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Free-Radical Polymer Science Structural Cancer Model: A Review

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Abstract and Figures

Polymer free-radical lipid alkene chain-growth biological models particularly for hypoxic cellular mitochondrial metabolic waste can be used to better understand abnormal cancer cell morphology and invasive metastasis. Without oxygen as the final electron acceptor for mitochondrial energy synthesis, protons cannot combine to form water and instead mitochondria produce free radicals and acid during hypoxia. Nonuniform bond-length shrinkage of membranes related to erratic free-radical covalent crosslinking can explain cancer-cell pleomorphism with epithelial-mesenchymal transition for irregular membrane borders that "ruffle" and warp over stiff underlying actin fibers. Further, mitochondrial hypoxic conditions produce acid that can cause molecular degradation. Subsequent low pH-activated enzymes then provide paths for invasive cell movement through tissue and eventually blood-born metastasis. Although free-radical crosslinking creates irregularly shaped membranes with structural actin-polymerized fiber extensions as filopodia and lamellipodia, due to rapid cell division the overall cell modulus (approximately stiffness) is lower than normal cells. When combined with low pH-activated enzymes and lower modulus cells, smaller cancer stem cells subsequently have a large advantage to follow molecular destructive pathways and leave the central tumor. In addition, forward structural spike-like lamellipodia protrusions can leverage to force lower-modulus cancer cells through narrow openings. By squeezing and deforming even smaller to allow for easier movement through difficult passageways, cancer cells can travel into adjacent tissues or possibly metastasize through the blood to new tissue.
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Volume , Article ID ,  pages
http://dx.doi.org/.//
Review Article
Free-Radical Polymer Science Structural Cancer Model:
AReview
Richard C. Petersen
Department of Biomaterials and Biomedical Engineering, e University of Alabama at Birmingham, SDB 539, 1919 7th Avenue South,
Birmingham, AL 35294, USA
Correspondence should be addressed to Richard C. Petersen; richbme@uab.edu
Received December ; Accepted  December 
Academic Editors: S. Fukushige, K. Jung, and S.-Y. Shieh
Copyright ©  Richard C. Petersen. is is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Polymer free-radical lipid alkene chain-growth biological models particularly for hypoxic cellular mitochondrial metabolic waste
can be used to better understand abnormal cancer cell morphology and invasive metastasis. Without oxygen as the nal electron
acceptor for mitochondrial energy synthesis, protons cannot combine to form water and instead mitochondria produce free radicals
and acid during hypoxia. Nonuniform bond-length shrinkage of membranes related to erratic free-radical covalent crosslinking can
explain cancer-cell pleomorphism with epithelial-mesenchymal transition for irregular membrane borders that “rue and warp
over sti underlying actin bers. Further, mitochondrial hypoxic conditions produce acid that can cause molecular degradation.
Subsequent low pH-activated enzymes then provide paths for invasive cell movement through tissue and eventually blood-
born metastasis. Although free-radical crosslinking creates irregularly shaped membranes with structural actin-polymerized ber
extensions as lopodia and lamellipodia, due to rapid cell division the overall cell modulus (approximately stiness) is lower than
normal cells. When combined with low pH-activated enzymes and lower modulus cells, smaller cancer stem cells subsequently have
a large advantage to follow molecular destructive pathways and leave the central tumor. In addition, forward structural spike-like
lamellipodia protrusions can leverage to force lower-modulus cancer cells through narrow openings. By squeezing and deforming
even smaller to allow for easier movement through dicult passageways, cancer cells can travel into adjacent tissues or possibly
metastasize through the blood to new tissue.
1. Introduction
Cancer Fundamentals. Cancer is a pathological condition
related to malignant uncontrolled rapid cell growth prolifer-
ation, invasive cell movement into adjacent tissues, and occa-
sional metastatic spread through blood and lymph to more
distant locations []. Conversely, benign tumors represent
uncontrolled cell growth that does not invade other tissues
[]. Cancers are the result of progressive accumulations in
genetic mutations through cell interactions with carcinogens
such as tobacco, sunlight, radiation, infectious microbes, or
certain chemicals/material []. Some genetic changes can
be added by being passed along from one generation to
another to increase cancer risk []. Although normal cells
have limits to replication or the number of cell divisions
to control growth by apoptosis cell death when necessary
with a cascade of caspase enzymes, cancer cells can develop
almost limitless uncontrolled growth aer at least four genetic
mutations [], Figure .
Cancer cells can thus become less prone to death so that
unneeded cells develop to form extra tissue known as tumors
[]. However, benign tumors that localize and do not
invade adjacent tissue are not cancer []. Benign tumors are
furtheroenencapsulatedbyconnectivetissue[, ]. Con-
versely, malignant tumors invade adjacent tissues and can
enter the blood stream to attack other organs by metastasis [,
, ]. In order to become motile and invade into other tissues,
major genetic cell traits change involving mutation through a
process termed the epithelial-mesenchymal transition (EMT)
[]. Several important cellular alterations occur during EMT
to include loss of integrin attachment between cytoskeleton
actin bers with adjacent cells and subsequent motility with
invasiveness []. Further, EMT is characterized by large vari-
ations in cell shape that comprise loss of spheroid rounding
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Loss of normal growth control
Normal
cell division
cell division
Cell suicide or apoptosis
Cell damage
no repair
Cancer
First
mutation mutation mutation
Second
ird
Fourth or
later mutation
Uncontrolled growth
F : Normal cells control growth by programmed death
known as apoptosis. Cancer cells conversely build genetic muta-
tions that can result in uncontrolled growth aer at least the
fourth mutation. (With permission from the National Institutes of
Health/Department of Health and Human Services).
