Estres oxidativ y su efecto sobre calidad seminal / The oxidative stress effect on semen quality

La membrana espermática tiene ácidos grasos insaturados que la tornan vulnerable al ataque de sustancias oxígeno reactivas. Los espermatozoides poseen sistemas protectores, pero un desbalance entre pro y antioxidantes produce “estrés oxidativo". Objetivo: estudiar en semen de hombres infértiles, el efecto del estrés oxidativo sobre la membrana y núcleo espermático. Se analizaron muestras seminales de 142 hombres infértiles. Se efectuó espermograma, se seleccionaron 83 muestras sin aglutinación ni hiperviscosidad y con concentración espermática mayor a 5 x 10 6 /ml. Se estudió la membrana espermática con Test Hipoosmótico, la condensación cromatínica con Azul de Anilina y el ADN con Naranja de Acridina. Para estrés oxidativo se aplicó el Test MOST (movilidad traslativa final/movilidad traslativa inicial) que evalúa la pérdida de movilidad de los espermatozoides luego de ser incubados por 4 hs. en baño de agua a 40 °C. Se agruparon las muestras en G1: MOST mayor o igual a 0.40 (normal) y G2: MOST menor a 0.40 (anormal). El análisis estadístico demostró diferencia significativa (p<0.003) en las 3 pruebas funcionales. El estrés oxidativo altera estructuras espermáticas esenciales.

The spermatozoids have in its plasmatic membrane high concentrations of unsaturated fatty acids, vulnerable to the attack of the reactive oxygen substances. The male gamete has protective systems, the distortion between pro and antioxidizers produces “oxidative stress". Objective: to study samples of semen from infertile men, the effect of oxidative stress on the sperm membrane and the nucleus. 142 semen samples were analyzed from infertile men. The sperm study was evaluated according to OMS and 83 samples without agglutination and hiperviscosity with concentration of spermatozoids more of 5 x 10 6 /ml were chosen. The sperm membrane was studied with the Hipoosmotic Test, the maturity of chromatina with blue aniline and the nuclear AND with acridine orange. The EO was evaluated with the MOST test (motility end/motility initial) measuring the loss of motility of the spermatozoids incubated at 40ºC in a water bath for 4 hours, separating the samples in : G1 : MOST major or equal to 0.40 (normal) G2 : MOST under to 0.40 ( abnormal). The statistic analysis of the three tests showed significant difference (p<0.003). The oxidative stress affects essential structures.

[Effect of tobacco consumption on the spermatogenesis in males with idiopathic infertility]. / Efecto del tabaquismo sobre la espermatogenesis en hombres con infertilidad idiopatica.

OBJECTIVES: Our objective was to relate the tobacco with seminal spermatogenic parameters such as sperm morphology, concentration of spermatozoids and germinal cells in samples of semen from men with idiopatic infertility. METHODS: A prospective study was carried out on a population of 131 men with idiopatic infertility that attended the Reproduction Service of Centenario Hospital in Rosario from may 2004 to june 2006. Sperm study according to WHO was carried out evaluating germinal cells and sperm morphology with Papannicolaou. The concentration of spermatozoids was determined by means of a subjective method with Neubauer camera. The studied population was divided in the three groups: G1: smokers more of 20 cigarettes/day, G2: smokers under 20 cigarettes/day, G3 non smokers. The smokers had had the habit for over a year. RESULTS: Results were analyzed with the student's t-test. Statistically significant differences between G1 vs G3 (p<0.001) and G2 vs G3 (p<0.005) were found for the three variables. No significant difference was found between the groups of smokers G1 vs G2 (p>0.1). CONCLUSIONS: The results show that tobacco alters sperm concentration and morphology with an increase of immature forms, demonstrating an altered spermatogenesis process. The consumption of tobacco should be evaluated to carry out the integral study of infertile man.

Efecto del tabaquismo sobre la espermatogenesis en hombres con infertilidad idiopatica / Effect of tobacco consumption on the spermatogenesis in males with idiopathic infertility

Objetivo: Relacionar en hombres con infertilidad idiopática, el tabaquismo con parámetros seminales espermatogénicos, tales como morfología espermática, concentración de espermatozoides y células germinales en muestras de semen fresco. Métodos: Se realizó un estudio prospectivo en 131 hombres con infertilidad idiopática que asistieron al Servicio de Reproducción del Hospital Centenario de Rosario desde mayo de 2004 a junio de 2006. Se realizó espermograma según normas OMS evaluando células germinales y morfología espermática con tinción Papanicolaou. La concentración de espermatozoides se determinó por el método subjetivo con cámara de Neubauer. La población en estudio se dividió en 3 grupos G1 fumadores de más de 20 cig/día, G2 fumadores de menos de 20 cig/día y G3 hombres no fumadores. Los fumadores lo hacían al menos desde hace un año. Resultados: Los resultados obtenidos se analizaron mediante la prueba t Student, encontrándose diferencia estadísticamente significativa para las 3 variables entre G1 vs G3 (p<0.001) y G2 vs G3 (p<0.005). .No se encontró diferencia significativa entre los grupos de fumadores G1vsG2 (p>0.1). Conclusiones: El tabaco altera la concentración y morfología espermática con aumento de formas espermáticas inmaduras que manifiestan un proceso espermatogénico alterado. El consumo de tabaco debe ser evaluado al realizar un estudio integral del hombre infértil (AU)

