Abnormalities in extrinsic and intrinsic innervation and in the abundance and distribution of interstitial cells of Cajal are known to contribute to impairment of gastrointestinal motility in diabetes. Abnormalities in smooth muscle function in relation to diabetes have not been fully explored. The proposed studies are intended to fill this gap in our knowledge. Our previous studies showed that sustained smooth muscle contraction and relaxation are regulated by the RhoA/Rho kinase and cGMP/PKG pathways, respectively. Our preliminary studies in diabetic animal models (NOD and db/db mice) and human diabetic smooth muscle have shown that hyperglycemia causes upregulation of the RhoA/Rho kinase pathway and downregulation of the cGMP/PKG pathway leading to sustained increase in smooth muscle contraction and decrease in its ability to relax. The focus of this proposal is to define the molecular mechanisms that trigger these changes in smooth muscle signaling and identify their contribution to motility dysfunction in diabetes. 1) Microarray and RNA hybrid analysis of control and diabetic gastric and colonic smooth muscle revealed a functional link between miR-133a and RhoA expression that led us to hypothesize that a decrease in miR-133a expression in diabetic smooth muscle mediates the increase in RhoA expression leading to upregulation of the RhoA/Rho kinase pathway and sustained increase in muscle contraction (Specific Aim 1). 2) Sequence analysis of the human and mouse PDE5 promoter region in gastric and colonic smooth muscle identified a link to transcription factor NFATc4 that led us to hypothesize that O- GlcNAcylated phospholamban mediates sequential activation of calcineurin and NFATc4 and increase in PDE5 expression leading to downregulation of the cGMP/PKG pathway and inhibition of muscle relaxation (Specific Aim 2). 3) Preliminary studies identified a link between miR-21 and a decrease in cystathionine- ?-lyase (CSE) expression in diabetic smooth muscle that led us to hypothesize that a decrease in endogenous H2S formation attenuates inhibitory S-sulfhydration of RhoA and PDE5 and promotes upregulation of the RhoA/Rho kinase and downregulation of the cGMP/PKG pathways (Specific Aim 3). The novel concepts underlying these hypotheses have been validated by preliminary studies in smooth muscle of human and diabetic NOD and db/db mice and support the notion that specific alterations in smooth muscle signaling are driven by high glucose levels. A novel approach involving intraperitoneal injection of miR-133a, miR-21, and other specific inhibitors so as to block selectively each pathway provided evidence that these specific alterations in smooth muscle signaling contribute to motility dysfunction in diabetes. Completion of these studies will expand our understanding of overall motility dysfunction in diabetes and offer avenues for the development of novel therapies.

Public Health Relevance

Gastrointestinal motility disorders are common in patients with diabetes and exert an adverse effect on their metabolic and nutritional status. Abnormalities in smooth muscle function in relation to diabetes have not been adequately explored. The long-term goal of this project is to understand how diabetes affects smooth muscle function and gain insights into molecular mechanisms that alter smooth muscle function.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
2R01DK028300-34A1
Application #
8755767
Study Section
Clinical, Integrative and Molecular Gastroenterology Study Section (CIMG)
Program Officer
Grey, Michael J
Project Start
1984-04-01
Project End
2018-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
34
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Virginia Commonwealth University
Department
Physiology
Type
Schools of Medicine
DUNS #
City
Richmond
State
VA
Country
United States
Zip Code
23298
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Nalli, Ancy D; Rajagopal, Senthilkumar; Mahavadi, Sunila et al. (2015) Inhibition of RhoA-dependent pathway and contraction by endogenous hydrogen sulfide in rabbit gastric smooth muscle cells. Am J Physiol Cell Physiol 308:C485-95
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Liu, Miao; Shen, Shanwei; Kendig, Derek M et al. (2015) Inhibition of NMDAR reduces bladder hypertrophy and improves bladder function in cyclophosphamide induced cystitis. J Urol 193:1676-83
Anderson Jr, Charles D; Kendig, Derek M; Al-Qudah, Mohammad et al. (2014) Role of various kinases in muscarinic M3 receptor-mediated contraction of longitudinal muscle of rat colon. J Smooth Muscle Res 50:103-19
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Hurst, Norm R; Kendig, Derek M; Murthy, Karnam S et al. (2014) The short chain fatty acids, butyrate and propionate, have differential effects on the motility of the guinea pig colon. Neurogastroenterol Motil 26:1586-96
Bala, Vanitha; Rajagopal, Senthilkumar; Kumar, Divya P et al. (2014) Release of GLP-1 and PYY in response to the activation of G protein-coupled bile acid receptor TGR5 is mediated by Epac/PLC-ε pathway and modulated by endogenous H2S. Front Physiol 5:420
Huang, Jiean; Nalli, Ancy D; Mahavadi, Sunila et al. (2014) Inhibition of Gαi activity by Gβγ is mediated by PI 3-kinase-γ- and cSrc-dependent tyrosine phosphorylation of Gαi and recruitment of RGS12. Am J Physiol Gastrointest Liver Physiol 306:G802-10
Nalli, Ancy D; Kumar, Divya P; Al-Shboul, Othman et al. (2014) Regulation of Gβγi-dependent PLC-β3 activity in smooth muscle: inhibitory phosphorylation of PLC-β3 by PKA and PKG and stimulatory phosphorylation of Gαi-GTPase-activating protein RGS2 by PKG. Cell Biochem Biophys 70:867-80

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