with formation of spindle-shaped or broblast-like cells and
irregular membrane borders especially at the invasive tumor
edge []. EMT may also consist of possible long growth pro-
cesses []. During EMT, cells dedierentiate from the normal
tissue phenotype toward the more primitive mesenchymal
stem cell []. To better understand EMT by comparison
between carcinoma cells derived from the epithelium with
smooth normal cell membranes tightly bound together, a
classical stellate mesenchymal stem cell with membrane
extensions is isolated free in bone marrow extracellular space
and surrounded by advancing dierentiated preosteoblastic
stem cells, Figure .
A more complete list of cancer causing risk factors
includes the following.
() Age increases cancer risk probably by additive eects
of genetic mutation and exposures [, , ].
() Tobacco is the most widely known cause of death
and directly related to an extremely high level of
cancer mortality. About – percent of all cancer
deaths are related to tobacco. Cancers that increase
in risk with smoking are found in the lung (highest
percent of cancer-related deaths), mouth, throat, lar-
ynx, esophagus, stomach, pancreas, bladder, kidney,
cervix, and in the blood with myeloid leukemia [].
() Ultraviolet (UV) light from the sun or other sources
for tanning can initiate aging and eventually cancer
[, , , ].
() Ionizing radiation from radioactive sources such as
nuclear explosions increases cancer for leukemia, the
thyroid, breast, lung and stomach, and radon gas
formed in the earth to increase lung cancer. Other
sources of radiation include X-rays and cosmic rays
from space [, , , ].
F:Mesenchymalstemcellwithmorelightlystainednucleus
in bone marrow central to dierentiating preosteoblastic stem cells
above and below the stellate cell acquired during histomorphometry
analysis.
() Chemicals and materials from a multitude of various
sources can increase cancer risk from arsenic, asbes-
tos, styrene, benzene, benzidine, cadmium, nickel,
or vinyl chloride. Carcinogenesis by chemicals was
shown to occur by an initiation step with a primary
application and then an irritation step that promotes
cancer [, , , ].
() Infectious microbes like viruses and bacterium can
even exist as a subclinical reservoir for long periods
[, , , ].
() Hormones such as estrogen increase the severity for
breast cancer while estrogen reduced and androgen
increasedprostatecancerseverity[, , , ].
() Familial DNA genetic alterations where added and
passed on from one generation to another [, ].
() Alcohol with more than drinks per day for a long
time increases the risk for cancer of the liver, mouth,
throat, larynx, esophagus, and breast [, , ].
() Diet with high levels of fat can increase cancer risk for
thecolon,uterus,andprostate.Antioxidantnutrients
have alternatively shown decreased risks for cancer [,
, , ].
() Sedentary life style with lack of physical activity is
related to overweight factors leading to cancer of the
colon, esophagus, breast, kidney, and uterus [, , ].
() Hypoxia and ischemia from low molecular oxygen
concentrations resulting in mitochondrial free radi-
calsandacidaremoregeneralbiologicalconditions
thatexistforcancercellularactivity[, , ].
Subsequent tumors exhibit anaerobic metabolism
that produces energy without oxygen in hypoxic
microenvironments [, ]. Conversely, normal
tissues use oxygen for aerobic mitochondrial energy
synthesis [, ].
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2. Hypoxic Free Radicals and Polymerization
Mitochondria create electrons from dierent fuel sources
of the body that ow through a series of protein enzymes
and other mobile electron carriers [, ]. ese electrons
through the mitochondrial electron transport chain combine
with molecular oxygen and protons to form water [, ].
However, imperfect formation of water by mitochondria
during hypoxic low O
2
states produces free-radical reactive
oxygen species (ROS) as superoxide (O
2
∙−
), hydrogen perox-
ide (H
2
O
2
), and hydroxyl radicals (
OH) [, ]. Other
reactive free-radical species can also be formed as well [].
By denition, a free radical is a molecule that contains an
unpaired electron which is highly unstable and seeks out
another electron for a stable covalent bond pair [, ].
Subsequent hypoxic free-radical species are then involved
in damage to lipids, proteins, and DNA [, , , ].
As a result, hypoxic low-oxygen concentrations of tumors
emphasize the strong inuence that free radicals play during
the development of cancer such that reactive oxygen species
are considered oncogenic [, , ].
In terms of similarities with a basic chemistry under-
standing for molecular pathobiology, polymer science unsat-
urated alkene chain-growth free-radical covalent bond for-
mation with increased liquid/resin viscosity by molecu-
lar crosslinking toward solid structure produces irregular
shrinkage patterns []. Further, from polymer science free
radicals polymerize unsaturated resins to produce increased
material modulus (or stiness) with higher density measured
as reduced bulk volume percent from shorter interatomic
bond distances [, ]. In fact, the most character-
istic subsequent manifestations of overall polymerization
crosslinking is material shrinkage with some internal stresses
and warpage [, ]. A highly common free-radical
cure method in past has been a resin system with styrene
monomer and dibenzoyl peroxide, depicted in Figure ,
to illustrate free-radical reactive secondary sequence by
crosslinking across a carbon-carbon (C=C) vinyl double
bond [].
