Embryonic Stem Cells and Therapeutic Cloning

Biology 3201

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• Password: NLbiol

Somatic Cell Reproduction

• A somatic cell is any cell in a multicellular organism excluding gametes (sperm and egg)
• Examples of somatic cells include: skin cells, liver cells, and guard cells.
• Eukaryotic somatic cells reproduce by mitosis.
• Mitosis is the division of genetic material and the cell's nucleus.

Genetic Material Revisited

• Chromosomes are duplicated chromosomes with two chromatids held together by a centromere.
• Chromatin is made up of DNA and proteins.
• DNA is found in the nucleus of a eukaryotic cell.

Terms

• DNA: a double-stranded molecule that governs heredity.
• Chromatin: the uncondensed form of genetic material made of DNA and proteins.

Chromosome

• Chromosomes are well-packaged DNA, necessary for transport during mitosis.
• Humans normally have 23 pairs of chromosomes, one set from each parent.
• 22 pairs are called autosomes containing DNA that codes for various functions.
• One pair are sex chromosomes that carry important information to determine sex.

Human Karyotypes

• Male karyotypes display an X and Y chromosome.
• Female karyotypes display two X chromosomes.

The Cell Cycle

• Cell cycle: A continuous series of cell growth and division for a cell.
• The cell cycle consists of two stages: Growth Phase (Interphase), and Division Phase (mitosis and cytokinesis).

Growth Phase (Interphase)

• Also known as Interphase.
• Consists of three segments:
  • G1 or Gap 1 phase: Cell undergoes rapid growth and normal metabolic activities
  • S Phase: The cell replicates (duplicates) the DNA.
  • G2 or Gap 2 Phase: The centrioles replicate in preparation for cell division

Division Phase

• This is the stage where the cell divides into two new cells (daughter cells).
• Consists of two Stages
  • Mitosis: The process of dividing the genetic material and the nucleus.
  • Cytokinesis: The process of dividing the cytoplasm

Purpose of Mitosis

• To ensure genetic continuity by creating a copy of the DNA, maintaining the same number of chromosomes from one cell to the next.
• To create new cells for growth and repair.
• Mitosis occurs in somatic cells (Ex: skin cells, muscle cells, bone cells), but not the gonads (ovaries and testes).

Stages of Mitosis

• Prophase

  • Chromatin condenses to reveal Double Stranded (DS) chromosomes.
  • Centrioles begin to move to the opposite ends of the cell.
  • The nuclear membrane breaks down and disappears.
  • The centrioles grow filaments called spindles.

• Metaphase

  • Chromosomes line up at the middle of the cell.
  • Spindles attach to the chromosomes at the Centromeres.

• Anaphase

  • The Spindles pull the chromosomes apart and to the opposite ends of the cell.
  • There are now Single Stranded (SS) chromosomes at each end of the cell.

• Telophase

  • The chromosomes uncoil and become chromatin again.
  • The nuclear membrane begins to form around each set of chromosomes.
  • The Spindle fibres disappear.
  • Cytokinesis begins. The cytoplasm is divided equally between the two new daughter cells. In animal cells the cell membrane pinches together in the middle; in plant cells a cell plate is formed that eventually closes off from each other.

Meiosis

• Meiosis: A type of cell division that results in gametes (sperm and egg) being created with half the number of chromosomes (haploid – n) as the parent cell.
• Sperm and egg must have only half the chromosome number in order for a fertilized egg (zygote) to develop normally into the proper diploid (2n) organisms.
• If sperm and egg were created by mitosis, there would be too many chromosomes in the resulting zygote.

Purpose of Meiosis

• To ensure genetic continuity by creating the correct chromosome number for the offspring.
• To create genetic variability (diversity) through independent assortment and crossing over.

Stages of Meiosis I

•  Prophase I: Homologous chromosomes line up next to each other as pairs (tetrads). Crossing over of non-sister chromatids occurs when synapsis of homologous chromosomes occurs.
• Metaphase I: Homologous chromosomes pair up at the middle of the cell. The spindle from one end of the cell attaches to one pair of sister chromatids while a spindle from the other end attaches to the other pair of sister chromatids.
• Anaphase I: the spindle pulls homologous chromosomes to opposite ends of the cell.
• Telophase I: The nuclear membrane reappears around each set of separated chromosomes. The spindle disappears. Cytokinesis (division of the cytoplasm) occurs.

