Biomedical Engineering Reference
In-Depth Information
consistent with the notion that EGFR-dependent pathways (such as those involved in cell motil-
ity and organogenesis) must be spatially restricted. However, in cells overexpressing EGFRs or
when receptor/ligand endocytosis was blocked, those two signals spread over the entire cell
even though EGF stimulation was localized—a result that is consistent with dysregulated cell
motility and EGFR overexpression that are observed in tandem in carcinoma cells. hus, ligand-
independent propagation of EGF signaling may be the mechanism underlying the dysregulated
cell motility that characterizes tumor invasion and metastasis.
his example illustrates how rapidly prototyped microluidic systems can be used as inexpen-
sive perfusion chambers for studies in which fast changes of the cell environment are required.
Unlike in open-air perfusion chambers, in which the low patterns can be erratic due to convec-
tion and evaporation at the air-liquid interface, in microchannels, the low proile is parabolic
and the concentration proile can be easily modeled, yielding more quantitative analysis of cell
response as a function of analyte concentration changes.
Another example in which local signal transduction is manifested dramatically is in the
immunological synapse
. Cells of the immune system such as T cells cannot react to free anti-
gen—they can only “see” it once it has been processed and presented to them via a molecule
called major histocompatibility complex (MHC) present on the surface of antigen-presenting
cells (APCs). he immunological synapse is the approximately 75 μm
2
contact that forms
(through MHC+ antigen) between a lymphocyte (a type of white blood cell in the vertebrate
immune system, including natural killer cells, T cells, and B cells) and an APC. Recent studies
have shown that the microscale protein structure of the contact point is important in modulating
T-cell activation. Hence, a team led by Lance Kam from Columbia University used microstamp-
ing to simulate the presence of proteins on the APC membrane in diferent spatial arrangements
(colocalized versus segregated), as shown in
Figure 6.19
. Naïve CD4
+
T cells were seeded on top
of patterns of antibodies against CD3 and CD28 (surrounded by ICAM-1), and interleukin-2
(IL-2) secretion was measured by luorescence microscopy (the cells were treated with an IL-2
capture reagent so IL-2 was detected on the cell surface). Naïve CD4
+
T cells attached to the
features of anti-CD3 antibodies, forming a stable synapse. When anti-CD28 was presented in
the cell periphery, surrounding an anti-CD3 feature, this “segregated pattern” enhanced IL-2
secretion compared to having these signals combined at the center of the immunological syn-
apse. Overall, these experiments show that costimulation geometries modulate IL-2 secretion
(a)
(b)
100
Colocalized
Segregated
CD3
COL
SEG
80
Pattern
IL-2
60
40
20
DIC
0
0 500
IL-2 (f luorescence units)
1000
25 µm
FIGURE 6.19
The.immunological.synapse..(a).CD4
+
.cells.are.seeded.on.surfaces.that.simulate.the.
microscale. organization. of. ligands. presented. by. APC. cells;. patterns. of. anti-CD3. and. anti-CD28.
antibodies.are.shown.in.red,.where.IL-2.secretion.is.shown.in.green;.the.remainder.of.the.surface.
was.coated.with.ICAM-1.(omitted.for.clarity);.(b).histogram.of.IL-2.secretion.for.a.CD3-only.pattern.
(CD3),.the.colocalized.pattern.(COL),.and.the.segregated.pattern.(SEG)..(From.Keyue.Shen,.V..Kaye.
Thomas,.Michael.L..Dustin,.and.Lance.C..Kam,.“Micropatterning.of.costimulatory.ligands.enhances.
CD4+.T.cell.function,”.
Proc. Natl. Acad. Sci. U. S. A.
.105,.7791-7796,.2008..Copyright.(2008).
National.Academy.of.Sciences,.U..S..A.)
