Transcript Research Background

The Effects of Ginseng Pectins on Tumor
Cell Migration and Their Mechanisms
Yuying Fan
26th, Nov, 2012
Outlines:
1st, Research Background
2nd, Research Strategies
3rd, Research Progress
Research Background-panax ginseng
C.A.meyer
• Ginseng (Ginnsuu), belongs to the panax genus in the
family Araliaceae.
American Ginseng (Panax quinquefolius)
Asian (Japanese/Korea/China) Ginseng (Panax ginseng)
Ginseng
Prince Ginseng (Pseudostellaria heterophylla)
Siberian Ginseng (Eleutherococcus senticosus)
Research Background-classification
of asian ginseng
fresh ginseng: raw product
panax
ginseng
white ginseng (WG): fresh ginseng which has been dried
red ginseng (RG): heated by steaming or sun-drying
sun ginseng (SG): steaming at a higher temperature than RG
Research Background-structures
of ginseng polysaccharides
flower, fruit
stipe
leaf
 chemical compounds: ginseng
polysaccharides, ginsenosides,
ginseng peptides and other small
molecules.
stalk
root tip
taproot
rootlet
radicula
 Ginseng polysaccharides=neutral
polysaccharides (starch-like
glucans)+acidic substances
(ginseng pectin)
Research Background-activities of
ginseng polysaccharides
 It has been shown that ginseng polysaccharides have many
bioactivities, including immunomodulation, anti-tumor, antifatigue, hpyerglycemic, anti-atheroscloresis, anti-radiation…
immunomodulation
tumor cell
proliferation
antifatigue
ginseng
polysacc
harides
hpyerglycemic
antitumor
tumor cell
apoptosis
tumor cell
metastasis
Research Background-tumor
metastasis
Research Background-cell migration
Polarization
Cell polarization refers to the tendency of migrating cells to have a
distinct, stable front and rear. The polarity is reinforced and often
even arises from environments that provide a directional cue. These
directional cues can be chemotactic, (induced by chemoattractants or
morphogens), haptotactic (caused by varying concentrations of
substrate), mechanotactic (breakdown of cell-cell contacts, as in
wound healing), electrotactic (induced by electric fields), and
'durotactic', (due to differences in pliability), or combinations of any
of these.
The result is a defined cell front and a rear. The leading edge is
usually characterized by intense actin polymerization that
generates a protrusive structure, and by adhesion to the
substratum. The trailing edge is characterized by stable bundles
and the release and disassembly of adhesions. The central part of
the cell usually contains the nucleus and microtubules (which
exhibit different degrees of polarization depending on the cell
type).
Major mechanisms that
control nuclear and
microtubule positioning
during cell migration.
The small GTPase
Cdc42 acts through
MRCK and myosin II to
control the distance and
position of the nucleus
with respect to the
centrosome (MTOC).
Cdc42 also controls the
Par3/Par6/aPKC
module to control
microtubule dynamics
through the
dynein/dynactin
complex.
Protrusion
Protrusion is the de novo formation of membrane extensions, or
protrusions, in the direction of migration, i.e. the leading edge. It
has three major components: the expansion of the plasma
membrane, the formation of an underlying backbone that
supports membrane extension, and the establishment of contacts
with the substratum, which provides traction for the movement
of the rest of the cell body and signals that regulate actin
polymerization.
The protrusion is produced by local actin polymerization. One
kind of protrusion is flat and fan-like, the edge of which is often
called the lamellipodium and within which actin is polymerizing
and often branched. Spike-like filopodia are another kind of
protrusion; these structures comprise polymerized actin filaments
that are arranged into long parallel bundles. These two forms of
protrusion are thought to serve different roles: filopodia act as
mechanosensory, exploratory devices, whereas lamellipodia
provide wide surfaces that generate traction for forward
movement.
Actin polymerization results from the nucleation of new filaments
and the availability and addition of new monomers. The Arp2/3
complex is a molecular heptamer that attaches itself to the side of a
pre-existing
actin
filament
and
nucleates
de
novo
actin
polymerization at a fixed angle. This is essential for branching in
lamellipodia, and also important in the formation of the base of
filopodia. Monomers are provided by severing of older filaments
and their dissolution into actin monomers. This is mediated by an
actin-filament severing protein, cofilin.
Scheme depicts the major mechanisms and
molecules implicated in regulating and enabling
actin polymerization/depolymerization and
organization. The Arp2/3 complex induces
branching by attaching to the side of a pre-existing
filament and promoting de novo polymerization.