During similar structure-related hypoxic free-radical
pathobiology, cell pleomorphism occurs as part of cancer cell
EMT by irregular invaginated lipid membranes containing
extensive folds associated with possible new lipid polymer-
ization chemistry [, ] in the presence of membrane
rues []. EMT pleomorphism further includes extreme
dierences in cell shapes/sizes [] related to increased overall
tumor mass density from a stier (or higher modulus)
denser stroma of brotissue [, , ]. In advanced carcino-
mas EMT-dedierentiated desmoplastic stroma produces a
harder denser overall tumor mass related to a more aggressive
gradeofcancerthatintimebecomesanacellularcollagenous
extracellular matrix []. Relative to free-radical covalent bond
formation to produce material structure, increasing density
surface oxidation crosslinking of rubber reduces oxygen
diusion into the deeper subsurface layers [].
Pertinent to cancer pathophysiology, capillary distance
has been measured in tissue showing a respective decrease
in both oxygen concentration and pH [, , ], where
increasing hypoxic mitochondrial electron-transport free
radicals would be expected to produce some biologic struc-
ture from covalent bonding. As increased biologic structure
limits O
2
diusion more, a new fall in pH with lower O
2
would be expected by interfering even further with capillary
diusion of molecules. Because oxygen is fundamentally
critical to prevent cell death, tumors found over . mm
away from blood vessels fail to grow [, , ]whichis
the approximate distance for O
2
diusion through living
tissue before a zero concentration develops [, , ]. e
low pH eventually becomes an overall result of lactic acid
production by anaerobic mitochondria glycolysis energy
synthesis [, ]. On the other hand, throughout ecient
aerobic cellular respiration in the mitochondria, oxygen is
fundamental during energy synthesis to form water from
the electron respiratory transport chain and proton gradient
[, ]. Part of mitochondrial respiratory aerobic energy
synthesis includes one of the most well-studied enzymes ever
with cytochrome c that was identied in earliest National
Cancer Institute research, Figure .eenzymecytochrome
c is part of the electron transport chain and localizes in
the intermembrane space but can leak into the cytosol
to participate in cell death by apoptosis with the caspase
enzymes [].
During hypoxia, cells switch to anaerobic glycolysis to
produce lactic acid [], but with alternate aerobic respiration
energy synthesis reactive oxygen species with other free
radicals and also protons start to form [, ]. However,
cancer cells can adapt to hypoxia by high glucose uptake with
anaerobic glycolysis and lactic acid production to minimize
free radical formation from the mitochondria [, , , ].
Subsequent hypoxic cancer conditions then not only produce
large amounts of free radicals [, ]butalsocontribute
to acid formation for a much lower pH microenvironment
[, , , ]. rough a mechanism known as the Warburg
eect [, , ], cancer cells continue to produce energy by
aerobic glycolysis even with O
2
present [, , ]possibly
as mentioned before as an adaptive mechanism to help
limit hypoxic free-radical formation by the mitochondria
[]. Free radicals may represent a potentially more serious
threat than lower pH since free-radical molecular structure
through the rapid chemistry of covalent crosslinking limits
oxygen diusion progressively as a primary source for cancer
pathology through continual increasing levels of free radicals
and acids from oxidatively stressed mitochondria.
In addition to chronic infections with widespread in-
ammatory cells, chronic inammation has been recognized
as a major common cause of cancer promotion that further
involves free radicals [, , , , ]. Sources of chronic
inammation can include microbial/viral infections or
toxic/allergic substances and also obesity [, , , , ].
Dysplasia occurs in chronic inammation and also in benign
growths before premalignant cell proliferation [, , ].
Cells that invade through the basement membrane are then
considered malignant [, , ]. Pleomorphism occurs
as a sign of dysplasia with cells displaying large variations
in cell sizes and shape morphologies with unusually large
deeply stained nuclei [, , ]. Mitotic cellular division is
also high such that the increased possibility of malignant
transformation through EMT can occur by benign cells
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(a) (b)
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F : (a) Dibenzoyl peroxide commonly known as benzoyl peroxide or BPO dissociates into two molecules to provide (b) benzoyl free
radicals (R
). (c) In liquid form, styrene monomer with vinyl-group C and C (C=C) double pi (𝜋) bond upper le of molecule is attacked by
R
to add a benzoyl group on one of the carbon atoms. (d) Subsequent styrene radical formation depicted by e on the opposite vinyl carbon
atom can now enter into a reaction with a new styrene molecule C=C vinyl group. (e) A second styrene molecule can add the benzoyl-styrene
radical on the C=C vinyl group for the growing chain to form a free radical again as e on the opposite vinyl carbon atom. Free-radical
additions through the liquid styrene monomer C=C vinyl groups continue the polymerization process by the reactive secondary sequence
method a multitude of times to eventually form a solid.
