Unscheduled DNA synthesis assay in mammalian spermatogenic cells: An update

Sotomayor, Rene E.; Sega, Gary A.

doi:10.1002/1098-2280(2000)36:4<255::aid-em1>3.0.co;2-o

Cited by 41 publications

(17 citation statements)

References 81 publications

(65 reference statements)

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“…The increased chromatin dissociation after H 2 O 2 treatment is most likely due to oxidant-induced DNA breaks. Owing to the condensed nature of the chromatin and the loss of most of their cytoplasm, mature spermatozoa lack appreciable levels of DNA repair (65)(66)(67). Knowing that OH • damages DNA by inducing base modification and strand breaks (41), we used the quantitative PCR assay to examine if the slight difference in percentage of abnormal DD that we saw between young and old animals was indeed due to increased susceptibility to oxidative DNA modifications with age.…”

Section: Zubkova Effect Of Age On Spermatozoal Chromatin Fertil Stementioning

confidence: 77%

Changes in spermatozoal chromatin packaging and susceptibility to oxidative challenge during aging

Zubkova

Wade

Robaire

2005

Fertility and Sterility

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Objective: Our goal was to test the hypothesis that spermatozoal chromatin packaging changes with age and that aging affects the susceptibility of spermatozoal DNA to oxidative damage. Design: Laboratory study. Setting: Academic facility. Patient(s): Young (4 months) and old (21 months) Brown Norway rats. Intervention(s): Spermatozoa were collected from the cauda epididymidis and were incubated in saline or H 2 O 2 . Main Outcome Measurement(s): Thiols levels, chromatin condensation, DNA susceptibility to acid-induced DNA denaturation, and DNA damage were evaluated using monobromobimane, chromomycin A3 (CMA3), acridine orange, and polymerase chain reaction, respectively. Result(s): Spermatozoa from old rats had 25% fewer disulfides but similar levels of free thiols as compared with young. The CMA3 staining was decreased by 13% with age. Levels of chromatin denaturation and DNA damage were similar in control groups. After exposure to oxidant, free thiols became oxidized by about 20% irrespective of age, but CMA3 staining changed little. The acridine orange assay, however, showed a trend for greater chromatin denaturation in spermatozoa from old rats after oxidant treatment. Furthermore, the DNA from spermatozoa of old rats was significantly more susceptible to developing DNA breaks and modification after oxidative challenge.

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Section: Zubkova Effect Of Age On Spermatozoal Chromatin Fertil Stementioning

confidence: 77%

Changes in spermatozoal chromatin packaging and susceptibility to oxidative challenge during aging

Zubkova

Wade

Robaire

2005

Fertility and Sterility

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“…spermatogonia) germ cells have a greater capacity for repair, making them less susceptible to stress-induced abnormalities? Spermatids formed from secondary spermatocytes must have a minimal capacity for DNA repair (Sega 1979, Sotomayor & Sega 2000 as they are silenced both transcriptionally and translationally during the condensation of the chromatin during sperm head remodelling. Furthermore, as sperm cannot respond by inducing either apoptosis or DNA repair, any DNA damage induced in these post-meiotic spermatids could persist.…”

Section: Discussionmentioning

confidence: 99%

A single, mild, transient scrotal heat stress causes DNA damage, subfertility and impairs formation of blastocysts in mice

Paul¹,

Murray²,

Spears³

et al. 2008

Reproduction

235

168

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Infertility represents a major clinical problem and 50% of cases are attributable to the male partner. Testicular function is temperature dependent, and in both man and mouse the position of the testes in the scrotum ensures that they are kept at between 2 and 8 8C below core body temperature. We used a mouse model to investigate the impact of a single, transient, mild, scrotal heat stress (38, 40 or 42 8C for 30 min) on testicular function, sperm DNA integrity and embryo survival. We detected temperature-dependent changes in testicular architecture, number of apoptotic cells and a significant reduction in testis weight 7 and 14 days after heat stress at 42 8C. We report for the first time that DNA strand breaks (g-H2AX-positive foci) were present in spermatocytes recovered from testes subjected to 40 or 42 8C. Fertility of heat-stressed males was tested 23-28 d after treatment (sperm at this time would have been spermatocytes at time of heating). Paternal heat stress at 42 8C resulted in reduced pregnancy rate, placental weight and litter size; pregnancies from the 40 8C group had increased resorptions at e14.5. Abnormalities in embryonic development were detected at e3.5 and in vitro fertilisation with sperm recovered 16 h or 23 d after scrotal stress at 42 8C revealed a block in development between the 4-cell and blastocyst stages. This study has provided evidence of temperature-dependent effects on germ cell DNA integrity and highlighted the importance of an intact paternal genome for normal embryo development. Reproduction (2008) 136 73-84

