Extracts from BBC web page - Friday, 11 October, 2002, Frozen egg birth brings IVF hope

The first British woman to become pregnant using her own frozen eggs has given birth to a healthy baby girl. Helen Perry, 36, from Ludlow, Shropshire, became pregnant using an egg which had been taken from her ovary six months earlier, frozen, stored then thawed and replanted.

Dr Gillian Lockwood, whose team made the breakthrough, says: "The technology ... will work just as well for the Bridget Jones generation who want to freeze their eggs to keep their reproductive options open.. On ITV1's Tonight with Trevor McDonald on Friday, she explains: "I think that egg freezing may come to be seen as the ultimate kind of family planning."

The reproductive lifespan

Ovarian cycles Reproductive potential

Pregnancy

Lactation

Male

Puberty

Female Menopause Age

Menstrual cycle lengths

16 14 12 % of cycles 10 8 6 4 2 0 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 C ycle len g th (d ays)
Note: Variations in cycle length are normal! Particularly: just after menarche (1st menstrual period) approaching the menopause

Menstruation is an external indicator of ovarian events controlled by the hypothalamicpituitary axis Feedback

Hypothalamus

GnRH (gonadotrophin
Pituitary releasing hormone) LH FSH (gonadotrophins)

Ovaries

Roles of the ovary Steroids 1. Gametes (ova) (oestradiol, 2. Hormones progesterone).

MENSTRUATION Reproductive tract Other targets

What are oestrogens?

OH

Oestradiol-17
HO

CH3

Ovaries Testes Placenta

Brain Answer: Steroids with characteristic effects, esp. on female reproductive tract. Some are more potent than others. Breast ` Cardovascular system

Bone

Reproductive tract

GnRH pulse Hypothalamus generator

Pulsatile activity of GnRH neurones

GnRH
Pituitary LH FSH

Pulses of GnRH

Pulses of LH

Ovaries

Plasma LH

24 hours

Natural suppression Before puberty Lactation Diet induced Anorexia Malnutrition Exercise
Hypothalamic neurones GnRH pulse generator

Modulation during menstrual cycle Diet Stress?

Frequency and amplitude of GnRH pulses

Pituitary

Changing patterns of LH and FSH

Timing events in the menstrual cycle.

1. Onset of menstruation

Day 1

Day 1

12

16

20

24

28

Menstruation

Timing events in the menstrual cycle.

2. LH surge
LH
Days before Days after

Day 1

Day 1

Follicular phase

Luteal phase

12

16

20

24

28

Menstruation

OVULATION

Animated ovarian events

Key events in the ovarian cycle

LH
Day 1 1. Follicular growth

12
Oestra diol

16

20

24

28

Menstruation

OVULATION

Animated ovarian events

Key events in the ovarian cycle

2. Ovulation 1. Follicular growth

LH

Day 1

12
Oestra diol

16

20

24

28

Menstruation

OVULATION

Animated ovarian events

Key events in the ovarian cycle

2. Ovulation 1. Follicular growth

LH
3. Luteal function

Day 1

12
Oestra diol

16

20

24

28

Menstruation

OVULATION

Progester one (and oestradiol

Animated ovarian events

Key events in the ovarian cycle

2. Ovulation 1. Follicular growth

LH
3. Luteal function 4. Luteal regression

Day 1

12
Oestra diol

16

20

24

28

Menstruation

OVULATION

Progester one (and oestradiol

The follicle is the fundamental element of the ovary:

Blood vessels

Theca

Granulosa cells
Antrum

Cumulus cells Oocyte Zona pellucida

(non-cellular glycoprotein coat)

Oocyte

Cumulus cells

Cytoplasmic bridges from cumulus cells to oocyte for transport

Zona pellucida
(non-cellular glycoprotein coat)

Where do follicles come from?