F : Image depicting cytochrome c which is a small soluble
globular protein used in the mitochondrial electron transfer chain
that carries electrons as radicals with other mobile transporter
carriers ultimately to combine with oxygen and protons to form
water [, , ]. (With permission from the National Institutes of
Health/Department of Health and Human Services).
unrestrained from normal process in the dysplastic states
[, , ]. In addition, inammatory cells mediate cancer
progression through free radicals that can mutate DNA,
produce epigenetic alternations with DNA methylation,
increase cell proliferation, oxidize lipids, crosslink proteins,
and promote angiogenesis [, ].
3. Other Important Fundamentals in
the Study of Cancer
() Angiogenesis or blood vessel development was shown
to be an important part of tumor expansion that could
be used as a pharmaceutical chemotherapeutic anti-
cancer target. Later research showed the pathology
that tumors produce factors to encourage angiogene-
sis with new abnormal blood vessels to support cancer
growth. Tumors cannot increase to more than - mm
in diameter without growing blood vessels to bring in
oxygen and nutrients [, ].
() Apoptosis is a normal cell physiologic death that helps
to prevent unwanted cell growth. However, tumors on
the other hand contain cancer cells with mutations
thatpreventnormalapoptosis.Cellslackingapoptosis
death regulatory pathways are then more resistant
to death in addition to chemotherapeutic cancer
cytotoxic agents [, ].
Scientica
() Tumors secrete protease enzymes that degrade col-
lagen protein to provide exits for entering the blood
stream and metastasizing to more distant tissue sites
[, , , ].
() DNA methylation can turn a gene o [, , ].
Hydrogen abstraction has been commonly identied
as part of a free-radical lipid peroxidation break-
down process that produces characteristic biomark-
ers. Since breaking a carbon-carbon bond requires
much less energy than breaking a hydrogen-carbon
bond [, ], highly reactive methyl and other small
acyl-free radicals could be considered as sources for
DNA methylation during periods of lipid peroxida-
tion or during hypoxia with the generation of free-
radicalelectronsandacid[].
() Tumor suppressor genes inhibit cell proliferation
and are targets of carcinogenesis when such genes
are inactivated. For example, proteins from several
viruses bind to a tumor suppressor gene to stop
manifestation of downstream functional activities to
subsequently promote cancer. e most common
tumor suppressor gene is the p gene that initiates
apoptosis through the p protein [, ].
() Protooncogenes code for cell growth or proliferation
and can mutate to become oncogenes to cause cancer,
Figure . Common proto-oncogenes can code for
growth factor proteins, cytoplasmic enzymes, mem-
brane receptors with tyrosine kinase enzymes, or
transcription factors in the nucleus for cell division
[, ].
4. Irregular Membrane Ruffling by
Free-Radical Crosslinking
In order to better simplify cancer fundamentals into a molec-
ular biology polymer approach for an easier mechanistic
understanding theory of cellular and tissue changes during
carcinogenesis, gures in imaging from the National Cancer
Institute are helpful. A typical National Cancer Institute
consensus illustration, Figure , depicts the basic morphol-
ogy that a cancer cell has irregular borders with membrane
wrinkling as ruing when compared to the identical normal
rounded cell. In addition, Figure shows how an oncogene is
activated by a cancer causing agent so that instead of normal
cell division, oncogenic cellular DNA damage causes cancer.
Cancer cell nuclei are also misshapen into strange unusual
irregular shapes compared to the normal rounded oval nuclei
with smooth borders. For a plausible explanation regarding
the membrane irregular borders with ruing in cancer cells,
free radicals need some understanding. As a constant source,
free radicals produced during hypoxia are found at high
levels in cancer cells [, ]. Further, excessive free radicals
as reactive oxygen species attack unsaturated fatty acids
found as lipids in cell membranes with carbon-carbon (C=C)
double bonds [, ].
One of the most characteristic features for free-radical
covalent bonding in a liquid to increase viscoelastic solid
Normal cell
Cancer cell
Activated
oncogena
Cancer causing
agents
Potrooncogena
DNA
strand
F : National Cancer Institute illustration shows the stages
of how a normal round cell is converted to a cancer cell with
shape irregularities for both the nuclear and plasma cell membranes
when an oncogene becomes activated. (With permission from the
National Institutes of Health/Department of Health and Human
Services).
structure with increased modulus and density proper-
ties becomes most apparent in materials that polymerize
by electron-pair chain-growth polymerization as the lin-
ear/volumetric cure shrinkage [, , , ]. In fact,
free radicals are engineered for specic materials science
application to crosslink molecules with consequent cure
shrinkage and possibly warpage as one the most distinguish-
ing material problems of extensive polymer electron-pair
bonding [, , ]. Because shrinkage is not necessarily
perfect with inhomogeneous material, nonuniform electron
pairing in curing in addition to increasing the modulus
can create residual internal stresses to produce warpage of
materialsthatweakensparts[, ]. Warpage is particularly
accentuated during free-radical polymer curing with thin
lm coatings of variable thickness without good support
[]. As covalent single sigma (𝜎) bonds form by replacing
C=C pi (𝜋) bonds during reactive secondary sequence chain
growth [, ], polymer chains draw together from more
distant van der Waals intermolecular attraction forces and
chain-entanglement equilibrium distances to closer covalent
distances with increasing chain entanglement that reduce
bulk volume by linear/volumetric cure shrinkage [, 
]. Free-radical double-bond conversion to single bonds
is thermodynamically favorable and forms an exothermic
polymerization even without extra energy added at room
temperature []. Consequently, solutions of unsaturated
lipids that undergo thermoset free-radical chain growth will
Scientica
Cluster of normal intermediate cells.
