The research question: Whether mitochondrial dysfunction might be responsible for age-related decline in female fertility.

Study design: Review of the literature on decline in fertility with advanced female age, and relation between ageing and mitochondrial dysfunction was performed in 2000 when the idea was conceived.

Findings: In humans, the fertility in female declines slowly from the age of 30 years Embryo implanting ability and survival start declining gradually after 30 years of age, but by more than two thirds after 40 years and in younger women with reduced ovarian reserve. While decline in the frequency of intercourse is one of the reasons, reduction in the quality of either the embryos arising from ageing oocytes due to higher incidence of oocyte aneuploidy or the older uterus have been implicated as the probable causes. Controversy still exists about which one of these is the main cause or whether both of them play a role together. While the suboptimal quality of the ageing oocytes and/or older uterus may be responsible for the age-related decline in female fertility, it is not clear why the functional quality of the uterus or the oocytes declines with increasing age. Recent researches have indicated the role of oxygen radical damage to the mitochondria in the somatic cells leading to mitochondrial dysfunction as the possible cause of the ageing process. In this article, a possible role of age-related mitochondrial dysfunction in the ageing oocytes and/or the uterus has been proposed as the cause of age-related decline in female fertility.

Implications: Research in to this area would be useful to explore the possibility. If decline in fertility with advanced female age is found to be associated with mitochondrial dysfunction preventive measures that might retard the process of mitochondrial dysfunction associated with ageing could be taken in advance. N. B. The hypothesis was originally conceived and the article was written by the author in 2000 and has been included here as it was written in 2000.

Keywords: female fertility, ageing oocyte, mitochondrial dysfunction

In humans, the fertility in female declines slowly from the age of 30 years.^1,2 Embryo implanting ability and survival start declining gradually after 30 years of age, but by more than two thirds after 40 years and in younger women with reduced ovarian reserve.³ While decline in the frequency of intercourse is one of the reasons, reduction in the quality of either the embryos arising from ageing oocytes due to higher incidence of oocyte aneuploidy or the older uterus have been implicated as the probable causes. Controversy still exists about which one of these is the main cause or whether both of them play a role together.^1–6 While the suboptimal quality of the ageing oocytes and/or older uterus may be responsible for the age-related decline in female fertility, it is not clear why the functional quality of the uterus or the oocytes declines with increasing age. Recent researches have indicated the role of oxygen radical damage to the mitochondria in the somatic cells leading to mitochondrial dysfunction as the possible cause of the ageing process. In this article, a possible role of age-related mitochondrial dysfunction in the ageing oocytes and/or the uterus has been proposed to be the cause of the age-related decline in female fertility.

Older uterus or ageing oocytes

A substantial drop in the ongoing pregnancy rate per embryo transfer has been observed in women undergoing assisted reproduction, 48.8% in women aged <30 years to 13.6% in women aged >42 years). Embryo implantation rate also declines in a linear fashion, from 29% in women <34 years to approximately 5% at age 42. The reduced implantation rate in older women is apparently independent of the magnitude of their stimulation response. Though oocyte factors are held primarily responsible for the decline in fertility in older women, diminished endometrial receptivity has also been suggested as a contributor.⁷ It has been suggested from studies performed in women undergoing in vitro fertilisation (IVF) treatment and IVF with oocyte donation, that senescence affects both the ovary (oocytes and granulosa cells) and the uterus.^1,8 A defective vasculature of the uterus has been indicated as the possible cause of an increased miscarriage rate in women aged >40 years.¹ With ovum donation, an increased rate of pregnancy loss after the completion of implantation has been observed in women >40 years. A retardation of steroid synthesis suggests that the mechanism(s) responsible for placenta formation and functioning in the uterus is affected by age.⁹

A negative correlation between age and fecundity has been shown in the mouse. It has been attributed to the appearance of amorphous material beneath the basal lamina of the endometrial epithelium that could impair implantation.¹⁰ Elevated levels of epidermal growth factor might also have a physiological role in fertility decline in ageing mice possibly via uterine hypertrophy.¹¹ In cycling women >40 years, a disturbance in follicular recruitment but not in luteal function or endometrial maturation has been observed and its possible role in the decline in fertility with ageing suggested.¹² Elsewhere, it has been suggested that age-related changes in the ovary account for most of the decline in fertility. Oocytes, which originate in the fetal life, decline in numbers and quality with age. The endocrine function of the ovary also declines with age leading to dysfunction of the neuro endocrine axis. The latter may be further affected by primary changes in the hypothalamus and pituitary due to ageing, although there is no such evidence in humans. The uterine vasculature may change due to the age-related endocrine dysfunction thereby losing its ability to support implantation and growth of the embryo.¹³ The frequency of chromosomal anomalies in abortuses increases in parallel with the age-related rise in the incidence of spontaneous abortions.¹⁴ That oocyte aneuploidy is one major reason for low pregnancy rates in older women, is suggested by the higher pregnancy rate in this group where young donor oocytes are used.¹⁵ Majority of the studies support this view.