Male
Spermatogonia

Female
Primordial germ cells (oogonia)

Continuous replacement in adult

Mitoses stop in fetal life

Meiosis All enter MEIOSIS

Mitoses

This means there is Continuous pool a fixed, limitedsperm ofproduction oocytes.

Spermatozoa

Arrested in 1st meiotic division Primary oocytes in primordial follicles

~7 m

(Fig adapted from Johnson & Everitt, 2000)

No. of germ cells (millions)

Continuous loss 99.9% by Atresia Ovulation (post-puberty) Puberty BIRTH

3 6 9 months 1 10

~ 300,000
20 years 40

Conception

Age from conception

Growth of follicles:
Antral follicle Graafian follicle

Primordial follicle

Oocyte Granulosa cells Thecal cells

Antrum (fluid filled space)

Ovulation

Lets look at follicular growth first There are a number of questions to ask

How many follicles reach this point? Normally 1

Ovulation

Menstruation

Many! 30-50
How many follicles are growing at the start of the cycle?

Ovulation

Menstruation

Many! 30-50
How many follicles are growing at the start of the cycle?

Why is only 1 selected and becomes dominant?

When do follicles start growing?

Ovulation

2-3 months earlier!

Menstruation

What controls follicular growth?

Hypothalamus

GnRH (gonadotrophin
Pituitary
LH

releasing hormone)

Steroid feedback

Ovaries

FSH

(gonadotrophins)

Oestradiol (E2)

Reproductive tract Other targets

What controls follicular growth?

OVULATORY FOLLICLE

??????
Gonadotrophin independent

FSH
+ LH Ovulation

Menstruation

OVULATORY FOLLICLE

FSH
+ LH Ovulation OESTRADIOL
Menstruation

As each follicle grows, it produces increasing amounts of oestradiol.

How is oestradiol production controlled ?

Theca

Granulosa cells

LH
Androgens (Note: the production of androgens is a normal part of ovarian physiology)
Androgens are converted (aromatized) to oestradiol by the granulosa cells

FSH Inhibin (protein)

OESTRADIOL (steroid)

_
Increasing amounts of oestradiol.

Hypothalamus (gonadotrophin GnRH releasing hormone) FSH

Pituitary

Ovaries

LH

Oestradiol (E2)

Reproductive tract Other targets

Increasing negative feedback

Hypothalamus (gonadotrophin GnRH releasing hormone)

Pituitary INHIBIN
(suppresses FSH)

Increasing amounts of oestradiol.

Ovaries

Decreased FSH

Oestradiol (E2)

Reproductive tract Other targets

As the follicles grow, FSH levels fall due to the negative feedback,
FSH Oestradiol

12

16

20

24

28

Many follicles at the start of the cycle

Why is only 1 selected and becomes dominant?

Ovulation

Menstruation

Hypothalamus

GnRH
Pituitary

Ovaries

FSH

Large follicles: less Small follicles: Population of growing follicles dependent on FSH very dependent Oestradiol (E2) on FSH

Hypothalamus

GnRH
Pituitary

Ovaries

FSH

Large follicles: less Small follicles: dependent on FSH very dependent Oestradiol (E2) on FSH

Hypothalamus Increasing negative GnRH feedback Pituitary

Oestradiol

INHIBIN

Ovaries

Decreased FSH FSH

Insufficient FSH

Large follicles: less Small follicles: dependent on FSH very dependent Oestradiol (E2) on FSH Growth factors Oestradiol + +

Dominant follicle

FSH secretion suppressed

FSH
Oestradiol

Dominant follicle(s) can survive

Insufficient FSH to keep smaller follicles going they become atretic.