Normal cell
Cytoplasm
Cytoplasm
Cell
membrance
Nucleolus
Nucleolus
Nucleus
Nucleus
Chromatin
Chromatin
clumps
Strain artifact
Cancer cell
Note the 1) relatively large amount of cytoplasm,
2) clear nuclear stain, 3) fine chromatin granules in the nucleus,
4) small nucleolus, 5) the smooth nuclear boarder,
6) stain artifact in one cell where another
small white blood cell overlaps.
N
uc
l
eo
l
us
N
uc
l
eus
C
hromati
n
c
lum
p
s
Strain artifact
Note 1) large size of the nucleus compared to the total cell size,
2) the dark staining of the nucleus, 3) the larger size of the chromatin
clumps in the nucleus, 4) the large nucleolus,
5) irregular nuclear boarder.
(a)
Normal cell
Normal and cancer cells
Multinucleated
Multipolar mitosis
Multiple, large nucleoli
Variable cell shapes
Coarse
Lagging chromosomes
Chromatin
and sizes
Arrested mitosis
Small amount
of cytoplasm
(b)
F : (a) Graphics and description comparing normal round cells and cancer cells with both irregular nuclear and plasma cell membranes
(With permission from the National Institutes of Health/Department of Health and Human Services). (b) e round normal and cancerous
characteristics with warped irregular borders are identied. (With permission from the National Institutes of Health/Department of Health
and Human Services).
also produce linear/volumetric cure shrinkage without added
energy [, , , ].
Cytoskeleton actin bers provide tensile strength and
support to the cell [] so that the composite plasma cell
membrane with lipid oils and phosphate groups oers
conditions that maintain separate mediums to accentuate
free-radical polymerization warping particularly as an outer
veil thin lm. Hydrocarbon lipid molecules drawn together
at a rounder border would require some invagination to
wrinkleinwardespeciallywhencombinedwithcouplingto
underlying rigid bers resulting in the possible explanation
for common ruing irregular membrane appearances of
cancer cells depicted in consensus National Cancer Institute
Figures , (a), (b),and(a)(d). Lipid oil with an unsat-
urated linoleic/oleic fatty acid combination for free-radical
crosslinking has previously demonstrated a wrinkling and
warpage during a solidication polymerization process [].
As well as irregular plasma cell membrane borders on the
outer periphery of a cancer cell, the nuclear membranes are
misshapen and nuclear to cytoplasm ratios increase [, ].
In addition to normal cell round morphology changing by
EMT to irregular membrane border patterns with invagina-
tions in cancer, other common ndings are presented as basic
consensusforchangesincancercellsthroughtheNational
Scientica
Normal
(a)
Cancer
(b)
(c) (d)
F : Normal cells on the le and cancer cells with more spike-like membrane extensions on the right in culture from human connective
tissue. At a magnication of x, the cells were illuminated by darkeld amplied contrast technique. (a) Normal cells compared to (b)
cancer cells. (c) Normal Cells compared to (d) cancer cells (with permission from the National Institutes of Health/Department of Health
and Human Services).
Cancer Institute with two separate illustrations, Figures (a)
and (b),as.
() darker staining nucleus,
()coarsechromatinandclumping,
() more irregular nuclear border,
() less cytoplasm and larger nuclei,
() multinucleated,
()variablepleomorphismincellsizesandshapes.
Cell cultures that show normal cells with smoother mem-
brane outlines compared to cancer cells with more irregular
membranes help to document the plasma cell membrane
spike-type extensions that form deeper invaginated irregular
bordersaspartoftheEMTwithtransformationtocancer,
Figures (a)(d).
Another more common intracellular organelle mem-
brane feature possibly related to covalent C=C double-bond
crosslink shrinkage and thin-lm warpage [] includes the
most distinctive inner membrane features of the mitochon-
dria with the high convolutions called cristae [, , 
], Figures (a) and (b).econvolutedmitochondrial
cristaethatmakeuptheinnermembraneservetocreatean
impermeable conning larger surface area for important bio-
logic electron interactions during aerobic respiratory energy
synthesis [, , ]. Subsequent electron ow during
energy synthesis [, , ] should further be an extreme
source for free-radical lipid crosslinking through C=C double
bonds to provide the structural convolutions noted in the
mitochondria. e mitochondrial interior inside the inner
membrane is a gel of approximately % protein []. Most
of the mitochondrial protein studies thus far are not bers
but rather soluble or globular enzymes [, , ]that
would not provide cytoskeleton-type support like structural
bers in a composite []. During crosslink shrinkage by high
concentrations of free radicals from the electron transport
chain, the lipid hydrocarbons would accentuate warpage into
convolutions of the thin-lm inner membrane without the
more stable inuence from reinforcing stacked planar bers
provided by the plasma cell membrane or nuclear membrane.
Proteins contain ionizable groups on the terminal car-
boxylic acid end, the terminal amino end, and many side
chain groups to function as buers []. Proteins also gen-
erally exist with a negative charge at physiologic pH . [].
