The Cell

Dr. J Leonard Pascual (JPL)

1
1 /9
Castaneda, Catalan, Catubag, Cayabyab, Cengca
1
13 August 2014
The Cell
LECTURE
I. How are specimens prepared for light
microscopy?
II. The Cell
a. Definition
b. Two Basic Compartments
c. Components of the Cell
d. Structure-Function Correlations of the Cell
LABORATORY
I. Staining of Histologic Structures
II. Slides
LECTURELECTURE
I. SPECIMEN PREPARATION
1) Fixation
To preserve tissue morphology and
prevent cell digestion
Use of fixative
A snapshot of what it is
2) Dehydration
To remove cell water (to prevent decay)
Use of ethyl alcohol
3) Clearing
Use of lipid solvent like benzene or
xylene
Tissue becomes transparent
White spots = where fats were once
located
4) Embedding
Baked in paraffin (a white colorless soft
solid) at 58-60 C
Makes tissue more firm for sectioning
5) Sectioning
Use of microtome (cutting tool used to
cut extremely thin slices of material)
Tissue is cut for Light Microscope (LM) :
3-8m & Electron Microscope (EM):
0.02-1 m; section = resolution
6) Staining
Hematoxylin: Basic dye (basophilic)
Purplish Blue
Eosin: Acidic dye (acidophilic) Bright
Pink
a. Staining of Proteins: high carboxyl group
content would be stained with basic dye
(hematoxylin) and those high in amino
group, acidic dye
b. Staining of Nucleic Acids: NAs have
phosphoric acid (strongly acidic) content
which causes varying pH; they are
basophilic (absorbs hematoxylin) at
alkaline, neutral and moderately acid pH
and acidophilic (absorbs eosin) at very
acidic pH
c. Staining of Mucopolysaccharides and
Mucoproteins: they contain sulfate
groups (very acidic), their affinity to dyes
same as nucleic acids
d. Staining of Fats and Carbohydrates: H&E
procedure usually dissolves or does not
stain these substances, thus cannot be
seen properly; if structure is large
enough, it is seen as unstained spot in
cytoplasm
Table 1. Staining Properties
Structure
being
noted
Philicity Dye
Absorbed by
structure
Dye
Color
Acidic Basophilic Hematoxylin Blue or
Purple
Basic Acidophilic Eosin Red or
Pink
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
2 /10
II. The Cell
A. DEFINITION
Cell - the smallest unit of protoplasm capable of an
independent existence
Cell Tissue Organ Organ System
B. TWO BASIC COMPONENTS
1. Nucleus
Repository of the cells genetic material
(DNA)
Tells the cell what to do; How long its
lifespan will be
Parts of the Nucleus:
a. Chromatin
Nucleic acids (Deoxyribonucleic acid, DNA):
genetic material of the nucleus, resides in the
chromosomes
(1) Condensed inactive, stainable;
Heterochromatin
(2) Extended in transcription, not visible with LM;
Euchromatin
Only a fraction of the total complement of units of
heredity (genes) is actively involved, at any given
time, in directing protein synthesis. This portion is in
the euchromatin, whereas the genes that are not
being expressed are in the heterochromatin.
Histone proteins package DNA into
nucleosomes and for gene regulation
Gene sequence of base pairs in the DNA
that contain information for synthesis of
nucleic acid or protein
1) Transcription (nucleus) = from
DNA -> mRNA -> tRNA -> rRNA
(codons)
2) Translation (cytoplasm) = in action
b. Nuclear envelope: barrier, separates nucleus
from cytoplasm
Participates in chromatin organization
Controls movement of macromolecules
between the nucleoplasm and
surrounding cytoplasm
In EM: 2 parallel membranes
separated by perinuclear cisterna (10-
30 nm space)
- Outer membranes: may have
ribosomes, continuous with
rough endoplasmic reticulum
- Inner and outer membranes:
continuous with each other at
nuclear pore complex (intricate
complex of proteins)
Nuclear pore: serve as avenues of communication
between the nucleoplasm and the cytoplasm
c. Nucleoli
Deeply stained by basic dyes due to
ribonucleoprotein
Often surrounded by an intensely
stained rim of nucleololus-associated
chromatin
Usually large in growing cells that are
making large amounts of tissue proteins
d. Nuclear Matrix scaffold/skeleton; 3-
dimensional filamentous protein network
2. Cytoplasm: contains organelles (metabolically
active, essential) and inclusions suspended in
fluid called cytosol; where protein synthesis
occurs
C. COMPONENTS OF THE CELL
1. Organelles metabolically active, essential
a. Endoplasmic Reticulum
Continuous or extensive system of
membrane-bounded canaliculi
Visible in histological sections ONLY if
intact cells in tissue culture are stained
with a lipophilic anionic
In EM, it appears as branching tubular
profiles of varying length; the tubules
may expand into broad flat saccules
called cisternae (fluid-filled cavities)
Functional and Morphological Regional
Differentiation:
(1) Rough Endoplasmic Reticulum (RER)
Contains ribosomes
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
3 /10
(manufacture proteins) on the
outer surface of its limiting
membrane
Most abundant in glandular cells
that secrete protein; antibody-
producing plasma cells; and liver
cells that produce most blood
proteins
(2) Smooth Endoplasmic Reticulum (SER)
Lacks adherent particles
Synthesis of lipids (fatty acids, et.
