Investigating the effect of intracameral cefuroxime on oxidative stress and apoptosis in the rat cornea / Investigando o efeito de cefuroxima intracameral sobre o estresse oxidativo e a apoptose na córnea de ratos

ABSTRACT PURPOSE: We examined the effect of intracameral administration of cefuroxime on oxidative stress and endothelial apoptosis in rat corneal tissue. METHODS: In total, 30 rats were divided into 3 groups of 10 rats each (intracameral administration of cefuroxime 0.1 mg/0.01 mL (cefuroxime group); intracameral administration of balanced salt solution 0.01 mL (control group); or absence of intracameral injection (sham group). Corneal endothelial apoptosis was assessed by immunohistochemical analysis using caspase-3 and caspase-8. Total oxidant status, total antioxidant status, oxidative stress index, and paraoxonase and arylesterase levels were examined in corneal endothelial tissue and serum. RESULTS: Paraoxonase levels in the serum were significantly different between the sham and cefuroxime groups (p=0.027). A significant difference was also observed in total oxidant status levels between the cefuroxime and balanced salt solution groups (p=0.023). In addition, there were significant differences in total antioxidant status levels in corneal tissue between the cefuroxime and sham groups (p<0.001) and between the cefuroxime and balanced salt solution groups (p<0.001). Furthermore, significant differences were also observed in oxidative stress index levels between the cefuroxime and balanced salt solution groups (p=0.001) and between the cefuroxime and sham groups (p=0.026). According to the immunohistochemical staining results, a significant association with caspase-3 activity existed between the cefuroxime and balanced salt solution groups (p=0.007), while no significant difference was found with caspase-8 activity (p=0.541). Caspase-3 activity exhibited a significant relationship between the sham and balanced salt solution groups (p=0.018), but no relationship was found with caspase-8 activity (p=0.623). CONCLUSION: Immunohistochemical examination revealed that intracameral cefuroxime increased apoptosis when compared to the sham and balanced salt solution groups. Moreover, intracameral cefuroxime increased oxidative stress in the cornea and simultaneously induced apoptosis.

RESUMO OBJETIVO: Examinamos o efeito da administração intracameral da cefuroxima sobre o estresse oxidativo e a apoptose endotelial no tecido corneano de ratos. MÉTODOS: No total, 30 ratos foram divididos em 3 grupos de 10 ratos cada (administração intracameral de cefuroxima 0,1 mg/0,01 mL (grupo cefuroxima), administração intracameral de solução salina balanceada 0,01 mL (grupo controle) ou ausência de injeção intracameral (grupo sham)). A apoptose endotelial da córnea foi avaliada por análise imuno-histoquimica usando caspase-3 e -8. O status oxidante total, o status antioxidante total, o índice de estresse oxidativo e os níveis de a paraoxonase e arilesterase foram investigados no tecido endotelial da córnea e no soro. RESULTADOS: Os níveis de paraoxonase no soro foram significativamente diferentes entre os grupos sham e cefuroxima (p=0,027). Foi também observada uma diferença significativa nos níveis de estado oxidante total entre os grupos cefuroxima e solução salina balanceada (p=0,023). Além disso, houve diferenças significativas nos níveis de status antioxidante total no tecido da córnea entre os grupos cefuroxima e sham (p<0,001) e entre os grupos cefuroxima e solução salina balanceada (p<0,001). Diferenças significativas também foram observadas nos níveis do índice de estresse oxidativo entre os grupos cefuroxima e solução salina balanceada (p=0,001) e entre os grupos cefuroxima e sham (p=0,026). De acordo com os resultados de coloração imuno-histoquimica, houve associação significativa com a atividade da caspase-3 entre os grupos cefuroxima e solução salina balanceada (p=0,007), enquanto não houve diferença significativa com a atividade da caspase-8 (p=0,541). A atividade da caspase-3 exibiu uma relação significativa entre os grupos sham e solução salina balanceada (p=0,018), mas nenhuma relação foi encontrada com a atividade da caspase-8 (p=0,623). CONCLUSÃO: O exame imuno-histoquímico revelou que a cefuroxima intracameral aumentou a apoptose quando comparada com os grupos sham e solução salina balanceada. Além disso, a cefuroxima intracameral aumentou o estresse oxidativo na córnea e induziu simultaneamente a apoptose.

Esterasa leucocitaria como prueba diagnóstica eficaz, económica y rápida ante un proceso infeccioso articular de rodilla / Leucocyte esterase as a reliable diagnostic, cost-effective and fast in knee infections

