Metachromatic staining of human sperm nuclei after reduction of disulphide bonds

Barrera, Clydes; Mazzolli, Alicia B.; Pelling, Claus; Stockert, Juan C.

doi:10.1016/s0065-1281(11)80366-1

Cited by 27 publications

(31 citation statements)

References 39 publications

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“…The spermatozoa of infertile men can also exhibit incomplete nuclear sulfhydryl group oxidation (8,9), the reaction leading to the formation of stabilizing disulfide cross-links. These sperm abnormalities (histone/protamine ratio and sulfhydryl group status) can potentially result in defective chromatin compaction (10) and in an increased susceptibility to DNA damage (11).…”

Section: Conclusion(s)mentioning

confidence: 99%

Sperm head morphology is related to high deoxyribonucleic acid stainability assessed by sperm chromatin structure assay

Zini

Phillips²,

Courchesne³

et al. 2009

Fertility and Sterility

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Section: Conclusion(s)mentioning

confidence: 99%

Sperm head morphology is related to high deoxyribonucleic acid stainability assessed by sperm chromatin structure assay

Zini

Phillips²,

Courchesne³

et al. 2009

Fertility and Sterility

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“…The structure of this insoluble, inactive chromatin is therefore very different from that found in the genetically active somatic nucleus. In fact, after release from the testis, the sperm chromatin is not only refractory to in situ enzymatic treatment [10,11], but also reveals a progressive decrease in Feulgen stainability, as well as a variable reduction in accessibility and binding capacity to many dyes and fluorochromes [12][13][14][15][16][17][18]. In the past decade, the variations in accessibility to DNA have been analyzed in order to establish hypothetical correlations between sperm nuclear instability and conditions of subfertility and infertility in mammals, including man [11,[19][20][21][22].…”

Section: Introductionmentioning

confidence: 99%

Effect of Deoxyribonucleic Acid Protamination on Fluorochrome Staining and in Situ Nick-Translation of Murine and Human Mature Spermatozoa1

Bianchi

Manicardi

Bizzaro

et al. 1993

Biology of Reproduction

240

140

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A major event in enhancing sperm chromatin stability is the replacement of the histones by protamines during spermiogenesis. In this study, we present results indicating that chromomycin A3 (CMA3) can be used to show protamine deficiency in sperm chromatin. Fixed chromatin of mature mouse spermatozoa showed high fluorescence after treatment with ethidium bromide (EB), but was completely unstained after treatment with CMA3. The same chromatin was found to be highly resistant to in situ nick-translation. In contrast, a substantial fraction of human spermatozoa were positive for CMA3. The accessibility of CMA3 to the DNA of human sperm was eliminated if the slides were previously treated with protamine in situ. This treatment did not affect the accessibility of EB to the chromatin. Individual human sperm samples revealed a substantial frequency of spermatozoa with endogenous nicks, which was found to be the same as the frequency of spermatozoa responding positively to CMA3 staining. Treatment of preparations with protamines prevented the identification of the endogenous nicks. These data as a whole suggest that CMA3 could represent a useful tool for the detection of protamine deficiency in sperm chromatin. Furthermore, confirmation of experiments relating sensitivity to nick translation and positivity to CMA3 may allow an indirect in situ visualization of nicked and partially denatured DNA, which could correlate with certain forms of male factor infertility.

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“…De esta manera, utilizando dicho colorante es posible clasificar a los espermatozoides de acuerdo al grado de compactación de la cromatina. Por otro lado, la reducción de los grupos disulfuro mediante agentes reductores como ditiotreitol (DTT) o mercaptoetanol (ME), permite que las proteínas se relajen y que los grupos fosfato del ADN queden disponibles y accesibles a la unión de los colorantes (Barrera et al, 1993). Esto permite, utilizar al DTT o al ME como controles positivos de la tinción con AT.…”

Section: Revisión Del Temaunclassified

“…Esto permite, utilizar al DTT o al ME como controles positivos de la tinción con AT. Esta tinción se ha utilizado en varias especies para detectar defectos en la condensación de la cromatina (humano: Barrera et al, 1993;Erenpreiss et al, 2001;Erenpreisa et al, 2003;Tsarev et al, 2009;toro: Beletti y Mello, 1996;Beletti et al, 2005;conejo: Beletti y Mello, 2004;padrillo equino: Naves et al, 2004;Sardoy et al, 2008;Carretero et al, 2012a) y actualmente se ha empleado para evaluar la cromatina en caninos, felinos y porcinos (Monachesi y Carretero, 2014;Allera et al, 2016;Arraztoa et al, 2016).…”