al)
Mostly found in steroid-secreting
endocrine cells
In liver, it is mostly involved in
synthesis of VLDL (very low-
density lipoprotein) - cholesterol
carrier
Involved in the detoxification and
metabolism of lipid-soluble drugs
Break down stored glycogen to
form free glucose
Sarcoplasmic Reticulum: in
muscle, storing and releasing
Ca
2+
during muscle contraction
b. Ribosomes
Consists of RNA and proteins
Attached in Rough ER and freely floating
in free cytoplasm
1) In Rough ER: synthesize proteins
for incorporation into cell
membranes or lysosomes, or for
export from the cell
2) In cytoplasm: make soluble
proteins
*ribosomal units can switch back and
forth between these two functions
depending on the type of protein they
are making at a given time
Sites where amino acids are assembled
in synthesis of proteins. To carry out this
function,
1) Ribosomes must occur in clusters
or 10 or more linked to one mRNA
or polyribosomes.
2) When chain of amino acids is
formed long enough, they detach
themselves and are found free
within the RER.
3) Vesicle containing newly
synthesized protein detaches from
RER and proceeds to Golgi
apparatus for processing
c. Golgi Complex
Essential for secretion; found in
secretory cells
It processes, concentrates and
packages proteins from RER into
secretory granules for discharge from
the cell.
Controls traffic of small vesicles involved
in the recycling of membrane between
organelles and cytoplasm and cell
surface
Revealed in LM only by staining with
silver. In EM, it appears as a stack of 4
to 10 lamellae of cisternae
Functional polarity (cis and trans faces)
is evident
Vesicles bud off from cisterna and fuse
with the next one on the stack
cis - "receiving" area; trans -
"packaging"/ shipping
d. Mitochondria flexible, shaping-changing rods
Provides energy for motion and
biosynthesis
Principal biochemical activity: oxidative
phosphorylation
The oxidation of the nutrients, like
glucose and fatty acids; energy
thus generated is used to
synthesize ATP; ATP (cells
currency of Energy) broken down
for energy release
Its density in a cell reflects the cells
energy requirement (kung saan mas
busy, doon mas marami)
Self-replicating organelles (via binary
fission) with a limited life-span
Origin from endosymbiosis of ancient
bacteria
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
4 /10
- Similarities with bacteria
1) Own protein synthesis machinery
(ribosomes) more like bacteria than
eukaryotes (sensitivity to
Streptomycin)
2) Inner membrane of mitochondria
contains unusual phospholipid
characteristic of bacterial
membranes.