Resumen: Antecedentes: La infección articular es un reto ortopédico por la complejidad diagnóstica y sus efectos devastadores al no tratarse oportunamente. Se cuenta con diversos estudios de diagnóstico: cultivo, VSG, PCR, conteo de leucocitos, entre otros, pero ninguno es preciso, tardan más de 30 minutos en realizarse y requieren una infraestructura compleja. En este estudio se determina la sensibilidad y especificidad de la esterasa leucocitaria para la detección de un proceso infeccioso articular en población mexicana. Material y métodos: de Noviembre de 2015 a Abril de 2016 se obtuvo líquido sinovial de pacientes con diagnóstico de infección articular con o sin implante y sin infección con patología degenerativa de rodilla. Se evaluó la muestra mediante el test de esterasa leucocitaria COMBI-SCREEN 11SYS con lectura colorimétrica a los dos minutos, determinando positivo para infección: dos cruces, el resto de la muestra fue enviado a cultivo. Resultados: Se aplicó el test a 64 muestras de líquido sinovial de rodilla, 19 diagnosticadas con infección articular y 45 sin infección. Se obtuvo una sensibilidad 100%, especificidad 88.24% VPP 68.42% y VPN 100%, índice de concordancia kappa 0.753 mediante el programa IBM SPSS Statistics 22, Python versión 2.7. Conclusiones: La esterasa leucocitaria es una prueba rápida, económica y eficaz para detectar un proceso infeccioso contra un proceso inflamatorio con alta probabilidad de acierto. Este estudio presentó un índice de concordancia kappa de 0.753, demostrando ser reproducible, por lo que se recomienda implementarse en los servicios de urgencias a nivel nacional.

Abstract: Background: The articular infection represents a challenge due to its complexity and its devastating effect when not treated promptly. We have various diagnostic studies: cultures, ESR, CRP, count of leukocytes, among others but none is specific, it takes more than 30 minutes to complete and require complex infrastructure. In this study we determine the sensitivity and specificity of the leukocyte esterase for detection of an infectious process joint in Mexican population. Material and methods: From November 2015 to April 2016 was obtained synovial fluid from patients with diagnosis of knee infection with or without implant and without infection with degenerative pathology of the knee. It assessed the sample through the COMBI-SCREEN 11SYS leukocyte esterase with reading colorimetric test at two minutes determining positive for infection: two crosses, the remainder of the sample was sent to culture. Results: We perform the test in 64 samples of synovial fluid of knee joint 19 diagnosed with infection and 45 without infection. Was obtained a sensitivity 100%, specificity of 88.24%, PPV 68.42% and PNV 100%, kappa index 0.753 using the program IBM SPSS Statistics 22, Python ver. 2.7. Conclusions: Leukocyte esterase is a fast, economical and effective to detect an infectious process against one inflammatory with high probability of success. This study showed an index of concordance 0.753 Kappa, proving to be reproducible so recommend be implemented in the emergency department at the national level.

Esterasa leucocitaria como prueba diagnóstica ante un proceso infeccioso articular de rodilla / Leukocyte esterase as a diagnostic tool for an infectious disease of the knee

Resumen: Antecedentes: La infección articular es un reto ortopédico por su complejidad diagnóstica y efectos devastadores al no tratarse oportunamente. Contamos con diversos estudios de diagnóstico: cultivo, VSG, PCR, conteo de leucocitos, entre otros, pero ninguno es preciso, tardan más de 30 minutos en realizarse y requieren de infraestructura compleja. En este estudio determinamos la sensibilidad y especificidad de la esterasa leucocitaria para la detección de un proceso infeccioso articular en población mexicana. Material y métodos: Durante Noviembre de 2015 a Abril de 2016, se obtuvo líquido sinovial de pacientes con diagnóstico de infección articular con o sin implante, y de otros sin infección, con patología degenerativa de rodilla. Se evaluó la muestra mediante el test de esterasa leucocitaria COMBI-SCREEN 11SYS con lectura colorimétrica a los dos minutos; se determinó positivo para infección con dos cruces; el resto de la muestra fue enviado a cultivo. Resultados: Realizamos el test en 64 muestras de líquido sinovial de rodilla, 19 diagnosticadas con infección articular y 45 sin infección. Se obtuvo una sensibilidad de 100%, especificidad de 88.24%, VPP de 68.42% y VPN de 100%; índice de Kappa de .753. Conclusiones: La esterasa leucocitaria es una prueba eficaz para detectar un proceso infeccioso contra uno inflamatorio con alta probabilidad de acierto. Este estudio presentó un índice de concordancia Kappa de 0.753, con lo que demostró ser reproducible, por lo que recomendamos implementarlo en los servicios de urgencias a nivel nacional.

Abstract: Background: Articular infection is an orthopedic challenge due to its difficult diagnosis and devastating results. Various diagnostic studies exist: culture, ESR, CRP, count of leukocytes, among others, but none is specific, they all take more than 30 minutes to complete, and require complex infrastructure. In this study, we determine the sensitivity and specificity of the leukocyte esterase for detection of an infectious process joint in Mexican population. Material and methods: During November 2015 to April 2016, we obtained synovial fluid from two groups of patients: one with a diagnosis of synovial joint infection with or without implant, and the control group, without infection but with degenerative pathology of the knee. We evaluated the sample using the leukocyte esterase test COMBI-SCREEN 11SYS with colorimetric reading at two minutes; two crosses determined positive for infection; the remainder of the sample was sent for culture. Results: We performed the test in 64 samples of synovial fluid, 19 diagnosed with articular infection and 45 without it. We obtained a sensitivity of 100%, specificity of 88.24%, PPV of 68.42%, and NPV of 100%; Kappa index of .753. Conclusions: Leukocyte esterase is an effective test to detect an infectious process against an inflammatory one with a high probability of success. This study presented an index of agreement Kappa of 0.753, proving to be reproducible.