Since proteins maintain a negative charge intracellularly and
are further the major blood buer as part of the plasma [],
the ideal medium is available to sequester radicals from the
mitochondrial electron transport chain and buer protons
Scientica
(a)
Intermembrane
Space
Matrix
Proteins
Inner Membrane
Outer
membrane
DNA
Cristae
Ribosome
Granules
I
ntermem
b
rane
Spac
e
M
atr
i
x
Protei
ns
I
nn
e
r M
e
m
b
r
a
n
e
O
uter
me
m
b
r
a
n
e
D
N
A
C
ristae
R
i
boso
m
e
u
le
s
(b)
F : Mitochondrial inner membrane cristae (a) D illustration upper le shows spheroidal mitochondria in red located peripherally
circumferential around the nucleus and SEM below showing convoluted cristae. (b) Mitochondrion illustration with tortuous cristae ((a) with
permission from the National Institutes of Health/Department of Health and Human Services).
fromtheprotongradientintheformofthemultipleprotein
enzymes that help synthesize energy aerobically by oxygen
with water as the nal product. In fact, amino acid side
chains on hemoglobin protein [] and even in short peptides
[, ] are known to sequester radicals with tyrosine,
histidine, and cysteine residues [, ]. Further, radicals
can be delocalized from a side chain into the peptide bond
[, ]. Although the spin density for radical delocalization
from a side chain into the peptide bond atoms is small
for pentapeptides, projections could be large depending on
peptide bond conformational changes or electrostatics [].
In addition, proteins could increase radical delocalization
through the much higher numbers of peptide bonds than
a small peptide molecule so that radicals from the electron
mitochondrial transport chain could be stabilized for even
longer periods than normal radical intermediates studied
thus far. So, instead of radicals disseminating into dierent
pathologies constantly, much longer induction periods would
be made available by proteins acting as antioxidants, but nev-
ertheless could still become oversaturated eventually to act as
an electron pool or source for free-radical damaging proper-
ties. Still, free-radical crosslinking of the mitochondrial inner
membrane would help explain the extreme convolutions that
structure into an impermeable medium.
5. Cell Movement and Lamellipodia
A state-of-the-art scanning electron microscope (SEM)
shows the intricate details for irregular plasma cell membrane
borders that form ruing and an extensive network of struc-
tural spike-like ridges that project long distances, Figure (a).
A more complete conception of an SEM D-enhanced
National Cancer Institute image shows how a cancer cell and
its long processes called lamellipodia move on a cellular tissue
surface, Figure (b). Carcinogenesis requires cell movement
with EMT and cell shape changes for metastasis and invasion
[, , ]. Subsequent cell movement involves reactive oxygen
species that includes H
2
O
2
as common denominators for
the formation of the hydroxyl-free (
OH) radical which in
turn creates protrusions at the cell edges [, ]. Cell move-
ment is directed through extracellular chemical gradients by
chemotaxis [, , ]. Free radicals that form as a constant
source from reactive oxygen species including H
2
O
2
have
been shown to act as key chemotactic factors to regulate
chemoattractants that bind to cell membranes with actin
polymerization for cell migration toward H
2
O
2
and other
reactive oxygen species []. Further, the traveling cell
is polarized by microtubules radiating from the centrosome
near the nucleus to the outer cell edges [, ]togrow
actin protrusions with adhesions between the extracellular
matrix that contract in the forward direction [, , , ].
e cell protrusions are long lamellipodia extensions and
short focal adhesive lopodia constructed from actin bers
that polymerize at the advancing forward edge [, , ].
Conversely, depolymerization of actin occurs away from the
advancing cell edge [, , ]. Cytoskeleton microtubule
and actin bers are polarized positively near the cell mem-
branetoelongate[, ] and negatively by microtubules
toward the organizing centrosome center near the nucleus
[, ]. A strong long-range static electric eld develops
on the mitochondria and also on the microtubules that lie
in close contact [] resulting in a possible delocalization
mechanism for the electron transport chain during periods of
mitochondrial oxidative stress. Polymerization of actin bers
at the plus end forms protrusions that contain focal adhesions
with the extracellular matrix so contractions provide forward
movement [, , , ]. In addition, depolymerization
occursatthenegativeendsoftheactinbersontherear
edge of cell movement for release of focal adhesions and in
addition making actin monomers available to be recycled for
polymerization at the forward positive actin protrusive end
[, , ].
Scientica
(a) (b)
F : (a) SEM of a breast cancer cell that clearly shows both the so petals of membrane ruing and much longer lamellipodia spiking
extensions. (b) A scanning electron microscopic D-enhanced NIH image of cancer cells and lamellipodia spike processes on a cellular tissue
surface (with permission from the National Institutes of Health/Department of Health and Human Services).
Free-radical crosslinking and agglomeration with weaker
secondary bonding previously described account for atoms,
molecules, and larger molecular chains being drawn together
[, , , ]toprovideapossiblemechanismfor
forward movement in the contraction process. Lipid per-
oxidation products with low molecular weights and a C=C
double bond have demonstrated high crosslinking with both
unsaturated fatty acids []andprotein[]. Further, a
polyunsaturated fatty acid with C=C double bonds has
been shown to activate cells as a chemoattractant to migrate
during reorganization of actin ber in lamellipodia using a
strong free-radical oxidant as peroxinitrate [], with strong
free-radical crosslinking indication for contraction through
covalent bond shrinkage []. In addition to lipid free-radical
crosslinkingacrossC=Cdoublebondsasareactivesecondary
sequence, amino acid side chains of protein are modied
at about  percent of the residue by the hydroxyl-free
radical formed from H
2
O
2
[]. Further, proteins are found
to agglomerate or crosslink most notably through the amino
acid tyrosine [, ] with metal-catalyzed reactions [].
