Understanding how male age affects couple infertility

Date: 2008

In press: National Fertility and Adoption Directory, The American Fertility Association 2008





            Gentlemen: The Clock ticks for you.  You may have thought only your wife had a biological clock for pregnancy--  that you were fertile up until your golden years and any lack of a pregnancy, or miscarriages, or children with birth defects, were the fault of an aging woman who had ignored the ring of the alarm.  Not so!  --as now evidenced by clinical data and a recent study on the deterioration of sperm DNA integrity with age of man.  While it has become socially acceptable to father children at an older age, this increased age of fatherhood has been correlated with an increased time to establish a pregnancy or no pregnancy.   Since 1980, U.S. birth rates have increased up to 40% for men 35-49 years and have decreased up to 20% for men under 30.1


What might be the defect(s) that causes a decreased fertility potential for older men?  While many studies have been done examining sperm under the light microscope for concentration, motility and morphology, a pioneering technique, the Sperm Chromatin Structure Assay (SCSA® ),  was developed2.3  to measure, in situ, the integrity of sperm DNA —the male genome required for development of the embryo.  In this study, semen samples obtained from men attending an infertility clinic had significantly higher levels of sperm DNA damage/fragmentation than men known to have fathered a child within the past two years. 


Sperm DNA fragmentation measures

 In a comprehensive study,4,5 200 couples with no known infertility factors were enrolled in a male factor infertility study.   Monthly semen samples were obtained for the first 3 months or up to the time of biochemical or clinical pregnancy.  Pregnancies were recorded over the first 12 months.  Sperm were analyzed by the SCSA in which  small aliquots of frozen/thawed raw semen ejaculates are treated for 30 sec with an acid solution that denatures the DNA at sites of DNA strand breaks,  followed by staining DNA with acridine orange (AO).  AO stains native, double stranded DNA green and fragmented DNA red.  These samples were then measured for the amount of green and red fluorescence in a powerful instrument called a flow cytometer (FCM).  In contrast to light microscope observations, the FCM provides non-biased instrument measurements on thousands of cells, thereby making robust statistical data.  The computer calculated percent of sperm with fragmented DNA is the % DNA Fragmentation Index (% DFI).    The men who had a <15% DFI had the shortest time to establish a pregnancy, those with >15 and <30% DFI had the next longest time period while those with >30% had the longest time to pregnancy or no pregnancy. This latter group also had the highest level of miscarriages.  Thus, sperm DNA integrity had a strong correlation to man’s potential for fatherhood.


Age of man and loss of fertility potential

The first study6 on the relationship between AGE of non-smoking, healthy men and sperm DNA integrity showed that among all the sperm genomic endpoints measured, age had the strongest effects on sperm DNA integrity.  A gradual upward trend in the average frequency of sperm with DFI was observed, beginning in the early reproductive years.  A healthy 20 year old man typically has about 5% DFI.  The statistical odds of a healthy man to reach the threshold for negative pregnancy outcome (30% DFI, see below) is age 50. Statistical data showed that a 50 yr old has a one-third chance of  reaching this 30%DFI threshold by age along.   Men, however, have a heterogeneous biological clock in contrast to a narrow time window for women.   In this age study, men in their 50’s ranged from excellent DFIs (5%) to very poor ones (73%).  Even men in their 20s and 30s had abnormal DFI values, suggesting they too might experience diminished fertility and/or abnormal pregnancy outcomes, consistent with clinical experiences.  This factor is likely related to harmful environmental exposures as discussed below.



Sperm DNA Fragmentation and Pregnancy Outcomes

Sperm DNA fragmentation measurements using SCSA or other DNA fragmentation assays7 have been shown to be predictive for male sub/infertility.  Data from > thousand donors8 show that semen samples with DFI values of <27-30% have a higher probability of successful pregnancies by natural means (7.0-8.7 fold), intrauterine insemination (IUI, ~ 8 fold), and IVF/ICSI 1.5-2.0 fold.9 Many infertile couples have no detectable cause for their infertility.   In some cases, the likely cause of their infertility was due to fragmented sperm DNA which can not be identified by routine infertility measures.  In one study, 9 of 13 couples having a high % DFI were pregnant within three months after using donor sperm. 10  Males of couples experiencing repeated spontaneous abortions (>2) were shown to have a 4 times higher level of sperm DNA fragmentation than men who were sperm donors.11


Environmental toxins and male infertility 

Data suggest that environmental insults are a major factor for high levels of sperm DNA fragmentation in younger men.  The world is awash with man- made chemicals.  While chemical X alone may not have a deleterious effect, or chemical Y by itself, the mixture of X and Y and the multitude of other environmental chemicals may have a serious effect.  