Degenerative changes and chromosomal abnormalities in the ageing oocyte have been implicated as the causes of the decline in female fertility with age.^4–22 A study, performed on women undergoing IVF, concluded that the reduction in quality of the embryos arising from ageing oocytes is the cause for age-related decline in female fertility. The blastocyst formation rate ([blastocysts/embryos on day 2] 100) and the blastocyst expansion rate ([expanded blastocysts/blastocysts] 100) according to the woman’s age on the day of IVF were determined. With increase in age, the number of retrieved oocytes decreased, without alteration of the cleavage rate. In women above age 30 years, preimplantation development to blastocysts declined due to an arrest at the morula stage. A negative linear relationship between blastocyst expansion rate and woman’s age was found, where the blastocyst stage was reached. Increasing age led to a drastic decrease in women having at least one expanded blastocyst (<30 years, 82%; >40 years, 36%) because of reduction in gamete production and embryo development. A high delivery rate per oocyte retrieval (25.8%) was found in women above age 40 years after embryo transfer at the blastocyst stage.⁵ This study indicated a poor quality embryo up to the stage of blastocyst as the cause of reduction in fertility with increasing age. Although majority of the studies have suggested a decrease in the quality of the oocytes as the main cause of the age-related decline in female fertility, contribution by the uterus also remains a possibility.

Embryonic development and the mitochondria

Normal mitochondrial function in the embryo to produce adequate energy is essential for mitochondrial protein synthesis, DNA synthesis and embryonic growth.²³ In a study to determine the actual and potential activities of the cytochrome system in cleavage-stage mouse embryos, three major shifts in the mode of ATP production during preimplantation stages were detected. The first, between the two-cell and late four-cell stages; the second, between the eight-cell and late morula stages; and the third, between the late morula and late blastocyst stages.²⁴ The importance of ooplasmic factors has been postulated for the continued development of the zygote, particularly during early cleavage, when transcription of the embryonic genome is minimal.^25,26 In recent years, mitochondrial DNA (mtDNA) has been a subject of interest due to the subtle role it may have in early development.²⁷ Sperm mitochondria, carrying potentially harmful paternal mtDNA are eliminated from the embryo at an early stage, probably by a ubiquitin-dependent mechanism.^28,29 Therefore the mtDNA in the fetus comes from the oocyte.^27–31 As a consequence, any dysfunction of the oocyte mitochondria may have profound detrimental effects on the development of the embryo.

Ageing and mitochondrial dysfunction

Mitochondrial dysfunction has been attributed as the cause of the ageing process.^32,33 Oxidative damage appears to play a critical role in causing mitochondrial dysfunction of ageing.³⁴ The contributing factors include the intrinsic rate of proton leakage across the inner mitochondrial membrane (a correlate of oxidant formation), reduced membrane fluidity, and decreased levels and function of cardiolipin, which supports the function of many of the proteins of the inner mitochondrial membrane. Acetyl-L-carnitine, a high-energy mitochondrial substrate, appears to reverse many age-associated cellular dysfunctions, partly by increasing cellular ATP production.³⁵ With increasing age the mitochondrial function declines with an increase in the production of reactive oxygen species (ROS) and free radicals in the mitochondria. Up to a concentration range ROS may induce stress responses of the cell by altering genetic expression in order to increase energy metabolism to rescue the cell. However beyond this threshold, ROS may elicit apoptosis by induction of mitochondrial membrane permeability transition and release of cytochrome c. Recent researches have established a pivotal role of mitochondria in the early phase of apoptosis in mammalian cells.³⁶ Observations in transgenic Drosophila melanogaster support the hypothesis that oxidative injury might directly cause the ageing process.³⁷ The “free radical production hypothesis of aging” states that a decrease in oxygen radical production per unit of oxygen consumption near critical DNA targets (mitochondria or nucleus) increases the maximum life span of extraordinarily long-lived species like birds, primates and man.³⁸ It has been hypothesized that oxygen free radicals may damage the mtDNA thereby inhibiting mitochondrial division leading to age-related reduction in mitochondrial numbers, a deficient energy production with decrease in protein synthesis and deterioration of physiological performance.^37–41 mtDNA is 20 times more susceptible to mutation than nuclear DNA⁴² due to its location close to the site of reactive oxidative species production, and a lack of protective histones in mtDNA.⁴³ Contrary to the popular belief, mitochondria can efficiently repair oxidative damage to their DNA.⁴⁴

Ageing oocyte - a mitochondrial dysfunction?