While most cancer cells studied through the National
Institute of Health gures presented demonstrate extensive
spike protrusions that could greatly interfere with movement
and prevent leakage through a small tissue pore, the more
aggressive cancer cells have a smaller membrane area that
allows the lower modulus cells to more easily squeeze through
and penetrate small openings in the endothelium [].
Nevertheless, membrane protrusions are most frequent on
the leading edge of cancer cells during metastasis movement
[]. Formation of the leading edge with cytoskeleton actin
bers that shape lamellipodia [, ]isonlyabout
nanometers thick [, , ]. In addition, the actin bers
orient along the axis of the membrane protrusions so that the
modulus is highest to resist deformations in the lamellipodia
planeandmuchlowerintheperpendicularplane[].
As a result, the leading lamellipodia edge can push with
leverage through the smallest openings and still deform
on the sides to start squeezing through extremely narrow
spaces for escape into new tissue []. Fiber polarizations
fromthenegativecentrosomeendnearthenucleustothe
positively charged plasma cell membrane side []are
thought to be responsible for the forward polymerization
of the actin bers during cell movement [, , ]. As
such, electrons conducted from microtubules into actin bers
by highly charged mitochondria [] overstressed under
hypoxic conditions should accumulate at accentuated levels
through the electron transport chain deprived of oxygen
that could help account for a portion of the free radical
polymerization mechanisms with actin bers. In fact, actin
has been shown to restructure under free radical conditions
with H
2
O
2
to increase cell motility []. Regarding H
2
O
2
ability to polymerize actin by free-radical chain lengthen-
ing mechanisms, H
2
O
2
hasproventobeanexceptional
initiator for free-radical polymerization in polyester resin
formulations []. By related free-radical reactive oxygen
species biological chemistry, actin polymerization has also
been observed in macrophages exposed to oxidized low-
density lipids []. Further, high levels of H
2
O
2
and other
reactive oxygen species are found in various cancer cells [].
6. Nucleus Changes with Free-Radical
DNA Methylation
Cancer initiation and progression have been linked to
increasing free-radicals following oxidative stress and
reactive oxygen species that are continuously produced by
mitochondria during cell metabolism [, , ]. Cancer
cells are characterized by increased reactive oxygen species
andaccumulationwhilethemitochondriaareconsidered
the major source for reactive oxygen species []. Subsequent
hypoxic mitochondrial free-radical species are then involved
in damage to lipids, proteins, and DNA [, , , , , ].
As a result, hypoxic low oxygen concentrations of tumors
 Scientica
emphasize the strong inuence that free radicals play
during the development of cancer such that reactive oxygen
species are considered oncogenic [, , , ]. A major
role for free radicals produced by hypoxia or ischemia
through mitochondrial metabolism includes crosslinking
with either simple electron pairing or as extensive reactive
secondary sequence C=C double-bond chain growth. A
major eect of free-radical crosslinking molecules is a
characteristic volumetric shrinkage and warpage [, 
, , ]. By similar free-radical crosslink chemistry,
biologic crosslinking could explain the coarse or clumping
chromatin depicted in Figures (a)-(b),couplingofDNAto
DNA or DNA to protein [], abnormal cell shrinkage with
warpage and irregular membrane borders [, , , , ],
membrane ruing [], protein agglomeration with insoluble
accumulation [, , ], and actin polymerization [].
Also, as the nuclear membrane structures, a greater amount
of the cell is constrained within the nucleus [, , ].
In addition to genetic DNA alterations or changes in
the DNA sequence [, ], epigenetics that alter the gene
slightly further inuence cancer growth and metastasis [,
]. Hypoxia is already accepted as a general condition that
promotes tumors [, , ], supports cancer recurrence
[], intensies malignancy [, , ], increases metastases
[, , ], and inhibits chemo/radio therapies [,
]. e
main epigenetic change is by DNA methylation that can be
linked to hypoxic-free radical environments with acyl-radical
breakdown products [, , , ]. Molecular break-
down is further a well-known biomarker when free radicals
accumulate with unsaturated lipid fatty acids []espe-
cially when combined with lower pH acid []thatoccurs
during hypoxia in mitochondrial metabolism []. Further,
thermodynamics for bond dissociations studied for lipid
peroxidation favors acyl radical formation over hydrogen
bond dissociation as a key source for DNA methylation [].
Hypermethylation of DNA subsequently causes gene silenc-
ing of particular importance for tumor suppressor genes [,
]. Oxidation of DNA is another risk factor for mutation
that modies bases to favor cancer []. Subsequent long-
term stages of hypoxic-related free radicals particularly seen
with chronic inammation then account for advanced chro-
mosomal damage with DNA base substitutions particularly
through purine replacement by a pyrimidine termed a G T
transversion, DNA crosslinks, and chromosomal breaks [].