Stages of Meiosis II

• Prophase II: Nuclear membrane disappears. Spindle forms. Chromosomes become visible.
• Metaphase II: Chromosomes line up at the middle of the cell. Spindle attaches to centromere of each chromosome.
• Anaphase II: Spindle pulls one chromatid from each chromosome to opposite ends of the cell.
• Telophase II: Nuclear membrane reappears around each group of chromatids (now called SS chromosomes). The spindle disappears. Cytokinesis occurs (division of the cytoplasm). Two new cells are created each having the haploid number of chromosomes.

Oogenesis (Egg Formation)

• A diploid (2n) cell called an oogonium undergoes meiosis in the ovaries of females.
• Before meiosis begins, the oogonium replicates (double) its chromosomes so that it has 46 DS chromosomes (or 92 sister chromatids).
• During the first meiotic division, the homologous chromosomes separate, and cytokinesis occurs unevenly, creating a large cell called a primary oocyte and a small cell called a polar body.
• The primary oocyte has 23 DS chromosomes.
• During the second meiotic division, the resultant cells divide again, but once again, cytokinesis is unequal.
• The result is a single haploid (n) ovum (egg) and multiple polar bodies.
• Only one mature egg is formed. The three polar bodies die because they do not have enough cytoplasm to support them.

Spermatogenesis (Sperm Formation)

•  A diploid (2n) cell called a spermatogonium undergoes meiosis in the testes of males.
• Before meiosis begins, the spermatogonium replicates (double) its chromosomes so that it has 46 DS chromosomes (or 92 sister chromatids).
• The first meiotic division produces two new cells each with 23 DS chromosomes.
• The second meiotic division creates four haploid sperm cells each with 23 SS chromosomes.
• The four spermatids differentiate into mature spermatozoa or sperm.

Comparing Sperm and Egg

• Physical structure: Sperm are small with a head and tail; eggs are large non-motile cells.
• Size: Sperm are smaller in size than eggs (thousands of times smaller).
• Energy reserves: Sperm rely on fructose; eggs use reserves in the oocyte.
• Mitochondria: Sperm have mitochondria; egg mitochondria are depleted in the three polar bodies.
• Number produced: Billions of motile sperm are produced continuously; a female produces one non-motile egg approximately per cycle.

Reproductive Strategies

• Two fundamental types of reproduction are sexual and asexual reproduction.

Sexual Reproduction

• Involves the fusion of haploid (n) nuclei from two parents forming a diploid (2n) zygote.
• This process of sperm nuclei meeting with the egg nucleus.
• Offspring resemble the parents but are not identical to them.

Asexual Reproduction

• Involves the making of genetically identical copies (clones) of offspring.
• It occurs through mitotic divisions.
• There are several different methods of reproduction (modes) that occur, for example, vegetative reproduction, budding, binary fission, spore production and fragmentation.

Budding

• Outgrowth that occurs on an organism and later breaks off and grows into new organisms.
• Examples include: yeast, and hydra

Binary Fission

• DNA is divided through mitosis.
• The organism then splits to create two complete new organisms.
• Examples include: bacteria.

Spore Production

• Mitotic cell divisions create a series of spores.
• Spores grow into new organisms.
• Examples of spore production include fungi (i.e. rhizopus or bread mold).

Fragmentation

• Piece of an organism breaks off that later grows into a full organism
• Examples: cuttings or slips of plants and the regrowth of pieces on organisms such as planaria and starfish.

Parthenogenesis

• An organism produces haploid (n) unfertilized eggs that grow into adults.
• Examples: insects (balsam wooly aphid, wasps); whiptail lizards

Vegetative Reproduction

• Produce new plants from vegetative parts of the original plant, such as leaves, stems, and roots.
• Examples include strawberry runners.

Viral Reproduction

• All viruses are composed of strands of genetic material (RNA or DNA) surrounded by a protein coat (capsid).
• Viruses containing RNA are called Retroviruses.
• Viruses act as parasites. They rely on host cells for survival.

Viral Lifecycle (Lytic Cycle)

• A short cycle about 30 minutes.
• The virus injects its DNA/RNA into a host cell, taking over cell machinery.
• New virus cells are produced.
• The host cell is destroyed.
• Examples include: T4 Bacteriophages

Viral Lifecycle (Lysogenic/Provirus Cycle)

• Viral DNA/RNA enters a cell and becomes part of the cell's DNA.
• Called a provirus when this occurs.
• Viral DNA is reproduced with cell DNA.
• The virus can exist in this form for years without harming the host cell.
• When active, the provirus separates from the host cell DNA and undergoes the Lytic Cycle.
• The host cell is NOT usually destroyed.
• Examples include: HIV and Herpes.

