Protective effect of nitroglycerin ointment and dimethyl sulfoxide on necrosis of skin flaps in rats.

OBJECTIVE: To compare the protective effect of nitroglycerin ointment 2% and Dimethylsulfoxide (DMSO) in dorsal flaps of the rat. METHODS: A blind, experimental study was conducted in 24 male Wistar rats, with a mean weight of 320 (286-376) grams. Group 1: Control. Petrolatum jelly (Vaseline), n = 8, Group 2: Nitroglycerin (NTG) ointment 2% (Nitro-Bid, Altana Co.) n = 8, and Group 3: DMSO gel 90% (Neogen corp. Lexington KY, 40611), n = 8. RESULTS: A total of 24 rats were operated on in the 6-month period of this study. Using a non-parametric Mann-Whitney U-test analysis, a statistically significant p was obtained between the control group and 2% NTG ointment, both in the area of necrosis and in the healthy area (p = 0.026). In contrast, the comparison between DMSO [CH3) 2SO] and the control group (p = 0.180) and between both study groups, with a p = 0.18, was not significant. CONCLUSIONS: Our study concluded that there is a protective effect of 2% NTG ointment for flap survival in relation to the control group (petrolatum). DMSO administered topically did not show a protective effect, compared to the control group.

OBJETIVO: Comparar el efecto protector del ungüento de nitroglicerina 2% y el dimetilsulfoxido 90% en colgajos dorsales en ratas. MÉTODOS: Se realizó un estudio experimental ciego en 24 ratas Wistar macho, con un peso medio de 320 gramos. Grupo 1: Control. Petrolato n = 8, Grupo 2: Nitroglicerina unguento al 2 % (Nitro-Bid, Altana Co.), n = 8, Grupo 3. Dimetilsulfóxido al 90% (Neogen corp. Lexington KY.), n = 8. RESULTADOS: Un total de 24 ratas fueron operadas en el período de 6 meses de este estudio. Mediante un análisis no paramétrico de la prueba U de Mann Whitney, se obtuvo una p estadísticamente significativa entre el grupo control y la pomada de nitroglicerina al 2%, tanto en el área de necrosis como en el área sana (p = 0.026). Por el contrario, la comparación entre DMSO y el grupo control (p = 0.180) y entre ambos grupos de estudio, con una p = 0.18, no fue significativa. CONCLUSIONES: Nuestro estudio concluyó que existe un efecto protector de la pomada de nitroglicerina al 2% para la supervivencia del colgajo en relación al grupo control (vaselina). El DMSO administrado por vía tópica no mostró un efecto protector, en comparación con el grupo de control.

Caracterización del polirribosilribitol fosfato por cromatografía de exclusión molecular de alta eficacia con detección ultravioleta / Characterization of polyribosylribitol phosphate by high efficiency size exclusion chromatography with ultraviolet detection

Haemophilus influenzae tipo b es un importante patógeno del hombre causante de varias de las enfermedades invasivas en niños menores de cinco años, contra el cual fueron autorizadas las vacunas glicoconjugadas a partir del polirribosilribitol fosfato. Quimi-Hib® es la primera y única vacuna contra este patógeno que utiliza el polisacárido obtenido por síntesis química. El Ingrediente Farmacéutico Activo es producido por el Centro de Ingeniería Genética y Biotecnología y se obtiene a partir de su conjugación al toxoide tetánico. En el presente reporte se hizo una caracterización del polirribosilribitol fosfato mediante la técnica de cromatografía de exclusión molecular de alta eficacia con detección ultravioleta a 215 nm. En el estudio se evaluaron tres lotes y se determinó el perfil de elución en una columna SuperdexTM 75 10/300 GL Increase con un porciento de pureza de 77,42 ± 8,97 y una masa molar promedio de 7.381 Da ± 210,93. La principal impureza presente en el polirribosilribitol fosfato es el dimetilsulfóxido, disolvente utilizado en la reacción de activación con el éster N-hidroxisuccinimidilo del ácido β-maleimidopropiónico. El polirribosilribitol fosfato se purificó por filtración con un Amicon Ultra-15 de 2.000 Da hasta una pureza de 99,1 por ciento y se conjugó al toxoide tetánico. El rendimiento de la reacción de conjugación con el polisacárido purificado fue de 30,0 por ciento 1,77 el cual no muestra diferencias significativas con el control que fue 33,7 por ciento ± 3,57 demostrándose que el dimetilsulfóxido no afecta el desempeño de la reacción de conjugación(AU)

Haemophilus influenzae type b is an important human pathogen causing some invasive diseases in children less than five years of age. Glycoconjugate vaccines based on polyribosylribitol phosphate have been licensed against this bacterium. Quimi-Hib® is the first and only vaccine against this pathogen using the chemically synthesized polysaccharide. The Active Pharmaceutical Ingredient is produced by the Center for Genetic Engineering and Biotechnology and is obtained from its conjugation to tetanus toxoid. In the present report a characterization of polyribosylribitol phosphate was performed by high performance molecular exclusion chromatography with ultraviolet detection at 215 nm. Three batches were evaluated in the study and the elution profile was determined on a SuperdexTM 75 10/300 GL Increase column with a purity percentage of 77.42 ± 8.97 and an average molecular weight of 7,381 Da ± 210.93. The main impurity present in polyribosylribitol phosphate was dimethylsulfoxide, the solvent used in the activation reaction with N-hydroxysuccinimidyl ester of β-maleimidopropionic acid. Polyribosylribitol phosphate was purified by filtration using a 2,000 Da cut-off Amicon Ultra-15 to a purity of 99.1 percent and conjugated to tetanus toxoid. The yield of the conjugation reaction with the purified polysaccharide was 30.0 percent ± 1.77 which shows no significant difference with the control which was 33.7 percent ± 3.57 demonstrating that dimethylsulfoxide does not affect the performance of the conjugation reaction(AU)

Effect of cryoprotectant on the motility, viability, fertilization, and DNA integrity of naleh fish Barbonymus sp. (Cyprinidae) sperm / Efeito do crioprotetor na motilidade, viabilidade, fertilização e integridade do DNA no sêmen de peixes naleh Barbonymus sp. (Cyprinidae)

