Human Fertility
Master Medical Biology
Lecture 1: Oogenesis
- Development of female gamete
- Lengthy process (from early female embryogenesis to
menopause = approx. 50 years)
- Male contribution to zygote: DNA; Female: DNA,
organelles, RNA, protein
- Highly complex developmental program of the oocyte to get
fertilized: including cytoplasmic maturation and nuclear
maturation
- More than 99% of the oocytes is lost during oogenesis
Primordial germ cells
(precursors of oocyte) are
generated from the proximal
epiblast during gastrulation
and are part of the extra
embryonic mesoderm.
- The primordial germ cells migrate from the primitive gonads
(part of embryo) to the wall of the yolk sac (extra-embryonic tissue) for several
weeks and then they migrate back to the embryo and to the gonads that are forming
at that time.
When the primordial germ cells migrate back to the forming
gonads, they will be multiplied by mitosis.
Generation of primordial follicles
- In the gonads syncytia are formed by synchronized
mitotic division of gonocytes with incomplete
cytokinesis leading to syncytia.
- Somatic pre-granulosa cells invade the syncytia
- Granulosa cells will break down the syncytia and will
form single oocytes surrounded by a single layer of
the helper cells → the granulosa cells.
- From this stage on they are called primordial follicles.
These cells rest in the prophase stage of meiosis I
In humans:
- Primary germ cells first differentiate to oogonia that
multiply by mitosis
- At week 6-7 of embryonic development this results in
100.000 oogonia
- Differentiation of oogonia to primary oocytes and start of meiosis I
Steps:
1. Oogonia (2n, 2c)
2. DNA replication starts (2n,4c)
3. Condensation of chromatids (pachytene)
4. Alignment of homologous chromosomes (diplotene)
1
, Human Fertility
Master Medical Biology
5. Crossing over (diakinesis): exchange of sister chromatids
Meiosis I arrest for 12-50 years: start in the beginning of puberty
This process will lead to about 7 M primordial oocytes arrested in the
prophase of meiosis 1
The oocyte only shows 1 strong signal per pair of chromosomes, due to close
localization of sister chromatids (FISH analysis).
Reduction in number of oocytes during life:
- From 7 M to 2 M oocytes at birth.
- At six months of gestation, you get mitosis of your precursor oocyte → oocytes
2
, Human Fertility
Master Medical Biology
Prenatal reduction in number of germ cells:
- Massive germ cell apoptosis takes place in pachytene of meiosis 1 and during
formation of primordial follicles in mammals
- To prevent survival of compromised cells (deficient synapsis/crossing-over),
damaged mitochondria or to protect genomic integrity??
Explanation for the massive loss of oocytes during fetal development: LINE-1 genotoxicity
- LINE-1 = long interspersed element-1, a 7 kb retro-transposon
- Accounts for approximately 20% of the human genome (not all intact:
mutated/truncated)
- Overexpression and insertion lead to DNA damage and cell death.
- Two open reading frames (ORF)
o ORF1: encodes RNA-binding protein (chaperone function)
o ORF2: encodes endonuclease and reverse transcriptase
The Line-1 sequences are normally methylated
- Genomes are protected from L1 genotoxicity by multiple epigenetic mechanisms:
DNA-methylation → inactive
- Epigenetic remodelling of primordial germ cells includes genome-
wide removal of DNA methylation creating permissive conditions
for L1 expression (de-methylation is necessary to produce
functional germ cells) → endo-nuclease activity + reverse
transcriptase activity from ORF2
- L1 reverse transcription products activate complement system
and immune cells to eliminate oocytes by apoptosis. Inhibited by
azidothymidine (AZT)
o Reverse transcriptase produces eventually ss DNA, and all mammalian cells
are hypersensitive for ss DNA because this is an alarm function for viruses →
complement system activation.
3
, Human Fertility
Master Medical Biology
- L1 endonuclease activity creates DNA breaks (DSBs) that activates DNA damage
checkpoint through checkpoint kinase 2 (Chk2) leading to oocytes elimination.
FOA (Fetal oocyte attrition): acts as quality control for the genome to prevent transmission
of damaged genetic material.
Mouse model:
DNA methylation levels go down over time and LINE-1 levels
increase, leading to a decrease of the oocyte pool.
What is the fate of oocytes surviving the fetal attrition?
- Growing
- Multiple layers of granulosa cells
- Tertiary follicle: fluid filled cavity & new cell type will
surround the follicle → Theca cells
- Graafian follicle: end stage
o Ready for ovulation
The sequence of follicle maturation within the human ovary:
Other mechanisms important for the size of the
ovarian pool:
Life history of ovarian follicles:
- Initial recruitment is induced by cytokines and
EGFa (active pre-birth)
- It is a continuous process that starts when
primordial follicles are formed, long before the
onset of puberty
- Follicles develop unto the antral stage
independent of Follicle Stimulating Hormone
4
Master Medical Biology
Lecture 1: Oogenesis
- Development of female gamete
- Lengthy process (from early female embryogenesis to
menopause = approx. 50 years)
- Male contribution to zygote: DNA; Female: DNA,
organelles, RNA, protein
- Highly complex developmental program of the oocyte to get
fertilized: including cytoplasmic maturation and nuclear
maturation
- More than 99% of the oocytes is lost during oogenesis
Primordial germ cells
(precursors of oocyte) are
generated from the proximal
epiblast during gastrulation
and are part of the extra
embryonic mesoderm.
