CRS Laboratory Definitions

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JUNIOR ONCOFERTILITY SATURDAY ACADEMY
Module 1 Regulation of Ovarian Function
The Oncofertility Saturday Academy is a
collaborative effort between Northwestern
University and The Young Womens Leadership
Charter School. This research was supported by
the Oncofertility Consortium (NIH 1 UL1
RR024926-01, 1 U54 HD041847, and R25/RCA133836A)
as part of the NIH Roadmap Interdisciplinary
Research Consortia.
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JOSA Module 1 Schedule Saturday February 14,
2009 Chicago Campus
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JOSA Module 1 Faculty
Teresa K. Woodruff, PhD
Susan Barrett, PhD Post-Doctoral Fellow
Miranda Bernhardt PhD Student
Jennifer Hirshfeld-Cytron Medical Fellow
Sarah Kiesewetter Research Technologist
Betty Kong MD/PhD Student
Ariella Shikanov Post-Doctoral Fellow
Jennifer Jozefik Research Technologist
Min Xu, MD, PhD Research Assistant Professor
Tyler Wellington Histologist
Candace Tingen PhD Student
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JOSA Module 1 Guiding Questions and Outcomes
OSA Guiding Question How can we preserve the
fertility of cancer patients?
Learning Outcomes Define oncofertility and how
the field emerged. Identify female reproductive
anatomical structures of different species (e.g.
human, rat, mice, frog, monkeys). Explain the
ovarian cycle (process of follicular development,
ovulation, corpus luteum formation). Describe how
novel strategies (ovarian protection, follicle
culture, cryopreservation, vitrification, ovarian
autografts) can preserve fertility. Define the
structure, source, and function of hormones
involved in the female reproductive system.
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JOSA Module 1 Vocabulary
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Oncofertility a new discipline that makes
connections between oncology and reproductive
medicine, providing viable fertility preservation
options for people with cancer and other
fertility threatening diseases. Reproductive
Science a branch of science that deals with the
mechanism and regulation of reproductive
processes and the diagnosis and treatment of
reproductive disorders Ovarian Vocabulary Egg
(or oocyte) a haploid (possessing half the
number of chromosomes found in other cells in the
body) female reproductive cell or gamete Ovary
female reproductive organ that produces eggs and
the female sex hormones Ovarian Follicle
functional unit of the ovary, contains an egg
surrounded by somatic cells that produce hormones
such as estrogen Zygote diploid cell formed by
the union of sperm and egg the product of
fertilization Gamete haploid sex cell the egg
or a sperm, which join in fertilization to form a
zygote Granulosa Cell one of the
estrogen-secreting cells of the epithelial lining
of a graafian follicle or its follicular
precursor Theca Cells an enveloping sheath of
cells surrounding the granulosa cells of the
follicle, a site of androgen production in
females Vesicular (Graafian or Antral) follicle
a mature liquid-filled follicle within the ovary
that houses a developing egg
Theca Cells
Antral Follicle
Oocyte
Granulosa Cells
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JOSA Module 1 Vocabulary
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Biomaterials Vocabulary Alginate a linear
co-polymer that appears as a viscous gum and is
abundant in the cell walls of brown
algae Bioengineering biological or
medical application of engineering principles or
engineering equipment called also biomedical
engineering Biomaterial a natural or
synthetic material (as a metal or polymer) that
is suitable for introduction into living tissue
especially as part of a medical device (as an
artificial joint) Crosslinking the process of
chemically bonding two or more molecules by a
covalent bond Hydrogel a gel composed of
one or more polymers suspended in
water Polymerization a chemical reaction in
which two or more molecules combine to form
larger molecules that contain repeating
structural units Follicle Culture
Vocabulary Culture the act or process of
cultivating living material in prepared nutrient
media In vitro outside the body and in an
artificial environment Tissue Culture the
process or technique of making tissue grow in a
culture medium outside the organism
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JOSA Module 1 Vocabulary
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Histology Vocabulary Cryostat A cold
temperature chamber containing a microtome used
to prepare thin sections from frozen tissues. The
tissues are often maintained at -20 degrees
Celsius. Embedding The process of placing a
tissue into a preserving material (such as
paraffin wax) that provides a support matrix for
further manipulation of the tissue, such as
sectioning Fixation The preservation of a
tissue sample to retain the structure it had in
the body. Generally uses a chemical fixative,
like formaldehyde. Hematoxylin Eosin (HE)
Staining A commonly used morphologic stain for
tissue sections Histology The study
of cells and tissues at the microscopic level,
branch of anatomy dealing with tissue
structure Microtome A machine designed for
cutting uniform (and often very thin) sections
of a specimen, such as a paraffin-embedded
tissue like an ovary Morphology The form and
structure of an organism or any of its
parts Sectioning Using a microtome or cryostat
to make thin sections of tissue ( 5 microns)
that are placed on microscope slides for further
analysis
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How do researchers perform preliminary studies
for human research?
Human
Mouse
What differences are there between the mouse and
human reproductive tracts? Why do researchers
use mice for research?
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How do researchers perform preliminary studies
for human research?
Fill in the blanks with the following
words Ovary Liver Intestines Kidney Bladder Uter
us Spleen
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How does a scientist obtain ovarian follicles for
a study?
Method Using 2 needles/syringes, like a knife
and fork, isolate individual follicles from the
ovary.
Draw each type of follicle and label the
following cell types Oocyte, Granulosa Cells,
Theca Cells
Secondary Follicle 2 cell layers
Antral Follicle multiple layers visible antrum
Primary Follicle 1 cell layer
Questions to Think About
How many follicles are in a mouse ovary? A human
ovary? Why would a researcher want to grow a
follicle by itself?
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What are biomaterials?

• Alginate Procedure
• Fill 1mL syringe with blue alginate
• Pour 1-2 inches calcium solution into a dish
• Holding syringe 4-6 inches above the calcium
  solution, slowly allow drops of alginate to fall
  into the solution
• Allow alginate droplets to cross-link for 2
  minutes
• Carefully locate an alginate bead and remove it
  from the solution
• Place bead in micro-centrifuge tube containing
  calcium solution to take home

Questions to Think About
What biomaterials can you name? Where does
alginate come from? Where might you find alginate
in your home? Why is alginate a good biomaterial
for ovarian follicle culture?
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How are biomaterials used for in vitro follicle
cultures?
Follicle Encapsulation
50 mM CaCl2
2. Pipette individual follicles into 2-3mL
droplets of alginate on a mesh
1. Mechanically isolate individual follicles
3. Gel beads in 50mM CaCl2 for 2 min before
transferring to 96 well plates for culture
Questions to Think About
What happens when a follicle is cultured in just
a petri dish? What does the alginate provide
that the follicle needs to grow?
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How do researchers look at follicle morphology?
Histology Procedure
3. Sectioning Molded paraffin blocks are sliced
into 4 micrometer sections, floated on a water
bath, and placed on glass microscope slides
4. Staining Tissue slides are deparaffinized
and re-hydrated, then dipped into nuclear and
cytoplasmic stains to show mophology
Questions to Think About
Why would someone need to look at the morphology
of a follicle?
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JOSA Module 1 Wrap-up Discussion
OSA Guiding Question How can we preserve the
fertility of cancer patients? Module Guiding
Question How do we test fertility preservation
methods? Laboratory Questions How do
researchers perform preliminary studies for human
research? How does a scientist obtain ovarian
follicles for a study? What are biomaterials? How
are biomaterials used for in vitro follicle
cultures? How do researchers look at follicle
morphology?