Naleh fish Barbonymus sp. is a commercial freshwater fish, which is indigenous to Aceh, Indonesia. The population of this species has declined over the years as a result of habitat perturbations and overfishing. Hence, the crucial need to develop a cryopreservation method to support breeding programs. This involved the use of a cryoprotectant as an important component. The objective of this study, therefore, was to explore the best cryoprotectant for naleh fish spermatozoa, and a total of five types were tested. These include the DMSO, Methanol, Ethanol, Glycerol, and Ethylene Glycol at a similar concentration of 10%, which were individually combined with 15% egg yolk, and every treatment was performed in three replications. Conversely, Ringer's solution was adopted as an extender, and the sperm was cryopreserved in liquid nitrogen for 15 days. The results showed significant influence on sperm motility and viability, as well as egg fertility of naleh fish (P <0.05), although the DMSO provided the best outcome, compared to others at 47.17%, 50.13%, and 45.67%, respectively. Furthermore, DNA fragmentation had not occurred in the fresh and cryopreserved sperm samples, indicating the protective effect of tested cryoprotectants. It is concluded that the 10% DMSO and 15% egg yolk is the best cryoprotectant for naleh fish spermatozoa.(AU)

O peixe naleh Barbonymus sp. é um peixe comercial de água doce, originário de Aceh, Indonésia. Durante vários anos, as perturbações provocadas no seu habitat e a pesca predatória determinaram o declínio da sua população, cuja preservação deve apoiar-se em um programa de reprodução controlada, com o emprego de espermatozoides criopreservados. O presente trabalho realizou um estudo comparativo de cinco crioprotetores: dimetilsultóxido, metanol, etanol, glicerol e etileno glicol. Todos os crioprotetores foram testados na concentração de 10%, combinados a 15% de gema de ovo. Cada tratamento foi efetuado em triplicatas. A solução de ringer foi utilizada como extensor e o esperma foi criopreservado em nitrogênio líquido por 15 dias. Os resultados obtidos revelaram a existência de influência significante (P<0,05) na viabilidade e motilidade espermática bem como na fertilidade dos ovos do peixe naleh, em que o dimetilsulfóxido apresentou o melhor resultado com os valores de 47,17%, 50,13% e 45,67%, respectivamente. Por outro lado, a fragmentação do DNA não ocorreu nas amostras de esperma fresco e criopreservado, indicando o efeito protetor dos crioprotetores testados. A conclusão obtida foi que o dimetilsulfóxido e 15% de gema de ovo foram o melhor crioprotetor para os espermatozoides do peixe naleh.(AU)

Effect of cryoprotectant on the motility, viability, fertilization, and DNA integrity of naleh fish Barbonymus sp. (Cyprinidae) sperm / Efeito do crioprotetor na motilidade, viabilidade, fertilização e integridade do DNA no sêmen de peixes naleh Barbonymus sp. (Cyprinidae)

Naleh fish Barbonymus sp. is a commercial freshwater fish, which is indigenous to Aceh, Indonesia. The population of this species has declined over the years as a result of habitat perturbations and overfishing. Hence, the crucial need to develop a cryopreservation method to support breeding programs. This involved the use of a cryoprotectant as an important component. The objective of this study, therefore, was to explore the best cryoprotectant for naleh fish spermatozoa, and a total of five types were tested. These include the DMSO, Methanol, Ethanol, Glycerol, and Ethylene Glycol at a similar concentration of 10%, which were individually combined with 15% egg yolk, and every treatment was performed in three replications. Conversely, Ringer's solution was adopted as an extender, and the sperm was cryopreserved in liquid nitrogen for 15 days. The results showed significant influence on sperm motility and viability, as well as egg fertility of naleh fish (P <0.05), although the DMSO provided the best outcome, compared to others at 47.17%, 50.13%, and 45.67%, respectively. Furthermore, DNA fragmentation had not occurred in the fresh and cryopreserved sperm samples, indicating the protective effect of tested cryoprotectants. It is concluded that the 10% DMSO and 15% egg yolk is the best cryoprotectant for naleh fish spermatozoa.(AU)

O peixe naleh Barbonymus sp. é um peixe comercial de água doce, originário de Aceh, Indonésia. Durante vários anos, as perturbações provocadas no seu habitat e a pesca predatória determinaram o declínio da sua população, cuja preservação deve apoiar-se em um programa de reprodução controlada, com o emprego de espermatozoides criopreservados. O presente trabalho realizou um estudo comparativo de cinco crioprotetores: dimetilsultóxido, metanol, etanol, glicerol e etileno glicol. Todos os crioprotetores foram testados na concentração de 10%, combinados a 15% de gema de ovo. Cada tratamento foi efetuado em triplicatas. A solução de ringer foi utilizada como extensor e o esperma foi criopreservado em nitrogênio líquido por 15 dias. Os resultados obtidos revelaram a existência de influência significante (P<0,05) na viabilidade e motilidade espermática bem como na fertilidade dos ovos do peixe naleh, em que o dimetilsulfóxido apresentou o melhor resultado com os valores de 47,17%, 50,13% e 45,67%, respectivamente. Por outro lado, a fragmentação do DNA não ocorreu nas amostras de esperma fresco e criopreservado, indicando o efeito protetor dos crioprotetores testados. A conclusão obtida foi que o dimetilsulfóxido e 15% de gema de ovo foram o melhor crioprotetor para os espermatozoides do peixe naleh.(AU)

Avaliação da toxicidade de solvente de extratos vegetais com ação antiviral em sêmen caprino refrigerado / Evaluation of solvent toxicity of plant extract with antiviral action in refrigerated goat semen

Background: Caprine Arthritis Encephalitis Virus have been detected in sperm of breeding goats causing economic losses. In order to control the virus, researches aiming to identify natural extracts with potential antiviral effects are performed. However, aqueous or ethanolic extracts must be diluted in dimethyl sulfoxide (DMSO), which is a substance with unknown effects in sperm quality when present in diluting media. Therefore, this study aimed to evaluate sperm viability of refrigerated caprine semen diluted in media containing DMSO. This was performed to provide data that aid in researches involving the use of this component with natural extracts that may inactivate the caprine lentivirus in sperm.Materials, Methods & Results: The experiment was performed at the Laboratory of Seminal Technology in Embrapa Goats and Sheep in the city of Sobral, Brazil. Sperm viability was assessed in caprine semen refrigerated in two dilution media with crescent concentrations of DMSO. Sperm samples of five goats seronegative for the caprine lentivirus were pooled and diluted in minimal essential medium (MEM) enriched with glucose at 0.01 M added of crescent concentrations of DMSO (0%, 1.5%, 1.75%, 2.0%, 2.25% and 2.5%). The same breeders provided the pool of sperm to test Tris added 2.5% of egg yolk and the same concentrations of DMSO previously mentioned. Treatments were refrigerated at 7°C and evaluated up until four h after DMSO addition. Individual progressive motility (MIP), sperm vigor (V), percentage of spermatozoa reactive to hypoosmotic test (HO) and morphologically normal (NOR) were evaluated. IPM, vigor and NOR remained within normal standards for the caprine species in all treatments test. Percentage results of spermatozoa reactive to hypoosmotic was higher in Tris yolk with values ranging between 34.66% to 46.33%. Sperm vigor was positively correlated (r = 0.85) with IPM in the MEM diluted pool of sperm.[...]

