Ovarian hormones

bioassay and pharmacology of ovarian hormones

1. Follicle stimulating
hormone [FSH] in female
stimulates ovarian follicle
development. The follicle
secretes estrogen when it
becomes mature it is
called grafian follicle.
Lutenizing hormone[LH]
in female acts on the
mature grafian follicle to
induce ovulation (LH
peak). It also stimulates the
Corpus lutum (derived
from the ruptured grafian
follicle after ovulation) to
secrete progesterone.
Hypothalamus
GnRF [+]

Anterior Pituitary GlandAnterior Pituitary Gland

2. Hormonal inter-relationship in the control of
the female reproductive system

3. 1) Follicular phase
•During the first half of this stage FSH secretion Slightly &
acts on one of the ovaries promoting growth of several
follicles each contains one ovum. After about a week, one of
the follicles out grows the others & secretes increasing
amount of estrogen & matures to form grafian follicle while
the other small follicles degenerate.
•By -ve feed back mechanism of estrogen secretion
FSH secretion regression of the other small follicles,
while the dominant follicle can continue its growth due to
its local concentration of estrogen which its sensitivity
to FSH through number & sensitivity of FSH receptors
on it. The secreted estrogen stimulates proliferation of
endometrium.
A brief Account on Hormonal A brief Account on Hormonal
Changes in the Menstrual CycleChanges in the Menstrual Cycle

4. 2) Ovarian Phase
The LH secretion peaks due to +ve feed back
effect of estrogen secreted from the
dominant follicle resulting in rupture of the
developed follicle & expulsion of ovum.
3) Luteal Phase
The remaining part of the ruptured follicle
develops into corpus luteum, which secretes
progesterone.

5. If fertilization occurs:
Fertilized ovum Secrete (hCG)
Human chronic Gonadotrophins
With LH activity Support corpus
Lutum secrete Progesterone
Relax uterus & thickness and
vascularity of endometrium.
i.e. prepare it for implantation of
fertilized ovum & Protect it.
If no fertilization
occurs:
Degeneration of corpus
luteum sudden drop of
progesteron loss of
endometrium support
necrosis of endometrium
menstrual cycle
N.B.
Progesterone is secreted during 9 months of pregnancy,
firstly by corpus lupus then by placenta.
This effect is stimulated by LH. The secreted
progesterone promotes secretory function of
the endometritm. Then

6. Plasma
concentrations of
ovarian hormones
and gonadotrophis
in women during
normal menstrual
cycle.

7. Ovarian Hormones
Bioassay
of
Estrogen
s
# Injection of estrogen the thickness of the
vaginal wall, leucocytes cannot pass& only squamus
cornified cells are identified in the vaginal smear in
estrous stage.
# The number of animals with +ve response is directly
proportional to the estrogen dose.
# The wall of vagina in diestrous stage is very thin&
allows the passage of leucocytes into the vaginal
lumen. So leucocytes appear in the vaginal smear.
1) Vaginal smear methodVaginal smear method
Principle:

8. Vaginal Smear MethodVaginal Smear Method

9. We use ovarictomized rats to avoid endogenous
estrogen interference.
4 groups of animals in the diestrous stage are used (2T &
2S)
Choose 2 SM doses of T estrogen (T1 &T2) & 2
SM doses of S (S1& S2) provided that:
S2/S1=T2/T1
Each dose is injected in a group of animals & the vaginal
smear is made on the third day & examined for the
presence of squamus cornified cells.
Plot a relation between log dose& % of
animals with +ve response in each group of
S&T.
Determine relative potency (R.P)
Procedures:

10. S2
T2
T1
S1
%

+
v
e

5
0
%

r
e
s
p
o
n
s
e
XYLog dose
R.P =X/Y
N.B.
# It is an in vivo method
# 4-point assay
# Objective method (accurate)
# No cross-over test can be done
# this test can be used for screening &
bioassay of estrogen.

11. 2) Uterus weight method2) Uterus weight method
Principle:
* Estrogen uterus weight of ovarectomized female rats.
* The in weight is directly proportional to the dose of estrogen.
Ovarictomized or immature female rats to avoid endogenous
estrogen interference.
4 groups of the animals are used.
Choose 2 SM doses of S estrogen (S1& S2) & 2 SM doses of
T estrogen (T1&T2) provided that: S2/S1=T2/T1
Each dose is injected in a group of animals in the 3rd,
4th,5th & 6th days after ovarectomy.
Rats are killed on the 8th day & the uterus is isolated, dried&
weighed.
Procedures:
Plot a relation between log dose & % in uterus weight &
determine R.P

12. S2
T2
T1
S1
%

i
n

U
t
e
r
u
s
W
e
i
g
h
t
X Y
Log dose
R.P =X/Y
N.B.
* It is an in vivo test
* 4-point assay
* Objective method (accurate)
* No cross-over test can be carried out because
the animals are killed

13. ## Endometrial proliferation method
# Progesterone causes proliferation of the
endometrium of immature of ovarictomizes
rabbits.
# The degree of proliferation is directly
proportional to the dose of progesterone.
# Proliferation occurs through 4 phases
Principle:
Bioassay
of
Progesterone

14. Endometrial
proliferation
method for
Progesterone

15. Procedures:
Choose 2 SM doses of S progesterone( S1&S2) & 2 SM doses of
T progesterone (T1&T2) provided that S2/S1=T2/T1
Each dose of progesterone is injected S.C in a group of animals
daily for 5 days
Compare the degree of proliferation in T&S
We use immature or ovarictomized female rabbits to avoid
interference by endogenous progesterone.
Bring 4 groups of animals
Inject all animals with estrogen for 6 days (for development of
uterus to prepare it for progesterone action)
Kill the animal, isolate the uterus & make section through it
Plot a relation between log dose& degree of proliferation the
determine R.P.

16. S2
T2
T1
S1
D
e
g
r
e
e
o
f

P
r
o
l
i
f
e
r
a
t
i
o
n
XYLog dose
R.P =X/Y
N.B.
# It is an in vivo test.
# 4-point assay.
# No cross-over test can be
carried out because the animals
are killed.

17. Prepared by: