, UCVA, K1, K2, avgK, AKf, and AKb values significantly affected visual acuity at the 6-month follow-up. Conclusions: Accelerated CXL is an effective treatment for the prevention or even reversal of keratoconus progression. The preoperative K values and apexes of the anterior and posterior cornea were found to affect visual acuity at 6 months after accelerated CXL. Both AKb steepening and AKf flattening appear to be important factors in the stabilization of keratometric values and improvement of visual outcomes.

RESUMO Objetivo: O objetivo do estudo é relatar os resultados do sexto mês após o tratamento de crosslinking acelerado (CXL) em pacientes com ceratocone progressivo e determinar os fatores que afetam a melhora da acuidade após o tratamento. Métodos: Neste estudo retrospectivo, foram incluídos 35 olhos de 34 pacientes com ceratocone progressivo que se submeteram CXL. Acuidade visual não corrigida (UCVA) e melhor acuidade visual corrigida (BCVA) foram registradas. Medidas topográficas foram obtidas utilizando uma câmara Scheimpflug rotativa no pré-operatório e no 1º, 3º e 6º meses após a cirurgia. Os valores de ceratometria (K) mais plana (K1), K mais curva (K2), médio de K (avgK), astigmatismo topográfico (Cyl), ápice anterior da ceratoscopia (AKf), ápice posterior da ceratoscopia (AKb), índice anterior de simetria (SIf), índice posterior de simetria (SIb) e ponto mais fino da córnea (ThkMin) foram avaliados. Resultados: A média UCVA e BCVA melhoraram, enquanto valores de K ficaram estáveis 6º mês. Houve uma diminuição estatisticamente significativa na AKf e um aumento estatisticamente significativo na AKb (p=0,04, p=0,01, respectivamente). O ponto mais fino da córnea diminuiu significativamente (p=0,001). Na análise de correlações, além da UCVA e BCVA pré-operatórias; valores K1, K2, avgK, AKf e AKb pré-operatórios influenciaram significativamente a acuidade visual no 6º mês de acompanhamento. Conclusões: CXL acelerado é uma forma eficaz de tratamento na prevenção ou no mesmo inversão da progressão do ceratocone. A acuidade visual no 6º mês após CXL acelerado foi afetada a partir dos valores de K e dos ápice anterior e posterior da córnea. Encurvamento do AKb e aplanamento do AKf parecem ser fatores importantes na estabilização dos valores ceratométricos e na melhora da acuidade visual.

Photodynamic action of protoporphyrin IX derivatives on Trichophyton rubrum

Abstract: BACKGROUND: Dermatophytes are filamentous keratinophilic fungi. Trichophyton rubrum is a prevalent infectious agent in tineas and other skin diseases. Drug therapy is considered to be limited in the treatment of such infections, mainly due to low accessibility of the drug to the tissue attacked and development of antifungal resistance in these microorganisms. In this context, Photodynamic Therapy is presented as an alternative. OBJECTIVE: Evaluate, in vitro, the photodynamic activity of four derivatives of Protoporphyrin IX by irradiation with LED 400 nm in T. rubrum. METHOD: Assays were subjected to irradiation by twelve cycles of ten minutes at five minute intervals. RESULT: Photodynamic action appeared as effective with total elimination of UFCs from the second irradiation cycle. CONCLUSION: Studies show that the photodynamic activity on Trichophyton rubrum relates to a suitable embodiment of the photosensitizer, which can be maximized by functionalization of peripheral groups of the porphyrinic ring.

Desenvolvimento e testes in vitro de um novo corante para terapia fotodinâmica em Periodontia / Development and in vitro tests of a new dye for photodynamic therapy in Periodontics

