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In the Name of ALLAH, the Most Gracious, the Most Merciful, and Peace and Blessings be upon His Prophet Mohamed and his conscience followers, ever!

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Title: In the Name of ALLAH, the Most Gracious, the Most Merciful, and Peace and Blessings be upon His Prophet Mohamed and his conscience followers, ever!


1
In the Name of ALLAH, the Most Gracious, the Most
Merciful, and Peace and Blessings be upon His
Prophet Mohamed and his conscience followers,
ever!
2
REPRODUCTION IN HUMANS SPERMATOGENESIS, OOGENESIS,
CONCEPTION, IMPLANTATION, INTRODUCTION TO IVF
  • Ahmed M. Isa, Ph.D., HCLD (ABB), REM (ACE)
  • Assistant Professor , Head of IVF Lab
  • Department of Obstetrics Gynecology
  • King Khalid University Hospital
  • King Saud University

3
Sexual Reproduction in Humans
  • In general, sexual reproduction is the formation
    of a new individual following the union of two
    gametes, one from each parent.
  • In humans and the majority of eukaryotes, plants
    and animals, the two gametes differ in structure
    and function ("an-isogamy") and are contributed
    by two different parents.

4
Sexual Reproduction in Humans
  • These two different parents are
  • The father or the male, who produces the sperm,
    through a process called Spermatogenesis.
  • The mother or the female, who produces the egg,
    through a process called Oogenesis.

5
Production of Gamets
  • Spermatogenesis
  • The production of Sperms takes place in the two
    Testes.
  • Each testis is packed with seminiferous tubules
    (laid end to end, they would extend for more than
    20 meters long) where spermatogenesis occurs.

6
Male Reproductive System or the Semen Factory
7
Seminal Fluid Components
  • Sperms From the Epididymis. Normally, 2-5 of
    the volume.
  • Seminal Vesicle Secretion 65-75. amino acids,
    citrate, enzymes, flavins, fructose (energy
    source), phosphorylcholine, prostaglandins
    (suppress female immune system), proteins,
    vitamin C

8
Seminal Fluid Components
  • 3. Prostate Gland Secretion 25-30, acid
    phosphatase, citric acid, fibrinolysin, prostate
    specific antigen(PSA), proteolytic enzymes, zinc
    (about 13540 µgm/ml. Zinc helps to stabilize the
    sperm DNA-containing chromatin).

9
Seminal Fluid Components
  • 4. Bulbo-Uretheral Glands Secretion lt1,
    galactose, mucous (increase sperm mobility.
    Contributes to the cohesive jelly-like texture of
    semen.), pre-ejaculate
  • Cowpers fluid (a lubricant), and sialic acid.

10
Spermatogenesis
  • Steps of spermatogenesis
  • The walls of the seminiferous tubules consist of
    the germinal epithelium that gives rise to the
    diploid spermatogonia, which are the precursors
    of the sperm.
  • At puberty, Spermatogonia divide by mitosis to
    either produce more spermatogonia, or
  • differentiate into 1ry spermatocytes (2N).

11
Spermatogenesis
  • Each 1ry spermatocyte differentiates into 2
    haploid secondary spermatocytes (1st Meiosis).
  • Each 2ry spermatocyte differentiates into two
    haploid spermatids (2nd Meiosis).
  • Spermatids (4 from each 1ry spermatocyte) develop
    into sperms, losing most of their cytoplasm in
    the process, and developing their long tails.

12
T.S. of Rat Testis (humans is similar)
13
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14
Spermatogenesis
  • With 22 pairs of autosomes and an average of two
    crossovers between each pair, the variety of
    genetic material combinations in the resulting
    sperms are countless.
  • In humans, spermatogenesis from start to end
    takes about 64 days before a sperm is ready for
    ejaculation in the epididymis.

15
Sperm Structue
  • Sperm is a lot more than a flagellated cell. It
    is consisted of
  • A head (5µm by 3µm), which has
  • an acrosome on its tip, and
  • a nucleus contains a haploid set of chromosomes
    in a compacted state.
  • A midpiece containing the mitochondria and a
    single centriole.
  • A tail (midpiece and tail are 50µm long).

16
Diagram of an L.S. of a spermatozoan
17
Sperm Ultra-structure
  • This electron micrograph shows the sperm cell of
    a bat.
  • Note the orderly arrangement of the mitochondria
    in the sperm mid-piece.

