Riboflavin and ultraviolet light a therapy as an adjuvant treatment for medically refractive Acanthamoeba keratitis: report of 3 cases.

PURPOSE: To present the first 3 cases of Acanthamoeba keratitis (AK), unresponsive to medical treatment, that were successfully treated with a novel adjunctive therapy using ultraviolet light A (UVA) and riboflavin (B2). DESIGN: Interventional case series. PARTICIPANTS: Two patients with confirmed AK and 1 patient with presumptive AK, which were all refractive to multidrug conventional therapy. INTERVENTION: Two treatment sessions involving topical application of 0.1% B2 solution to the ocular surface combined with 30 minutes of UVA irradiation focused on the corneal ulcer. MAIN OUTCOME MEASURES: Clinical examination by slit lamp, confocal microscopy, and histopathology, when available. RESULTS: All patients in these series showed a rapid reduction in their symptoms and decreased ulcer size after the first treatment session. The progress of the clinical improvement began to slow after 1 to 3 weeks of the first application and was then renewed after the second application. All ancillary signs of inflammation mostly resolved after the second treatment session. The ulcers in all patients continued to decrease and were closed within 3 to 7 weeks of the first application. Two patients developed dense central corneal scars, and penetrating keratoplasty was performed for visual rehabilitation. Histopathologic examination of the excised tissue revealed no Acanthamoeba organisms. The remaining patient had no symptoms or signs of infection, both clinically and by confocal microscopy, and was left with a semitransparent eccentric scar that did not affect visual acuity. CONCLUSIONS: The adjunctive use of UVA and B2 therapy seems to be a possible alternative for selected cases of medication-resistant AK.

Proteoglycan synthesis by human corneal explants submitted to laser in situ keratomileusis (LASIK).

PURPOSE: To evaluate the acute effects of laser in situ keratomileusis (LASIK) upon the synthesis of proteoglycans (PGs) and collagen fibril organization in human corneal explants. METHODS: Human corneas that had been rejected for transplants were obtained at Banco de Olhos of Hospital São Paulo. For each eye pair, one cornea was submitted to refractive surgery, and the other was used as its matched control. After surgery, the corneas were excised from the eyes and immediately placed in a Ham F-12 nutrient mixture containing (35)S-sulfate for the metabolic labeling of PGs. After 24 h incubation, PGs were extracted and identified by a combination of agarose gel electrophoresis and enzymatic degradation with protease and specific glycosaminoglycan lyases. Histopathological and birefringence analysis were performed in fixed tissue slices. RESULTS: A marked decrease in (35)S-sulfate incorporation in PGs was observed in corneal explants that received LASIK, especially concerning dermatan sulfate-PGs, with keratan sulfate- and heparan sulfate-PG synthesis reduced to a lower degree. Only low molecular weight PGs were present in the corneas, both before and 24 h after LASIK. No sign of wound healing processes were observed, but a marked change in corneal birefringence was seen following LASIK treatment. CONCLUSIONS: Laser application led to decreased PG biosynthesis in human corneal explants, with marked changes in the collagen fibril organization, as revealed by changes in the tissue birefringence.

[Study of ocular fungal microbiota in patients with Hansen's disease and in individuals who deal with them]. / Estudo da microbiota fúngica da conjuntiva ocular em portadores de hanseníase e seus comunicantes.

PURPOSE: To study the conjunctival fungal flora of patients with leprosy who live in the colony hospital and of those who deal with "Hansen" patients. METHODS: Sixty-one eyes (61 patients) with Hansen's disease and twenty-five healthy eyes (25 individuals) of individuals who deal with patients were investigated. RESULTS: Conjunctival fungi growth occurred in 19.67% and 20% of the patients and healthy individuals, respectively. There was no significant difference (p<0.05) regarding fungi percentage of fungi isolated from the conjunctiva between both groups. Candida ssp was the most frequent fungus isolated from the conjunctiva of patients with Hansen's disease, whereas Penicillium spp was the most frequent fungus isolated from the group of people who deal with these patients. CONCLUSION: The most frequent yeast isolated from conjunctival samples of leprosy patients was Candida, which could be explained by cellular immunodeficiency of these patients, associated with dry eye and the prolonged use of antibiotics.