A terapia fotodinâmica antimicrobiana (Antimicrobial Photodynamic Therapy - aPDT) tem sido utilizada em Periodontia para redução bacteriana de bolsas periodontais na forma de tratamento adjuvante da raspagem e alisamento radicular. Seu efeito em cirurgias periodontais é pouco estudado. Pesquisas recentes têm procurado descobrir novos fotossensibilizadores para aumento da efetividade dessa terapia. O objetivo do presente estudo foi testar in vitro um novo corante à base de corantes fenotiazínicos para uso em aPDT, quanto à sua biocompatibilidade e efetividade em eliminar bactérias. Foram realizados testes para avaliar a desmineralização e desgaste causados pelo corante em esmalte e dentina bovinos, bem como o tempo de aplicação ideal para se conseguir efeitos semelhantes ao ácido cítrico; comportamento óptico dos novos corantes em relação aos corantes convencionais; teste de biocompatibilidade de fragmentos de raiz humana, tratados pelo corante, em culturas de fibroblastos gengivais humanos; efetividade dos novos corantes sozinhos ou associados ao laser na eliminação de S. aureus e E. coli. Nos experimentos de desmineralização e biocompatibilidade foi utilizado um laser vermelho (660nm, 30mW, 45J/cm2, 30s) e nos experimentos com bactérias laser vermelho (660nm, 100mW, 45J/cm2, 12s). As análises estatísticas foram realizadas com testes paramétricos e não paramétricos (p<0,05). Os resultados mostraram que o novo corante promoveu desmineralização em esmalte (perda de dureza 55,5% e desgaste 10,33μm) e dentina (perda de dureza 65,9% e desgaste 8,37μm) bovinos de forma semelhante ao ácido cítrico, sendo 180s, o tempo mais adequado. Nos ensaios ópticos, observou-se que o novo corante à base de azul de toluidina (NCTBO) reduziu a banda de absorção para 575nm e teve uma fotodegradação mais rápida, enquanto que à base de azul de metileno (NCMB) o pico manteve-se em 660nm com fotodegradação lenta. O crescimento de fibroblastos gengivais humanos sobre superfícies de raiz...

The antimicrobial photodynamic therapy (aPDT) is used in Periodontics for reduction of bacteria in periodontal pockets as an adjuvant treatment to scaling and root planning. Recent researches aim at finding new photosensitizers in order to improve its effectiveness. The aim of this study was to test in vitro the biocompatibility and efficiency of eliminating bacteria of a new phenothiazine-based dye for aPDT in Periodontics. The following tests were performed: magnitude of demineralization and wear in bovine dentin and enamel fragments; the ideal time of application to obtain results similar to citric acid conditioning; optic behavior of new dye compared to conventional dyes; biocompatibility of human root fragments treated by the new dye in human gingival fibroblasts culture; effectiveness of the new dye in elimination of S. aureus and E. coli. In the experiments of demineralization and biocompatibility a red laser (660nm, 30mW, 45J/cm2, 30s) was applied, while in the microbiology experiments the parameters were changed (660nm, 100mW, 45J/cm2, 12s). The statistical analysis were done with parametric and non-parametric tests (p<0.05). The results showed that the new dye promoted demineralization in bovine enamel (surface hardness loss 55.5% and wear 10.33μm) and dentin (surface hardness loss 65.9% and wear 8.37μm). The demineralization was similar to citric acid and the ideal time of application was 180s. The optic tests showed that the toluidine blue-based new dye (TBOND) diminished the absorption band to 575nm and had a faster photodegradation. The methylene blue-based new dye (MBND) maintained a peak of absorption in 660nm and had a slower photodegradation. Growth of human gingival fibroblasts onto human root surfaces was similar to those treated with regular dyes (3.07 x 2.6 cells after 72h). The TBOND when used alone, reduced E. coli and S. aureus in 59.6% e 54.6%, respectively. When associated to laser, the reduction of E. coli was 52.6%. The MBND...

Cell accumulation and antileishmanial effect of exogenous and endogenous protoporphyrin IX after photodynamic treatment / Acumulación celular y efecto anti- Leishmania de la protoporfirina IX exógena y endógena después del tratamiento fotodinámico