Formins such as mDia1 and mDia2 induce
processive actin polymerization by attaching
themselves to the barbed end of the filament. The
formins compete with actin capping proteins, such
as gelsolin and capZ (not depicted). Ena/VASP
serves a dual role as an adaptor protein and a
'leaky' capper, which enables actin polymerization
through other molecules that are part of a complex
(not shown). Myosin II (in green, between filaments)
cross links antiparallel actin mediates contraction
due to its ATPase-dependent motor domain, and its
cross-linking, ATPase-independent activities. Cofilin
(grey arrowhead) severs actin filaments, promoting
the formation of new barbed ends. It also generates
actin monomers to be incorporated to the filaments
via profilin/formin (treadmilling). Another small
protein, thymosin β4, sequesters actin monomers
and maintains the pool of available monomers.
Some of the signaling proteins that control these
molecules are also shown. They include: Rho-kinase
(ROCK), which is under the control of RhoA and
activates LIMK to activate cofilin, blocks the
myosin phosphatase, promoting myosin activation,
and phosphorylates and activates myosin directly;
PAK also activates LIMK but it inhibits MLCK,
inactivating myosin II; the WAVE/Scar-Abi complex
and WASP, which activate the Arp2/3 complex, are
under the control of Rac and Cdc42, respectively;
finally, formins are also activated by small GTPases
directly (mDia1 by RhoA; mDia2 by Cdc42).
adhesion
Cell adhesion (the physical interaction of a cell with another cell or
with the extracellular matrix (ECM)) is essential for cell migration
and tissue integrity. Cell-cell adhesion maintains epithelial tissues,
supports functional contacts between specialized cells, and can
facilitate directed migration (for example, radial glia can guide
neuronal progenitor cells to specific layers of the brain). Cellmatrix adhesion is the best-studied form of adhesion that mediates
cell migration, and is the focus of this outline.
Scheme depicts the process of adhesion maturation. Nascent adhesions (on the right) are connected to the actin cytoskeleton
by an actin linkage that includes talin, vinculin and α-actinin; other signaling adaptors are also recruited to these complexes,
including FAK and paxillin. These adhesions generate signals that activate Rac, promoting actin polymerization and
preventing myosin II engagement in the lamellipodium. These signals are also required for the dissolution of some adhesions
(turnover) as the cell advances. Some adhesions mature into focal adhesions; this process is characterized by an enlargement
and elongation of the complexes as well as the appearance of new players, like tensin. Some components have conformations
that are tension sensitive (like talin, vinculin, and p130Cas, not shown here). Rho activation is thought to generate focal
adhesions and actin bundling due to increased myosin II activity. Other events that occur at adhesions are internalization of
the integrins, which is mediated by dynamin; and microtubule targeting, which may contribute to adhesion disassembly.
Cell body translocation and rear retraction
For a cell to migrate, protrusion of the front and translocation of the
cell body must be followed by retraction of the rear of the cell. Cell
body translocation immediately follows protrusion and is
independent of actin polymerization. In keratocytes, the cell body
'rolls' behind the front protrusion. This movement is propelled by a
coordinated contraction of the actomyosin cytoskeleton, and thus
depends on myosin II. Myosin-mediated contraction and
microtubule motors (e.g. dynein) also control the translocation of
the nucleus. On the other hand, rear retraction requires the
coordinated contraction of the actin cytoskeleton and disassembly
of the adhesions at the trailing edge. Several mechanisms converge
to promote adhesion disassembly: actomyosin contraction,
microtubule-induced adhesion relaxation, endocytosis of adhesion
receptors, and proteolytic cleavage of focal adhesion proteins.
At the rear of the cell, adhesions disassemble; this process is mediated by at least
four major mechanisms: 1) actomyosin contraction, controlled by RhoA - Rhokinase - Myosin II; 2) protein dephosphorylation and inactivation, mediated by
phosphatases, e.g. calcineurin; 3) proteolysis of adhesion components, mediated by
calpain; 4) microtubule targeting; the identity of the disassembly signal is not
known.
Research Background
relationship of
structure and activity
？
mechanism
development and application of ginseng better
Research Strategies
STEP:1
ginseng root
amylase treat
ginseng pectin
（pectins mixture）
scratch/
transwell
STEP:2
ginseng pectin
ion exchange/molecular gel
filtration chromatograph
fractions with different
structures （fine structure）
scratch / transwell / cell adhesion / cell
spreading / cell morphology / cytoskeleton /
phosphorylation of signaling molecular
tumor cell migration
structure-activity relationship
and mechanism
Research Progress
Ginseng
pectin
inhibited
tumor
migration (specificity and selective)
cell
Mechanisms(HG and RG-I pectins have
synergy effect on migration, cell
morphology,
cytoskeleton,
focal
adhesion,
phosphorylation
of
signaling molecules → cell adhesion
and spreading → cell migration,
without galectin-3）