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(2쪽) These days, the efforts to reproduce the native
cell motility have been. Glioblastoma multiform is the most malignant tumor in adults, and
its remarkable tendency to wildly infiltrate through normal neural tissue make complete
resection impossible. As a consequence, this feature confounds therapeutic attempts at
local control in malignant gliomas. Histological evidence has shown that glioma cell invasion
in the brain occurs following the orientation of thin, elongated anatomic structures, such
as capillaries, white matter fibers, and unmyelinated axons. Unfortunately, standard assays devised
to study glioma cell motility do not incorporate such topographical cues guiding cell adhesion
and traction in vivo. 3D Nanofibers offer a unique advantage as glioma cell migration
models: the ability to individually investigate the influence of chemical and mechanical effects
on cell behavior. Today, I’d like to review three research articles on glioma cell migration
on 3D nanofiber scaffold mimicking white matter tract. Let’s begin.
(3쪽) First, I will briefly introduce the overview
of three journals. The first Journal published on Neoplasia at 2011 is about analyzing the
motility, gene expression, and sensitivity to migration inhibitors of glioma cells cultured
on nanofibers mimicking the neural topography. This research contribute to enable cell migration
analysis using nanofiber scaffolds by reproducing native mechanisms of migration. Second one
is from Biomaterial at 2013. To improve the mechanical properties of nanofiber, they fabricated
various types of materials. These materials are used to make nanofiber via core-shell
electrospinning that permit systematic study of mechanical and chemical influences on cell
adhesion and migration. From this research, they enable further examination of the complex
interplay of mechanics, chemistry, and topography in regulating brain tumor behaviors.
(4쪽) Lastly, the third article is from Integrative
Biology at 2013. This research paper is focused on importance of understanding and eventual
arrest of cancer metastasis by investigating the mechanistic influence of the tumor microenvironment
on cancer cell migration and membrane blebbing. So, they suggested that the influence of the
mechanistic environment on the invasion dynamics of glioma cells add to the understanding of
how biophysical components influence glioma cell migration and blebbing dynamics.
(5쪽) Let’s look through in detail. This is detail
review for Journal #1 (Neoplasia, 2011). By using their nanofiber scaffolds, they found
that glioma cell morphology and migration depend on fiber alignment. To check out the
effect of nanofibers, the morphology and behavior of glioma cells cultured on three-dimensional
nanofiber scaffolds versus conventional two-dimensional surfaces were analyzed. Cells cultured on
aligned nanofibers adopted a fusiform morphology, usually with a leading process following an
individual fiber. In contrast, cells on randomly oriented fibers remained relatively rounded.
In neither case did we see obvious lamellipodia or fan-shaped morphologies that were typical
of these cells cultured on TCPS. (6쪽)
Next, to determine whether migration of glioma cells on nanofiber scaffolds reproduced this
key molecular feature of three-dimensional migration, the effect of inhibitors targeting
myosin II and actin polymerization on cell migration was assessed. Migration of U251
glioma cells out of aggregates seeded on aligned nanofibers was significantly inhibited by
the myosin II inhibitor Blebbistatin. Cell motility on aligned nanofibers was highly
dependent on myosin II activity as in other three-dimensional models. Results from acto–myosin
II disruption suggested that cell migration on nanofibers reproduced more closely the
molecular features observed on three-dimensional migration rather than those observed for migration
on rigid two-dimensional surfaces. From these data, they suggested that glioma cell migration
on aligned nanofibers is myosin II dependent.
(7쪽) Lastly, they used microarray analysis to compare
U251 glioma cells cultured on aligned versus randomly oriented nanofibers, resulting significantly
different gene expression. Strikingly, there was a remarkable up-regulation of genes related
to JAK/STAT signaling, recently highlighted as a central regulator of malignant progression
in high-grade gliomas. Both STAT3 inhibitors, stattic and LLL12, significantly inhibited
the migration of U251 cells cultured on aligned Nanofibers. These results revealed a possible
role for STAT3 in the migration of glioma cells in response to topographical cues and
demonstrated the advantages of 3-D nanofiber scaffolds as a culture model to investigate
pathways involved in cancer cell migration.
(8쪽) Let’s move on to the next journal article
of Biomaterials. Aligned electrospun nanofibers are particularly interesting as neural guides
because of their topographical similarity to white matter. Additionally, aligned electro-spun
nano-fibers reproduce the morphological and molecular signatures of glioma migration ex
vivo. In this study, aligned, core-shell electrospun nanofibers were fabricated using coaxial electrospinning
to mimic the topography of the native in vivo environment, including white matter tracts.
Specific to GBM migration, several brain mimetic hydrogels, including hyaluronic acid, have
been employed. To examine the influence of mechanics, various core materials were used
(i.e., gelatin, PDMS, and PES) with an identical shell (i.e., poly(ε-caprolactone) (PCL)).
Patient-derived GBM (OSU-2) cells obtained from tumor tissue were cultured on scaffolds,
resulting elongated along the nanofibers, exhibiting a bipolar, spindle shaped morphology.
This morphological behavior displayed sensitivity to intermediate nanofiber moduli especially.
