1. IL-1B and IL-1RA Expression in Glioma Stimulated Microglia Cells
Sara Maass, Thuy-Dung Ngo, and Victor Suarez
Advisor: Dr. Salvatore Coniglio
New Jersey Center for Science, Technology, & Mathematics
Kean University
Introduction
Glioblastoma is an extremely deadly brain cancer.
Glioblastoma tumors recruit macrophages which in turn
promote cell invasion and immune escape. In the brain
macrophages are called microglia. Macrophages play an
important role in the bodies immune system. There are
two major types of macrophage states: M1 and M2. The
M1 response induces inflammation while M2 reduces
inflammation and promotes tumor spread. Interleukins
also play a role in the immune response. The IL-1 family
induces fever and inflammation. IL-1B tends to be pro-
inflammatory while IL-1RA acts as an antagonist of IL-1B
function and therefore down regulates inflammation. In
this study, we examined the effect of glioma cell
conditioned media on IL-1B family member genes in
microglia using Quantitative Real Time PCR(QRT-PCR)
and immunofluorescence. The role of CSF-1R and CCR1
signaling in this process was also assessed using small
pharmacological inhibitors of these receptors. The
expression of CCR1 on glioma cells and
microglia/macrophages was determined using
immunofluorescence.
Methods
Results
Results cont.
Conclusion
Using Quantitative Real Time PCR (QRT-PCR)
and immunofluorescence we have observed that
conditioned media from the murine glioma cell
line GL261 is able to induce both IL-1B and IL-
1RA expression in microglia. CSF-1R and CCR1
inhibition enhances IL-1B while repressing IL-1RA
expression in glioma stimulated microglia. This
follows the idea that the CSF-1 pathway causes
microglia to become immuno suppressive.
We would like to thank Dr. Salvatore Coniglio
and the fellow research students in our lab for all
of their help and support. Thank you Dr. James
Merritt for providing the CCR1 antagonists. Also,
thank you to New Jersey Center for Science,
Technology, & Mathematics and Kean University
for giving us this research opportunity.
Figure 1: Expression of CCR1 receptor in GL261
glioblastoma cells using immunofluorescence staining.
4 ug/ml aCCR12ary Alone
Figure 2: Expression of CCR1 Receptor in THP1
Macrophages differentiated with PMA express using
immunofluorescence staining.
OVERLAY
Figure 3: Expression of CCR1 in MG Microglia
using immunofluorescence staining.
Phalloidin (Actin) CCR1
Future Works
Acknowledgements
We would like to assess the consequences of the
shift in IL-1 family expression on tumor associated
macrophages/microglia polarity by measuring
changes in M1 vs M2 markers.
QRT-PCR
2ary Alone 4 ug/ml aCCR1
Immunofluorescence
Figure 4: QRT-PCR analysis of JnJ CSF-1R inhibitor in untreated
and glioma conditioned cells. A) IL-1B expression increased
with JnJ. B) IL-1RA expression reduced with JnJ. C) IL-18
expression slightly increased.
0123456
IL-1B
JnJ: - - +
Glioma Cond. Media
A IL-1RA
JnJ: - - +
Glioma Cond. Media
B
0123456
Untx Untx
00.20.40.60.811.2
IL-18
JnJ: - - +
Untx Glioma Cond. Media
C
IL-1B IL-1RA
+ GL261 Cond. Media + GL261 Cond. Media
DMSO DMSO CCR1 inhibitors DMSO DMSO CCR1 inhibitors
Figure 5: QRT-PCR analysis of CCR1 inhibition. CCR1
inhibition enhances IL-1B and represses IL-1RA gene
expression in glioma stimulated microglia.
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