Reproduction in Mosses - Alternation of Generations

• There is a switching between the gametophyte (n) and sporophyte (2n) generations during the life cycle of an organism.
• Within the antheridium of a male moss, sperm (n) are created.
• Sperm swim from the antheridium to the archegonium of a female moss containing an egg(n).
• Fertilization occurs.
• A zygote (2n) forms.
• The sporophyte generation (2n) now becomes dominant.
• The sporangiophore and sporangium grow from the fertilized female plant.
• Spores undergo meiosis inside the sporangium and become haploid (n) spores.
• Spores are released into the environment by wind, water, etc., where they grow into new moss plants

Reproduction in Cnidarians (e.g., jellyfish) - Alternation of Generations

• There is a switching between sexual and asexual reproduction in Cnidarians.
• The polyp stage can undergo asexual reproduction (budding); the medusa stage can undergo sexual reproduction.

Reproduction in Flowering Plants

• These plants are known as angiosperms (covered seed plants).
• These plants undergo sexual reproduction.
• These plants have evolved to be independent of water for reproduction; they use structures in flowers.
• Perfect flowers contain both male and female structures, while imperfect flowers only contain either male or female structures.

Reproductive Structures of a Flowering Plant (Pistil)

• The female reproductive structure of a flower.
• Made up of the stigma, style, and ovary.
• Stigma: sticky portion on top of the style to trap pollen.
• Style: slender tube leading from the stigma to the ovary (passageway to ovary).
• Ovary: contains the eggs (haploid n) formed through meiosis in the ovule.

Reproductive Structures of a Flowering Plant (Stamen)

• The male reproductive structure of a flower made up of the anther and filament.
• Anther: reproductive structure of a flower that produces the pollen. Pollen is created through Meiosis.
• Creates pollen (male sex cells) for fertilization.
• Filament: slender stalk that holds up the anther.

Creation of Egg an Pollen within Flowering Plants

• The diploid (2n) cell within the pollen sac of the anther undergoes meiosis to produce four haploid (n) cells called microspores.
• Each microspore undergoes mitosis to create two haploid nuclei known as the tube nucleus and the generative nucleus.
• The outer wall of the microspore hardens and is called a pollen grain.
• Within the ovule of the flower, a diploid (2n) cell called the megasporongium undergoes meiosis to produce four haploid (n) cells.
• Three of the cells die; the remaining cell (megaspore) undergoes mitosis three times to create a total of eight haploid structures in the ovule. One of these becomes the egg.

Sexual Reproduction/Fertilization in the Flowering Plants

• Pollen grains reach the stigma and stick to the style.
• The tube nucleus begins to travel through the style toward the ovary.
• The generative nucleus divides by mitosis into a generative and tube nucleus.
• Sperm nuclei make their way down through the pollen tube and into the ovule.
• One of the sperm fertilises the egg and produces a diploid (2n) zygote.
• The other sperm joins with two polar nuclei to form a triploid (3n) structure known as endosperm.
• The ovule becomes the seed and eventually a new plant. The endosperm provides food for the developing seed.

Male Reproductive System

• Function: To create and deliver sperm to an egg (ovum) for the purposes of fertilization.

• Scrotum: Sac that holds testicles and helps maintain proper temperature for sperm production.

• Testicles: Two oval-shaped structures that create sperm (haploid n).

• Epididymis: Oval structure on top of the testis. It stores sperm and where sperm become motile.

• Vas Deferens: Hollow tube leading from the testis to the urethra. It carries sperm from the testes to the penis.

• Cowper's Gland: Adds an alkaline (basic) fluid to sperm to neutralize the acidity of the female reproductive tract.

• Prostate Gland: Adds fluid to sperm

• Seminal Vesicles: Glands that secrete a mucus-like fluid with the sperm, containing fructose for energy.

• Urethra: Opening through which sperm and urine exit the male body.

• Penis: The male reproductive organ.

• Seminiferous Tubules: Tubules in the testes where sperm are produced.

• Interstitial Cells: Cells around seminiferous tubules that release testosterone.

• Semen: Male reproductive fluid containing sperm and other fluids.