7. Metastasis
Although a cancer tumor can invade adjacent tissue directly,
most cancer deaths occur by metastasis to distant tissue
through the blood [, , ]. Metastasis takes place through
an extremely biocomplex sequence of events in several stages,
Figures (a) and (b). Biocomplexity is a hallmark of cancer
that involves both a low pH acidic environment [, ]and
free radicals [, , ] produced from hypoxic conditions
[, , , ]. Metastasis further involves cell motility
withEMTcellshapechanges[] and also destructive tissue
breakdown by pH-activated protein enzymes [, , ]. Fur-
ther, focal contacts develop between the extracellular matrix
and lopodia that extend as short spikes o the advancing
lamellipodia lengthening arms []. Microtubules polarized
by electric elds inuenced through close association with
mitochondria [] have capability to conduct electrons or
transport free radical species rapidly to the plasma cell
membrane. Free radicals available as a result of low oxygen
concentrations leaking from the electron transport chain can
then help to provide adhesive bonds by electron pairing and
secondary bonding forces during cell migration through focal
contacts with the lopodia. In fact, unsaturated lipids that
have C=C double bonds with reactive secondary sequence
capability []areknowntoinuencecelladhesionbyfocal
contacts with the extracellular matrix [].
e rst advanced invasive stage involves attachment of
cancer cells from the primary tumor to the basement mem-
brane of local blood vessel endothelial cells []. Endothelial
cells compose a single layer that wraps around to form a
tube inside the blood vessels which are further depicted in
Figure (b).enextstageinvolveslocalbreakdownofthe
endothelial cell barrier by cancer cell protease enzymes lled
with both free radicals and acids [, , ]. As a related
biocomplex event, the oxidation of the amino acid cysteine
by reactive oxygen species H
2
O
2
includes disulde crosslinks
thatareinvolvedwithmetastasisandreducedenzymeactivity
[].
Cancer cells with greater metastasis potential have a lower
modulus and lower viscosity than normal cells with the
ability to deform more in addition to the pleomorphic smaller
sizes with less cell membrane area [, , ]. Cell stiness
increases with organized actin bers of the cytoskeleton
and during cancer transformation actin bers are disrupted
into irregular networks that results in a lower modulus
or less sti, more deformable cell []. Conversely, overall
increased tumor tissue density is a risk factor for cancer [,
, ]. Increasing stroma density is further characterized
by increased collagen deposition [] that provides better
traction forces with focal adhesions to promote cell migration
for metastasis [, ]. Also, cells tend to migrate toward
stier substrates []. rough a similar reference to cell
structure, cancer cell pseudopods with high modulus actin
bers [] provide sti leverage movement through narrow
gaps to invade other tissue, Figure .But,inopposition
again with another molecular biology reverse structural-
related mechanism, tissue degradation from cancer hypoxic-
associated lower pH with protease enzymes removes intercel-
lularadhesiontomoreeasilyreleasecellsfromtheprimary
tumor and to carve out space for invading cancer cells [].
Consequently, the smaller cancer cells with lower moduli
can subsequently be freed from the primary tumor to move
through small openings [] produced by the common
protease breakdown and enter the blood stream. On the other
hand, as larger cancer cells invade, metastasis then most
commonly progresses when cells get trapped in capillaries to
spread into new tissue [].
8. Protease Enzymes
Cancers revert by EMT to more primitive less dierentiated
cells with increased acid production [, ]. Subsequent
Scientica 
Tumor cells
1. Attachment
2. Local breakdown
3. locomotion
4. secondary tumor
(metastasis)
Attachment sites
Blood vessel
(a)
How cancer spreads
Attachment
Local breakdown
Blood vessel
Secondary tumor
(b)
F : (a) Schematic drawing for the stages of metastasis ()
attachment () local breakdown with lamellipodia penetration into
a blood vessel () locomotion with lamellipodia to exit from blood
vessel()secondarytumor.(b)Oncemetastaticcellsareattachedto
the vessel wall basement membrane (a physical barrier that separates
tissuecomponents),cancercanbreakthroughwithstilamellipodia
on the leading edge and the help of protease enzymes. Cancer cells
then move through the blood stream enabling them to spread to
other parts of the body. A secondary tumor may subsequently form
at another site in the body. (With permission from the National
Institutes of Health/Department of Health and Human Services).
protease enzymes activated by acid due to related hypoxic
conditions are responsible for degrading structural bers of
the extracellular matrix [, , ]. As the extracellular matrix
is broken down under hypoxic conditions at a much lower
pH than normal, cancer cells can initiate paths through the
extracellular matrix for cancer cell migration to spread and
F : SEM of a cancer cell movement through a man-made
hole involves arms or pseudopodia, termed lamellipodia, enabling
them to migrate to other parts of the body. Locomotion cell
motility is integral to the entire process of invasive metastasis. (With
permission from the National Institutes of Health/Department of
Health and Human Services).
eventually detach into areas occupied by dying cells [, ].
Cancer cells can eventually even use the surrounding dead
cells for a nutrition source during rapid cell proliferation
by intracellular means with oncogene expression of enzymes
linked to membrane receptors []. Further, in an unusual
biocomplexity, cancer cells are able to induce stromal cells
such as broblasts to produce H
2
O
2
reactive oxygen species
and lactate acid through metabolic anaerobic glycolysis to
increase invasiveness [, , 