Avaliação da toxicidade de solvente de extratos vegetais com ação antiviral em sêmen caprino refrigerado / Evaluation of solvent toxicity of plant extract with antiviral action in refrigerated goat semen

Background: Caprine Arthritis Encephalitis Virus have been detected in sperm of breeding goats causing economic losses. In order to control the virus, researches aiming to identify natural extracts with potential antiviral effects are performed. However, aqueous or ethanolic extracts must be diluted in dimethyl sulfoxide (DMSO), which is a substance with unknown effects in sperm quality when present in diluting media. Therefore, this study aimed to evaluate sperm viability of refrigerated caprine semen diluted in media containing DMSO. This was performed to provide data that aid in researches involving the use of this component with natural extracts that may inactivate the caprine lentivirus in sperm.Materials, Methods & Results: The experiment was performed at the Laboratory of Seminal Technology in Embrapa Goats and Sheep in the city of Sobral, Brazil. Sperm viability was assessed in caprine semen refrigerated in two dilution media with crescent concentrations of DMSO. Sperm samples of five goats seronegative for the caprine lentivirus were pooled and diluted in minimal essential medium (MEM) enriched with glucose at 0.01 M added of crescent concentrations of DMSO (0%, 1.5%, 1.75%, 2.0%, 2.25% and 2.5%). The same breeders provided the pool of sperm to test Tris added 2.5% of egg yolk and the same concentrations of DMSO previously mentioned. Treatments were refrigerated at 7°C and evaluated up until four h after DMSO addition. Individual progressive motility (MIP), sperm vigor (V), percentage of spermatozoa reactive to hypoosmotic test (HO) and morphologically normal (NOR) were evaluated. IPM, vigor and NOR remained within normal standards for the caprine species in all treatments test. Percentage results of spermatozoa reactive to hypoosmotic was higher in Tris yolk with values ranging between 34.66% to 46.33%. Sperm vigor was positively correlated (r = 0.85) with IPM in the MEM diluted pool of sperm.[...](AU)

Vitrification of bovine preantral follicles with dimethylsulfoxide and sucrose plus alfa-tocopherol / Vitrificação de folículos pré-antrais bovinos com dimetilxulfoxido e sacarose adicionado de alfa-tocopherol

The objective of this study was to evaluate the vitrification of bovine preantral follicles with dimethylsulfoxide (D) and sucrose (S) plus α-tocopherol 5mmol/L (T5) or 10mmol/L (T10) and, evaluate the thawed with minimal essential medium (m) with or without sucrose (s). Ovaries of cows were collected from slaughterhouse for the experiment I (n=66) and II (n=51). In the laboratory ovarian fragments were randomly assigned either to fresh control and 8 vitrification treatments (Controle and Dm; Dms, DSm; DSms; DST5m; DST5ms; DST10m; DST10ms). Ovarian fragments were placed in vitrification solution (5 min) and immersed in liquid nitrogen (-196°C), after a week, the fragments were thawed and analyzed. In the experiments I, preantral follicles were morphologically observed for histological evaluation, (normal; degenerated and developing of stage). In the experiment II, preantral follicles were mechanically isolated from ovarian tissue and examined with trypan blue, where dead and live corresponded to stained or non-stained. The treatments DSm, DSms and DST10m were effective in preserving the morphology in situ. However, the viability of isolated preantral follicles after vitrification remained high only in treatment DST10m. Thus, DST10m preserves survival rates and morphological integrity during vitrification of bovine preantral follicles.

Os objetivos deste estudo foram avaliar a vitrificação de folículos pré-antrais bovinos com dimetilsulfóxido (D) e sacarose (S) adicionando α-tocoferol 5mmol/L (T5) ou 10mmol/L (T10) e, avaliar o aquecimento com meio essencial mínimo (m) com ou sem sacarose (s). Ovários de fêmeas bovinas foram coletados de abatedouro, para o experimento I (n= 66) e II (n= 51). No laboratório fragmentos ovarianos foram distribuídos aleatoriamente para o controle fresco e 8 tratamentos de vitrificação (Controle e Dm; Dms, a DSm; DSms; DST5m; DST5ms; DST10m; DST10ms). Os fragmentos ovarianos foram colocados na solução de vitrificação (5 min) e imersos em nitrogênio líquido (-196°C). Após uma semana os fragmentos foram aquecidos e analisados. No experimento I, folículos pré-antrais foram observados morfologicamente para avaliação histológica (normal, degenerados e estádio de desenvolvimento). No experimento II, folículos pré-antrais foram mecanicamente isolados do tecido ovariano e examinados com o azul de trypan, observando mortos e vivos corados e não corados respetivamente. Os tratamentos a DSm, DSms e DST10m foram eficazes na preservação da morfologia in situ. No entanto, a viabilidade de folículos pré-antrais isolados após a vitrificação manteve-se elevada apenas no tratamento DST10m. Assim, DST10m preservou as taxas de sobrevivência e integridade morfológica durante a vitrificação de folículos pré-antrais bovinos.

Vitrification of bovine preantral follicles with dimethylsulfoxide and sucrose plus alfa-tocopherol / Vitrificação de folículos pré-antrais bovinos com dimetilxulfoxido e sacarose adicionado de alfa-tocopherol

The objective of this study was to evaluate the vitrification of bovine preantral follicles with dimethylsulfoxide (D) and sucrose (S) plus α-tocopherol 5mmol/L (T5) or 10mmol/L (T10) and, evaluate the thawed with minimal essential medium (m) with or without sucrose (s). Ovaries of cows were collected from slaughterhouse for the experiment I (n=66) and II (n=51). In the laboratory ovarian fragments were randomly assigned either to fresh control and 8 vitrification treatments (Controle and Dm; Dms, DSm; DSms; DST5m; DST5ms; DST10m; DST10ms). Ovarian fragments were placed in vitrification solution (5 min) and immersed in liquid nitrogen (-196°C), after a week, the fragments were thawed and analyzed. In the experiments I, preantral follicles were morphologically observed for histological evaluation, (normal; degenerated and developing of stage). In the experiment II, preantral follicles were mechanically isolated from ovarian tissue and examined with trypan blue, where dead and live corresponded to stained or non-stained. The treatments DSm, DSms and DST10m were effective in preserving the morphology in situ. However, the viability of isolated preantral follicles after vitrification remained high only in treatment DST10m. Thus, DST10m preserves survival rates and morphological integrity during vitrification of bovine preantral follicles.(AU)