Each mitochondrion contains 2-10 copies of mtDNA in all human tissues, except oocytes and platelets, which contain only one copy per mitochondrion.⁴² During germ-line development in early bovine embryogenesis, the number of mitochondria increases 100-fold, from 1000 per oogonium to 100 000 per oocyte, while the number of mtDNA increases only 10-fold, from 10 000 to 100 000.⁴⁵ As a result each mitochondrion contains 1 mtDNA, instead of the usual 5-10.^45,46 Only a small number of mtDNA molecules replicate and give rise to the whole cytoplasmic genotype during the late stage of oogenesis.⁴⁷ This bottleneck may be a cause of the accumulation of oxidative damage to mtDNA during the ageing process,⁴⁸ which also occurs in oocytes.⁴⁵ Ageing of the ooplasm has been considered to be responsible for producing an abnormal meiotic spindle¹⁷ with spindle abnormalities as the source of incorrect seggregation of chromosomes/chromatids at meiosis I.⁴ To overcome this problem, the possibility of using manufactured oocytes is being investigated, where the nucleus of immature oocyte from an older woman is transplanted into the cytoplast of a younger woman.^49,50 The oocytes originate in the fetal life and enter the first meiotic division that remains arrested until ovulation. In contrast to the continuous production of primary spermatocytes in the male after puberty, no primary oocytes form after birth. The first meiotic division in the primary oocytes completes just before ovulation, forming secondary oocytes.⁵¹ Therefore the age of the secondary oocytes in a woman above 40 years will be just above the woman’s age, as these were originated before her birth. This means that the oocytes would have been exposed to the ageing process for that period, during which oxidative damage to the oocyte mitochondria might lead to mitochondrial dysfunction. This may affect the functional quality of the oocyte or cause oocyte aneuploidy due to interference with the meiosis, which would have profound detrimental effect on fertilisation, protein synthesis and cell division in the fertilised ovum, migration of the fertilised ovum to the uterine cavity, implantation and subsequent development of the embryo. As normal mitochondrial function is essential for embryonic development, dysfunction of the oocyte mitochondria may be responsible for the age-related decline in female fertility. Mitochondrial dysfunction may also lead to suboptimal quality of the uterus, thereby decreasing fertility.

Fertility in the female declines with age. This has been attributed to the decrease in frequency of intercourse, and reduction in the quality of an older uterus or the embryos resulting from ageing oocytes. Although an older uterus may impair implantation or increase miscarriage rates due to a decreased receptivity of the endometrium or defective vasculature, majority of the studies indicated ageing oocytes as the main contributing factor. The etiology for the suboptimal function of the older uterus or ageing oocytes is not clear. Oxidative damage to mitochondria leading to mitochondrial dysfunction is considered to be the cause of age-related changes in somatic cells. The latter are associated with a decrease in protein synthesis and deterioration in physiological performance. There is evidence that age-related changes also occur in the oocyte mitochondria. The mtDNA in the embryo is inherited from the oocyte mitochondria, any dysfunction of which might have profound detrimental effect on the embryonic development. Therefore, mitochondrial dysfunction associated with increase in age might have a role for the age-related decline in female fertility.

None.

The authors declare there is no conflict of interests.

None.