- The primordial germ cells migrate from the primitive gonads
(part of embryo) to the wall of the yolk sac (extra-embryonic tissue) for several
weeks and then they migrate back to the embryo and to the gonads that are forming
at that time.
When the primordial germ cells migrate back to the forming
gonads, they will be multiplied by mitosis.
Generation of primordial follicles
- In the gonads syncytia are formed by synchronized
mitotic division of gonocytes with incomplete
cytokinesis leading to syncytia.
- Somatic pre-granulosa cells invade the syncytia
- Granulosa cells will break down the syncytia and will
form single oocytes surrounded by a single layer of
the helper cells → the granulosa cells.
- From this stage on they are called primordial follicles.
These cells rest in the prophase stage of meiosis I
In humans:
- Primary germ cells first differentiate to oogonia that
multiply by mitosis
- At week 6-7 of embryonic development this results in
100.000 oogonia
- Differentiation of oogonia to primary oocytes and start of meiosis I
Steps:
1. Oogonia (2n, 2c)
2. DNA replication starts (2n,4c)
3. Condensation of chromatids (pachytene)
4. Alignment of homologous chromosomes (diplotene)
1
, Human Fertility
Master Medical Biology
5. Crossing over (diakinesis): exchange of sister chromatids
Meiosis I arrest for 12-50 years: start in the beginning of puberty
This process will lead to about 7 M primordial oocytes arrested in the
prophase of meiosis 1
The oocyte only shows 1 strong signal per pair of chromosomes, due to close
localization of sister chromatids (FISH analysis).
Reduction in number of oocytes during life:
- From 7 M to 2 M oocytes at birth.
- At six months of gestation, you get mitosis of your precursor oocyte → oocytes
2
, Human Fertility
Master Medical Biology
Prenatal reduction in number of germ cells:
- Massive germ cell apoptosis takes place in pachytene of meiosis 1 and during
formation of primordial follicles in mammals
- To prevent survival of compromised cells (deficient synapsis/crossing-over),
damaged mitochondria or to protect genomic integrity??
Explanation for the massive loss of oocytes during fetal development: LINE-1 genotoxicity
- LINE-1 = long interspersed element-1, a 7 kb retro-transposon
- Accounts for approximately 20% of the human genome (not all intact:
mutated/truncated)
- Overexpression and insertion lead to DNA damage and cell death.
- Two open reading frames (ORF)
o ORF1: encodes RNA-binding protein (chaperone function)
o ORF2: encodes endonuclease and reverse transcriptase
The Line-1 sequences are normally methylated
- Genomes are protected from L1 genotoxicity by multiple epigenetic mechanisms:
DNA-methylation → inactive
- Epigenetic remodelling of primordial germ cells includes genome-
wide removal of DNA methylation creating permissive conditions
for L1 expression (de-methylation is necessary to produce
functional germ cells) → endo-nuclease activity + reverse
transcriptase activity from ORF2
- L1 reverse transcription products activate complement system
and immune cells to eliminate oocytes by apoptosis. Inhibited by
azidothymidine (AZT)
o Reverse transcriptase produces eventually ss DNA, and all mammalian cells
are hypersensitive for ss DNA because this is an alarm function for viruses →
complement system activation.
3
, Human Fertility
Master Medical Biology
- L1 endonuclease activity creates DNA breaks (DSBs) that activates DNA damage
checkpoint through checkpoint kinase 2 (Chk2) leading to oocytes elimination.
FOA (Fetal oocyte attrition): acts as quality control for the genome to prevent transmission
of damaged genetic material.
Mouse model:
DNA methylation levels go down over time and LINE-1 levels
increase, leading to a decrease of the oocyte pool.
What is the fate of oocytes surviving the fetal attrition?
- Growing
- Multiple layers of granulosa cells
- Tertiary follicle: fluid filled cavity & new cell type will
surround the follicle → Theca cells
- Graafian follicle: end stage
o Ready for ovulation
The sequence of follicle maturation within the human ovary:
Other mechanisms important for the size of the
ovarian pool:
Life history of ovarian follicles:
- Initial recruitment is induced by cytokines and
EGFa (active pre-birth)
- It is a continuous process that starts when
primordial follicles are formed, long before the
onset of puberty
- Follicles develop unto the antral stage
independent of Follicle Stimulating Hormone
4