Introduction: Photodynamic therapy (PDT) using 5-aminolevulinic acid-induced protoporphyrin IX (ALA-PpIX) constitutes an interesting alternative for cutaneous leishmaniasis treatment. Objective: To evaluate the production of PpIXbased on the administration of ALA and MAL and the effect of ALA-PDTat cellular level on non-infected and infected THP-1 cells using Leishmania ( Viannia ) panamensis or Leishmania ( Leishmania ) infantum (syn Leishmania chagasi ) parasites. Materials and methods: Protoporphyrin IX (PpIX) production and mitochondrial colocalization were evaluated by confocal microscopy. Cell toxicities were evaluated after treatment with the compounds, followed by light irradiation (597-752 nm) at 2.5 J/cm 2 fluency using a colorimetric MTT assay for THP-1 cells and a standard microscopic analysis of parasites. Results were expressed as compound concentration activity against 50% of cells or parasites (CC 50 or IC 50 ). Results: ALA or MAL induced an endogenous PpIX with a red fluorescence localized mainly in the mitochondria inside human cells. ALA and MAL-PDT induced a similar range of toxicities on THP-1 cells (CC 50 0.16±0.01mM and 0.33±0.019 mM, respectively) without any apparent inhibition of intracellular parasites in the infected cells as compared to untreated controls. Exogenous PpIX-PDT was toxic to THP-1 cells (CC 50 0.00032±0.00002 mM), L. (L.) infantum (IC 50 0.003±0.0001 mM) and L. (V.) panamensis (IC 50 0.024±0.0001 mM) promastigotes. Conclusions: Despite the effectiveness of exogenous PpIX on promastigotes and the production of PpIX by human infected cells, treatment with ALA or MAL before irradiation was unable to completely destroy L. (L.) infantum or L. (V.) panamensis intracellular amastigotes.

Introducción. El tratamiento fotodinámico con ácido 5-aminolevulínico como inductor de la protoporfirina IX (ALA-PpIX) constituye una alternativa interesante en el tratamiento de la leishmaniasis cutánea. Objetivo. Evaluar la producción de protoporfirina IX (PpIX) a partir de la administración de ALA o MAL y el efecto de la PDT con ALA a nivel celular en células THP-1 no infectadas e infectadas con Leishmania ( Viannia ) panamensis o Leishmania ( Leishmania ) infantum (syn. Leishmania chagasi ). Materiales y métodos. La producción de protoporfirina IX y su ‘colocalización´ mitocondrial se evaluaron mediante microscopía ‘confocal´. Se evaluó la toxicidad celular después del tratamiento con los compuestos y la aplicación de irradiación de luz (597-752 nm) en una fluencia de 2,5 J/cm 2 mediante el empleo de la prueba colorimétrica con metil-tiazol-tetrazolio (MTT) en las células, y de métodos microscópicos estándar en los parásitos. Los resultados se expresaron como la concentración del compuesto activo en el 50 % de las células o parásitos (CC 50 o CI 50 ). Resultados. El ácido aminolevulínico o el metil-5-aminolevulinato indujeron la protoporfirina IX endógena en células humanas, y se observó fluorescencia de color rojo en las mitocondrias. La actividad del ácido aminolevulínico y del metil-5-aminolevulinato utilizados con terapia fotodinámica fue similar en las células THP-1 (CC 50 0,16±0,01 mM y 0,33±0,019 mM, respectivamente) y, aparentemente, no inhibió los parásitos en las células infectadas, en comparación con los controles. El tratamiento exógeno con protoporfirina IX y terapia fotodinámica fue tóxico para las células THP-1 (CC 50 0,00032 ±0,00002 mM) y para los promastigotes de L. (L .) infantum (IC 50 0,003±0,0001 mM) y L. ( V .) panamensis (CI 50 0,024±0,0001 mM). Conclusiones. A pesar de la ‘fotoactividad´ del tratamiento con protoporfirina IX en promastigotes y de su producción después del tratamiento con ácido aminolevulínico y metil-5-aminolevulinato en las células infectadas con Leishmania , no se observó daño en los amastigotes presentes en las células de L. ( L .) infantum o L . ( V .) panamensis .