Correlation between the serum and tissue levels of oxidative stress markers and the extent of inflammation in acute appendicitis

OBJECTIVES: To determine the serum and tissue levels of markers of impaired oxidative metabolism and correlate these levels with the histopathology and Alvarado score of acute appendicitis patients. METHOD: Sixty-five acute appendicitis patients (mean age, 31.4±12.06 years; male/female, 30/35) and 30 healthy control subjects were studied. The Alvarado score was recorded. Serum samples were obtained before surgery and 12 hours postoperatively to examine the total antioxidant status, total oxidant status, paraoxonase, stimulated paraoxonase, arylesterase, catalase, myeloperoxidase, ceruloplasmin, oxidative stress markers (advanced oxidized protein products and total thiol level) and ischemia-modified albumin. Surgical specimens were also evaluated. RESULTS: The diagnoses were acute appendicitis (n = 37), perforated appendicitis (n = 8), phlegmonous appendicitis (n = 12), perforated+phlegmonous appendicitis (n = 4), or no appendicitis (n = 4). The Alvarado score of the acute appendicitis group was significantly lower than that of the perforated+phlegmonous appendicitis group (p = 0.004). The serum total antioxidant status, total thiol level, advanced oxidized protein products, total oxidant status, catalase, arylesterase, and ischemia-modified albumin levels were significantly different between the acute appendicitis and control groups. There was no correlation between the pathological extent of acute appendicitis and the tissue levels of the markers; additionally, there was no correlation between the tissue and serum levels of any of the parameters. CONCLUSIONS: The imbalance of oxidant/antioxidant systems plays a role in the pathogenesis acute appendicitis. The Alvarado score can successfully predict the presence and extent of acute appendicitis. .

A novel approach for the identification of isozymes using inhibitors.

Carboxylesterase isozymes of a termite and a fungus, have been identified by exploiting their differential sensitivity towards organophosphate inhibitors. The inhibition curves obtained by plotting pI (negative logarithm of inhibitor concentration) versus per cent inhibition are used to distinguish the isozymes. The termite and its associated fungus possess 2 and 4 isozymes respectively. Each of the purified isozyme gave a single sigmoidal inhibition curve with its characteristic I50 value. The pattern of the inhibition curve of the crude extract is found to mimic that of the mixture of the purified isozymes in both termite and fungus. This method provides not only identification of isozymes but also serves as a criterion of homogeneity.

Effect of soman administration on beta-esterases in blood, liver microsomes and brain regions of rats.

Activities of enzymes cholinesterase (ChE) and carboxylesterase (CaE) were assayed in serum, liver microsomes and three regions of brain, viz; cerebrum, cerebellum and brain stem (with mid brain) in male albino rats at 0.5 and 2 h periods after administration of 1/2 LD 50 dose of soman (0.22 mg/kg) intraperitoneally in olive oil as vehicle. At 0.5 h, in serum, ChE activity declined to 33% of its initial level whereas CaE activity was almost completely inhibited. However, in the liver microsomes at this period, ChE activity was greatly inhibited (18% of initial level) whereas CaE activity was nearly unaffected. At 2 h period, both the enzymes in the serum were almost completely inhibited. In the brain regions (excepting in cerebellum), both the enzymes were nearly similarly inhibited (by 55% to 65% of the initial level) at both the periods. The time related differential response of these two beta-esterases in acute soman intoxication probably occurred in the peripheral tissues like blood and liver but not in the CNS.

Relación entre la actividad de esterasas y la resistencia a malatión en una cepa de Tribolium Castaneum / Relation activity from enterase and t.castaneum malathion resistant

Se estudiaron comparativamente las esterasas de las cepas de Tribolium castaneum ULP (suceptible) y ML (resistente a malatión). La actividad de esterasas que hidrolizan acetato de * y ß naftol presentes en homogenatos de T. castaneum fue mayor en la cepa susceptible ML, mientras que la actividad frente al acetato de tiofenilo fue similar en ambas cepas. La distribución subcelular de la actividad esterástica frente al PTA mostró diferencias en ambas cepas. En los homogenatos de insectos de la cepa susceptible se observó que a mayor velocidad de centrifugación el sobrenadante resultó considerablemente más activo en la hidrólisis de acetato de tiofenilo. Se supone que ese resultado se debe a la presencia de un inhibidor endógeno no dializable, de peso molecular tal que puede ser parcialmente precipitado a 10.000g y en mayor medida a 100.000g. Los zimogramas de esterasas no inhibibles por eserina separadas por electroforesis en gel de poliacrilamida y reveladas con * y ß naftilacetato, mostraron, en todos los casos, una única banda que fue clasificada como aril o acetil esterasa. Los resultados descriptos encuadran en la teoría de la aliesterasa mutada, postulada para casos de resistencia al malatión, descriptos en otas especies de insectos