Effects of topical human amniotic fluid and human serum in a mouse model of keratoconjunctivitis sicca.

PURPOSE: To compare the effects of topical human amniotic fluid (HAF), topical human serum (HS), and topical artificial tears in a mouse model of dry eye. METHODS: Thirty C57BL/6 mice were divided into 3 treatment groups: HAF, HS, and preservative-free artificial tears. Dry eye was induced by an injection of botulinum toxin B (BTX-B) into the lacrimal gland. Tear production and ocular surface fluorescein staining were evaluated in each mouse at 6 time points during a 4-week period. Goblet cell density was assessed in stained histological sections. Apoptotic keratocytes were evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling test assay. RESULTS: A significant decrease in tear production was observed 3 days after BTX-B injection in all groups. At week 1, the HAF and HS groups had improved tear production compared with the control group (P < 0.001 and P = 0.003, respectively). HAF had a significantly improved fluorescein staining score compared with the HS (P = 0.043) and control (P = 0.007) groups at week 2. Goblet cell density was significantly decreased in the control group compared with the HAF and HS groups (P < 0.001). No difference in the amount of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive keratocytes was observed among the groups. CONCLUSION: HAF was superior to HS and artificial tears for improving corneal staining within 2 weeks of therapy in this induced mouse model of keratoconjunctivitis sicca. Clinical studies are needed to ascertain the benefits of these therapies in patients with ocular surface disorders associated with dry eye.

[Topical amniotic fluid: a potential new alternative for ocular surface diseases]. / Líquido amniótico tópico: uma potencial nova alternativa para doenças da superfície ocular.

Amniotic fluid bathes the fetus during intrauterine life and is in permanent contact with the fetal ocular surface in this important period of development. It contains a series of growth factors that may have multiple effects on the wound healing process. These factors are thought to accelerate the recovery of corneal sensitivity and nerve regeneration after keratorefractive procedures, and also may control scar formation and balance the ocular surface after topical application. Hundreds of different proteins have been identified in the human amniotic fluid, and the role of each still not quite understood. The outcomes obtained so far with amniotic fluid application to ocular surface diseases suggest a promising therapy. Research is underway to identify the effects of specific factors of the amniotic fluid in ocular inflammation. The purpose of this review is to report the properties and current utilizations of amniotic fluid as well as to summarize the recent studies related to the use of this fluid for ocular surface diseases.

Antimicrobial efficacy of riboflavin/UVA combination (365 nm) in vitro for bacterial and fungal isolates: a potential new treatment for infectious keratitis.

PURPOSE: To demonstrate the antimicrobial properties of riboflavin/UVA (365 nm) against common pathogens. METHODS: One group of bacteria (Pseudomonas aeruginosa [PA], Staphylococcus aureus [SA], and Staphylococcus epidermidis [SE]) was tested by using Kirby-Bauer discs with (1) empty disc (Control - C); (2) riboflavin 0.1% (B2); (3) riboflavin 0.1% previously activated by UVA (B2'); (4) UVA alone (UVA); (5) group 2+additional UVA exposure (UVA+B2); and (6) group 3+additional UVA exposure (UVA+B2'). In addition, another group of microbes was tested with the same approach: methicillin-resistant S. aureus (MRSA), multidrug-resistant P. aeruginosa (MDRPA), drug-resistant Streptococcus pneumoniae (DRSP), and Candida albicans (CA). The mean growth inhibition zone (GIZ) in square millimeters was measured around the discs. The mean standard deviation (MSD) was calculated to be 3.65 when alpha = 0.01. A mean deviation (MD) > MSD indicates a significant difference. RESULTS: In the first group, the GIZ was significantly greater after UVA (MD = 14.30), UVA+B2 (MD = 39.61), and UVA+B2' (MD = 40.45) when compared with C, B2, and B2'. UVA alone was less effective than UVA+B2 (MD = 25.31) and UVA+B2' (MD = 26.15). The second group demonstrated increased GIZ in UVA (MD = 6.98), UVA+B2 (MD = 17.80), and UVA+B2' (MD = 21.15) when compared with C, B2, and B2'. UVA alone was less effective against the second group of bacteria than was UVA+B2 (MD = 10.82) and UVA+B2' (MD = 14.17). CA did not show any GIZ after treatment. CONCLUSIONS: Riboflavin/UVA was effective against SA, SE, PA, MRSA, MDRPA, and DRSP, but was ineffective on CA and has the potential for use in treatment of microbial keratitis in the future.