1. Pellicer A, Simon C, Remohi J. Effects of aging on the female reproductive system. Hum Reprod. 1995;10 Suppl 2:77–83.
2. Navot D, Drews MR, Bergh PA, et al. Age–related decline in female fertility is not due to diminished capacity of the uterus to sustain embryo implantation. Fertil Steril. 1994;61(1):97–101.
3. Hull MG, Fleming CF, Hughes AO, et al. The age–related decline in female fecundity:a quantitative controlled study of implanting capacity and survival of individual embryos after in vitro fertilization. Fertil Steril. 1996;65(4):783–790.
4. Dailey T, Dale B, Cohen J, et al. Association between nondisjunction and maternal age in meiosis–II human oocytes. Am J Hum Genet. 1996;59(1):176–184.
5. Janny L, Menezo YJ. Maternal age effect on early human embryonic development and blastocyst formation. Mol Reprod Dev. 1996;45(1):31–37.
6. Marcus SF, Brinsden PR. In–vitro fertilization and embryo transfer in women aged 40 years and over. Hum Reprod Update. 1996;2(6):459–468.
7. Rosenwaks Z, Davis OK, Damario MA. The role maternal age in assisted reproduction. Hum Reprod. 1995;10 Suppl 1:165–173.
8. Saranti L, Allali F, Olivennes F, et al. Results of oocyte donation in women with different indications, ages and therapeutic strategies. Contracept Fertil Sex. 1997;25(7–8):643–646.
9. Cano F, Simon C, Remohi, et al. Effects of aging on the female reproductive system:evidence for a role of uterine senescence in the decline in female fecundity. Fertil Steril. 1995;64(3):584–589.
10. Shimizu K, Yamada J. Relationship of decrease in fecundity with advancing age to structural changes in mouse endometrium. J Anat. 2000;196:111–114.
11. Tsutsumi O, Taketani Y, Oka T. Evidence for the involvement of epidermal growth factor in fertility decline in aging female mice. Horm Res. 1993;39 Suppl 1:32–36.
12. Batista MC, Cartledge TP, Zellmer AW, et al. Effects of aging on menstrual cycle hormones and endometrial maturation. Fertil Steril. 1995;64(3):492–499.
13. Fitzgerald C, Zimon AE, Jones EE. Aging and reproductive potential in women. Yale J Biol Med. 1998;71(5):367–381.
14. Nasseri A, Grifo JA. Genetics, age, and infertility. Maturitas. 1998;30(2):189–192.
15. Hassold TJ, Jacobs P A. Trisomy in man. Ann Rev Genet. 1984;18:69–97.
16. Balmaceda JP, Bernardini L, Ciuffardi I, et al. Oocyte donation in humans:a model to study the effect of age on embryo implantation rate. Hum Reprod. 1994;9(11):2160–2163.
17. Battaglia DE, Goodwin P, Klein NA, et al. Influence of maternal age on meiotic spindle assembly in oocytes from naturally cycling woman. Hum Reprod. 1996;11:2217–2222.
18. Lim AS, Tsakok MF. Age–related decline in fertility:a link to degenerative oocytes? Fertil Steril. 1997;68(2):265–271.
19. Navot D, Bergh PA, Williams MA, et al. Poor oocyte quality rather than implantation failure as a cause of age–related decline in female fertility. Lancet. 1991;337(8754):1375–1377.
20. Sauer MV. The impact of age on reproductive potential:lessons learnt from oocyte donation. Maturitas. 1998;30(2):221–225.
21. Te Velde ER, Scheffer GJ, Dorland M, et al. Development and endocrine aspects of normal ovarian aging. Mol Cell Endocrinol. 1998;145(1–2):67–73.
22. Vermeulen A. Environment, human reproduction, menopause, and andropause. Environ Health Perspect. 1993;101 Suppl 2:91–100.
23. Oerter D, Bass R. Embryonic development and mitochondrial function. 1. Effects of chloramphenicol infusion on the synthesis of cytochrome oxidase and DNA in rat embryos during late organogenesis. Naunyn Schmiedebergs Arch Pharmacol. 1975;290(2–3):175–189.
24. Ginsberg I, Hillman N. Shifts in ATP synthesis during preimplantation stages of mouse embryos. J Reprod Fertil.1975;43(1):83–90.
25. Liu L, Day Y, Moor RM. Nuclear transfer in sheep embryos:the effect of cell–cycle coordination between nucleus and cytoplasm and the use of in vitro matured oocytes. Mol Reprod Dev. 1997;47(3):255–264.