The susceptibility of Streptococcus mutans to antibacterial photodynamic therapy: a comparison of two different photosensitizers and light sources

Streptococcus mutans is the main etiological agent for dental caries. Recently, photodynamic therapy (PDT) has been introduced as a new modality in bacterial decontamination. Objective: This in vitro study was carried out to evaluate the susceptibility of Streptococcus mutans to antibacterial PDT using two different photosensitizers and light sources. Material and Methods: Standard suspensions of S. mutans were exposed to laser light at 662 nm and Radachlorin® or LED 630 nm in combination with Toluidine blue O (TBO). Radiation-only groups, photosensitizer alone, and groups with no treatment were used as controls. Bacterial suspension from each treatment was subcultured onto the surface of Mueller-Hinton agar plates and bacterial growth was assessed. The results were analyzed by analysis of variance and Tukey test (p<0.05). Results: PDT with TBO and Radachlorin® significantly reduced S. mutans viability, whereas no difference was observed between two groups of PDT. In the groups treated just with the photosensitizer or irradiated alone, no significant reduction of S. mutans colonies was observed. Conclusion: S. mutans colonies were susceptible to either 662 nm laser or LED light in the presence of Radachlorin® and TBO respectively with no priority. .

Study of photodynamic, sonodynamic and antioxidative influence on HeLa cell line.

Photodynamic treatment (PDT) in combination with sonodynamic treatment (SDT) can be used as suitable methods to treat malignant and benign diseases or combat resistant bacteria. Both methods affect the production of reactive oxygen species (ROS). On the other hand, antioxidants are useful for cell protection against ROS. This work was aimed to study the effect of PDT and SDT treatments on the HeLa cell line using antioxidant Pronalen Sensitive Skin® as a protection from free radicals in the cells. We evaluated the effect of sensitizer ClAlPcS2 using battery of in vitro methods, including MTT assay, kinetic production of ROS, mitochondrial membrane potential change, type of cell death and microscopic analysis. Ultrasound treatment was observed to increase the production of ROS, only in combination with PDT, particularly at higher concentrations of ClAlPcS2. The added antioxidant acts as protection against free radicals and has potential as a dietary supplement against aging or free radicals. The results of study suggested that ClAlPcS2 could be used as a potential photosensitizer for treatment of a specific type of cancers.

Modulatory effect of different riboflavin compositions on the central corneal thickness of African keratoconus corneas during collagen crosslinking

A pilot investigation to transfer the established corneal collagen crosslinking [CXL] procedure in European eyes into clinically affected African eyes and to optimize the treatment by adapting the riboflavin composition. CXL was performed in 15 eyes [11 patients] with advanced stages of keratoconus in the Eye Clinic of Bafoussam in the West Region of Cameroon. The following six riboflavin compositions with different portions of active swelling additives were applied-Solution 1 [0.5% methylhydroxypropylcellulose [MHPC]], solution 2 [1.0% MHPC], solution 3 [1.7% MHPC], solution 4 [5% dextran], solution 5 [10% dextran] and solution 6 [no active swelling ingredient]. The central corneal thickness [CCT] was measured by ultrasound pachymetry before and after de-epithelialization and at least every 10 min during CXL. The application of the riboflavin solutions resulted in the following mean final CCT values: 172 +/- 15% using solution 1 [60 min/n = 5]; 183 +/- 8% using solution 2 [60 min/n = 5]; 170% using solution 3 [60 min/n = l]; 80% using solution 4 [45 min/n = l]; 99% using solution 5 [45 min/n = l] and 150 _ 13% using solution 6 [50 min/n = 2]. The combination of riboflavin compositions with swelling and stabilizing effects on the corneal stroma seems necessary in African eyes with advanced keratoconus. Further studies are required to confirm these primary results

Corneal thickness changes during corneal collagen cross-linking with UV-A irradiation and hypo-osmolar riboflavin in thin corneas / Alterações na espessura da córnea durante "cross-linking" do colágeno com irradiação UV-A e riboflavina hipo-osmolar em córneas finas