Section: Revisión Del Temaunclassified

“…Por otra parte, distintos autores han utilizado diferentes soluciones para fijar los extendidos: metanol (Barrera et al, 1993), etanol-ácido acético (3:1) más etanol 70% (Beletti y Mello, 2004;Beletti et al, 2005), etanolacetona (1:1) (Gagnon 1999;Erenpreiss et al, 2001;Erenpreisa et al, 2003;Tsarev et al, 2009) pero muchas de estas sustancias pueden ser altamente tóxicas si son inhaladas durante su manipulación (Macedo y Nava Muñoz, 2000). En CSA, equino, felino, canino y porcino, la fijación simple con etanol 96° durante 2 minutos fue efectiva (Sardoy et al, 2008;Carretero et al, 2009;2010a, 2010b2012a;Monachesi y Carretero, 2014;Allera et al, 2016;Arraztoa et al, 2016), dado que se encontraron patrones de coloración similares a los observados en otras especies (Barrera et al, 1993;Gagnon 1999;Erenpreiss et al, 2001;Erenpreisa et al, 2003;Beletti y Mello, 2004;Beletti et al, 2005;Tsarev et al, 2009). Todas las características mencionadas hacen que el AT sea una técnica sencilla de realizar, económica y rápida para determinar el grado de condensación de la cromatina de los espermatozoides.…”

Section: Revisión Del Temaunclassified

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Evaluación de la calidad del ADN espermático en camélidos sudamericanos y otras especies domésticas

Carretero¹,

Giuliano²,

Neild³

2017

SPERMOVA

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RESUMENLas anormalidades en el material genético pueden expresarse por defectos en la maduración o condensación nuclear, rupturas del ADN y anormalidades cromosómicas. Los parámetros de rutina utilizados para evaluar el semen (movilidad espermática, concentración, viabilidad, funcionalidad de membranas y morfología) no reflejan la calidad del ADN espermático. Esto cobra importancia para las técnicas de reproducción asistida, particularmente en la inyección intracitoplasmática de un espermatozoide en la que espermatozoides con genomas anormales pueden alcanzar el material genético del ovocito. Se postula que tres factores pueden estar involucrados en la etiología del daño del ADN: estrés oxidativo, aberraciones en el empaquetamiento de la cromatina y apoptosis. Varias técnicas han sido utilizadas para estudiar los defectos del ADN. A modo general, las técnicas de evaluación de ADN pueden ser divididas en 2 grupos: aquellas que evalúan el grado de condensación o compactación de la cromatina y aquellas que miden el grado de fragmentación del ADN. Muchas de estas técnicas son laboriosas, costosas y algunas dependen de enzimas cuya actividad y accesibilidad a las roturas de ADN pueden ser irregulares. En este artículo se comenta sobre el desarrollo y puesta a punto de varias de las técnicas de evaluación de la cromatina espermática en camélidos sudamericanos (CSA) y otras especies domésticas. Palabras clave: calidad de ADN, camélidos sudamericanos, espermatozoides ABSTRACTAbnormalities in sperm genetic material can be expressed as maturation or nuclear condensation defects, DNA damage or breakdown and chromosomal abnormalities. Routine semen evaluation (sperm motility, concentration, viability, membrane function and morphology) does not reflect sperm DNA quality. This becomes important when applying assisted reproductive techniques, especially intracytoplasmic sperm injection, in which sperm with abnormal genomes can reach the oocyte's genetic material. Three factors have been suggested to be involved in the etiology of DNA damage: oxidative stress, chromatin packaging anomalies and apoptosis. Various techniques have been used to study DNA defects. In general, DNA evaluation techniques can be divided into two groups: those that evaluate the degree of chromatin condensation or compactation and those that measure the degree of DNA fragmentation. Many of these techniques are laborious, costly and some depend on enzymes whose activity and accessibility to DNA fragments can be irregular. In this article, the development and setting up of various techniques to evaluate sperm chromatin in South American Camelids (SACs) and other domestic species is commented on.

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Metachromatic staining of human sperm nuclei after reduction of disulphide bonds

Cited by 27 publications

References 39 publications

Sperm head morphology is related to high deoxyribonucleic acid stainability assessed by sperm chromatin structure assay

Sperm head morphology is related to high deoxyribonucleic acid stainability assessed by sperm chromatin structure assay

Effect of Deoxyribonucleic Acid Protamination on Fluorochrome Staining and in Situ Nick-Translation of Murine and Human Mature Spermatozoa1

Evaluación de la calidad del ADN espermático en camélidos sudamericanos y otras especies domésticas

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