(9쪽) Migration was sensitive to nanofibers with
different mechanical properties. The fastest migration speeds were observed for fibers
of intermediate modulus. Further increases in the modulus reduced migration speed, although
these speeds were still significantly higher than those observed with the softest core-shell
nanofiber. Interestingly, in no case was cell migration completely inhibited, further demonstrating
that aligned topographical features of the appropriate scale are sufficient to promote
cell migration. To further evaluate the mechanisms behind the observed migration responses in
fibers with different mechanical moduli, they investigated the expression of molecular markers
for cell adhesion (Focal Adhesion Kinase, FAK) and migration (Myosin Light Chain 2,
MLC2). Consistent with single cell migration results, the active forms of FAK and MLC2
were overexpressed by cells on fibers as FAK and MLC2 are overexpressed in GBM tissue.
(10쪽) The authors on third article is focused on
the mechanistic influence of aligned nanofibers on cell shape, migration and blebbing dyanimics
of glioma cells as you can see on the title. They used DBTRG-05MG cells, which means Denver
Brain Tumor Research Group-05MG cell line. The cells mainly adopted 3 major morphologies
on STEP fibers. First, the cells showed spindle shape on single fibers, and- rectangular shaped
on parallel fibers, and polygonal shape at orthogonal fiber intersections. On flat control
substrates, however, DBTRG-05MG cells mostly showed spread configurations with pronounced
focal adhesions around the circumference of the cells. Using the STEP platform, the polarization,
extension of protrusions at the leading edge, and retraction of the trailing edge of a single
cell on a single nanofiber were captured using time lapse images.
(11쪽) They investigated the difference in migration
speeds of cells on SD, versus SS. SD means 2 layers of orthogonally arranged fibers and
SD is a single parallel layer of fibers. After then, they also checked the role of structural
stiffness within SS to check the effect of ECM stiffness. Glioma cells spread more and
migrated significantly slower on double suspended (SD) fibers when compared to cells on parallel
single suspended (SS) fibers and on flat fibers. Nanofiber length significantly increased cell
migration and cells migrated the fastest on 10 mm nanofibers among the three lengths test.
(12쪽) Lastly, they addressed a question that a phenomenon
that interlinks cytoskeleton organization, migration, and apoptosis is blebbing. The
dynamics of blebbing in cancerous cells requires more attention as it has been associated with
invasiveness, ability to escape apoptosis, and motility. So they thought understating
of glioma migration as blebs are being increasingly associated with tumor cell behaviors. So,
they used time lapse imaging to observe the membrane blebbing. DBTRG-05MG cells demonstrated
relatively continuous blebbing on flat surfaces and exhibited an interesting reversible blebbing-non-blebbing
phenomenon on suspended fibers. As cells spread along the STEP nanofibers (both SS and SD)
and increased in spread area, both bleb size and their occurrence count decreased. Blebbing
cells had significantly lower migration than cells that were not blebbing.
(13쪽) Here is discussion section. From first journal,
they addressed that a major problem in studying cell motility in vitro is the difficulty of
reproducing the native behavior of these tumor cells. A topographically complex environment
for cell culture, using biocompatible scaffolds formed by electrospun submicron-sized fibers
was developed. Cell motility in nanofibers reproduced, at least in part, molecular features
of three-dimensional motility such as myosin II dependence and low sensitivity to disruption
of stress fibers. An important finding is the observation that cell motility in nanofiber
scaffolds was STAT3-dependent and could be specifically disrupted with low, subtoxic
concentrations of STAT3 inhibitors. (14쪽)
The core-shell spinning methodology from Biomaterial journal provides a novel means of isolating
the effects of mechanical or chemical factor on native GBM cell behaviors. This work addresses
a crucial issue in brain cancer biology by providing tunable and well defined platforms
that mimic a major GBM migration highway allowing examination of biophysical and biochemical
cues regulating tumor cell behaviors. These biomaterials could be broadly applied to study
behaviors of neural cells in white matter mimetic microenvironments with potential implications
for stem cell therapy, studies of neural regeneration and development, and myelination.
(15쪽) Using the STEP platform which used non-electrospinning
platform, the structural stiffness of the fibers was altered by the choice of fiber
length on Integrative Biology journal. As structural stiffness decreased with fiber
length, cell migration increased. Blebbing and its influence in cancer requires more
investigation as the study shows that cell spreading can significantly reduce blebbing.
For better prognosis of cancer, a better understanding of migration and blebbing dynamics is necessary.
(16쪽) This is conclusion pages. Until now, I introduced
efforts to understand the mechanisms gradually developed by mimicking the native behavior
of cell motility in vivo. This biomimetic material platform should have broad applicability
for examining and further evaluating fundamental questions in the biology. Specifically, it
should be possible to extend these studies to other tumor cell types that disperse in
vivo along anatomic structures, such as pancreatic, prostate, or head and neck tumors that use
perineural migration for metastasis. In all these cases, the topographically complex nature
of nanofiber scaffolds could provide considerable advantages over other models to study three-dimensional
cell migration. Neoplasia & Biomaterials
Junghwa Cha