Líquido amniótico tópico: uma potencial nova alternativa para doenças da superfície ocular: [revisão] / Topical amniotic fluid: a potential new alternative for ocular surface diseases: [review]

O líquido amniótico banha o feto durante a vida intra-uterina e está em contato permanente com a superfície ocular durante este importante período do desenvolvimento. Ele contém uma série de fatores de crescimento que podem ter diversos efeitos sobre o processo cicatricial. Estes fatores aceleram a recuperação da sensibilidade corneana e regeneração nervosa após procedimentos cerato-refrativos, além de controlar a formação de cicatriz e o equilíbrio da superfície ocular após sua aplicação tópica. Centenas de diferentes proteínas têm sido identificadas no líquido amniótico humano e o papel de cada uma continua desconhecido. Os resultados obtidos até o momento sobre a aplicação de líquido amniótico em doenças de superfície ocular sugerem uma terapia promissora. Pesquisas estão sendo realizadas para identificar os efeitos dos fatores específicos do líquido amniótico sobre a inflamação ocular. O objetivo desta revisão é relatar as propriedades e utilizações atuais do líquido amniótico, bem como apresentar os recentes estudos relacionados ao uso deste líquido e doenças da superfície ocular.

Amniotic fluid bathes the fetus during intrauterine life and is in permanent contact with the fetal ocular surface in this important period of development. It contains a series of growth factors that may have multiple effects on the wound healing process. These factors are thought to accelerate the recovery of corneal sensitivity and nerve regeneration after keratorefractive procedures, and also may control scar formation and balance the ocular surface after topical application. Hundreds of different proteins have been identified in the human amniotic fluid, and the role of each still not quite understood. The outcomes obtained so far with amniotic fluid application to ocular surface diseases suggest a promising therapy. Research is underway to identify the effects of specific factors of the amniotic fluid in ocular inflammation. The purpose of this review is to report the properties and current utilizations of amniotic fluid as well as to summarize the recent studies related to the use of this fluid for ocular surface diseases.

Estudo da microbiota fúngica da conjuntiva ocular em portadores de hanseníase e seus comunicantes / Study of ocular fungal microbiota in patients with Hansen's disease and in individuals who deal with them

OBJETIVO: Investigar a microbiota fúngica de pacientes portadores de hanseníase residentes no hospital-colônia e seus comunicantes. MÉTODOS: Foram estudados por meio da cultura em meio de ágar-Saboraud-dextrose, o material do fórnice conjuntival, de sessenta e um pacientes portadores de hanseníase, residentes no hospital-colônia de Goiânia e 25 indivíduos que residiam ou trabalhavam com os referidos pacientes, no mínimo há cinco anos. RESULTADOS: Nos portadores de hanseníase foram isolados fungos da conjuntiva de 12 pacientes (19,67 por cento), sendo o gênero Candida o mais isolado, e no grupo controle de 5 indivíduos (20 por cento), em que o gênero mais isolado foi Penicillium spp. CONCLUSÃO: Candida foi o gênero predominante na flora conjuntival dos portadores de hanseníase: fato que poderia ser explicado pela imunodeficiência celular que estes pacientes apresentam, além do olho seco e uso prolongado de antibiótico.

PURPOSE: To study the conjunctival fungal flora of patients with leprosy who live in the colony hospital and of those who deal with "Hansen" patients. METHODS: Sixty-one eyes (61 patiets) with Hansen's disease and twenty-five heathy eyes (25 individuals) of individuals who deal with patients were investigated. RESULTS: Conjunctival fungi growth occurred in 19.67 percent and 20 percent of the patients and healthy individuals, respectively. There was no significant difference (p<0.05) regarding fungi percentage of fungi isolated from the conjunctiva between both groups. Candida ssp was the most frequent fungus isolated from the conjunctiva of patients with Hansen's disease, whereas Penicillium spp was the most frequent fungus isolated from the group of people who deal with these patients. CONCLUSION: The most frequent yeast isolated from conjunctival samples of leprosy patients was Candida, which could be explained by cellular immunodeficiency of these patients, associated with dry eye and the prolonged use of antibiotics.