12

16

20

24

28

Polycystic ovaries
The classical picture of PCO: a string of follicles, 2-8 mm in diameter

Section of ovary showing multiple follicles in PCO

Ultrasound of ovary showing multiple follicles

Theca

Granulosa cells

LH
Androgens (Note: the production of androgens is a normal part of ovarian physiology)
Androgens are converted (aromatized) to oestradiol by the granulosa cells

FSH
OESTRADIOL (steroid)

4. The disturbed steroid feedback may re-inforce the abnormal LH/FSH secretion

1. Raised LH, lowered FSH

3. The high LH induces high androgen secretion from the theca HIRSUTISM

2. .. leads to disturbed follicle growth ANOVULATION Disturbed cycles

4. The disturbed steroid feedback may re-inforce the abnormal LH/FSH secretion

1. Raised LH, lowered FSH

3. The high LH induces high androgen secretion from the theca HIRSUTISM

2. .. leads to disturbed follicle growth ANOVULATION Disturbed cycles

4. The disturbed steroid feedback re-inforces the abnormal LH/FSH secretion

1. Raised LH, lowered FSH

3. The high LH induces high androgen secretion from the theca

2. .. leads to disturbed follicle growth ANOVULATION Disturbed cycles

HIRSUTISM

What causes ovulation?

12

16

20

24

28

What causes ovulation?

LH

12

16

20

24

28

What effects does it have?

What causes the LH surge?

12

16

20

24

28

NOT HUMANS!
What causes the LH surge? Reflex ovulation

0
Mating

12

16
LH

20

24

28

Neuroendocrine reflex

Oestradiol

12

16

20

24

28

For most of the cycle, negative feedback operates

Hypothalamus

_
Pituitary LH FSH

GnRH

Inhibited by oestradiol

Oestradiol

Ovary

BUT, with high levels of E2 maintained for long enough

Hypothalamus

GnRH

Pituitary

Oestradiol

LH LH FSH surge Ovary

BUT, with high levels of E2 maintained for long enough

Hypothalamus

Increased GnRH

Pituitary

GnRH Increased sensitivity to GnRH

Oestradiol

LH LH FSH surge Ovary

How does the LH surge affect the follicle?

About 36 h between LH surge and oocyte release..

Oocyte: Completion of the 1st meiotic division (unequal division; extrusion of 1st polar body) 2nd meiotic division starts but becomes arrested before completion. Microvilli across the zona pellucida are withdrawn. Loosening of cumulus cells

Enzyme induction in the follicle wall

Transformation of ruptured follicle into corpus luteum (CL) Ruptured follicle becomes solid corpus luteum Thecal cells and blood vessels invade Granulosa cells hypertrophy and terminally differentiate (luteinisation). Steroid secretion changes Progesterone

+ Oestradiol

Follicular phase: Oestradiol domination

Luteal phase: Progesterone domination

What maintains the CL?

Oestradiol Progesterone
Why does the CL degenerate at the end of the cycle?

12

16

20

24

28

OVULATION

What maintains the CL?

8 hr between LH pulses

Hypothalamus

GnRH Pituitary

CL

LH (low levels)

Progesterone + E2

CL very sensitive to LH

What maintains the CL? Steroid negative feedback keeps LH and FSH levels relatively low
Progesterone + E2 Reproductive tract etc

Hypothalamus

GnRH Pituitary

CL

LH (low levels in luteal phase)

CL very sensitive

Hypothalamus

Towards the end of the cycle, the sensitivity to LH reduces.

Pituitary

GnRH

The low levels of LH are insufficient to keep the CL going

LH

Progesterone + E2

CL degenerates

Hypothalamus

GnRH

As CL degenerates steroid negative feedback reduces ..

Pituitary

FSH + LH

Progesterone + E2

Hypothalamus

GnRH

As CL degenerates steroid negative feedback reduces .. New wave of follicles stimulated by rising Progesterone FSH and LH +E
2

Pituitary

FSH + LH

Oestradiol Progesterone

12

16

20

24

28

OVULATION

Oestradiol Progesterone

12

16

20

24

28

OVULATION

Oestradiol Progesterone

12

16

20

24

28

OVULATION

Oestradiol Progesterone

12

16

20

24

28

OVULATION

Other changes in the cycle

a) Outer muscle layer the myometrium b) Inner glandular mucosa the endometrium

Uterine changes in the menstrual cycle.