PURPOSE: To evaluate the thinnest corneal thickness changes during and after corneal collagen cross-linking treatment with ultraviolet-A irradiation, using hypo-osmolar riboflavin solution in thin corneas. METHODS: Eighteen eyes of 18 patients were included in this study. After epithelium removal, iso-osmolar 0.1% riboflavin solution was instilled to the cornea every 3 minutes for 30 minutes. Hypo-osmolar 0.1% riboflavin solution was then applied every 20 seconds for 5 minutes or until the thinnest corneal thickness reached 400 µm. Ultraviolet-A irradiation was performed for 30 minutes. During irradiation, iso-osmolar 0.1% riboflavin drops were applied every 5 minutes. Ultrasound pachymetry was performed at approximately the thinnest point of the cornea preoperatively, after epithelial removal, after iso-osmolar riboflavin instillation, after hypo-osmolar riboflavin instillation, after ultraviolet-A irradiation, and at 1, 6 and 12 months after treatment. RESULTS: Mean preoperative thinnest corneal thickness was 380 ± 11 µm. After epithelial removal it decreased to 341 ± 11 µm, and after 30 minutes of iso-osmolar 0.1% riboflavin drops, to 330 ± 7.6 µm. After hypo-osmolar 0.1% riboflavin drops, mean thinnest corneal thickness increased to 418 ± 11 µm. After UVA irradiation, it was 384 ± 10 µm. At 1, 6 and 12 months after treatment, it was 372 ± 10 µm, 381 ± 12.7, and 379 ± 15 µm, respectively. No intraoperative, early postoperative, or late postoperative complications were noted. CONCLUSIONS: Hypo-osmolar 0.1% riboflavin solution seems to be effective for swelling thin corneas. The swelling effect is transient and short acting. Corneal thickness should be monitored throughout the procedure. Larger sample sizes and longer follow-up are required in order to make meaningful conclusions regarding safety.

OBJETIVO: Avaliar as alterações da espessura mínima da córnea durante e após o cross-linking do colágeno corneano com radiação ultravioleta A e solução hipo-osmolar de riboflavina em córneas finas. MÉTODOS: Dezoito olhos de 18 pacientes foram incluídos neste estudo. Após a remoção do epitélio, solução iso-osmolar de riboflavina 0,1% foi instilada a cada 3 minutos por 30 minutos. Solução hipo-osmolar de riboflavina 0,1% foi então aplicada a cada 20 segundos por 5 minutos ou até que a espessura mínima da córnea atingisse 400 µm. Irradiação UVA foi feita durante 30 minutos. Durante a irradiação, riboflavina iso-osmolar 0,1% foi aplicada a cada 5 minutos. Paquimetria ultrassônica foi realizada no ponto mais fino da córnea antes da cirurgia, após a remoção do epitélio, após a instilação de riboflavina iso-osmolar, após a instilação de riboflavina hipo-osmolar, após a irradiação com UVA e após 1, 6 e 12 meses do tratamento. RESULTADOS: Antes da cirurgia, a espessura mínima da córnea era de 380 ± 11 µm. Após a remoção do epitélio, este valor foi reduzido para 341 ± 11 µm e após 30 minutos de riboflavina iso-osmolar, caiu para 330 ± 7,6 µm. Após a riboflavina hipo-osmolar, a espessura mínima da córnea aumentou para 418 ± 11 µm. Após a irradiação com UVA, era de 384 ± 10 µm. Após 1, 6 e 12 meses do tratamento este valor era de 372 ± 10, 381 ± 12,7 e 379 ± 15 µm, respectivamente. Não foram observadas complicações no intra ou no pós-operatório precoce ou tardio. CONCLUSÕES: A solução de riboflavina hipo-osmolar 0,1% parece ser eficaz para edemaciar córnea finas. Este efeito é transitório e de curta duração. A espessura da córnea deveria ser monitorada durante todo o procedimento. Maior número de casos e seguimento prolongado são necessários para tirarmos conclusões quanto à segurança.

Antimicrobial susceptibility of photodynamic therapy (UVA/riboflavin) against Staphylococcus aureus / Suscetibilidade antimicrobiana da terapia fotodinâmica (UVA/riboflavina) contra Staphylococcus aureus

PURPOSE: To assess S. aureus in vitro viability after the exposure to ultraviolet light A (UVA) and riboflavin (B2). METHODS: Samples of S. aureus in 96 well plates (in triplicate) were exposed to riboflavin (B2) and ultraviolet light A (365 nm wavelength) at a power density of 3 mW/cm², 8 mm spot diameter, for 30 minutes. Control groups were prepared as well in triplicate: blank control, ultraviolet light A only, riboflavin only and dead bacteria Control. The bacterial viability was measured using fluorescent microscopy. In order to investigate the occurrence of "viable but non-culturable" microorganisms after treatment, the cell viability was also investigated by plate culture procedure onto a broth medium. Statistical analysis was performed using the triplicate values from each experimental condition. RESULTS: No difference was observed among the treatment group and the control samples (p=1). CONCLUSION: The combination of riboflavin 0.1% and ultraviolet light A at 365 nm did not exhibit antimicrobial activity against oxacillin susceptible S. aureus.