18
  • In average, a normal adult man manufactures about
    100 million sperms each day.
  • As they are produced, they are moved into the
    epididymis where they undergo further maturation.
  • The acidic environment of the epididymis keeps
    the mature sperm inactive.
  • Animated Spermatogenesis
  • http//wps.aw.com/bc_martini_eap_4/40/10469/2680
    298.cw/content/index.html

19
Hormones of Spermatogenesis
  • Testosterone
  • The Interstitial cells in each testis function as
    an endocrine gland. Its principal hormone,
    testosterone, is responsible for
  • Sperm production.
  • Secondary sex characteristics of men.
  • The Interstitial Cells of Leydig lie between the
    seminiferous tubules.

20
LH
  • Interstitial cells are, in turn, the targets for
    a hormone often called interstitial cell
    stimulating hormone (ICSH), also called the
    luteinizing hormone (LH).
  • Prolactin
  • From the A. Pituitary, up-regulates LH receptors
    expression on the ICL thus stimulates
    Testosterone production.

21
Interstitial Cells of Leydig (7)
  • LH, from the Anterior Pituitary Gland, stimulates
    the ICL to secrete the Testosterone.
  • Prolactin, also from the A. Pituitary,
    upregulates the LH receptor expression on the ICL.

22
FSH
  • Follicle-stimulating hormone (named for its role
    in females, like the LH).
  • Matures the Seminiferous tubules.
  • Acts directly on spermatogonia to stimulate sperm
    production (aided by the LH needed for
    testosterone synthesis).

23
Production of Gamets
  • Oogenesis
  • In contrast to males, the initial steps in eggs
    production occur early prior to the girls birth.
  • Diploid ovarian stem cells called oogonia, that
    arise from the ovarian Germinal Epithelium,
    divide by mitosis to produce more oogonia and
    primary oocytes.
  • The 20 weeks old female fetus already possesses
    all the primary oocytes that she will ever have
    4-7 million eggs.

24
Oogenesis (Cont.)
  • At birth, only 12 millions of the1ry oocytes
    remain, each has begun the first meiotic division
    and has stopped, at prophase I, or the Germinal
    Vesicle stage, GV (1st Arrest).
  • Only at puberty, those primary oocytes resume
    development, usually one or a few, at each
    menstrual cycle.

25
Oogenesis (Cont.)
  • They grow and complete meiosis l, forming a
    larger haploid secondary oocyte and a small polar
    body, each bears one set of chromosomes.
  • In humans (and most vertebrates), first polar
    body degenerates.
  • The secondary oocyte immediately proceeds to
    meiosis II, but again stops at metaphase II (2nd
    Arrest), and known as Mll oocye, i.e. metaphase
    stage of meiosis II.

26
Oogenesis (Cont.)
  • Only if fertilization occurs will meiosis II ever
    be completed.
  • Entry of the sperm triggers the completion of
    meiosis II,
  • where the secondary oocyte ejects the second
    polar body, and becomes a fertilized egg with 2
    pronuclei, its own and the sperms.
  • The 2 pronuclei fuse in one at the zygot stage

27
Oogenesis (cont.)
  • Egg maturation to the MII stage takes place
    within the follicle, a fluid-filled envelope of
    cells surrounding the developing egg.
  • The ripening follicle also serves as an endocrine
    gland. Its cells make a mixture of steroid
    hormones collectively known as Estrogens.
    Estrogens are responsible for the development of
    the secondary sexual characteristics of girls at
    puberty and maintains them thereafter.

28
Female Reproductive System
29
Section of the ovary
  • 1. Germinal epithelium. 2. Central stroma.
  • 3. Peripheral stroma.
  • 4. Bloodvessels.
  • 5. Vesicular follicles in their earliest
    stage.
  • 6, 7, 8. More advanced follicles.
  • 9. An almost mature follicle.
  • 9'. Follicle from which the ovum has escaped.
  • 10. Corpus luteum.

30
Oogenesis a simplified graph for one chromosome
31
Oogenesis
  • Animated oogenesis, and
  • Animated comparison between spermatogenesis and
    oogenesis

32
Ovulation
  • Occurs about two weeks after the onset of
    bleeding in a regular 28-day menstruation cycle.
  • In response to an LH surge, the follicle
    discharges the secondary MII oocyte.
  • The oocyte is swept into the open end of the
    fallopian tube, then it moves slowly along it,
    then down into the uterus.