Microbiota fúngica conjuntival: estudo comparativo entre pacientes com AIDS, pacientes infectados pelo HIV e pacientes HIV-negativos antes da era do HAART / Fungal microbiota: comparative study between patients with acquired immuno-deficiency syndrome, patients infected with human immunodeficieny virus and immunocompetent individuals before the HAART era

Foi pesquiasda a presença de fungos na conjuntiva ocular de 23 pacientes com AIDS, 24 pacientes infectados pelo HIV, ambos de controle ambulatoraial e 48 indivíduos HIV negativos, durante um ano, nos meses de fevereiro, abril, julho e outubro. O fungo mais isolado foi Penicillium sp, seguido de Aspergillus sp, Candida sp e Rhodotorula sp. Embora sem diferença significante, em todas as coletas foi verificados um crescimento maior de fungos na conjuntiva dos portadores de AIDS, seguido pelo grupo infectado pelo HIV e em menor número nos individuos HIV-negativos

Efeito agudo da cirurgia refrativa sobre a síntese de proteoglicanos em explantes de córnea humana / Acute effect of refractive surgery upon proteoglycan synthesis in human cornea explants

Objetivo: Avaliar o efeito agudo (nas primeiras 24 horas) da cirurgia refrativa a laser sobre a síntese de proteoglicanos em explantes de córnea humana mantidos em cultura. Métodos: Foram utilizadas córneas do Banco de Olhos do Hospital São Paulo que foram rejeitadas para transplantes. Três tipos de procedimentos cirúrgicos foram analisados: ceratomileuse assistida por excimer laser in situ (LASIK), ceratectomia fotorrefrativa por excimer laser (PRK) e ceratotomia automatizada (AK). Para cada par de córneas, uma foi submetida à cirurgia e a outra foi mantida como seu controle pareado. Após o I procedimento cirúrgico, os botões corneais foram excisados e colocados I imediatamente em meio de cultura F-12 contendo 35S-sulfato para marcação I metabólica dos proteoglicanos (PGs). Após 24 horas de incubação a 37° C, em atmosfera contendo 2,5 por cento de C02, os PGs foram extraídos com hidrocloreto de guanidina 4 M e identificados por uma combinação de eletroforese em gel de agarose e degradação enzimática com mucopolissacaridases bacterianas específicas, vistos por radioautograma e quantificados em um contador de cintilações. Além disso, foram realizadas análise histológica das amostras corneais após os três tipos de procedimentos cirúrgicos e análise de birrefringência (microscopia de polarização) após a realização do LASIK. Resultados: Uma marcante diminuição na incorporação de 35S-sulfato foi observada em explantes de córnea humana submetidos a LASIK. Este efeito pode ser devido à morte celular. Em contraste, os grupos PRK e AK tiveram um efeito variável sobre a incorporação de 35S-sulfato. Isto pode ser decorrente da combinação de três eventos diferentes: maior disponibilidade de 35S-sulfato, morte celular (necrose ou apoptose), e/ou ativação de ceratócitos após o trauma cirúrgico. Previamente foi demonstrado que a remoção da camada epitelial leva a um aumento da incorporação de 35S-sulfato em proteoglicanos de dermatam sulfato (DS) em explantes de córnea humana, provavelmente devido a uma maior disponibilidade de 35S-sulfato nas células do estroma superficial e/ou estímulo dos ceratócitos superficiais. Em contraste, a ablação a laser em si pode levar à morte celular, seja por necrose ou por apoptose. Os exames histológicos não apresentaram alterações significativas na resposta cicatricial após os três procedimentos cirúrgicos (LASIK, AK e PRK), talvez porque essa análise (HE) tenha sido realizada 24 horas após as cirurgias. O estudo da birrefringência em LASIK demonstrou uma diminuição das medidas de retardos ópticos (RO). Conclusões: O laser causou uma diminuição na incorporação de 35S-sulfato após o LASIK, provavelmente devido à morte de ceratócitos. Após a realização de PRK e AK, houve uma grande variabilidade na incorporação de 35S-sulfato. A análise de birrefringência demonstrou uma diminuição da ordem molecular e do estado de orientação da agregação hierarquizada da córnea após o LASIK.