Autor(es):
Tavares, R. S. 
; Silva, A. F. ; Lourenço, B. ; Almeida-Santos, T. ; Sousa, A. P. ; Ramalho-Santos, J.
Data: 2013
Identificador Persistente: http://hdl.handle.net/10316/25635
Origem: Estudo Geral - Universidade de Coimbra
Assunto(s): ART fertility outcomes; chromatin status; Diff-Quik staining; human spermatozoa; sperm preparation techniques
Descrição
Sperm chromatin/DNA damage can be measured by a variety of assays. However, it has been reported that these tests may lose
prognostic value in Assisted Reproductive Technology (ART) cycles when assessed in post-prepared samples, possibly due to the normalizing
effect promoted by sperm preparation procedures. We have recently implemented a modified version of the Diff-Quik
staining assay that allows for the evaluation of human sperm chromatin status in native samples, together with standard sperm morphology
assessment. However, the value of this parameter in terms of predicting in vitro fertilization (IVF) and Intracytoplasmic
sperm injection (ICSI) outcomes after sperm selection is unknown. In this study, data from 138 couples undergoing in vitro fertilization
(IVF) or Intracytoplasmic sperm injection (ICSI) treatments showed that sperm chromatin integrity was significantly improved
after density gradient centrifugation and swim up (p < 0.001), but no correlations were found with fertilization or embryo development
rates (p > 0.05). However, sperm samples presenting lower percentages of damaged chromatin were associated with better
quality (Grade I) embryos in both ART procedures (p < 0.05) and clinical pregnancy among IVF couples (p < 0.05). Furthermore,
regression analysis confirmed the clinical value of Diff-Quik staining in predicting IVF (but not ICSI) clinical pregnancy (OR: 0.927,
95% CI: 0.871–0.985, p = 0.015), and a threshold value of 34.25% for this parameter was established. The proportion of IVF couples
achieving a clinical pregnancy was reduced 1.9-fold when the percentage of abnormal dark staining was ≥34.25% (p = 0.05). In
conclusion, the Diff-Quik staining assay provides useful information regarding ART success, particularly in IVF cycles, where some
degree of ‘natural’ sperm selection may occur; but not in ICSI, where sperm selection is operator dependent. This quick and low-cost
assay is suggested as an alternative method to detect sperm chromatin status in minimal clinical settings, when no other well-established
and robust assays (e.g. Sperm chromatin structure assay, terminal deoxynucleotidyl transferase-mediated dUDP nick-end
labelling) are available. This work was carried
out, in part, in order to fulfil the requirements for a Masters
Degree in Biology (A.F.S.) and a PhD Degree in Cell Biology
(R.S.T.), Department of Life Sciences, University of Coimbra,
Portugal. R.S.T. is a recipient of a PhD fellowship from FCT,
Portugal (SFRH/BD/46002/2008). CNC funding is supported by
FCT, Portugal (PEst-C/SAU/LA0001/2011).