Os objetivos deste estudo foram avaliar a vitrificação de folículos pré-antrais bovinos com dimetilsulfóxido (D) e sacarose (S) adicionando α-tocoferol 5mmol/L (T5) ou 10mmol/L (T10) e, avaliar o aquecimento com meio essencial mínimo (m) com ou sem sacarose (s). Ovários de fêmeas bovinas foram coletados de abatedouro, para o experimento I (n= 66) e II (n= 51). No laboratório fragmentos ovarianos foram distribuídos aleatoriamente para o controle fresco e 8 tratamentos de vitrificação (Controle e Dm; Dms, a DSm; DSms; DST5m; DST5ms; DST10m; DST10ms). Os fragmentos ovarianos foram colocados na solução de vitrificação (5 min) e imersos em nitrogênio líquido (-196°C). Após uma semana os fragmentos foram aquecidos e analisados. No experimento I, folículos pré-antrais foram observados morfologicamente para avaliação histológica (normal, degenerados e estádio de desenvolvimento). No experimento II, folículos pré-antrais foram mecanicamente isolados do tecido ovariano e examinados com o azul de trypan, observando mortos e vivos corados e não corados respetivamente. Os tratamentos a DSm, DSms e DST10m foram eficazes na preservação da morfologia in situ. No entanto, a viabilidade de folículos pré-antrais isolados após a vitrificação manteve-se elevada apenas no tratamento DST10m. Assim, DST10m preservou as taxas de sobrevivência e integridade morfológica durante a vitrificação de folículos pré-antrais bovinos.(AU)

Avaliação de diferentes crioprotetores e taxas de diluição na criopreservação seminal de Prochilodus brevis / Evaluation of different crioprotectants and dilution ratios on Prochilodus brevis sperm cryopreservation

Este estudo objetivou caracterizar o sêmen de Prochilodus brevis e avaliar os efeitos de diferentescrioproterores e taxas de diluição sobre a cinética e a morfologia do sêmen criopreservado. Inicialmente,amostras seminais de P. brevis (n = 40) foram analisadas quanto ao volume, pH, osmolaridade, concentração,motilidade e morfologia espermáticas. Para a criopreservação, o material coletado foi distribuído em 10 pools desêmen (n = 10), submetidos a seis tratamentos compostos pela combinação de glicose 5%, dois crioprotetores(DMSO ou MG) e três taxas de diluição (1:3, 1:6 ou 1:9 sêmen:diluidor). As amostras, in natura e pósdescongeladas,foram analisadas quanto à sua morfologia e cinética espermática utilizando o CASA. O sêmen deP. brevis apresentou características semelhantes a demais espécies de Prochilodus. O sêmen congelado comglicose e MG apresentou resultados significativamente superiores (P < 0,05) comparado àquele congeladoutilizando DMSO. As diluições 1:3 e 1:6 apresentaram melhores resultados para glicose + MG, enquanto 1:9 foimelhor para glicose + DMSO. Portanto, a associação glicose + MG, numa taxa de diluição de 1:3 ou 1:6, é amais indicada para a criopreservação do sêmen de Prochilodus brevis.(AU)

This study aimed to characterize Prochilodus brevis sperm and evaluate the effects of different diluentsand dilution ratios on kinetics and morphology of cryopreserved sperm. Initially, sperm samples of P. brevis(n = 40) were assessed for volume, pH, osmolality, spermatic concentration, motility and morphology. Forcryopreservation, the collected material was distributed in ten pools of semen (n = 10), submitted to sixtreatments composed by a combination of glucose 5% with two cryoprotectants (DMSO or MG) and threedilution ratios (1:3, 1:6 or 1:9 - sperm:diluent). In natura and post-thawed samples were assessed for spermaticmorphology and kinetics using CASA. The sperm of P. brevis showed similar characteristics to otherProchilodus species. Sperm cryopreserved in glucose plus MG presented significantly higher results (P < 0.05)compared to sperm frozen in DMSO. The dilution ratios of 1:3 and 1:6 yielded better results for glucose + MG,while 1:9 was the best dilution for glucose + DMSO. In conclusion, the association of glucose + MG, in adilution ratio of 1:3 or 1:6, is the most indicated for Prochilodus brevis sperm cryopreservation.(AU)

Avaliação de diferentes crioprotetores e taxas de diluição na criopreservação seminal de Prochilodus brevis / Evaluation of different crioprotectants and dilution ratios on Prochilodus brevis sperm cryopreservation

Este estudo objetivou caracterizar o sêmen de Prochilodus brevis e avaliar os efeitos de diferentescrioproterores e taxas de diluição sobre a cinética e a morfologia do sêmen criopreservado. Inicialmente,amostras seminais de P. brevis (n = 40) foram analisadas quanto ao volume, pH, osmolaridade, concentração,motilidade e morfologia espermáticas. Para a criopreservação, o material coletado foi distribuído em 10 pools desêmen (n = 10), submetidos a seis tratamentos compostos pela combinação de glicose 5%, dois crioprotetores(DMSO ou MG) e três taxas de diluição (1:3, 1:6 ou 1:9 – sêmen:diluidor). As amostras, in natura e pósdescongeladas,foram analisadas quanto à sua morfologia e cinética espermática utilizando o CASA. O sêmen deP. brevis apresentou características semelhantes a demais espécies de Prochilodus. O sêmen congelado comglicose e MG apresentou resultados significativamente superiores (P < 0,05) comparado àquele congeladoutilizando DMSO. As diluições 1:3 e 1:6 apresentaram melhores resultados para glicose + MG, enquanto 1:9 foimelhor para glicose + DMSO. Portanto, a associação glicose + MG, numa taxa de diluição de 1:3 ou 1:6, é amais indicada para a criopreservação do sêmen de Prochilodus brevis.