26. Van Blerkom J, Davies P, Merriam J, et al. Nuclear cytoplasmic dynamics of sperm penetration, pronuclear formation and microtubule organization during fertilization and early preimplantation development in the human. Hum Reprod Update. 1995;1(5):429–461.
27. Van Blerkom J. Developmental failure in human reproduction associated with preovulatory oogenesis and pre–implantation embryogenesis. In: Van Blerkom J, Molta P, editors. Ultrastructure of Human Gametogenesis and Embryogenesis. Kluwer, Dordrecht, Netherlands, Europe; 1989:125–180.
28. Cummins JM, Kishikawa H, Mehmet D, et al. Fate of genetically marked mitochondrial DNA from spermatocytes microinjected into mouse zygotes. Zygote. 1999;7(2):151–156.
29. Sutovsky P, Schatten G. Paternal contributions to the mammalian zygote:fertilization after sperm–egg fusion. Int Rev Cytol. 2000;195:1–65.
30. Allen JF. Separate sexes and the mitochondrial theory of ageing. J Theor Biol. 1996;180(2):135–140.
31. Enriquez JA, Martinez–Azorin F, Garesse R, et al. Human mitochondrial genetic system. Rev Neurol. 1998;26 Suppl:S21–S26.
32. Maftah A, Ratinaud MH, Dumas M, et al. Human epidermal cells progressively lose their cardiolipins during ageing without change in mitochondrial transmembrane potential. Mech Ageing Dev. 1994;77(2):83–96.
33. Rattan SI. Cellular and molecular determinants of ageing. Indian J Exp Biol. 1996;34(1):1–6.
34. Lenaz G, Cavazzoni M, Genova ML, et al. Oxidative stress, antioxidant defences and aging. Biofactors. 1998;8(3–4):195–204.
35. Shigenaga MK, Hagen TM, Ames BN. Oxidative damage and mitochondrial decay in aging. Proc Natl Acad Sci USA. 1994;91(23):10771–10778.
36. Lee HC, Wei YH. Mitochondrial role in life and death of the cell. J Biomed Sci. 2000;7(1):2–15.
37. Fukagawa NK. Aging:is oxidative stress a marker or is it causal? Proc Soc Exp Biol Med. 1999;222(3):293–298.
38. Barja G, Cadenas S, Rojas C, et al. A decrease of free radical production near critical targets as a cause of maximum longevity in animals. Comp Biochem Physiol Biochem Mol Biol. 1994;108(4):501–512.
39. Fleming JE, Miquet J, Cottrell SF, et al. Is cell aging caused by respiration–dependent injury to the mitochondrial genome? Gerontology. 1982;28(1):44–53.
40. Feuers RJ. The effects of dietary restriction on mitochondrial dysfunction in aging. Ann N Y Acad Sci. 1998;854:192–201.
41. Lenaz G, Bovina C, Formiggini G, et al. Mitochondria, oxidative stress, and antioxidant defences. Acta Biochim Pol. 1999;46(1):1–21.
42. Kagawa Y, Hayashi JI, Endo H. Gene therapy of mitochondrial diseases using human cytoplasts. Gene Therapy. 1997;4:6–10.
43. Tritschler HJ, Medori R. Mitochondrial DNA alteration as a source of human disorders. Neurology. 1993;43(2):280–288.
44. Bohr VA, Dianor GL. Oxidative DNA damage processing in nuclear and mitochondrial DNA. Bochimie. 1999;81(1–2):155–160.
45. Chen X, Prosser R, Simonetti S, et al. Rearranged mitochondrial genomes are present in human oocytes. Am J Hum Genet. 1995;57(2):239–247.
46. Robin ED, Wong R. Mitochondrial DNA molecules and virtual number of mitochondria per cell in mammalian cells. J Cell Physiol. 1998;136(3):507–513.
47. Hauswirth W, Laipis P. Transmission genetics of mammalian mitochondria:a molecular model and experimental evidence. In: Quagliarello E, editor. Achievements and Perspectives of Mitochondrial Research (Vol 2). Elsevier, Amsterdam, Netherlands, Europe; 1985:49–59.
48. Cortopassi GA, Shibata D, Soong NW, et al. A pattern of accumulation of somatic deletion of mitochondria DNA in aging human tissues. Proc Natl Acad Sci USA. 1992;89(16):7370–7374.
49. Tsai MC, Takeuchi T, Bedford JM, et al. Alternative sources of gametes:reality or science fiction? Hum Reprod. 2000;15:988–998.
50. Zhang J, Wang CW, Krey L, et al. In vitro maturation of human preovulatory oocytes reconstructed by germinal vesicle (GV) transfer. Fertil Steril. 1999;71(4):726–731.
51. Jones WR. Fertilization, implantation and early development of the embryo. In: Whitfield C R, editor. Dewhurst’s Textbook of Obstetrics and Gynaecology for Postgraduates. 5th edn. Blackwell Science, England, UK; 1995:64–72.