Reproductive Toxicology

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Reproductive Toxicology
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Question

• What was the 1st agent recognized to cause birth
  defects in an infant resulting from a mothers
  illness?
• Rubella

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What are At-Risk Populations?
Fetuses
Normal?
Neonates
Elderly
Healthy Adults
Immunocompromised or Diseased State
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I. Introduction

• Function Perpetuation of species
• Chemicals 50,000-60,000 chemicals in common
  use gt600 new chemicals per year
• Potential problems workplace
• endocrine-disrupting chemicals (DDT) affect
  wildlife, humans?
• environmental anti-estrogens (TCDD)

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II. Examples

• A. Lead spontaneous abortion, sperm
  abnormalities
• B. 1,2-dibromo-3-chloropropane male sterility
• C. Organic solvents low sperm count, abnormal
  sperm

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Reproductive functions susceptible to chemicals

• Endocrine functions
• Oogenesis
• Spermatogenesis
• Fertilization
• Implantation
• Early stages of development?

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III. Extent of problem

• A. Difficult to assess
• B. Estimated 1 in 5 couples involuntarily sterile
• C. Estimated over 1/3 of early embryos die
• D. About 15 of recognized pregnancies die
• (Cont)

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(Cont)

• E. for surviving fetuses
• about 3 have some developmental defect
• with increasing age about 2x that number become
  detectable.

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F. Why so difficult to assess?

• 1. Complexity
• 2. Unreliable lab test
• 3.Quality of human data
• What is normal?
• What is abnormal?

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IV. How do chemical affect general reproductive
biology?

• A. May alter
• production of gametes directly
• Production of gametes indirectly
• or interfere with process of fertilization.

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B. Sexual differentiation

• 1. Y chromosome influences male
• A. Mullerian inhibiting factor which suppresses
  development of Mullerian ducts.
• B. Testosterone
• 2. X or no Y female
• (cont)

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(Cont)

• 3. Nondisjunction
• Klinefelters syndrome XXY, male morphology
• Turners syndrome XO, female morphology
• Hermaphroditism XY/XX or XY/XO secondary sex
  characteristics different from genotype
• Chemicals anticancer drugs and radiation

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C. Gonadal Function 2 functions

• A. Endocrine
• 1. Ovary estrogens and progesterone
• 2. Testes testosterone, dihydrotestosterone and
  small amount of estrogen
• B. Production of gametes
• meiosis

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Barriers to Reproductive System

• There is a blood-testis barrier
• There is no blood-ovarian barrier

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Meiosis

• Many places in cell division to disrupt process
• Many antineoplastic agents target dividing
  cells, thus are reproductive toxicants.

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D. Spermatogenesis

• A. Sperm smallest human cell
• B. 2 types of spermatogonia figure x testes
• 1) produces other spermatogonia (diploid)
• 2) produces sperm (haploid) see figure
• 3) beginning at puberty, millions formed each day
• A) animals generally produce many more sperm/day
  than humans
• B) about 7 million/day/gm in young man after 50
  yrs drops to about ½
• (Cont)

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(Cont)

• C. Sertoli cells
• A. Form blood-testis barrier
• B. Secrete several proteins essential to
  spermatogenesis
• C. Leydig cells synthesize testosterone

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Casarett Doulls Toxicology
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E. Efferent ducts and epididymides

• A. Immature sperm subjected to changing chemical
  environment as they move through the ducts and
  epididymis
• B. Mature

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F. Ovarian Function

• 1. Oogenesis
• A. Between 300,000 and 400,000/ovary at birth
• B. Continuously die (atresia)
• C. Reduced to appr ½ by puberty
• D. Appr 25,000 by 30 yrs
• E. Depletion in the pool can lead to infertility
• F. About 400 primary follicles yield mature ova
• G. Remain in primary follicle stage until puberty
• H. Primary oocyte undergoes 2 divisions, one just
  before ovulation and the other after sperm fuses
  with egg

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G. Endocrinology of menstrual cycle (see figure)

• A. Estrogen
• B. Progesterone
• C. LH (luteinizing hormone) produced by
  pituitary
• D. FSH (follicle stimulating hormone) produced
  by pituitary
• (Cont)

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(Cont)

• E. Other animals
• 1) primates similar to humans
• 2) rodents estrus cycle and receptive to male
  only at specific time
• 3) rabbits ovulate as a reflex to copulation

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H. Hypothalamo-Pituitary-Gonadal Axis

• A. GnRH Gonadotropin releasing hormone ( or
  LHRH or FSHRH)
• B. Feedback mechanisms between testosterone or
  estrogen and the brain (decrease testosterone or
  estrogen increase LH and FSH)

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V. Toxicology of Reproductive System

• A. Barriers for the reproductive system
• 1. Ovary (?)
• 2. Blood-testes barrier figure
• 3. Placenta mostly sieve
• B. Biotransformation and metabolism
• 1. Testes cytochrome P450
• 2. Ovary not as well studied complex hormonal
  involvement cytochrome systems and mixed
  function oxidases

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IV. Evaluating Reproductive Function
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A. Males

• 1) Semen characteristics sperm count motility,
  semen volume, and morphology.
• 2) Lab animals produce more sperm than needed
  and may not be sensitive indicator
• 3) Humans may be reluctant to participate in
  study or not meet study design
• (Cont)

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(Cont)

• 4) In vitro oocyte penetration of zona-free
  hamster ova and human sperm only functional
  measure for sperm-egg interactions
• 5) Biochemical measures of enzymes in acrosome
  of sperm, but these tests are under development
• (Cont)

