Date of Award

January 2016

Document Type

Thesis

Degree Name

Medical Doctor (MD)

Department

Medicine

First Advisor

Robert A. Cowles

Abstract

Background: Serotonin (5-HT) is an important signaling molecule in the enteric nervous system, and it has been shown to be a powerful mediator of mucosal growth and enterocyte turnover. Vincristine is a commonly used chemotherapeutic agent that induces apoptosis in mitotically active cells, such as intestinal crypt cells. Deletion of the serotonin reuptake transporter (SERT) or the use of selective serotonin reuptake inhibitors (SSRIs) results in 5-HT excess and increased growth parameters, including villus height and crypt proliferation. We hypothesize that enhanced enteric 5-HT stimulates mucosal proliferation and mitigates vincristine-induced epithelial injury.

Methods: Wild type (WT) mice and mice lacking the serotonin reuptake transporter (SERT-KO) received a single intraperitoneal injection of vincristine (2 mg/kg) or saline and were euthanized 2, 4, 6, or 14 days later. In another group, WT mice received the SSRI citalopram (10 mg/kg) subcutaneously for 24 hours prior to vincristine injection and throughout the experiment. N=2 to 4 mice per condition. All mice were weighed daily. Terminal ileum was harvested, fixed, and stained. Outcome measures include body weight (%Basal), mucosal injury score, villus height (VH), crypt cell count (CCC), apoptotic index (AI), and crypt proliferation index (CPI).

Results: Vincristine caused weight loss in all groups (WT 87%, SERT-KO 90%, WT+SSRI 88%). Vincristine injection resulted in mild, but significant, mucosal injury as measured by an injury score (WT 0.8vs0.1*; SERT-KO 0.6vs0.2*), and 5-HT excess mitigated injury (SERT-KO 0.6vs0.8*; WT+SSRI 0.4vs0.8*). While vincristine caused a reduction in VH (WT 194vs172um*; SERT-KO 236vs214um*), SERT inhibition resulted in VH preservation (SERT-KO 214vs172um*; WT+SSRI 240vs172um*). Vincristine decreased CCC (WT 22vs20*; SERT-KO 29vs24*), and mice with excess 5-HT had overall higher CCC after injury (SERT-KO 29vs20*; WT+SSRI 26vs20*). Following vincristine injury, AI was significantly lower in mice with excess 5-HT compared to WT mice (SERT-KO 0.08vs0.12*; WT+SSRI 0.071vs0.12*). After injury, CPI was consistently elevated in WT+SSRI mice, whereas WT and SERT-KO mice exhibited a low CPI followed by a sharp increase. (*indicates p<0.05)

Discussion: Vincristine injured the intestinal mucosa in all groups of mice. These changes were predominantly focused in the crypt compartment, which is the site of active cellular proliferation and turnover. Vincristine’s injurious effects were mitigated by SERT inhibition, achieved by genetic and pharmaceutical means. Administration of SSRIs had a stronger effect that genetic knockout. This may be due to upregulation of alternative transporters in the knockout mice that partially clear serotonin in the absence of SERT. This work provides further evidence of serotonin’s ability to enhance enterocyte proliferation. Serotonin potentiation may be an important treatment target for promoting intestinal healing and regeneration.

Comments

This thesis is restricted to Yale network users only. This thesis is permanently embargoed from public release.

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