Abhishek Kulkarni 123 Annie Pineros 2 Sara Ibrahim 23 Marimar HernadezPerez 2 Sarah Tersey 1 Raghavendra Mirmira 1 and Ryan M Anderson 1 Affiliations Department of Medicine The University of Chicago Chicago IL 60637 ID: 919390
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Slide1
12/15-Lipoxygenase regulates Macrophage Migration during Islet Inflammation
Abhishek Kulkarni1,2,3, Annie Pineros2, Sara Ibrahim 2,3, Marimar Hernadez-Perez2, Sarah Tersey1, Raghavendra Mirmira1 and Ryan M. Anderson1. Affiliations:Department of Medicine, The University of Chicago, Chicago, IL 60637 Center for Diabetes and Metabolic Disease, Indiana University School of Medicine, Indianapolis, IN 46202Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, IN 46202
Slide2Background
Pancreatic islet inflammation is a shared characteristic of type 1 and type 2 diabetes pathogenesisMacrophage infiltration exacerbates pancreatic inflammation12/15-Lipoxygenase (12/15-LOX), an enzyme involved in polyunsaturated fatty acid metabolism, has also been implicated in islet inflammationTwo complementary model systems were used for this study: zebrafish and mice
Figure : Sources of pancreatic islet inflammation
Hypothesis: 12/15-LOX promotes macrophage migration to the islets
Slide3Metronidazole (X)
X
X
X
X
X
X
X
X
X
NTR-
β
cells
α
-cells
Macrophages
Methods
β-
cell injury assay
Upper chamber
Bottom chamber
Porous membrane
Peritoneal Macrophages (WT/12/15-LOX KO)
Conditioned islet media
Migrated Macrophages
4 hours
Wash, fix and stain
Quantify macrophages
Transwell
Migration Assay
12/15-LOX KO
Wild Type
Isolate peritoneal macrophages
Quantify expression of chemokine
receptors by flow cytometry
Receptor expression quantification
Quantify macrophages
in controls and 12/15-LOX inhibited zebrafish
Slide4Control
Control10μM ML355InsulinMacrophageInsulinMacrophageInsulin
Macrophage
Insulin
Macrophage
B
C
D
E
4ng 12-LOX Mo
A
H
G
F
Figure A-H:
Inhibition of 12/15-LOX prevents migration of macrophages in the islet in response to injured
β-
cells due to downregulation of CXCR3
Results
Slide5Conclusion and Future Direction
Inhibition of 12/15-LOX prevents macrophage infiltration in the pancreatic islets via downregulation of CXCR3 expressionThe effect of 12/15-LOX on macrophage migration as well as the mechanism is conserved across species Zebrafish prove to be excellent models for studying pancreatic islet inflammation in vivo due to their optical transparency and ease of genetic manipulationNext, the molecular mechanism of the regulation of CXCR3 expression by 12/15-LOX needs to be determinedIn vivo murine studies as well as in vitro migration assays with human monocytes in the presence of 12/15-LOX inhibitors would further consolidate the effects of 12/15-LOX on the cell migration in complex model systems
Slide612/15-LOX
CXCR3
12/15-LOX
CXCR3
CXCR3
CXCR3
HYPERGLYCEMIA
β
-cell
stress
ML355
Proposed Model/ Summary
β
-
cells
Macrophage