Download Characterization of the Satellite Glial Cell (SGC) in the extrinsic

Characterization of the satellite glial cell (SGC)
in the extrinsic sensory innervation of the gut in
rodent high-fat diet-induced obesity (DIO)
Grace England
STAR Mentor: Dr. Helen Raybould
11 August 2016
Obesity epidemic warrants scientific attention
•In the U.S., 39.4% of adults were obese in
2011-2012 (68.6% overweight or obese)1
•Canine obesity rates average 34-59% in
developed nations 2
•Feline obesity rates average 19-52% in
developed nations 3
•Obesity co-morbidities are often very
detrimental to quality of life.
•Therefore, the study of physiological regulation
of food intake is relevant for understanding
obesity onset and identifying treatments.
The vagal afferent pathway communicates information
about contents of the gut lumen to the hypothalamus
•Gut hormones target vagal afferent
terminals, which relay sensory
information to the brain.
•Anorexigenic hormones (eg leptin)
signal satiety.
•However, consumption of a high-fat
diet (HFD) leads to leptin resistance in
vagal afferent neurons (VAN).
Hypothalamus
Anorexigenic:
Cholecystokinin
Glucagon-like
peptide 1
Peptide YY3-36
Leptin
Leptin resistance is characterized by cellular and
electrophysiological changes in VAN
• Leptin is ineffective in communicating the “fed” state of
the gut to the brain.
• Changes in neuronal plasticity “lock” VAN in an orexigenic
phenotype, and hyperphagia ensues.4,5
Satellite glial cells (SGCs) envelop neuronal
cell bodies in 3D space
Neuron cell body
Satellite Glial Cells
http://philschatz.com/anatomy-book/resources/1210_Glial_Cells_of_the_PNS.jpg
NeuN IbaI DAPI
SGCs envelop VAN in 3D space
Iba1
DAPI
SGCs have macrophagic properties
• Derived from monocytic lineage & share characteristics of microglia in CNS
• Have two distinct macrophage phenotypes:
• M1:
classically-activated, pro-inflammatory
• M2:
alternatively-activated, anti-inflammatory
• Become activated in systemic inflammation6
• Respond to neuronal injury7
• Respond to CCK, leptin, and ghrelin in vitro8
Despite substantial investigation of SGC role in neuronal injury and pain, the potential role
of SGCs in diet-induced obesity and associated inflammation has not been well examined.
SGCs examination is relevant to DIO studies
Hypothesis: SGC in the nodose ganglion mediate phenotypic changes
in vagal afferent neurons (VAN) during diet-induced obesity (DIO).
Specific Aim: Characterize inflammatory phenotype of SGCs in
response to short- and long-term HFD
In particular, do SGCs express the leptin receptor (LepR)?
Experiments
• Short-term HFD
• 1-day HFD-induced inflammation in absence of obesity
•
Long-term HFD
• SGC phenotype & inflammatory response during
development (4 wk) and after establishment (9wk) of DIO
Process of long-term HFD study
4 wk.
Start
LF
9 wk.
x8
x4 per group
x4 per group
HF
x8
Perform immunohistochemistry (IHC)
Confocal Imaging & Quantitation of markers
Section frozen tissue
Food intake and body weight of HFD-fed rats increase at ~3 wks
*p < 0.05
***p < 0.001
2-way ANOVA
p-values are from Bonferroni post-tests
Markers analyzed with IHC
• Iba1 (ionized calcium-binding adapter molecule 1)
• SGC marker
• Arg1 (arginase 1)
• Alternatively-activated anti-inflammatory microglia (M2) marker
• iNOS (inducible nitric oxide synthase)
• Classically-activated pro-inflammatory microglia (M1) marker
• Leptin receptor
• Expressed on SGCs?
Iba1 is more highly expressed in rat NG of rats fed a HFD
for 1 day
1 day chow
1 day HFD
Iba1
DAPI
p = 0.2631
n= 2
1-day HFD induces trend toward pro-inflammatory
(M1) phenotype in SGCs
iNOS
Arg1
Chow
HFD
Leptin Receptor (LepR) co-localizes with Iba1 on SGCs
Iba1 only
Iba1
LepR
DAPI
Conclusions
•We have preliminary data demonstrating that SGCs respond to HFD-induced
inflammation, highlighting a potential role of the SGCs in VAN phenotype in DIO.
•HFD induces a trend toward a pro-inflammatory (M1) phenotype in NG SGCs.
•The leptin receptor is expressed on SGCs. This is the first evidence in vivo of
leptin receptor expression on SGCs in the nodose ganglion.
Future directions
• Conclude 9-wk HFD
• Continue quantitating inflammatory markers via IHC and qPCR
• Perform more extensive 3D analysis of SGC morphology
• Characterize leptin receptor expression on SGCs
• Repeat study to increase n
Acknowledgements
•STAR NIH grant
•Dr. Ingrid Brust-Mascher (Microscopy & Computer Imaging Lab VM:APC)
•The Raybould Lab!!
Elyse Wudeck, PhD student
Amy Gerety, PhD student
Dr. Helen Raybould
Questions?
FOOD
FOOD
FOOD
Hypothalamus
References
1. Ogden, C. L., Carroll, M. D., Kit, B. K., & Flegal, K. M. (2014). Prevalence of childhood and adult obesity in the United States, 2011-2012. Jama,
311(8), 806-814.
2. Raffan, E., Dennis, R. J., O’Donovan, C. J., Becker, J. M., Scott, R. A., Smith, S. P., ... & Summers, K. M. (2016). A Deletion in the Canine POMC
Gene Is Associated with Weight and Appetite in Obesity-Prone Labrador Retriever Dogs. Cell metabolism, 23(5), 893-900.
3. Van de Velde, H., Janssens, G. P. J., De Rooster, H., Polis, I., Peters, I., Ducatelle, R., ... & Verbrugghe, A. (2013). The cat as a model for human
obesity: insights into depot-specific inflammation associated with feline obesity. British Journal of Nutrition, 110(07), 1326-1335.
4. De Lartigue, G., de la Serre, C. B., Espero, E., Lee, J., & Raybould, H. E. (2011). Diet-induced obesity leads to the development of leptin resistance in
vagal afferent neurons. American Journal of Physiology-Endocrinology and Metabolism, 301(1), E187-E195.
5. De Lartigue, G., De La Serre, C. B., Espero, E., Lee, J., & Raybould, H. E. (2012). Leptin resistance in vagal afferent neurons inhibits cholecystokinin
signaling and satiation in diet induced obese rats. PLoS One, 7(3), e32967.
6. Feldman‐Goriachnik, R., Belzer, V., & Hanani, M. (2015). Systemic inflammation activates satellite glial cells in the mouse nodose ganglion and
alters their functions. Glia, 63(11), 2121-2132.
7. Hanani, M. (2010). Satellite glial cells in sympathetic and parasympathetic ganglia: in search of function. Brain research reviews, 64(2), 304-327.
8. Avau, B., Smet, B., Thijs, T., Geuzens, A., Tack, J., Vanden Berghe, P., & Depoortere, I. (2013). Ghrelin is involved in the paracrine communication
between neurons and glial cells. Neurogastroenterology & Motility, 25(9), e599-e608.