This study aimed to characterize Prochilodus brevis sperm and evaluate the effects of different diluentsand dilution ratios on kinetics and morphology of cryopreserved sperm. Initially, sperm samples of P. brevis(n = 40) were assessed for volume, pH, osmolality, spermatic concentration, motility and morphology. Forcryopreservation, the collected material was distributed in ten pools of semen (n = 10), submitted to sixtreatments composed by a combination of glucose 5% with two cryoprotectants (DMSO or MG) and threedilution ratios (1:3, 1:6 or 1:9 - sperm:diluent). In natura and post-thawed samples were assessed for spermaticmorphology and kinetics using CASA. The sperm of P. brevis showed similar characteristics to otherProchilodus species. Sperm cryopreserved in glucose plus MG presented significantly higher results (P < 0.05)compared to sperm frozen in DMSO. The dilution ratios of 1:3 and 1:6 yielded better results for glucose + MG,while 1:9 was the best dilution for glucose + DMSO. In conclusion, the association of glucose + MG, in adilution ratio of 1:3 or 1:6, is the most indicated for Prochilodus brevis sperm cryopreservation.

Histiocitoma de Células de Langerhans en canino: reporte de caso en Colombia / Langerhans Cell Histiocytoma in canine: a case report in Colombia / Histiocitoma das células de Langerhans em caninos: relato de caso na Colômbia

Resumen Se reporta el caso clínico de un paciente canino de 10 años de edad, macho, mestizo y esterilizado, que llega a la Clínica veterinaria de Antioquia, Medellín Colombia, con una masa de 5 cms de diámetro en la región lateral del prepucio. La masa de forma circular y de consistencia firme, a la evaluación histopatológica correspondió a un histiocitoma de las células de Langerhans. El paciente fue sometido a resección quirúrgica de la masa y tratado posteriormente con cefalexina 25 mg/kg cada 12 horas oral por 1 semana, prednisolona por 20 días y desinfección de la zona con clorhexidina cada 12 horas, además se aplica en la zona dimetilsulfóxido tópico cada 12 horas por 3 semanas. Debido a lo poco descrito este tipo de histiocitoma en dermatología, se considera de valor científico su reporte.

Abstract A 10–year-old crossbred neutered dog was submitted to Clínica Veterinaria de Antioquia (Medellín, Colombia) presenting a firm and circular mass of 5 cm in diameter in the lateral region of the foreskin. After histopathological evaluation, the mass corresponded to a Langerhans cell histiocytoma. The patient underwent surgical resection of the mass and was then treated with cephalexin (25 mg/kg every 12 hours, orally, for 1 week), prednisolone (20 days), disinfection of the area (every 12 hours using chlorhexidine), and applying dimethyl sulfoxide on the area (every 12 hours for 3 weeks). We consider this report has a scientific value because this type of histiocytoma is rarely described in dermatology.

Resumo Relatar o caso clínico de um paciente canino de 10 anos de idade, macho, mestiço e esterilizado, que chegou a Clínica Veterinária de Antioquia, Medellín, Colômbia, com uma massa de 5 cm de diâmetro na região lateral do prepúcio, a massa era de forma circular e de consistência firme, na avaliação histopatológica revelou-se um histiocitoma das células de Langerhans, o paciente foi submetido a resseção cirúrgica da massa e tratado posteriormente com cefalexina 25 mg/kg a cada 12 horas oral durante uma semana, prednisolona por 20 dias e desinfecção da região com clorexidina a cada 12 horas, além disto, se aplicou na região afetada dimetilsulfóxido tópico a cada 12 horas durante três semanas. Devido as poucas referencias escritas deste tipo de histiocitoma em dermatologia, considera-se de valor cientifico seu relato.

Cryopreservation and sperm storage of the white shrimp / Criopreservação e estocagem do sêmen do camarão branco

This study was performed to evaluate a protocol for sperm cryopreservation of the marine shrimp Litopenaeus schmitti, an important species in Brazilian commercial fisheries. No studies or protocols were found for cryopreservation of its spermatic mass. This paper provides information about the technique of semen storage based on protocols applied to other penaeids species. Two cryoprotectants, glycerol and dimetilsulfoxide (DMSO), were tested for sperm cells toxicity in two concentrations (5 and 10%), and two exposure times (10 and 30 minutes). Influence of liquid nitrogen storage in apparent sperm viability (ASV) was also tested in 1, 15 and 30 days of storage. The cryopreservation was performed in a two-step (2 and 0.5ºC min-1) freezing protocol. After freezing until the temperature -32C, sperm mass was immersed in liquid nitrogen (-196ºC). Thus, glycerol and DMSO was not toxic to sperm in both concentrations and equilibration times. For all toxicity tests ASV was up to 79.8% after exposure. Liquid nitrogen storage tests indicated no significant difference between glycerol 5 and 10%. Results were significantly different over timebetween days 15 and 30 of storage in liquid nitrogen, in which ASV was around 42.8% and 16.3%respectively. Thus, it is suggested glycerol 5% (10 minutes) as cryoprotectant for L. schmittispermatic mass cryopreservation. It is also suggested that the L. schmitti spermatic mass can be stored during 15 days in liquid nitrogen, using a two-step (2 and 0.5ºC min-1) freezing protocol.(AU)

Este estudo foi realizado ara avaliar um protocolo de criopreservação do sêmen do camarão marinho Litopenaeus schmitti, uma espécie importante para a pesca comercial no Brasil. Não foram encontrados estudos ou protocolos de criopreservação de sua massa espermática. Este estudo fornece informações sobre a técnica de armazenamento de sêmen com base em protocolos aplicados a outras espécies de peneídeos. Dois agentes crioprotectores, glicerol e dimetilsulfóxido (DMSO), foram testados quanto à toxicidade para as células espermáticas em duas concentrações (5 e 10%) e dois tempos de exposição (10 e 30 minutos). A influência da manutenção em nitrogênio líquido sobre a viabilidade espermática aparente (VEA) foi também testada em 1, 15 e 30 dias de armazenamento. Para criopreservação, utilizou-se um protocolo de congelamento em duas etapas (2 e 0,5ºC min-1). Após o resfriamento até a temperatura de -32°C, a massa espermática foi imersaem nitrogênio líquido (-196°C). Assim, glicerol e DMSO não foram tóxicos para o esperma em ambas as concentrações e tempos de equilíbrio. Para todos os testes de toxicidade a VEA foi de até79,8% após a exposição. Os testes de armazenamento em nitrogênio líquido não indicaram diferença significativa utilizando glicerol 5 e 10%, mas houve diferença significativa entre os dias 15(42,8%) e 30 (16,3%), para a VEA. Assim, sugere-se a utilização do glicerol 5 ou 10% (10 minutos) como crioprotetor para a criopreservação de massa espermática do camarão L. schmitti. Também é sugerido que a massa espermática do L. schmitti pode ser armazenada durante 15 dias emnitrogênio líquido, utilizando-se um protocolo de congelamento em duas etapas (2 e 0,5ºC min-1).(AU)