Endometrial depth

Oestradiol causes an increase in thickness (the proliferative phase)

More secretion from the glands hence the term secretory phase

12

16

20

24

28

Menstruation

OVULATION

Terminal differentiation of stromal cells decidualisation Characteristic spiral arteries

12

16

Menstruation

20 24 Optimal time for implantation

28

What causes the onset of menstruation?

Steroid levels fall

This is followed by the onset of menstruation

1. At end of the luteal phase, steroid production declines. 2. Loss of oedema and gradual shrinking of endometrial tissue. The spiral arteries become more highly coiled 3. Gradual reduction in blood flow to superficial layers leading to ischaemic hypoxia and damage to the epithelial and stroma cells. 4. 4 24 hours prior to menstrual bleeding, an intense constriction of spiral arteries occurs. 5. Individual arteries re-open at different times, tearing and rupturing the ischaemic tissues. 6. Bleeding into the cavity occurs via: 1. red cells diapedese between surface epithelial cells; 2. tears develop in the surface epithelium 3. pieces of weakened superficial endometrium crumble away 7. About 50% of degenerating tissues is resorbed and 50% is lost as 'menstrual bleeding'.

Onset of menstruation is rapid. Probably 95% of women have a total blood loss of less than 60 mls. This blood loss can represent a significant loss of iron (leading to anaemia) especially in women on marginal diets

Menstruation - WHY?

In preparation for pregnancy, the human uterine stromal cells go through complex changes and the stromal cells terminal differentiate - Decidualization. If implantation and pregnancy do not occur, this tissue is lost - and the uterus prepares itself again for another possible pregnancy.

Animated ovarian events

Key events in the ovarian cycle

2. Ovulation 1. Follicular growth

LH
3. Luteal function 4. Luteal regression

Day 1

12
Oestra diol

16

20

24

28

Menstruation

OVULATION

Progester one (and oestradiol

Nearly all pregnancies in a 6-day fertile window Day of intercourse

Ovulation?

Probability of clinical pregnancy following intercourse on a given day relative to ovulation (estimated from basal body temperature).

Cervical mucus
Abundant mucus - like raw egg white
Cervical mucus

Production of low viscosity mucus increases

Variable number of dry days

Thick, rubbery, high viscosity impenetrable to sperm.

12

16

20

24

28

Menstruation

OVULATION

With increasing oestradiol: 1. The mucus becomes more abundant - up to 30x more and its water content increases. 2. Its pH becomes alkaline. 3. Increased elasticity ("spinnbarkeit test") 5. Ferning pattern caused by the interaction of high concentrations of salt and water with the glycoproteins in the mucus.

Characteristic fernlike pattern as the mucus dries on a glass slide.

38 37.8 37.6 37.4 37.2 37 36.8 36.6 36.4 36.2 36

A small (0.5 oC) rise in BBT typically follows ovulation. LH

Basal body temperature

12

16

20

24

28

Menstruation

OVULATION

Anovulatory cycle?

Fertility

LH
Oestradiol Progesterone

Ovulation

12

16

20

24

28

OVULATION

Basal body temperature

Plasma oestradiol

Plasma progesterone Volume of cervical mucus and sperm penetration Uterine endometrium

There are a number of potential ways of trying to identify the fertile period..: a) Calendar Method - which is essentially based on the previous menstrual history. b) Temperature method - using a midcycle rise in body temperature as a sign when ovulation has occurred. c) Cervical changes - which can be detected by feeling the cervix and cervical mucus. d) Hormonal methods - using over-thecounter "kits" to assess urinary hormone levels.

Problem-based powerpoint presentation (using many of the same screens as this lecture) on the menstrual cycle can be found at: www.kcl.ac.uk/ip/stuartmilligan/ppt/pptpage.h tml