Spermatogenesis

• Process by which sperm are produced in the testes.
• Starts with primordial germ cells in the testes.
• Through various stages of division (mitotic) cells differentiate, producing spermatozoa.

Formation of Sperm Within a Male

• Spermatogenesis is controlled by hormones within the male.
• FSH (Follicle Stimulating Hormone): Hormone released by the anterior pituitary that causes spermatogenesis to begin.
• Inhibin: Hormone released by seminiferous tubules that signals to the hypothalamus to slow the production of releasing factors that control the release of FSH, keeping FSH and therefore sperm production in balance.

Other Male Reproductive Hormones

• LH (Luteinizing Hormone): Hormone released by the pituitary that stimulates the interstitial cells of the testicles to produce testosterone.
•  Testosterone: Major male hormone. Responsible for the development of secondary sex characteristics in a male (e.g., enlargement of penis and testicles; facial hair; lower voice/ increased muscle mass).

Female Reproductive System

• Function: To create ova and after fertilization, to provide a place of development for the fetus.
• Vagina: Opening
• Cervix: small opening at the top of the vagina and the base of the uterus to support the fetus and prevent foreign material from entering the uterus.
• Uterus: Large muscular chamber where the embryo attaches and the fetus develops.
• Fallopian Tubes (Oviducts): Tubes that lead from the ovaries (not attached) to the uterus; fertilization usually occurs in the fallopian tubes.
• Ovaries: Creates ova (eggs). Each egg contains the haploid (n) chromosome number
• Fimbriae: Finger-like projections on the opening of the oviduct that help sweep eggs from the ovary into the oviduct.
• Endometrium: The lining of the uterus. it contains rich blood supply to provide nutrients for a developing embryo.

The Female Menstrual Cycle

• The female menstrual cycle normally follows a 28-day pattern that functions in the release of a mature egg for the purposes of fertilization. The cycle can be from 20 – 45 days long.
• The cycle can be divided into stages.
  • Follicle Stage: Hormone called FSH causes the developing follicle in the ovary to produce estrogen causing uterine lining to build up in preparation for a pregnancy. This stage lasts from day 1 to 14 (approximately).
  • Ovulation : Day 14 hormone called LH is released from the pituitary gland causing the follicle to break open, to release the mature egg (ovulation).
  • Luteal Stage: Days 15 to 28 the left-over follicle (or corpus luteum) releases progesterone that keeps the uterine lining prepared for pregnancy
  • Menstruation: If no egg is fertilized, progesterone and estrogen fall which causes the uterine lining to be “shed” and passes through the vagina as blood-known as menstruation. New follicles mature, and the cycle repeats.

Sexually Transmitted Infections (STIs)

• Diseases transmitted mainly through sexual contact.
• Causes: Most STIs are caused by either bacteria or viruses.

HIV and AIDS

• AIDS: viral infection, attacks helper T-cells and compromises the immune response.
• Transmission: sexual contact with infected person; sharing needles/ breast feeding
• Cure: no cure. Many drugs to control the symptoms are available.

Chlamydia

• Bacterial infection
• Symptoms: Pain when urinating, discharge from penis, vaginal discharge, and fever.
• Transmission: Sexual contact with an infected person.
• Cure: Antibiotics

Hepatitis

• Viral infection that affects the liver.
• Symptoms: flu-like symptoms, jaundice, liver failure, and liver disease.
• Transmission: Sexual contact with an infected person.
• Cure: Vaccine

Genital Herpes

• Viral Infection;
• Symptoms: tingling or itching; blisters on genitals, buttocks, thighs or internal tissues; painful sores.
• Transmission: Sexual contact with an infected person.
• Cure: No cure; treatment with antiviral medications for symptoms.

Syphilis

• Bacterial STI;
• Symptoms/Action: Infection proceeds through three stages
• Stage 1: sores appear at infection site.
• Stage 2: rash appears on skin.
• Stage 3: Cardiovascular and nervous system becomes damaged resulting in mental disorders and/or heart disease.
• Transmission: Sexual contact with an infected person.
• Cure: Antibiotics

Gonorrhea

• Bacterial infection;
• Symptoms: Pain when urinating, discharge from penis, and vaginal discharge.
• Transmission: Sexual contact with an infected person.
• Cure: Antibiotics.

