Assessment of Hepatic Insulin Signaling in a Murine Model for

Alström Syndrome  

Ayla Kanber, Boston University The Jackson Laboratory

Summer Student Program Symposium August 10, 2015

What is Alström Syndrome?

-Recessive, progressive, fatal ciliopathy -1018 patients worldwide Clinical features: -Obesity -Type 2 Diabetes -Fatty Liver Disease -Hyperinsulinemia -Sensory vision and hearing loss -Dilated cardiomyopathy -Nephronophthisis -Liver steatosis -Excessive fibrosis in all tissues -Death from cardiac, renal, or liver failure by the third decade of life No treatment available for patients

Alms-Ntr Acetylated α-tubulin

4pi image: Localization of ALMS1 to basal bodies of ciliated cells ~Ciliated cells function throughout the body; normal cilia assembly in AS ~Exact function of protein is unknown ~Previous studies show that ALMS1 may be involved in docking proteins

hTERT-RPE1 ciliated cells

ALMS1 proteins are localized to the basal bodies of ciliated cells

cell Cilia

Adapted from:

The Alms1 mouse model recapitulates many of the clinical features observed in human patients

~Alms1Gt/GT mouse model created by a *gene trap in intron 13 of Alms1

~A gene trap is used to purposefully insert mutations randomly into the mammalian genome

Alms1Gt/GT mouse model and littermate control at 10 months old

Graphs of body weight (g) and plasma insulin values (time course)

Weeks Weeks

Bod

y W

eigh

t (g

)

Insu

lin (

ng/

ml)

Favaretto et al, 2014 Alms1-/- Control

Preliminary Findings: Obesity and hyperinsulinemia phenotypes present at ~8 weeks

old in mutant mice

Graph of ALT values (time course)

0

100

200

300

400

500

600

6W 12W 18W 24W 36W

Alms1-/- control

Weeks

IU/L

Preliminary Findings: ALT values are high in mutant mice at ~12 weeks, indicative of liver dysfunction

Extensive Portal Fibrosis in Postmortem ALMS liver specimen

J. Marshall

Normal liver

Trichrome stain

Alström Syndrome liver

Farrington et al., 2010 Clin Exp Gastroenterol

Trichrome stain

Main Goals �  Study livers of 6 week old mice, before the onset of

obesity, hyperinsulinemia, and other phenotypes �  Why? Liver failure is one of the common causes of

death for Alström Syndrome patients.

�  Determine whether or not liver insulin resistance occurs because of a defect in the insulin-signaling pathway

�  Determine if liver insulin resistance is a primary defect of Alström Syndrome

Mutant, A/A Control, WT

6M

Hepatosteatosis, an abnormal retention of lipid deposits, is expressed in 6 month old mutants

Progression of Liver Disease in Alström Syndrome from Hepatosteatosis to Hepatosteatitis

9M 12M 21M 24M

Blue fibers indicate lymphocyte infiltrations

Red collagen fibers indicate fibrosis

Comparison of Liver Weights at 6 weeks and 6 months old

0

1

2

3

4

5

6

6 weeks old 6 months old

Ave

rage

Liv

er w

eigh

t (g

)

Age *Male only

WT Alms1Gt/GT

~Retention of lipids in mutant mice at 6 months old causes fatty liver, and high liver weight compared to control mice

Mutant, IBA1, 8 months Control, IBA1, 8 months

Kupffer cells

Kupffer cells are activated in 8 month old mutants to repair liver damage

Control, GFAP, 8 months Mutant, GFAP, 8 months

Activated stellate cells

PV

Stellate cells are activated in 8 month old mutants to repair liver damage

Emamian, 2012

Confusing and Complicated AKT Pathway

The Insulin Signaling Pathway leads to the phosphorylation of AKT

Studying pAKT will show if there is a defect in this pathway in 6 week old mutant mice before obesity and hyperinsulinemia phenotypes

Jung et al., 2014

6 week old insulin-injected control and mutant mice express more phosphorylated AKT than

saline-injected mice

Western Blot Analysis

Hepatic insulin resistance is not a primary defect of Alström Syndrome!

Conclusion: The successful phosphorylation of AKT in 6 week old insulin-injected Alms1 mice shows that the mutant mice are not insulin resistant in the liver.

Rel

ativ

e pA

KT/β-

actin

insulin *p-value <0.05 is significant

WT Alms1Gt/GT

Future Research? �  Study the insulin signaling pathway downstream of

AKT where there could be defects that cause hepatic insulin resistance

�  Potential Therapeutic Intervention

Acknowledgements �  Special thanks to

Jürgen K. Naggert, Ph.D. Gayle Bouchard-Collin, M.S.

�  I would like to thank all of the Naggert/Nishina lab for welcoming me and supporting me during this program.

�  Thank you to the benefactors of the Summer Student Program. This Summer Student Fellowship was supported by the Beverly Coleman Endowment.

�  Thank you to the Jackson Laboratory.

�  Thank you Korstanje family and Summer Students for making this the best summer ever!