Seminario basado en artículo

1. Sofía Illatopa Marín
3rd semester | Medicine School |
Universidad Pontificia Bolivariana

2. INTRODUCTION
 Cancer is a term for many diseases in which abnormal cells divide without
control and crowd out normal cells.
 Cancer is becoming the leading cause of death in many countries around the
world and an issue of public health.

3. INTRODUCTION
 Cancer associated fibroblast
CAFs play a significant
functional role in cancer
progression by increasing
levels of transforming growth
factor

4. GENERAL OBJECTIVE
 Study the effect of ligustilide in
CAFs cells, in order to be able to
use this component in cancer
treatment

5. MTT CELL
PROLIFERATION
ASSAY
MATERIALS AND METHODS
Reduction of MTT,
by mitochondrial
enzyme -Succinate
DH in Formazan•It is used to
measure cell
survival and
proliferation.
The number of live
cells is proportional
to the amount of
formazan produced
Basis
The effect of ligustilide on
the growth of immune cells
can be determined by an
MTT cell proliferation assay.
Splenocytes were obtained
from wild-type mice and
cultured in 96-well U-
bottom plates
Utility

6. FLOW
CYTOMETRY
Flow cytometry is a
method that allows
obtaining information
on cell populations
from an individualized
study of a group of cells
On this group of
cells, it is induced
a laser with the
purpose of
studying its
dispersion and
reflexion
Basis The cells of the
study were
analyzed by Flow
cytometry to
determine the
number of
molecules on the
cellular surface
Utility
MATERIALS AND METHODS

7. WESTERN BLOT
Is a laboratory technique
used to detect an specific
protein in sample of
blood or tissue
Basis
CAFs treated with
different concentrations
of ligustilide (0, 15, 30
and 45μM) for various
durations were harvested
from a 6-well plate and
lysed in RIPA buffer to
isolate whole cell
proteins.
Utility
MATERIALS AND METHODS

8. CELL
IMMUNOflUORE
SCENCE
It is a technique that is
used for the detection of
subcellular structures that
allowed us to study with
the utilization of
antibodies coupled to
fluorophores
Basis
The cells were treated
with a certain process to
be analyzed at the
microscope to identify
subcellular structures
Utility
MATERIALS AND METHODS

9. RESULTS
FIG. 2 (A) T-cell
proliferation was
measured using the
intracellular dye
carboxyfluoresceinsuccini
midyl ester (CFSE).
CDA4+ and CD8+
revealed higher levels of
expresion with a
concentration of
ligustilide at 45 uM

10.  FIG 3. (A) with a
concentration of
ligustlide at 45 uM/24H
MyD88 showed an
elevated expression,
besides TAK at the
same concentration
had low levels of
expression.
 (B) MyD88 showed the
highest levels of
expression at a
concentration of
ligustilide at 45
uM/24h and TRAF6 was
the opposite

11.  FIG 3. (C) At the sample
with CAF, CAF+
ligustilide + CIL - 095,
DAPI and MERGE
showed expression, but
NFkB just showed a bit
of expression in +CIL -
095 and the highest
levels of the expression
were at the sample of
CAF+ Ligustilide
RESULTS

12. RESULTS
FIG 5. (A) GADPH (+/+) a 45 Um/24h showed high levels of expression and
alfa –SMA at the same concentration showed minimum levels (B) In the
sample of CAF, DAPI, alfa –SMA and merge showed expression and in the
sample were ligustilide was added, alfa –SMA it did not present a expression

13. DISCUSION
AUTHOR STATEMENT CONFIRMATION
N. Erez, M. Truitt, P. Olson, S.T.
Arron, D. Hanahan
Cancer-associated fibroblasts are
activated in incipient neoplasia to
orchestrate tumor-promoting
inflammation in an NFkappa B-
dependent manne
YES
S.Y. Han, C.X. Ma, M.H. Hu, Y.Y.
Wang, F.L. Ma, N. Tao, Z.H. Qin,
A polysaccharide from
Dictyophora indusiata inhibits the
immunosuppressive function of
cancer-associated fibroblasts
YES
T. Long, Z. Liu, J. Shang, X. Zhou,
S. Yu, H. Tian, Y. Bao,
Polygonatum sibiricum
polysaccharides play anti-cancer
effect through TLR4-MAPK/NF-
kappaB signaling pathways, Int.
YES

14. CONCLUSIONS
 It was found that ligustilide activates the NF-κB pathway and promotes the
conversion of CAFs to NFs
 Ligustilide reverses the immunosuppressive function of CAF cells through an effect on Toll-
like receptor 4
 The results revealed that ligustilide could serve as an agent in the prevention and
treatment of cancer.
 Ligustilide can reverse the inhibition of T – cells by CAFs.