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.

Ginseng

American Ginseng (Panax quinquefolius)

Asian (Japanese/Korea/China) Ginseng (Panax ginseng)

Prince Ginseng (Pseudostellaria heterophylla)

Siberian Ginseng (Eleutherococcus senticosus)

Research Background-classification of asian ginseng

panax ginseng

fresh ginseng: raw product

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

chemical compounds: ginseng polysaccharides, ginsenosides, ginseng peptides and other small molecules.

Ginseng polysaccharides=neutral polysaccharides (starch-like glucans)+acidic substances (ginseng pectin)

flower, fruit

stipe

leafstalk

radicula

rootlet

taproot

root tip

It has been shown that ginseng polysaccharides have many bioactivities, including immunomodulation, anti-tumor, anti-fatigue, hpyerglycemic, anti-atheroscloresis, anti-radiation…

ginseng polysaccharide

s

immunomodulation

anti-tumor

hpyerglycemic

anti-fatigue

tumor cell proliferation

tumor cell apoptosis

tumor cell metastasis

Research Background-activities of ginseng polysaccharides

Research Background-tumor metastasis

Research Background-cell migration

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.

PolarizationPolarization

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.

ProtrusionProtrusion

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).

adhesionadhesion

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). Cell-

matrix 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.

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.

Cell body translocation and rear retractionCell body translocation and rear retraction

At the rear of the cell, adhesions disassemble; this process is mediated by at least four major mechanisms: 1) actomyosin contraction, controlled by RhoA - Rho-kinase - 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

fractions with different structures （ fine structur

e）

ginseng pectin

ion exchange/molecular gel filtration chromatograph

structure-activity relationship and mechanism

ginseng pectin （ pectins mixture）

ginseng root

amylase treat

tumor cell migration

scratch/transwell

scratch / transwell / cell adhesion / cell spreading / cell morphology / cytoskeleton / phosphorylation of signaling molecular

STEP:1 STEP:2

Research Strategies

Research Progress