OBJETIVO: Avaliar a viabilidade celular de S. aureus in vitro após a exposição de riboflavina (B2) e luz ultravioleta A (UVA). MÉTODOS: Amostras de S. aureus colocadas em uma placa de 96 poços (em triplicata) foram expostas a riboflavina 0,1% (B2) e luz ultravioleta (comprimento de onda de 365 nm) poder de 3 mW/cm², 8 mm de diâmetro, por 30 minutos. Grupos controles foram também preparados em triplicata: controle branco, somente luz ultravioleta A, somente riboflavina e controle morto. A viabilidade bacteriana foi analisada usando microscópio de fluorescência. Para investigar a ocorrência de micro-organismos "viáveis porem não cultiváveis" a viabilidade celular foi avaliada utilizando-se placas de meio de cultivo bacteriano. Analise estatística foi realizada utilizando-se os valores obtidos em triplicata de cada grupo experimental. RESULTADOS: Nenhuma diferença foi observada entre o grupo tratamento e os grupos controle (p=1). CONCLUSÃO: A combinação riboflavina 0,1% e luz ultravioleta 365 nm de comprimento de onda não demonstrou atividade antimicrobiana contra S. aureus oxacilina sensível.

Susceptibility of planktonic cultures of Streptococcus mutans to photodynamic therapy with a light-emitting diode

The objective of this study was to evaluate the effect of photodynamic therapy with erythrosine and rose bengal using a light-emitting diode (LED) on planktonic cultures of S. mutans. Ten S. mutans strains, including nine clinical strains and one reference strain (ATCC 35688), were used. Suspensions containing 10(6) cells/mL were prepared for each strain and were tested under different experimental conditions: a) LED irradiation in the presence of rose bengal as a photosensitizer (RB+L+); b) LED irradiation in the presence of erythrosine as a photosensitizer (E+L+); c) LED irradiation only (P-L+); d) treatment with rose bengal only (RB+L-); e) treatment with erythrosine only (E+L-); and f) no LED irradiation or photosensitizer treatment, which served as a control group (P-L-). After treatment, the strains were seeded onto BHI agar for determination of the number of colony-forming units (CFU/mL). The results were submitted to analysis of variance and the Tukey test (p < 0.05). The number of CFU/mL was significantly lower in the groups submitted to photodynamic therapy (RB+L+ and E+L+) compared to control (P-L-), with a reduction of 6.86 log10 in the RB+L+ group and of 5.16 log10 in the E+L+ group. Photodynamic therapy with rose bengal and erythrosine exerted an antimicrobial effect on all S. mutans strains studied.

Photodynamic therapy - A strategic review.

Mechanical removal of the biofilm and adjunctive use of antibacterial disinfectants or various antibiotics have been conventional methods of the periodontitis therapy. There has been an upsurge of bacterial strains becoming resistant due to the injudicious use of antibiotics, recently. As a result there is pronounced interest and keenness in the development of alternate antimicrobial concepts. As the scientific community seeks alternatives to antibiotic treatment, periodontal researchers have found that photodynamic therapy (PDT) is advantageous to suppress anaerobic bacteria. Hence, PDT could be an alternative to conventional periodontal therapeutic methods. This review elucidates the evolution and use of photo dynamic therapy. The application of photosensitizing dyes and their excitation by visible light enables effective killing of periodontopathogens. Even though PDT is still in the experimental stages of development and testing, the method may be an adjunct to conventional antibacterial measures in periodontology. PDT application has an adjunctive benefit besides mechanical treatment at sites with difficult access. Necessity for flap operations may be reduced, patient comfort may increase and treatment time decrease. Clinical follow-up studies are needed to confirm the efficacy of the procedure.

Photodynamic effect of light-emitting diode light on cell growth inhibition induced by methylene blue.