Parts of the Mitochondria:
1) Membrane
a. Inner Membrane: cristae mitochondriales;
impermeable
b. Outer Membrane: contains
transmembrane channels
2) Spaces
a. Intercristal Space: occupied by the
mitochondrial matrix
b. Intracristal Space/Membrane Space
e. Lysosomes
Round, ovoid or highly irregular, electron-
dense bodies; heterogeneous
disintegrator bodies
Cellular digestion (histochemical
demonstration of hydrolase)
Work best in acidic conditions
Defense against bacteria
- abundant in phagocytic cells
(macrophage, et. al)
Apoptosis or cell death/suicide
Intracellular digestion
Types:
1) Primary: has not yet digested
2) Secondary: has digested or is
digesting
f. Peroxisomes
Small, spherical bodies
Contain oxidative enzymes and catalase
Neutralize free radicals (highly reactive
chemicals with unpaired electrons) that can
scramble the structure of cells
Oxidases convert free radicals to hydrogen
peroxide (still reactive and dangerous if
accumulated in cell)
Catalases convert hydrogen peroxide to
water
Oxidizes long chain of fatty acids
Present in nearly all cell types; they are
mostly found in metabolically active organs
like the liver and kidney (both active in
detoxification)
Form by budding off of the ER
2. Cytoplasmic Inclusions
Metabolically inert and dispensable
Passive components of the cytoplasm as
stored nutrients; inert-by products of
metabolism; accumulations of endogenous
or exogenous pigment
Usually not bounded by a membrane
Examples are glycogen, lipids, pigment
(lipofuscin, melanin) and crystals
3. Cytoskeleton
Structural support for the plasma membrane
and cellular organelles
Means for movement of cellular organelles
Movement of specialized structures such as
cilia and flagella
Muscle contractility
Three types of rods in cytoskeleton:
microfilaments, intermediate filaments and
microtubules (all are non-membranous)
4. Cell Membrane
AKA plasmalemma
A 2D viscous solutionallows for lateral
movement of unbound proteins
Tight junctions: prevent apico-
basal movement of cells (lock cells
in place/proper orientation)
Consists of bimolecular layer of mixed
phospholipids with the hydrophilic ends
oriented outward, and various embedded
integral proteins; Fluid mosaic model
Regulates traffic in and out of cell
Ions, gases, small uncharged
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
5 /10
polar molecules: allowed to pass
Large macromolecules, charged
molecules: not allowed in
Active Transport ion channels,
ion pumps
Have devices for cell attachment and cell-
to-cell communication
Receptors bind and
recognize specific molecules
(i.e. transmitters, hormones,
nutrients, etc.)
D. STRUCTURE-FUNCTION CORRELATIONS
OF THE CELL
1. Cell Motility (Mobile vs. Sessile)
Mobile: free cells; migrate as needed
Sessile: in close contact (ex.
Epithelia). Movement limited to
changes in surface configuration
via microvilli or cytoplasmic
streaming
2. Energy Production vs. Consumption
3. Electrical Conduction
4. Exocytosis
Release of secretory products (ex.
Glandular cells)
Secretory products may be
enclosed in granules,
released upon cells
reception of proper stimulus.
5. Digestion - through lysosomes
6. Transport through the cytoskeleton
7. Constantly Dividing vs. Permanent
Cells
Constant division: growth, renewal,
repair
Proliferation rates vary
o Rapid proliferation: cell
cycle
Two types of division: mitosis
(somatic cells), meiosis
(germ cells)
No cell division in postnatal life
(permanent)
Cardiac muscle, nervous
tissue
LABORATORY
I. STAINING HISTOLOGIC STRUCTURES
TYPES OF STAIN
1. H&E go together in most staining procedures.
a. Hematoxylin = Purplish blue
b. Eosin = Bright pink
2. Silver stain = Black stain; Silver staining aids
visualization of intracellular and extracellular
cellular components (DNA and proteins) by
deposition of metallic silver particles on the
targets (usually Reticular fibers, nerve fibers,
fungi); argentaffin/arygrophilic substances
stain black with silver
3. Orcein stain = commonly used for staining
elastic fibers that color dark brown
4. Giemsa stain = used to differentiate nuclear
and/or cytoplasmic morphology of platelets
(blue), RBCs (pink/red), WBCs (blue), and
parasites
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
6 /10
II. SLIDES
A. Plasmalemma
Smooth & sharply demarcated:
Fuzzy:
Ciliated:
Flagellated:
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
7 /10
Brush or striated:
Sterocillia:
B. Cytoplasm
Basophilic:
Acidophilic:
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
8 /10
C. Organelles
RER, Ribosomes:
Golgi Complex:
Fibrils:
Figure 1 Myofibrils of skeletal muscle
(http://www.lab.anhb.uwa.edu.au/mb140/corepages/
muscle/Images/skem0041he.jpg)
D. Inclusions
Secretory granules/droplets:
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
9 /10
Pigment granules
Fat vacuole
Figure 2: Slide 1
Phagocytosed matter
E. Nuclear Envelope can be found in all cells
F. Nucleolus
G. Heterochromatin can be found in all cells
1
The Cell
Castaneda, Catalan, Catubag, Cayabyab, Cengca
10 /10
H. Nucleoplasm can be found in all cells
I. Nuclear Number
J. Nuclear Size
K. Position of Nucleus
L. Nuclear Shape
Reference
Marieb, E. & Hoehn, K. (2011). Essentials of Human
Anatomy and Physiology (9
th
ed.). Pearson Education
Inc.
Pacual, J.L. The Cell. Power point