Criopreservação do espermatóforo e da massa espermática do camarão branco Litopenaeus schmitti post-mortem / Post-mortem spermatophore and sperm cryopreservation of the white shrimp Litopenaeus schmitti

This study was carried out in two phases; the first to evaluate the viability of two cooling protocols (A and B) using glycerol (10%) as a cryoprotectant and the second, the efficiency of two cryoprotectants (10% glycerol and DMSO) and times (30, 60 and 90 days of storage in liquid nitrogen) for the cryopreservation of post-mortem sperm mass and spermatophores of the shrimp Litopenaeus schmitti. The protocol A presented a cooling rate of 0.5C min-1 until reaching -32ºC and protocol B, 20ºC min-1 until reach -120ºC, after which the samples were transferred to liquid nitrogen (N2L). The sperm viability was assessed by smearing the semen stained with eosin-nigrosin. The curve resulted in higher mean sperm survival was Protocol A (50.9%). Therefore, for the second experiment, the protocol A was used, however there was no difference between the cryoprotectants for sperm mass, although the influence on survival time. Difference was observed between the cryoprotectants and the influence of time on sperm survival of cryopreserved spermatophores. Glycerol 10% was more efficient for cryopreservation of spermatophore, but for the masses sperm, both can be used.

O presente trabalho foi realizado em duas etapas; a primeira para avaliar a eficiência de dois protocolos de resfriamento (A e B) utilizando glicerol (10%) como crioprotetor e, a segunda, a eficiência de dois crioprotetores (glicerol e dimetilsulfóxido - DMSO a 10%) e o tempo (30, 60 e 90 dias de estocagem em nitrogênio líquido) para a criopreservação da massa espermática e espermatóforo de camarões post mortem da espécie Litopenaeus schmitti. O protocolo A apresentou velocidade de resfriamento de 0,5ºC min-1 até alcançar -32ºC e o protocolo B, 20ºC min-1 até alcançar -120ºC, sendo os materiais posteriormente transferidos para o nitrogênio líquido (N2L). A viabilidade espermática foi avaliada por meio do esfregaço de sêmen corado com eosina-nigrosina. A curva que resultou na maior média de sobrevivência espermática foi a do protocolo A (50,9%). Portanto, para o segundo experimento foi utilizado o protocolo A; entretanto, neste não houve diferença entre os crioprotetores para a massa espermática, contudo, o tempo influenciou na sobrevivência. Foi observada diferença entre os crioprotetores e a influência do tempo na sobrevivência espermática dos espermatóforos criopreservados. O glicerol a 10% foi mais eficiente para a criopreservação dos espermatóforos, porém, para as massas espermáticas, ambos podem ser utilizados.

Cryopreservation and sperm storage of the white shrimp / Criopreservação e estocagem do sêmen do camarão branco

This study was performed to evaluate a protocol for sperm cryopreservation of the marine shrimp Litopenaeus schmitti, an important species in Brazilian commercial fisheries. No studies or protocols were found for cryopreservation of its spermatic mass. This paper provides information about the technique of semen storage based on protocols applied to other penaeids species. Two cryoprotectants, glycerol and dimetilsulfoxide (DMSO), were tested for sperm cells toxicity in two concentrations (5 and 10%), and two exposure times (10 and 30 minutes). Influence of liquid nitrogen storage in apparent sperm viability (ASV) was also tested in 1, 15 and 30 days of storage. The cryopreservation was performed in a two-step (2 and 0.5ºC min-1) freezing protocol. After freezing until the temperature -32C, sperm mass was immersed in liquid nitrogen (-196ºC). Thus, glycerol and DMSO was not toxic to sperm in both concentrations and equilibration times. For all toxicity tests ASV was up to 79.8% after exposure. Liquid nitrogen storage tests indicated no significant difference between glycerol 5 and 10%. Results were significantly different over timebetween days 15 and 30 of storage in liquid nitrogen, in which ASV was around 42.8% and 16.3%respectively. Thus, it is suggested glycerol 5% (10 minutes) as cryoprotectant for L. schmittispermatic mass cryopreservation. It is also suggested that the L. schmitti spermatic mass can be stored during 15 days in liquid nitrogen, using a two-step (2 and 0.5ºC min-1) freezing protocol.

Este estudo foi realizado ara avaliar um protocolo de criopreservação do sêmen do camarão marinho Litopenaeus schmitti, uma espécie importante para a pesca comercial no Brasil. Não foram encontrados estudos ou protocolos de criopreservação de sua massa espermática. Este estudo fornece informações sobre a técnica de armazenamento de sêmen com base em protocolos aplicados a outras espécies de peneídeos. Dois agentes crioprotectores, glicerol e dimetilsulfóxido (DMSO), foram testados quanto à toxicidade para as células espermáticas em duas concentrações (5 e 10%) e dois tempos de exposição (10 e 30 minutos). A influência da manutenção em nitrogênio líquido sobre a viabilidade espermática aparente (VEA) foi também testada em 1, 15 e 30 dias de armazenamento. Para criopreservação, utilizou-se um protocolo de congelamento em duas etapas (2 e 0,5ºC min-1). Após o resfriamento até a temperatura de -32°C, a massa espermática foi imersaem nitrogênio líquido (-196°C). Assim, glicerol e DMSO não foram tóxicos para o esperma em ambas as concentrações e tempos de equilíbrio. Para todos os testes de toxicidade a VEA foi de até79,8% após a exposição. Os testes de armazenamento em nitrogênio líquido não indicaram diferença significativa utilizando glicerol 5 e 10%, mas houve diferença significativa entre os dias 15(42,8%) e 30 (16,3%), para a VEA. Assim, sugere-se a utilização do glicerol 5 ou 10% (10 minutos) como crioprotetor para a criopreservação de massa espermática do camarão L. schmitti. Também é sugerido que a massa espermática do L. schmitti pode ser armazenada durante 15 dias emnitrogênio líquido, utilizando-se um protocolo de congelamento em duas etapas (2 e 0,5ºC min-1).