33
Conception
  • Back again to the sperms!
  • The sperms are in the caudal epididymis
    approximately 64 days after the initiation of
    their spermatogenesis.
  • Sperm viability preservation during storage
    requires
  • Adequate testosterone levels.
  • Maintenance of the normal scrotal temperature,
    36c.

34
Conception
  • Sperm as an ejaculate component
  • The alkaline pH of semen activates the sperms and
    protects them from the relatively high acidic
    environment of the vagina.
  • The human, sperm can be found in the fallopian
    tube 5 minutes after insemination.
  • Of an average of 200 to 300 millions sperm
    deposited into the vagina, only few hundreds
    achieve proximity to the egg.

35
Conception
  • Fertilization starts with sperm capacitation,
    that is characterized by
  • acquiring hyper-motility.
  • binding to the zona pellucida.
  • undergoing the acrosome reaction to penetrate
    thorough into the oocyte.

36
Conception
  • At acrosome reaction, breaking down and merging
    of the plasma membrane and the outer acrosomal
    membrane take place, so the acrosin enzyme
    digests the zona to let the sperm head contents
    only into the oocyte.

37
Conception
  • The Egg and its Environment
  • The oocyte, at the time of ovulation, is
    surrounded by the sticky granulosa cells (the
    cumulus oophorus).
  • The zona pellucida, a none-cellular porous layer
    of glycoproteins (secreted by the oocyte),
    separates and protects the fragile oocyte from
    the surrounding environment.

38
Conception
  • Approximately 72 days are required to produce
    spermatozoa, a time period followed by storage in
    the epididymis prior to ejaculation.
  • Sperm enter the cervical mucus and then the
    fallopian tubes within minutes, but only a few
    hundred sperm or less reach the oocyte. The
    cervix serves as a reservoir of sperm for up to
    72 hours.
  • Capacitation, a process initiated during the
    sperm's passage through the cervix or during in
    vitro incubation in an appropriate medium, is
    characterized by the acquired ability of sperm to
    undergo the acrosome reaction to bind to the zona
    pellucida and to acquire hyperactivated motility.
  • The acrosome reaction is due to the modification
    and breakdown, followed by a merger, of the sperm
    cell membrane and the outer acrosomal membrane,
    allowing the release of enzymes and changes in
    the inner acrosomal membrane, necessary for
    fusion with the oocyte cell membrane.

39
Conception
  • The fimbriae at the end of the fallopian tubes
    sweep the ovaries surfaces and pick up the egg
    once it is out of its follicle.
  • The egg spends about 80 hours in the fallopian
    tube, 90 of which is at the junction of the
    ampulla and the isthmus.
  • It is in this location that fertilization and
    dispersion of the cumulus cells are completed.

40
Conception
The Fimbriae always scans the ovary surface for
any discharged mature eggs.
41
Conception
  • If fertilization is to happen, then it is a few
    minutes for the ovum sperm to meet! The ovum
    however can keep its readiness for about half a
    day then it starts to degenerate.
  • Within 23 minutes after ovulation, the oocyte is
    in the ampulla of the fallopian tube awaiting the
    sperms, that arrive within 5 minutes of their
    deposition into the cervix.
  • Tubal transport of the egg/embryo depends on the
    circular smooth muscles contractions and the
    cilia-induced flow.

42
Conception
  • The exact fertilizable life of the human mature
    oocyte is unknown, but the most estimated range
    is between 12 to 24 hours, at the most.
  • Detailed steps of fertilization
  • Cumulus oophorus expansion that helps
  • increase the chances of an encounter with sperms.
  • b. facilitate sperm passage through the cumulus
    cells, which is driven by its hyper-motility.

43
  • 2. The acellular zona pellucida has three major
    functions in the fertilization process
  • a. Activates the Sperm-Ligands (mostly species
    specific) that bind to the sperm.
  • b. Responds to the Acrosin to let the sperm cell
    into the oocyte cytoplasm.
  • c. Inactivate its ligands to prevent poly-spermia.

44
Finally they have met!
45
Conception
46
Conception
  • 3. Meiosis II resumes and is completed,
    approximately 3 hours after sperm cell
    penetration.
  • The 2nd polar body is released and leaves the egg
    with a haploid complement of chromosomes. But,
  • the addition of a chromosome-set from the sperm
    restores chromosome diploid number in the
    fertilized egg, that will start cleaving.