Objectives: Our objetive was to relate the tobacco with seminal spermatogenic parameters such as sperm morphology, concentration of spermatozoids and germinal cells in samples of semen from men with idiopatic infertility. Methods: A prospective study was carried out on a population of 131 men with idiopatic infertility that attended the Reproduction Service of Centenario Hospital in Rosario from may 2004 to june 2006. Sperm study according to WHO was carried out evaluating germinal cells and sperm morphology with Papannicolaou. The concentration of spermatozoids was determined by means of a subjective method with Neubauer camera. The studied population was divided in the three groups: G1: smokers more of 20 cigarettes/day, G2: smokers under 20 cigarettes/day, G3 non smokers. The smokers had had the habit for over a year. Results: Results were analyzed with the student’s t-test. Statistically significant differences between G1 vs G3 (p<0.001) and G2 vs G3 (p<0.005) were found for the three variables. No significant difference was found between the groups of smokers G1 vs G2 (p>0.1). Conclusions: The results show that tobacco alters sperm concentration and morphology with an increase of immature forms, demonstrating an altered spermatogenesis process. The consumption of tobacco should be evaluated to carry out the integral study of infertile man (AU)

Capacidad fecundante espermática luego de la recuperación de espermatozoides mediante la técnica de Swim-up / Spermatic fertilization capacity after swin-up spermatozoa recovery technique

OBJETIVO: Evaluar en pacientes infértiles, la calidad espermática pre y post swim up y compararlacon una población de hombres fértiles.MÉTODOS: De 55 pacientes que asistieron al Servicio de Reproducción del Hospital Centenario de Rosario a los que se efectuó espermograma y estudios funcionales espermáticos según normas OMS (1999), se seleccionaron30 muestras de semen con volumen mayor de 1.0 ml, concentración de espermatozoides mayor de 5.000.000/ml y que no presentaban hiperviscosidad. Se compararon los resultados obtenidos de: movilidad progresiva (MP), morfología (M), condensación de la cromatina (CC) e integridad de la cromatina (IC), en el semen fresco sin tratar (G2) y luego del swim-up (G3), con los valores obtenidos en muestras de semen frescode 15 hombres fértiles (G1). Se utilizó microscopio óptico para estudiar la movilidad espermática progresiva,tinción Hematoxilina-Verde Brillante para analizar la morfología, azul de anilina para evaluar la condensaciónde la cromatina y el fluorocromo naranja de acridina para analizar la integridad de la cromatina. La técnica de recuperación espermática utilizada fue el swim-up, que se realizó en medio HTF, incubando en tubo Falcon a 45 ° en estufa de 37 °C gaseada (atmósfera de CO2 5%) durante 1 hora. Luego se tomaronde la capa superior los espermatozoides que sobrenadaron,en los que se efectuaron las prácticas post swim-up. Se aplicó la prueba t - Student a los valores de los parámetros MP, M, CC e IC en los 3 grupos de pacientes, observándose diferencia significativa para los cuatro parámetros al comparar G1 vs. G2 y G2 vs. G3 (p0.1), lo cual indica que los parámetros estudiados MP, M, CC e IC, en los espermatozoidesrecuperados del swim-up, son semejantes a la población de hombres fértiles.RESULTADOS: Estos resultados muestran que mediante swim-up se pueden recuperar gametas con capacidad fecundante potencial semejante a la población fértil, para aplicar en técnicas de reproducción asistida de baja complejidad, como la inseminación intrauterina, donde la selección natural es aún viable