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“…Our results confirm and extend prior studies of the DNA repair-deficient window during spermiogenesis [13][14][15][16][17][18][19][20]. The results of our experiments showed that the frequencies of zygotes with chromosomal aberrations were significantly higher when a total dose of 28 mg/kg DEB was given over days 14 through 8 before mating than when it was given as a single dose 14 days before mating (Table 2 and Figure 3A).…”

Section: Discussionmentioning

confidence: 99%

“…These results show that the last two weeks of mouse spermiogenesis, which corresponds to the period of spermatogenesis that is thought to be DNA repair-deficient in prior studies [13][14][15][16][17][18][19][20], is also the sensitive window for induction of chromosomal aberrations in sperm after paternal exposure to DEB, and that both sperm and late spermatids are unable to repair DEB-induced damage.…”

Section: Chromosomal Aberrations Detected In Zygotes After Repeated Ementioning

confidence: 99%

“…Over fifty years of germ cell mutagenesis have shown that the majority of mutagens induce their highest effect during the last few weeks of spermatogenesis before fertilization [11,12]. The high sensitivity of the postmeiotic period to mutagenic exposure has been associated with the reduced DNA repair capacity of late spermatids and sperm as compared to early spermatids and other spermatogenic cell types [13][14][15][16][17]. All major DNA repair pathways seem to be less functional in late spermatids and sperm [17][18][19][20].…”

Section: Introductionmentioning

confidence: 99%

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DNA Repair Decline During Mouse Spermiogenesis Results in the Accumulation of Heritable DNA Damage

Marchetti¹,

Wyrobek²

2007

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The post-meiotic phase of mouse spermatogenesis (spermiogenesis) is very sensitive to the genomic effects of environmental mutagens because as male germ cells form mature sperm they progressively lose the ability to repair DNA damage. We hypothesized that repeated exposures to mutagens during this repair-deficient phase result in the accumulation of heritable genomic damage in mouse sperm that leads to chromosomal aberrations in zygotes after fertilization. We used a combination of single or fractionated exposures to diepoxybutane (DEB), a component of tobacco smoke, to investigate how differential DNA repair efficiencies during the three weeks of spermiogenesis affected the accumulation of DEB-induced heritable damage in early spermatids (21-15 days before fertilization, dbf), late spermatids (14-8 dbf) and sperm (7-1 dbf). Analysis of chromosomal aberrations in zygotic metaphases using PAINT/DAPI showed that late spermatids and sperm are unable to repair DEB-induced DNA damage as demonstrated by significant increases (P<0.001) in the frequencies of zygotes with chromosomal aberrations. Comparisons between single and fractionated exposures suggested that the DNA repair-deficient window during late spermiogenesis may be less than two weeks in the mouse and that during this repair-deficient window there is accumulation of DNA damage in sperm.Finally, the dose-response study in sperm indicated a linear response for both single and repeated exposures. These findings show that the differential DNA repair capacity of post-meioitic male germ cells has a major impact on the risk of paternally transmitted heritable damage and suggest that chronic exposures that may occur in the weeks prior to fertilization because of occupational or lifestyle factors (i.e, smoking) can lead to an accumulation of genetic damage in sperm and result in heritable chromosomal aberrations of paternal origin.

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Unscheduled DNA synthesis assay in mammalian spermatogenic cells: An update

Cited by 41 publications

References 81 publications

Changes in spermatozoal chromatin packaging and susceptibility to oxidative challenge during aging

Changes in spermatozoal chromatin packaging and susceptibility to oxidative challenge during aging

A single, mild, transient scrotal heat stress causes DNA damage, subfertility and impairs formation of blastocysts in mice

DNA Repair Decline During Mouse Spermiogenesis Results in the Accumulation of Heritable DNA Damage

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