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(Cont)

• 6) Tests using staining to detect DNA damage,
  but not validated as predictive
• 7) Endocrine profile tests to determine blood
  levels of pituitary hormones and testicular
  hormones. Presently indicate extreme levels
  only.
• (Cont)

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(Cont)

• 8) In vivo lab animals as indicators, but not
  necessarily same mechanism of action
• 9) Difficult to see in human studies due to low
  normal conception rate in humans

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B. Females

• 1) Oocyte and follicular toxicity oocyte and
  follicles are counted after exposure to agent
• 2) Reduction in oocytes leads infertility
• 3) Endocrine profiles

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C. For epidemiology study

• Measures of exposure should include dose
  (several), route of exposure, duration, time of
  exposure relative to conception and health
  outcome.

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D. Mechanisms of Action

• 1) normally dependent on neuroendocrine system,
  gonads, accessory organs.
• 2) receptor mechanisms many toxicants act as
  endogenous reproductive hormones which act
  through a membrane receptor or intracellular
  receptor (Ex estrogen xenobiotics such as DDT
  and Kepone). Bind to estrogen receptors. If
  binding occurs may act as agonist or antagonist
  (Tamoxifen)

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E. Hypothalamic-Pituitary Mechanisms

• (LH, FSH, Prolactin) control ovarian function
  and testicular function. Suppression of
  gonadotropin secretion suppresses gonadal
  function.

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F. Inhibition of Steroidogenesis

• Estrogens, progesterone, androstenedione, and
  testosterone are predominant steroids during
  reproductive years.

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Comparisons of potential animal models

• Are the affected organ systems similar?
• Is the disease progression and outcome similar?
• Are the mechanisms causing disease similar?

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Comparison of Reproduction and Development
Human Rhesus Guinea Pig Mice
Estrous Cycle 28 days 28 days 16 days 4-5 days
Gestation 38 weeks 23 weeks 65 days 19-21 days
Placental Shape Discoidal Bidiscoidal Discoidal Discoidal
Placental Membrane Hemo-monochorial Hemo-monochorial Hemo-monochorial Hemo-trichorial
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Comparison of Listeriosis
Humans Rh Monkey Guinea Pigs Mice
-Fetuses -Pregnant adult asymptomatic -abortions, stillbirth, neonatal deaths -pathology - Lm receptor- E. Cadherin -Fetuses -Pregnant adult asymptomatic -abortions, stillbirths, neonatal deaths -pathology ??? -Fetuses -Pregnant adult asymptomatic -abortions, stillbirths, neonatal deaths -Lm receptor E. Cadherin -not susceptible during pregnancy E. cadherin is NOT Lm receptor
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Mechanisms of susceptibility
Human Rhesus monkey Guinea pig Mouse
Oral exposure Oral exposure Oral exposure IV exposure
E-cadherin ? E-cadherin (transgenic mouse?)
Colonizes fetus Colonizes fetus Colonizes fetus ?
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Objective

• To develop the pregnant rhesus monkey as a
  surrogate for human listeriosis
• Obtain dose-response data for L. monocytogenes
  using the pregnant rhesus monkey

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Comparison of Listeria monocytogenes virulence

• In a mouse model for use in risk assessment

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Guinea Pig as a Surrogate Model for Listeriosis
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VI. Reproductive Toxicants
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A. Polycyclic Aromatic Hydrocarbons (PAHs)

• 1) formed in automobile exhaust, smoke
  emissions, cigarette smoke. Produced by fossil
  fuels.
• 2) several PAHs destroy oocytes in weanling and
  mature rats and mice.
• 3) Premature ovarian failure
• (Cont)

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(Cont)

• 4) Smoking produces a dose dependent decrease in
  age of spontaneous menopause.
• Female 1 pack/day 2 years before
  non-smoker
• ½ pack/day 1 year before
  non-smoker
• 5) cigarette smoke and nicotine suggested to
  impair reproduction and fetal development in
  experimental animals and humans. Not known
  whether from PAHs, nicotine, CO or other
  components of cigarette smoke.

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B. Alkylating Agents

• DNA conjugates, used in industry and treatment of
  neoplastic diseases
• 1) ex. 1,2-dibromo-3-chloropropane,
  antispermatic alkylates DNA. Incorrect DNA and
  RNA
• (Cont)

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(Cont)

• 2) ex. Cyclophosphamide mechanism of action
• metabolic activation of parent compound and
• formation of reactive metabolites.
• Detox through conjugation, but cell damage
  requires replacement of damaged macromolecules
  and DNA repair.
• (Cont)

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(Cont)

• Age dependent sensitivity to cyclophosphamide for
  gonadal toxicity between females and males.
• Young females more sensitive to oocyte
  destruction than older females.
• Young males more resistant to toxicity than older
  males.
• Rate of spermatogenesis in prepubertal testis is
  low. In female most sensitive population resting
  or small oocytes.

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C. Solvents

• 1) glycol ethers
• 2) industrial compounds with little or no
  mutagenic activity
• 3) apparently metabolized to active metabolite
  but no evidence for genetic or hormonal
  mechanism of action. MOA not known.
• (Cont)

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(Cont)

• 4) Mechanism studies and risk assessment
• 1) decision on reproductive risk likely made long
  before information available.

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D. Given the present state

• Given the present state of the science of
  reproductive biology and toxicology, it is
  difficult to predict differential species
  susceptibility, even assuming knowledge of
  mechanism of action.