Criopreservação do espermatóforo e da massa espermática do camarão branco Litopenaeus schmitti post-mortem / Post-mortem spermatophore and sperm cryopreservation of the white shrimp Litopenaeus schmitti

This study was carried out in two phases; the first to evaluate the viability of two cooling protocols (A and B) using glycerol (10%) as a cryoprotectant and the second, the efficiency of two cryoprotectants (10% glycerol and DMSO) and times (30, 60 and 90 days of storage in liquid nitrogen) for the cryopreservation of post-mortem sperm mass and spermatophores of the shrimp Litopenaeus schmitti. The protocol A presented a cooling rate of 0.5C min-1 until reaching -32ºC and protocol B, 20ºC min-1 until reach -120ºC, after which the samples were transferred to liquid nitrogen (N2L). The sperm viability was assessed by smearing the semen stained with eosin-nigrosin. The curve resulted in higher mean sperm survival was Protocol A (50.9%). Therefore, for the second experiment, the protocol A was used, however there was no difference between the cryoprotectants for sperm mass, although the influence on survival time. Difference was observed between the cryoprotectants and the influence of time on sperm survival of cryopreserved spermatophores. Glycerol 10% was more efficient for cryopreservation of spermatophore, but for the masses sperm, both can be used.(AU)

O presente trabalho foi realizado em duas etapas; a primeira para avaliar a eficiência de dois protocolos de resfriamento (A e B) utilizando glicerol (10%) como crioprotetor e, a segunda, a eficiência de dois crioprotetores (glicerol e dimetilsulfóxido - DMSO a 10%) e o tempo (30, 60 e 90 dias de estocagem em nitrogênio líquido) para a criopreservação da massa espermática e espermatóforo de camarões post mortem da espécie Litopenaeus schmitti. O protocolo A apresentou velocidade de resfriamento de 0,5ºC min-1 até alcançar -32ºC e o protocolo B, 20ºC min-1 até alcançar -120ºC, sendo os materiais posteriormente transferidos para o nitrogênio líquido (N2L). A viabilidade espermática foi avaliada por meio do esfregaço de sêmen corado com eosina-nigrosina. A curva que resultou na maior média de sobrevivência espermática foi a do protocolo A (50,9%). Portanto, para o segundo experimento foi utilizado o protocolo A; entretanto, neste não houve diferença entre os crioprotetores para a massa espermática, contudo, o tempo influenciou na sobrevivência. Foi observada diferença entre os crioprotetores e a influência do tempo na sobrevivência espermática dos espermatóforos criopreservados. O glicerol a 10% foi mais eficiente para a criopreservação dos espermatóforos, porém, para as massas espermáticas, ambos podem ser utilizados.(AU)

Lipoidoproteinosis: relato de un caso / Lipoidoproteinosis: case report

La lipoidoproteinosis, es una enfermedad hereditaria autosómica recesiva, de presentación infrecuente, con manifestaciones clínicas características desde etapas tempranas de la vida y hallazgos histopatológicosdados por un depósito de lípidos y proteínas anormales de tipo hialino en diferentes tejidos.Presentamos el caso de una mujer de 18 años, valorada en nuestra consulta por la aparición de llanto y habla disfónica, asociados a lesiones cutáneas y mucosas, con diagnóstico clínico e histopatológico de lipoidoproteinosis, con buena respuesta terapéutica al dimetil-sulfóxido.Resaltamos que se trata de un caso de infrecuente presentación, cuyo enfoque temprano y multidisciplinario son esenciales para su pronóstico...

Lipoidoproteinosis is a rare autosomal recessive hereditary disease, with typical clinical features since first years of life and histopathological findings of deposits of lipids and abnormal hyaline-like proteins in different tissues. We report the case of an 18 year-old woman, examined at our clinic for the presence of dysphonic weeping and speech, associated with cutaneous and mucosal lesions with clinical and histopathological diagnosis of lipoidoproteinosis, that presented a good therapeutic response to dimethyl sulfoxide. We emphasize that this is a case of rare presentation, whose early and multidisciplinary approach is essential for prognosis...

Topical treatment using amphotericin b and dmso for an atypically located equine cutaneous Pythiosis / Pitiose cutânea equina de localização atípica tratada topicamente com solução de anfotericina B e DMSO

Background: Cutaneous lesions by Pythium insidiosum infection are commonly observed in horses, especially in those living at fl ooded environments. Equine pythiosis is characterized by the development of tumoral masses that are frequently located at distal limbs, ventral abdômen, thorax, breast and face. The lesions are usually granulomatous, serosanguineous and ulcerated, most often destroyed by self-mutilation due to the intense pruritus. The proposed treatment includes surgical excision followed by antifungal drugs administration, which can be done systemically or topically. Amphotericin B and dimethyl sulfoxide (DMSO) in association has been successfully used for cutaneous pythiosis topical treatment due to the DMSO property to carry any substance through plasmatic membranes.Case: The present report concerns a 12-year-old mixed breed gelding presenting with self-mutilation of a tumoral mass located at the left fl ank. The owners reported that the horse had initially presented a small wound that had evolved to a 20-cm in diameter mass in 4 weeks. Tissue samples were collected, processed and stained by the Gomoris methenamine silver (GMS) method. The histopathological analysis revealed Pythium insidiosum hyphae in a granulomatous tissue, especially located at peripheral region, where kunkers were present. Surgical excision of the mass followed by cauterization was indicated

Background: Cutaneous lesions by Pythium insidiosum infection are commonly observed in horses, especially in those living at fl ooded environments. Equine pythiosis is characterized by the development of tumoral masses that are frequently located at distal limbs, ventral abdômen, thorax, breast and face. The lesions are usually granulomatous, serosanguineous and ulcerated, most often destroyed by self-mutilation due to the intense pruritus. The proposed treatment includes surgical excision followed by antifungal drugs administration, which can be done systemically or topically. Amphotericin B and dimethyl sulfoxide (DMSO) in association has been successfully used for cutaneous pythiosis topical treatment due to the DMSO property to carry any substance through plasmatic membranes.Case: The present report concerns a 12-year-old mixed breed gelding presenting with self-mutilation of a tumoral mass located at the left fl ank. The owners reported that the horse had initially presented a small wound that had evolved to a 20-cm in diameter mass in 4 weeks. Tissue samples were collected, processed and stained by the Gomoris methenamine silver (GMS) method. The histopathological analysis revealed Pythium insidiosum hyphae in a granulomatous tissue, especially located at peripheral region, where kunkers were present. Surgical excision of the mass followed by cauterization was indicated

Pitiose cutânea equina de localização atípica tratada topicamente com solução de anfotericina B e DMSO / Topical treatment using amphotericin B and DMSO for an atypically located equine cutaneous pythiosis