 A number of articles have reviewed how exposure to toxins in males can cause reproductive harm including infertility. 12,13,14 For example, various organic compounds, certain halogenated compounds and heavy metals, pesticides and phalate esters (chemicals leached from plastics into our diets) may all compromise male reproductive function.    Occupational activities involving exposure to chemicals, including hydrocarbons such as toluene, benzene and xylene, can cause abnormal sperm count, motility and morphology as well as fragmented DNA.   Lead exposure has long been related to reduced sperm motility and count and an increase in abnormal sperm morphology and male infertility.15


Whereas most of the previous studies have correlated occupational exposure or personal lifestyle factors to classic measures of semen quality (sperm count, motility, morphology) and hormonal changes, perhaps the most important change of interest is whether sperm DNA damage occurs. 

Exposure of mice to X rays, a variety of chemicals, and heat to the animal’s testes (similar to hot tubs) clearly had damaging effects to sperm DNA.  The data were highly dose responsive showing that damage to sperm DNA is reflective of environmental exposure.14 These animal data clearly suggested that humans would likewise by affected in a dose related manner.  Some examples follow.


Men exposed to various insecticides and pesticides showed significantly increased levels of sperm DNA fragmentation.16  A dramatic effect of exposure to organophosphorous pesticides showed that 3/4 pesticide operators, not using protective gear,  had DFI values >30%.17  Significantly higher levels of sperm DNA fragmentation were found in factory workers exposed to styrene which is used to make plastics, rubber, and resins.18


Residents of Teplice, Czech Republic, a town with heavy winter time air pollution, generated by burning soft brown coal, experienced a higher than normal rate of infertility and spontaneous miscarriages.19  Czech army conscripts, 18 years of age, provided semen samples in a two year longitudinal study that went through periods of clean summer air  and polluted winter air.  Sperm DNA fragmentation measured by the SCSA was the only measure to detect a statistically significant correlation between air pollution levels and semen quality in these young men.  One-fourth of these young men had %DFI >30, placing them in a statistical group known to be at an increased risk for infertility. 


Etiology of sperm DNA fragmentation

Sperm DNA damage is multifactorial and may be due to many environmental conditions such as chemotherapy, radiation, some prescription medications (e.g. cortisone), air pollution, smoking, pesticides, chemicals, heat (fever or hot tubs); other factors include assisted reproductive preparation protocols for sperm and various pathological conditions such as cryptorchidism, cancer, fever, age, infection, leukocytospermia and varicocele among others.14  Elevated levels of sperm DNA fragmentation have been significantly associated with a negative pregnancy outcome.5,10,8 


The most common factor in causing sperm DNA fragmentation and its consequence on pregnancy outcome is oxidative stress in response to reactive oxygen species (ROS).21,22  Simply stated, we need oxygen to live, but ROS activity is a negative consequence of this fact.  Aging is thought to be related to ROS activity and sperm DNA damage is not an exception.  Many of the environmental factors discussed here are related to increased oxidative stress.  Thus, many physicians and patients are well aware of the need to have a diet rich in antioxidants.  Even when unique antioxidants are added to sex lubricants, e.g., Pre-Seed, there is less DNA damage in the sperm available for fertilization and embryo development.23


Sperm DNA integrity preservation in a polluted world

While TIME WAITS FOR NO MAN,  a healthy lifestyle is an excellent option to preserve as well as possible the integrity of sperm DNA resulting in better pregnancy outcomes.  While many young men can not escape from their heavily polluted environments, maybe the time has arrived that young men should consider freezing some aliquots of sperm to be potentially used later for establishing a pregnancy.  All men may have an unexpected and even unknown exposure to reproductive toxins. Even cancer patients prior to therapy often have compromised sperm DNA integrity.   Potential grandparents may well be very happy to pay storage fees to gain the pleasure of having their own grandchildren.  Too futuristic??  Perhaps not—TIME may be running out in some parts of this polluted world.


1. Martin, J. A., Hamilton, B. E., Sutton, P. D., et al (2005) Births: final data for 2004. Natl. Vital Stat. Rep. 54, 1–116.

2. Evenson, D.P., Darzynkiewicz, Z. and Melamed, M.R. (1980)  Relation of mammalian sperm chromatin heterogeneity to fertility.  Science 240:1131-1133.

