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Identifying pathogenic stromal and acinar signaling for improved diagnosis and treatment of chronic pancreatitis

Komar, Hannah Marie, Komar
http://rave.ohiolink.edu/etdc/view?acc_num=osu1509529255511222

Abstract Details

2017, Doctor of Philosophy, Ohio State University, Biomedical Sciences.
Chronic pancreatitis (CP) is a devastating disease characterized by persistent inflammation and fibrosis of the pancreas leading to exocrine and endocrine insufficiency and increased risk of malignancy. Unfortunately, diagnosis of CP remains difficult and no curative therapeutic options exist, largely due to a limited understanding of the cellular and molecular mediators of CP pathology. Recent findings implicate pancreatic stellate cells (PSC) as prominent mediators of the inflammatory and fibrotic phenotype observed during CP. We hypothesized that the pro-survival Jak2/STAT3 and MAPK pathways play a role in PSC activation and proliferation and that these pathways would serve as therapeutic targets to reduce the pathogenic activity of these cells during disease. In vitro, cultured PSC demonstrated activation of both the Jak2/STAT3 and MAPK pathways and robust secretion of several soluble immunomodulatory factors including monocyte chemoattractant protein 1 (MCP-1), interleukin 6 (IL-6), and vascular endothelial growth factor (VEGF). Treatment of PSC with the small molecule Jak1/2 inhibitor Ruxolitinib reduced STAT3 phosphorylation and decreased cell growth. Treated cells remained adherent and did not display poly (ADP-ribose) polymerase (PARP) cleavage by western blot, suggesting the effects of Ruxolitinib on cell growth are not pro-apoptotic. Instead, western blot and fluorescent microscopy demonstrate a dose-dependent decrease in alpha-smooth muscle actin (aSMA), a marker of PSC activation. Treatment with a MAPK pathway inhibitor (MEK162) had no effect on growth or activation. These data suggest that the Jak/STAT pathway, and not the MAPK pathway, functions to regulate PSC growth and activation, thereby representing a viable therapeutic target. To examine this hypothesis in vivo, we utilized the caerulein-induced murine model of chronic pancreatitis. Ruxolitinib was administered at 90mg/kg twice daily by oral gavage during the final week of caerulein treatment. Results show a trend toward increased serum amylase and lipase and significantly reduced acinar cell loss and fibrosis by immunohistochemistry. Together these data suggest that inhibition of Jak/STAT signaling may limit caerulein-induced pancreatic damage in this model. To develop an experimental method to identify novel therapeutic and diagnostic biomarkers for CP, we hypothesized that proteomic analysis of pancreatic tissue from the caerulein-induced murine model of CP would elucidate candidate biomarkers of disease. To test this hypothesis, we utilized formalin fixed, paraffin embedded pancreatic tissue from the caerulein-induced murine model of CP. From this tissue, acinar cells were collected by laser capture microdissection (LCM). Following protein extraction, samples were analyzed by mass spectrometry (LC-MS/MS) to reveal 134, 303 and 331 proteins that were significantly changed by >2 fold in the 1,3 and 6 week treated animals, respectively. Notably, increased extracellular matrix (ECM)-associated proteins, including collagen, laminin, and fibronectin and decreased exocrine enzymes including lipase, amylase, and elastase were observed in caerulein-treated mice. These protein changes align with known pathological changes in CP. Furthermore, proteomic signatures in human CP tissue correlated with our results in murine tissue. These results suggest the validity of this method for the identification of disease-relevant biomarkers. Overall, these studies improve methods for experimental biomarker identification and suggest the efficacy of targeting Jak2/STA3 signaling in CP.
Gregory Lesinski, PhD, MPH (Advisor)
Aharon Freud, MD, PhD (Advisor)
William Carson, III, MD (Committee Member)
Traci Wilgus, PhD (Committee Member)
174 p.
Biomedical Research; Immunology
chronic pancreatitis; IL-6; Jak; STAT; stroma; inflammation; pancreatic stellate cell; proteomics; biomarkers

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Citations

  • Komar, Komar, H. M. (2017). Identifying pathogenic stromal and acinar signaling for improved diagnosis and treatment of chronic pancreatitis [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1509529255511222

    APA Style (7th edition)

  • Komar, Komar, Hannah. Identifying pathogenic stromal and acinar signaling for improved diagnosis and treatment of chronic pancreatitis. 2017. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1509529255511222.

    MLA Style (8th edition)

  • Komar, Komar, Hannah. "Identifying pathogenic stromal and acinar signaling for improved diagnosis and treatment of chronic pancreatitis." Doctoral dissertation, Ohio State University, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=osu1509529255511222

    Chicago Manual of Style (17th edition)

osu1509529255511222
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This open access ETD is published by The Ohio State University and OhioLINK.