Background: Cutaneous lesions by Pythium insidiosum infection are commonly observed in horses, especially in those living at flooded environments. Equine pythiosis is characterized by the development of tumoral masses that are frequently located at distal limbs, ventral abdômen, thorax, breast and face. The lesions are usually granulomatous, serosanguineous and ulcerated, most often destroyed by self-mutilation due to the intense pruritus. The proposed treatment includes surgical excision followed by antifungal drugs administration, which can be done systemically or topically. Amphotericin B and dimethyl sulfoxide (DMSO) in association has been successfully used for cutaneous pythiosis topical treatment due to the DMSO property to carry any substance through plasmatic membranes. Case: The present report concerns a 12-year-old mixed breed gelding presenting with self-mutilation of a tumoral mass located at the left flank. The owners reported that the horse had initially presented a small wound that had evolved to a 20-cm in diameter mass in 4 weeks. Tissue samples were collected, processed and stained by the Gomori's methenamine silver (GMS) method. The histopathological analysis revealed Pythium insidiosum hyphae in a granulomatous tissue, especially located at peripheral region, where kunkers were present. Surgical excision of the mass followed by cauterization was indicated as initial treatment, and due to financial reasons, the owners elected only the topical antifungal therapy to control the fungus infection after surgery. Flunixin meglumine was also administrated for five days aiming the control of pain and inflammation. The wound was cleaned with povidone-iodine solution and rinsed with a solution containing 50 mg of amphotericin B in 10 mL of sterile water and 10 mL of DMSO. This procedure was carried out twice a day. The wound healed fast due to an excellent centripetal epithelialization, and the horse was discharged after 64 days showing only 5% of the initial wound area. The owner reported by telephone the complete healing and hair growth 10 days after discharge. Discussion: Despite the atypical location of the tumoral lesion described at the present report, the history and clinical manifestations, especially the intense pruritus, showed similarity with other characteristic reports of equine cutaneous pythiosis. The diagnosis was confirmed by the histopathological examination showing hyphae structures, as described to be evidences of the presence of Pythium insidiosum in the tissue. The surgical procedure was the first step to provide remission of clinical signs, and one day after surgery the pruritus desapeared. After excision of the granulomatous tissue and cauterization, daily topical administration of amphotericin B associated with DMSO was effective in destroying the infectious agent, as observed by the excellent epithelization. A pink granulation tissue grew up providing an ideal surface for epithelial migration and the healing process progressed quickly. Centripetal epithelialization reduced the wound area until 3% of the initial area in 64 days of treatment, when the remaining wound was found almost completely healed and covered with hair. At the present report, the horse presenting pythiosis was only topically treated. The recommended therapy using amphotericin B and DMSO solution was effective, economically viable and low risk, considering that the systemic antifungal therapy usually suggested is expensive and extremely nephrotoxic. The atypical location of the lesion on the left flank shows that any anatomical region can be affected by the fungus, since the conditions for its development were present.

Topical treatment using amphotericin b and dmso for an atypically located equine cutaneous Pythiosis / Pitiose cutânea equina de localização atípica tratada topicamente com solução de anfotericina B e DMSO

Background: Cutaneous lesions by Pythium insidiosum infection are commonly observed in horses, especially in those living at fl ooded environments. Equine pythiosis is characterized by the development of tumoral masses that are frequently located at distal limbs, ventral abdômen, thorax, breast and face. The lesions are usually granulomatous, serosanguineous and ulcerated, most often destroyed by self-mutilation due to the intense pruritus. The proposed treatment includes surgical excision followed by antifungal drugs administration, which can be done systemically or topically. Amphotericin B and dimethyl sulfoxide (DMSO) in association has been successfully used for cutaneous pythiosis topical treatment due to the DMSO property to carry any substance through plasmatic membranes.Case: The present report concerns a 12-year-old mixed breed gelding presenting with self-mutilation of a tumoral mass located at the left fl ank. The owners reported that the horse had initially presented a small wound that had evolved to a 20-cm in diameter mass in 4 weeks. Tissue samples were collected, processed and stained by the Gomoris methenamine silver (GMS) method. The histopathological analysis revealed Pythium insidiosum hyphae in a granulomatous tissue, especially located at peripheral region, where kunkers were present. Surgical excision of the mass followed by cauterization was indicated

Background: Cutaneous lesions by Pythium insidiosum infection are commonly observed in horses, especially in those living at fl ooded environments. Equine pythiosis is characterized by the development of tumoral masses that are frequently located at distal limbs, ventral abdômen, thorax, breast and face. The lesions are usually granulomatous, serosanguineous and ulcerated, most often destroyed by self-mutilation due to the intense pruritus. The proposed treatment includes surgical excision followed by antifungal drugs administration, which can be done systemically or topically. Amphotericin B and dimethyl sulfoxide (DMSO) in association has been successfully used for cutaneous pythiosis topical treatment due to the DMSO property to carry any substance through plasmatic membranes.Case: The present report concerns a 12-year-old mixed breed gelding presenting with self-mutilation of a tumoral mass located at the left fl ank. The owners reported that the horse had initially presented a small wound that had evolved to a 20-cm in diameter mass in 4 weeks. Tissue samples were collected, processed and stained by the Gomoris methenamine silver (GMS) method. The histopathological analysis revealed Pythium insidiosum hyphae in a granulomatous tissue, especially located at peripheral region, where kunkers were present. Surgical excision of the mass followed by cauterization was indicated

Agentes crioprotetores intracelulares: características e utilização na criopreservação de tecido ovariano e oócitos / Intracellular Cryoprotectant Agents: Characteristics and Use of Ovarian Tissue and Oocyte Cryopreservation

Background: The application of cryopreservation in human and animal reproductive medicine has stimulated several studies about the effects of low temperatures and freezing processes on cells and tissues, in order to develop efficient protocols for gamete and embryo preservation. Moreover, cryopreservation is a fundamental tool for the establishment of animal germplasm banks, allowing the preservation of genetic material from several species and breeds or for further study and/or recovery of desirable characteristics. For the success of cryopreservation, the addition of an intracellular cryoprotectant agent in the freezing solution is indispensable. However, issues related to intracellular cryoprotectant agents used, e.g., their metabolism and potential toxicity, must be examined carefully so we can choose the cryoprotectant most suitable for a specific structure. Review: In this regard, this review introduces several aspects of cryopreservation, such as basic principles and methods used (slow freezing and vitrification), describing the fundamental steps of cryoprotectant agent’s exposure, cooling, storage, thawing or warming and removal of the cryoprotectant agent. The addition of an intracellular cryoprotectant to the freezing solution is essential, but does not guarantee the success of the cryopreservation protocol, due to its toxic effect, which requires a perfect balance between cryoprotectant concentration, temperature and exposure time to the structure which will be cryopreserved. Some studies attribute the toxicity of these agents mainly to the secondary metabolites formed when the cell resumes its activi ty and gradually begins to metabolize the cryoprotectant agent.[...]