The aim of this study was to propose the use of red light-emitting diode (LED) as an alternative light source for methylene blue (MB) photosensitizing effect in photodynamic therapy (PDT). Its effectiveness was tested against Staphylococcus aureus (ATCC 26923), Escherichia coli (ATCC 26922), Candida albicans (ATCC 90028) and Artemia salina. The maximum absorption of the LED lamps was at a wavelength of 663 nm, at intensities of 2,4,6 and 12 J.cm-2 for 10, 20, 30 and 60 min of exposure, respectively. Assays with and without LED exposure were carried out in plates containing MB at concentrations of 7 to 140.8 (micro) M for microorganisms and 13.35 to 668.5 (micro) M for microorganisms or microcrustaceans.The LED exposure induced more than 93.05%, 93.7% and 93.33% of growth inhibition for concentrations of 42.2 (micro)M for S.aureus (D-value=12.05 min) and 35.2 (micro)M for E.coli (D-value=11.51 min) and C.albicans (D-value=12.18 min), respectively after 20 min of exposure. LED exposure for 1 h increased the cytotoxic effect of MB against A.salina from 27% to 75%.Red LED is a promising light device for PDT that can effectively inhibit bacteria, yeast and microcrustacean growth.

Establishment of mus skin photo-damage model by 8-MOP plus UVA irradiation / 华中科技大学学报（医学）（英德文版）

To establish a simple and reliable animal model of skin photo-damage, 20 mice were treated with 8-MOP and exposed to UVA (UVA 320-400 nm) for 24 h. After irradiation, the structure of the epidermis and dermis, collagen fibers, elastic fibers were observed by using HE staining and Weigert technique and compared with the normal controls. The acanthosis and epidermis proliferation with accompanying hyperkeratosis and parakeratosis were observed. Inflammatory infiltration was noted in the dermis. The elastic fibers became coarse, irregularly arranged and clustered, with their number increased. The collagen fibers showed obvious degeneration and some amorphous materials could also be observed. The blood vessels were irregularly dilated and vascular walls were thickened, with infiltration of inflammatory cells. It is concluded that murine photodamage model can be quickly, conveniently and reliably established by means of 8-MOP/UVA.

Photodynamic inactivation of in vitro bacterial cultures from pressure ulcers

PURPOSE: To evaluate in vitro the antibacterial effect of diode laser light of wavelength 650 nm, in association with the photosensitive substance toluidine blue, on the bacteria in infected skin ulcers. METHODS: Samples were collected by means of swabs containing a medium for transporting infected material from skin ulcers. The material was inoculated into culturing medium containing azide blood agar for the growth of Gram-positive bacteria, and MacConkey agar for Gram-negative bacteria, and incubated for 48 hours. The results obtained from counting the colony-forming units were correlated and subjected to statistical analysis, adopting the significance level of p > or = 0.05. RESULTS: From analysis of variance (ANOVA), the result for the general mean was p = 0.0215. Using the t test with post-hoc test, the result for TBO vs. Control was p = 0.0186, and for TBO + Laser vs. Control it was p = 0.0039. CONCLUSION: There was a significant reduction in colony-forming units when the cultures were subjected to photodynamic therapy.

OBJETIVO: Avaliar in vitro o efeito antibacteriano do laser diodo com comprimento de onda de 650nn, associado a substancia fotossensível azul de toluidina sobre as bactérias de ulceras cutâneas infectadas. MÉTODOS: Foram coletadas amostras através de um swab com meio de transporte, de material infectado de úlceras cutâneas. Os materiais foram inoculadas em meios de cultura contendo ágar sangue azida para o crescimento de bactérias gram-positivas e agar Mac Conkey para as gram-negativas, e incubadas por 48 horas. Os resultados obtidos da contagem das unidades formadoras de colônias foram relacionados e submetidos a analise estatística adotando como nível de significância p > ou = 0.05. RESULTADOS: Os resultados da análise de variância ANOVA para a media geral foi p= 0,0215 e para o post hoc test teste t. TBO x Controle p=0,0186, TBO + Laser x Controle p=0,0039. CONCLUSÃO: Houve redução, significativa das unidades formadoras de colônias quando submetidas à terapia fotodinâmica.