Date of Award

January 2025

Document Type

Thesis

Degree Name

Medical Doctor (MD)

Department

Medicine

First Advisor

Leonel Rodriguez

Abstract

This project aims to identify tissue biomarkers that correlate with clinical presentations of disorders of the gut-brain interactions by analyzing mRNA expression of neuromodulators, neurotransmitters, and their receptors in mucosal tissue biopsy samples.This study recruited pediatric patients with Rome Criteria IV- diagnosed disorders of the gut-brain interactions, as well as asymptomatic controls, who were scheduled for esophagogastroduodenoscopy (EGD) or colonoscopy at Yale New Haven Hospital. Mucosal biopsy samples were collected from the gastric fundus, gastric antrum, and duodenum in those undergoing EGDs, and from the right and left colon in those undergoing colonoscopies. Tissue samples were frozen at-80 C until further processing. Total RNA was extracted and purified from each sample utilizing Qiagen’s RNeasy Mini Kit. NanoString technology was used for the analysis of gene expression, utilizing a customized cassette with 90 targets selected based on animal and human studies of known and potentially associated neurotransmitters, neuromodulators, and receptors affecting gastrointestinal motility and sensory functions. Sequence-specific mRNA probes were used to directly detect gene expression levels with a digital count of the target molecules. Digital counts were compared for each biopsy site between the groups using non-parametric testing. A total of 36 upper gastrointestinal (GI) biopsies from twelve patients, including two asymptomatic controls, were analyzed for neuromodulator gene expression. Our analysis demonstrates that neurotensin peptide expression was significantly elevated in the fundus of the stomach in symptomatic patients compared to controls (p = 0.030). No significant differences were found in the gastric antrum or duodenal samples. A total of 40 lower GI biopsies from 20 patients, including five asymptomatic controls, were analyzed for neuromodulator gene expression. Our analysis shows that there was a significant difference in the expression of neuropeptide Y receptor Y1 (p = 0.019), neuropeptide Y receptor Y2 (p = 0.044), vasoactive intestinal peptide (VIP) (p = 0.019), VIP exon 1 (p= 0.010), VIP exon 2 (P = 0.014), VIP exon 7 (p = 0.019), substance P receptor exon 5 (p = 0.034) in the right colon for symptomatic patients versus controls. When stratifying by Rome IV criteria-based diagnosis, there was a significant difference in the right colon between controls and patients with functional constipation (FC) in the following neurotransmitters: adrenoreceptor alpha 2A (p = 0.042), histamine receptor H1 (p = 0.042), neuropeptide Y receptor Y1 (p < 0.001), neuropeptide Y receptor Y2 (p = 0.007), neurotensin peptide (p = 0.012), Neurturin (p = 0.019), substance P receptor exon 5 (p = 0.029), VIP (p = 0.012), VIP exon 1 (p = 0.04), VIP exon 2 (p = 0.04), VIP exon 7 (p = 0.012). In contrast, no significant differences were found between controls and patients with irritable bowel syndrome (IBS) in the right colon. On the other hand, analysis of the left colon showed no significant differences in neurotransmitter expression between symptomatic and control patients overall. However, Rome-criteria based stratification revealed a significant increase in somatostatin receptor 4 expression in IBS (p = 0.016). Adrenoreceptor alpha 2A and neurotensin receptor type 1 were significantly altered in FC (p = 0.013 for both). This study highlights significant dysregulations in neurotransmitter and neuromodulator expression in the gastrointestinal tract mucosa of patients with disorders of gut-brain interactions (DGBIs). In the upper gastrointestinal tract, elevated neurotensin peptide levels in the gastric fundus were associated with DGBIs. Neurotensin has been implicated in modulating gastric motility, inhibiting acid secretion, and regulating appetite, and our results suggest that neurotensin dysregulation could be a feature of upper GI DGBI pathophysiology. In the lower GI tract, distinct differences in neurotransmitter expression were observed between the right and left colon. The right colon generally exhibited more pronounced deviations in the symptomatic state, which could potentially indicate greater involvement of the right colon in DGBI pathophysiology. Significant alterations were notably found in neuropeptide Y receptors, VIP, and substance P receptor in the right colon in patients with functional constipation, but not IBS. These results suggest that different DGBI subtypes are characterized by distinct neuromodulator profiles, which may provide insight into their pathophysiology and potential future avenues for uncovering specific biomarkers and therapeutic targets.

Comments

This thesis is restricted to Yale network users only. It will be made publicly available on 05/14/2027

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