Human Papillomavirus (HPV)

• Viral infection;
• There are over 100 varieties of HPV;
• Some varieties produce warts.
• Some varieties cause different types of cancer.
• Symptoms: Warts forming on different parts of the body; May lead to different types of cancer
• Transmission: Sexual contact or skin-to-skin contact
• Cure: Vaccines

Prenatal Screening

• Home Pregnancy Tests: Checking urine for human chorionic gonadotropin (hCG), a hormone released from a fertilized egg attached to the uterine wall
• Maternal Blood Tests: Shows how much human chorionic gonadotropin (HCG) is in the blood.
• Ultrasound: Use of sound waves beyond human hearing, to create a cross-sectional image of developing fetus.
• Fetoscopy: Direct observation of fetus through insertion of an endoscope into a small incision in the mother's abdomen.
• Amniocenthesis: Sample of amniotic fluid drawn to karyotype cells in fluid.
• Chorionic Villi Sampling; sample of chorionic villi is drawn for karyotyping

Reproductive Technologies

• Sterile: Men and women who are unable to have any children.
• Infertile: Men and women who have difficulty conceiving; usually at least a year of attempting pregnancy is needed to be considered infertile.
•  Causes of Male Infertility: Obstruction, low sperm count, inability to achieve erection or ejaculation, and high proportion of abnormal sperm.
• Causes of Female Infertility: Blocked oviducts; failure to ovulate; and endometriosis/damaged eggs.

Infertility Treatments

• Artificial Insemination: Sperm are concentrated, and placed in the vagina.
• In Vitro Fertilization (IVF): Stimulation of the ovaries and egg collection from the follicles.
• Surrogate Mothers: Another woman carries and delivers the child for an infertile couple. These couples use Al or IVF procedures.
• Superovulation: Hormone treatment to produce multiple eggs.
•  Cryopreservation: Preservation of oocytes, semen, and/or zygotes by freezing.

Reproductive Technologies That Reduce Reproductive Potential

• Methods to prevent conception; known as contraception or birth control.
  • Surgical sterilization: Tubal ligation (female) for women and vasectomy (male) for men; tying off the fallopian tubes or ductus deferens respectively.
  • Hormonal Contraception: Methods that alter the balance of reproductive hormones to inhibit implantation, ensuring that ovulation does not occur (e.g., birth control pills, vaginal ring, patch, intrauterine device, injectable).
  • Physical or Chemical Barriers: methods that physically block the sperm from fertilizing the egg/change environment, (e.g., condoms, diaphragm, cervical cap, spermicides, sponge).
  • Natural Methods: Rely on behavioral practices of individuals, including abstinence from intercourse, withdrawal method, and fertility awareness.

Stem Cells

• In animals, a cell that can differentiate into many different cell types is called a stem cell
• Stem cells are undifferentiated (unspecialized) cells having not yet begun to develop into specialized, differentiated body cells.
• They have two important features: they can divide and create copies of themselves; and, under laboratory conditions, they can be used to differentiate into other body cells.

Types of Stem Cells

• Embryonic stem cells: Derived from embryos after 4 or 5 days after fertilization; usually obtained as a by-product of the in vitro fertilization procedure. They are often pluripotent, meaning that they can develop into many types of cells in the human body.
• Adult stem cells: Found in humans of all ages, many different organs and tissues. They are often multipotent - that they are able to differentiate into only a range of cells within that specific cell type. (e.g. stem cells in bone marrows can differentiate into white blood cells, red blood cells or platelets)

Reproductive Cloning

• Organisms that are genetically identical are said to be clones of one another.
• Using the cells of mouse embryos, researchers were successful in producing cloned mice.
• Cloning of a sheep (Dolly) was accomplished in 1997 by using a cell from an adult donor.

Therapeutic Cloning

• Somatic cell nuclear transfer (SCNT)
• A scientist extracts the nucleus from an egg (a donor oocyte); This nucleus holds the genetic material.
• The nucleus of a somatic cell from a patient is taken and then extracted.
• This nucleus of the somatic cell is inserted in the enucleated oocyte.
• It is stimulated to divide and shortly thereafter forms a cluster of embryonic stem cells.
• The embryonic stem cells can be derived from this cluster of cells of the inner cell mass that are pluripotent.

Ethical Issues of Therapeutic Cloning

• Arguments for therapeutic cloning generally revolve around using cloned embryonic cells to treat diseases
• Arguments against the procedure often include whether it potentially leads to reproductive cloning or if it involves creation and destruction of human embryos.
• Whether or not the potential of the cells to become tumours should be factored in.