OBJECTIVES: To investigate sperm quality before and after swim up in infertile patients, and to compare it with a fertile men population. METHODS: Semen samples from 55 patients consulting at the infertility services of the Hospitals “Centenario” in Rosario and “Eva Perón” in Gro Baigorria were collected and analyzed accordingly with the WHO guidelines. 30 sperm samples with a volume higher than 1.0 ml, and spermatozoid concentration higher than 5.000.000/ml, not presenting hyperviscosity were selected. Outcome variables including progressive mobility (PM), morphology (M), chromatin condensation (CC) and chromatin integrity (CI), were compared in fresh semen samples, between patients without previous treatment (G2) and after swim up (G3) and 15 fertile men (G1). Sperm morphology was evaluated by brilliant green hematoxyllin stain ;progressive mobility with a subjective method accordingly to WHO (1999); chromatin condensation with aniline blue test; and chromatin integrity with acridine orange as fluorocrom. Swim up technique was based on Berger et al. (1985) with mHTF, heatingthe samples in a Falcon tube in a 45º angle in a 37ºC gas heater for one hour (5% C02 atmosphere). Following incubation 0.5 ml of the overlay containing sperm cells that swam up from the pellet were removed to process the recovered spermatozoids. Student’s t test was applied to compare PM, M, CC, and CI between the four groups. A significant difference was found between G1 vs G3 and G2 vs G3 (p 0.1). It showed that PM, M, CC and CI parameters in the recovered spermatozoids after swim up were similar to fertile population. CONCLUSIONS: Our results indicate that through the swim up procedure gametes with fertile ability similar to normal fertile population can be recovered to be applied in low complexity in vitro fertilization techniques such as intrauterine insemination, where the natural selection is still viable

[Spermatic fertilization capacity after swim-up spermatozoa recovery technique]. / Capacidad fecundante espermática luego de la recuperación de espermatozoides mediante la técnica de swim-up.

OBJECTIVES: To investigate sperm quality before and after swim up in infertile patients, and to compare it with a fertile men population. METHODS: Semen samples from 55 patients consulting at the infertility services of the Hospitals "Centenario" in Rosario and "Eva Perón" in Gro Baigorria were collected and analyzed accordingly with the WHO guidelines. 30 sperm samples with a volume higher than 1.0 ml, and spermatozoid concentration higher than 5,000,000/ml, not presenting hyperviscosity were selected. Outcome variables including progressive mobility (PM), morphology (M), chromatin condensation (CC) and chromatin integrity (CIl, were compared in fresh semen samples, between patients without previous treatment (G2) and after swim up (G3) and 15 fertile men (G1). Sperm morphology was evaluated by brilliant green hematoxyllin stain; progressive mobility with a subjective method accordingly to WHO (1999); chromatin condensation with aniline blue test; and chromatin integrity with acridine orange as fluorocrom. Swim up technique was based on Berger et al. ( 1985) with mHTF, heatingthe samples in a Falcon tube in a 45 degree angle in a 37 degree C gas heater for one hour (5% CO2 atmosphere). Following incubation 0.5 ml of the overlay containing sperm cells that swam up from the pellet were removed to process the recovered spermatozoids. Student's t test was applied to compare PM, M, CC, and CI between the four groups. A significant difference was found between G1 vs G3 and G2 vs G3 (p < 0.001). No significant differences were found between G1 and G3 (p > 0.1). It showed that PM, M, CC and C1 parameters in the recovered spermatozoids after swim up were similar to fertile population. CONCLUSIONS: Our results indicate that through the swim up procedure gametes with fertile ability similar to normal fertile population can be recovered to be applied in low complexity in vitro fertilization techniques such as intrauterine insemination, where the natural selection is still viable.

[Modified sperm stress test. Immune response and functionality of the human spermatozoid membrane]. / Estrés espermático modificado. Respuesta inmune y funcionalidad de la membrana del espermatozoide humano.

OBJECTIVE: Our purpose was to study, in semen samples of infertile patients, the relationship between the Modified Sperm Stress Test (MOST) and the presence of antisperm antibodies (ASA), macrophages concentration and the Hypoosmotic Swelling Test (HOST). METHODS: Semen samples from 42 men undergoing evaluation for infertility were examined according to WHO criteria. Twenty-five of them, whithout clumping non hyperviscosity, were selected. The MOST test was applied according to the author's original technique. ASA were determined with a direct mixed agglutination test, TAC II, developed and validate by our group. Macrophage concentration was evaluated with Neutral Red stain, and functional integrity of the sperm membrane with the Hypoosmotic Swelling Test using an hypoosmotic solution of 150 mOm/ml composed of equal parts of fructose and sodium citrate. RESULTS: The Chi square test was applied to the observational data obtaining the following results: There was a statistically significant association between the presence of ASA and altered MOST (p<0.001). In all samples with ASA, abnormal MOST values were obtained (MOST<0.39). Besides, there is a statistically significant association exists between the increased concentration of macrophages and abnormal MOST (p<0.01); and altered HOST was positively correlated with abnormal MOST (p<0.02). CONCLUSION: Results clearly demonstrate the high predictive power of MOST like a test of sperm resistance to the forced lipoperoxidation, offering conditions to become a good predictor of sperm performance. Understanding the sperm resistance to the ROS and their harmfull effects on the sperm functions, a proportion of infertile men can be succesfully treated.