3. Evenson, D.P., Larson, K., and Jost, L.K. (2002) The sperm chromatin structure assay (SCSA): clinical use for detecting sperm DNA fragmentation related to male infertility and comparisons with other techniques. Andrology Lab Corner.  J. Andrology 23: 25-43.

4.     Zinaman MJ, Brown CC, Selevan SG et al (2000) Semen quality and human fertility: a prospective study with healthy couples. J Androl.21:145-53

5. Evenson DP, Jost LK, Zinaman MJ et al. (1999) Utility of the sperm chromatin structure assay (SCSA®) as a diagnostic and prognostic tool in the human fertility clinic. Human Reproduction 14, 1039–1049

6. Wyrobek, A.J., Eskenazi, B., Young, al (2006)Advancing age in healthy men has differential effects on DNA strand damage, chromatin integrity, gene mutations, aneuploidies and diploidies in sperm. Proc. Natl. Acad. Sci.USA. 103: 9601-6

7. Erenpreiss J, Spano M, Erenpreisa J, et al  (2006) Sperm          chromatin structure and male fertility: biological and clinical aspects. Asian J Androl. 8:11-29.

8.  Bungum M, Humaidan P, Axmon A, et al (2007) A.Sperm DNA integrity assessment in prediction of assisted reproduction technology outcome.Hum Reprod. 22:174-9.

9. Evenson D, Wixon R (2006) Meta-analysis of Sperm DNA Fragmentation using the Sperm Chromatin Structure Assay. Reproductive BioMedicine 12:466-472.

10. Virro MR, Larson-Cook KL, Evenson DP. (2004) Sperm chromatin structure assay (SCSA®) related to blastocyst rate, pregnancy rate and spontaneous abortion in IVF and ICSI cycles. Fertil  Steril; 81:1289-95.

11. Carrell DT, Liu L, Peterson CM et al. (2003) Sperm DNA fragmentation is increased in couples with unexplained recurrent pregnancy loss. Archives of Andrology 49, 49–55.

12. Hauser R. (2006) The environment and male infertility: recent research on emerging chemicals and semen quality. Semin Reprod Med 24: 156-157.

13. Mendiola, J, “Torres-Cantero AM, Moreno-Grau, JM et al (2008)  Exposure to enviromental toxins in males seeking inferilty treatment: a case-controlled study.  Reproductive BioMedicine 16: 842-850.

14. Evenson DP and Wixon RL (2005) Environmental toxicants cause sperm DNA fragmentation as detected by the Sperm Chromatin Structure Assay (SCSA). Tox and Applied Pharmacology 532-537.

15.Wagner U, Schlebusch H, Van der Ven H, et al. Accumulation of pollutants in the genital tract of sterility patients. J of Clinical Chemistry and Clinical Biochemistry 28: 683-688.  

16. Bian Q,. Xu LC, Wang SL, Xia YK et al  Occup. Environ. Med., (2004), 61, 999.

17. Sanchez-Pena, L.C., Reyes, B.E., Lopez-Carrillo, L et al, (2004) Organophosphorous pesticide exposure alters sperm chromatin structure in Mexican agricultural workers. Toxicol. Appl. Pharmacol. 196, 108–113.

18.  Khargliore L, Naccarati A, Zanello A et al. (2002) Assessment of sperm DNA integrity in workers exposed to styrene. Hum Reprod. 17:2912-8.

19. Rubes J, Selevan SG, Evenson DP et a; (2005)  Episodic air pollution is associated with increased DNA fragmentation in human sperm without other changes in semen quality. Hum Reprod 20, 2776- 83.

20. Potts RJ, Newbury CJ, Smith G, et al. (1999) Sperm chromatin damage associated with male smoking.  Mutat Res 423:103-11.

21. Tremellen, K. (2008) Oxidative Stress and male infertility—a clinical perspective. Human Reproduction Update 14:243-258


22. Agarwal A, Saleh RA, Bedaiwy MA. (2003) Role of reactive oxygen species in the pathophysiology of human reproduction. Fertil Steril 79:829-843.

23.  Agarwal A, Deepinder F, Cocuzza M, et a;. (2007) Effect of vaginal lubricants on sperm motility and chromatin integrity: a prospective comparative study. Fertil Steril. 2007 May 15; 89 (2): 375-379.


Donald P. Evenson, Ph.D., HCLD

Disinguished Professor, Emeritus

South Dakota State Univeristy


President and Director

SCSA Diagnostics

Brookings, SD 57006