47
Embryo Cleaving Stages
48
Embryo stages From day-2 after insemination
until D-5 ( D-6, or D-7 in some cases). Day-1 is
the two-pronuclei stage and the one-cell zygote
stage (later on the day).
49
Female Reproductive Hormones, Ovulation, and
Conception in Humans
50
  • Offspring Genotypes possibilities are 50 to
    50 male to female.
  • However, Phenotype possibilities are just
    unlimited.
  • As Crossovers and hopefully balanced
    Translocations are also probable.

51
  • Sperm penetration of the zona pellucida depends
    on a combination of sperm motility, an acrosomal
    proteinase, and binding of sperm head receptors
    to zona ligands.
  • Binding of sperm head receptors and zona ligands
    produces an enzyme complex that induces the
    acrosome reaction, releasing enzymes essential
    for the fusion of the sperm and oocyte membranes.
  • Fusion of the sperm and oocyte membranes triggers
    the cortical reaction, the release of substances
    from the cortical granules, organelles just below
    the egg cell membrane.
  • The cortical reaction leads to the enzyme-induced
    zona reaction, the hardening of the zona and the
    inactivation of ligands for sperm receptors,
    producing an obstacle to polyspermy.
  • Cell division begins promptly after
    fertilization human gene expression begins
    between the 4- and 8-cell stages.

52
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53
Implantation
  • By definition, implantation is the process by
    which a blastocyst stage embryo
  • hatches out, in 1-2 days of its arrival to the
    uterine cavity,
  • attaches itself to the uterine wall,
  • 3. penetrates its epithelium, and gradually
  • 4. integrates its blood circulation with the
    mothers circulation, through the placenta.

54
Implantation
  • Endometrium Readiness Receptivity
  • Very critical for conception.
  • Normal endometrium is 1014 mm thick at
    implantation time, in mid-luteal phase.
  • By then, it has reached its maximum secretory
    activity, and has become rich in glycogen and
    lipids.

55
Implantation
  • The window of endometrial receptivity is much
    preferred at days 1620 of a 28-day menstrual
    cycle.
  • The blastocyst loosely adheres to the endometrial
    epithelium, a process called apposition, which
    most commonly occurs on the endometrium of the
    upper posterior wall of the uterus.

56
Implantation
  • The window of endometrial receptivity period is
    restricted to days 1620 of a 28-day normal
    natural cycle.
  • Endometrial receptivity is heralded by the
    progesterone-induced pinopodes formation that is
    the surface epithelial cells that lose the
    microvilli and develop smooth protrusions.

57
Implantation
  • The blastocyst loosely adheres to the endometrial
    epithelium, a process called apposition stage,
    which most commonly occurs on the endometrium of
    the upper posterior wall of the uterus.
  • Cytokines, growth factors, and their receptors
    have been identified in virtually all tissues
    associated with implantation.

58
Implantation
  • Possible types of interaction between implanting
    trophoblast and uterine epithelium
  • trophoblast cells intrude the uterine lining on
    their thorough path to the basement membrane.
  • endometrial lining lift itself off, to allow the
    trophoblast to intimate itself underneath the
    epithelium.
  • fusion of the trophoblast with uterine lining, on
    its way towards the basement membrane.

59
Stages of Embryo Implanta-tion within the
Endomet-rium
60
Implantation
  • In the second week after ovulation, the placenta
    is formed.
  • By this time, the trophoblasts at the
    implantation site have formed masses of
    cytotrophoblasts and syncytiotrophoblasts, and
    integration with maternal blood circulation has
    begun.
  • The walls of the spiral arteries are destroyed,
    as sinusoidal sacs are formed, lined with
    endovascular trophoblast.

61
Implantation
  • Goal Achieved!
  • The purpose of placental invasion is to remodel
    the uterine tissues and vasculature, establishing
    a structure that would allow and maintain a
    mother-fetus interchange, enough to sustain the
    fetus, until it becomes a baby ready to get out.
  • !! ?????? ????

62
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63
Introduction to In Vitro Fertilization or IVF
  • What is infertility?
  • Whose fault is it?!
  • It is a disease not a shame.
  • How to find out the reason?
  • How to propose the treatment?
  • Treatment ranges from 1) just counseling, 2) IUI
    with natural cycle, 3) IUI with ovulation
    stimulation, 4) conventional IVF, 5) IVF/ICSI,
    till finally, 6) IVF/ICSI/ PGD.
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