DM mellitus (DM) is a common disorder with a prevalence of 15.5 million sufferers in the United States (1). In 2007, complications arising from DM cost the US healthcare system $58 billion (1). One major complication of DM is neuropathies. Constipation is a key consequence of enteric neuropathy and is significantly more common in DM patients with a prevalence of 60% compared to 15% in the general population. DM-associated enteric neuropathy results from hyperglycemia-induced neuronal apoptosis. However, the mechanism by which hyperglycemia drives neuronal apoptosis in DM remains enigmatic. In light of the increasing appreciation of the role of toll-like receptors (TLR) in driving a variety of chronic disease processes, especially those associated with oxidative stress and advanced glycation products (AGEs), which is known to result from hyperglycemia, we hypothesized that TLRs play a role in DM-associated enteric neuropathy. In support of this notion, we have observed that upregulation of one particular TLR, namely TLR4 is associated with hyperglycemia-induced neuronal apoptosis. Moreover, genetic and pharmacological ablation of TLR4 prevents both hyperglycemia-induced neuronal apoptosis and DM-associated colonic dysmotility. Thus, we hypothesize that hyperglycemia-induced activation of TLR4 results in neuronal apoptosis and, consequently, drives the colonic dysmotility associated with DM. To test the hypothesis and further investigate the underlying mechanism(s) of hyperglycemia-induced neuronal apoptosis, we will perform experiments to establish the role of TLR4 in mediating hyperglycemia-induced enteric neuronal apoptosis and elucidate the mechanism involved. Using primary enteric neurons from WT/TLR4-/- mice and knock down/overexpression strategies in the enteric neuronal cell line developed in our laboratory, we will establish the necessity and sufficiency of TLR4 in modulating hyperglycemia-induced enteric neuronal damage. The mechanism of hyperglycemia induced TLR4 activation will be examined focusing on the role of Advanced Glycation end products (AGES) and oxidative stress. Further, we will characterize the signaling events involved in hyperglycemia-induced TLR4 activation in enteric neuronal apoptosis focusing on the downstream targets of TLR4 including NF-?B. Finally, we will determine the role of TLR4 on enteric neuronal apoptosis and colonic dysmotility in vivo. Our preliminary data indicate that streptozotocin (STZ) -induced DM results in enteric neuronal apoptosis and loss of mouse colonic myenteric neurons. These changes are ameliorated in TLR4-/- DM mice. Taken together these studies will not only elucidate a novel mechanism involving TLR4 in the pathogenesis of hyperglycemia-induced enteric neuronal apoptosis but also provide "proof of principle" for targeted therapies that could be used for the treatment or prevention of dysmotility associated with DM.

Public Health Relevance

The etiology and pathogenesis of the gastrointestinal complications of diabetes is largely unknown and thus treatment of this debilitating condition is a challenge to clinicians. In many patients the duration of diabetes correlates with the development of diabetic autonomic neuropathy as well as the presence of diabetic gastrointestinal complications such as diabetic constipation. In this proposal we will examine the mechanism of how hyperglycemia leads to loss of enteric neurons. Experiments outlined in this proposal will not only contribute to the understanding of the changes in enteric neurons in diabetes, but may also lead to new therapeutic targets for the altered gastrointestinal motility seen in diabetes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK080684-04
Application #
8310088
Study Section
Clinical and Integrative Gastrointestinal Pathobiology Study Section (CIGP)
Program Officer
Hamilton, Frank A
Project Start
2009-08-01
Project End
2014-07-31
Budget Start
2012-08-01
Budget End
2013-07-31
Support Year
4
Fiscal Year
2012
Total Cost
$297,375
Indirect Cost
$105,520
Name
Emory University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322
Nezami, Behtash Ghazi; Mwangi, Simon M; Lee, Jai Eun et al. (2014) MicroRNA 375 mediates palmitate-induced enteric neuronal damage and high-fat diet-induced delayed intestinal transit in mice. Gastroenterology 146:473-83.e3
Mwangi, Simon Musyoka; Nezami, Behtash Ghazi; Obukwelu, Blessing et al. (2014) Glial cell line-derived neurotrophic factor protects against high-fat diet-induced obesity. Am J Physiol Gastrointest Liver Physiol 306:G515-25
Zhang, Y; Bitner, D; Pontes Filho, A A et al. (2014) Expression and function of NIK- and IKK2-binding protein (NIBP) in mouse enteric nervous system. Neurogastroenterol Motil 26:77-97
Yarandi, S S; Srinivasan, S (2014) Diabetic gastrointestinal motility disorders and the role of enteric nervous system: current status and future directions. Neurogastroenterol Motil 26:611-24
Chandrasekharan, Bindu; Nezami, Behtash Ghazi; Srinivasan, Shanthi (2013) Emerging neuropeptide targets in inflammation: NPY and VIP. Am J Physiol Gastrointest Liver Physiol 304:G949-57
Chandrasekharan, Bindu; Jeppsson, Sabrina; Pienkowski, Stefan et al. (2013) Tumor necrosis factor-neuropeptide Y cross talk regulates inflammation, epithelial barrier functions, and colonic motility. Inflamm Bowel Dis 19:2535-46
Ravella, K; Al-Hendy, A; Sharan, C et al. (2013) Chronic estrogen deficiency causes gastroparesis by altering neuronal nitric oxide synthase function. Dig Dis Sci 58:1507-15
Tehrani, A Bonakdar; Nezami, B G; Gewirtz, A et al. (2012) Obesity and its associated disease: a role for microbiota? Neurogastroenterol Motil 24:305-11
Zhang, Yonggang; Liu, Jianjun; Yao, Shaohua et al. (2012) Nuclear factor kappa B signaling initiates early differentiation of neural stem cells. Stem Cells 30:510-24
Mwangi, Simon M; Usta, Yousef; Shahnavaz, Nikrad et al. (2011) Glial cell line-derived neurotrophic factor enhances human islet posttransplantation survival. Transplantation 92:745-51

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