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Characterization of a gp130 Signaling Receptor Polymorphism

Kercher, Katherine (2013) Characterization of a gp130 Signaling Receptor Polymorphism. Master's Thesis, University of Pittsburgh. (Unpublished)

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Abstract

We previously reported an association between human herpesvirus 8 (HHV-8) seroprevalence and increased prostate cancer risk among men on the Caribbean island of Tobago. More recently, we have found a single nucleotide polymorphism (SNP) in the IL-6 gp130 signaling receptor at position 148 associated with increased prostate cancer risk among HHV-8 seropositive men. The high risk genotype (Gly) was associated with increased prostate cancer risk among HHV-8 seropositive men (OR= 3.1) compared to the low risk genotype (Arg). This research aims to further explore the effect of this SNP on gp130 function.
The gp130 genotype at position 148 was determined in lymphoblastoid B cell lines (LCLs) derived from Tobago men (representing high and low risk genotypes) and two prostate cancer cell lines (PC3 and DU 145; high risk and low risk respectively). Growth curves were performed by Dr. Jill Henning for LCLs by using 25ng/mL of Interleukin-6 (IL-6), Interleukin-11 (IL-11), or Oncostatin M (OSM). It was discovered that IL-6 had an effect on the growth of LCLs, but IL-11 and OSM did not. I repeated growth curves on LCLs using a concentration of 10ng/mL, and found that there was still a difference in growth at this lower concentration. Levels of phosphorylated STAT3 were measured on cells treated with 10 or 100ng/mL IL-6, IL-11, or OSM for various times (2-60 minutes). Comparative IL-6-mediated downstream signaling between the two genotypes was analyzed in LCLs at 10 minutes post-treatment using the JAK/STAT Pathway PCR Array Plates (SABiosciences, Valencia, CA), and the Human IL-6 Pathway PCR Array Plates (Life Technologies, Grand Island, NY).
LCLs homozygous for the high-risk gp130 genotype grew significantly faster compared to LCLs homozygous for the low-risk genotype in response to IL-6 but not IL-11 or OSM. LCLs homozygous for both high-risk gp130 genotype as well as LCLs homozygous for the low-risk genotype both showed activation of STAT3 in response to IL-6 by 10 minutes post-treatment. DU 145 (low-risk genotype) cells showed STAT3 activation following IL-6 treatment while PC3 (high-risk genotype) failed to show any STAT3 activation even after 1hr of IL-6 treatment. Both cell lines showed STAT3 activation after OSM treatment while neither line showed activation following IL-11 treatment. RT-PCR analyses of JAK/STAT pathway genes and IL-6 pathway genes in LCLs following IL-6 treatment showed differential gene regulation between genotypes. For example, using the IL-6 pathway plates, the high-risk genotype showed a down-regulation of TP53BP2 (apoptosis stimulating protein of p53-2), while the low-risk genotype showed an up-regulation of this gene. This protein is known to inhibit cell growth and stimulate apoptosis, and is frequently suppressed in human cancers. This differential gene regulation of TP53BP2 is one example of a gene that is differentially regulated between the high- and low-risk genotypes. These data suggest that the G148R SNP of gp130 is involved in cell proliferation mediated by IL-6 downstream signaling.
The public health relevance observed by these results suggests that the resulting genotypes in the G148R polymorphism exhibit different biological affects upon treatment with cytokines that utilize the gp130 signaling receptor. The high-risk genotype could result in an increase in inflammation, which could ultimately contribute to the development or advancement of prostate cancer.


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Details

Item Type: University of Pittsburgh ETD
Status: Unpublished
Creators/Authors:
CreatorsEmailPitt UsernameORCID
Kercher, Katherinekak225@pitt.eduKAK225
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorJenkins, Frankfjenkins@pitt.eduFJENKINS
Committee MemberReinhart, Todd A.reinhar@pitt.eduREINHAR
Committee MemberFerrell, Robert E.rferrell@pitt.eduRFERRELL
Committee MemberBunker, Clareannbunkerc@pitt.eduBUNKERC
Date: 27 September 2013
Date Type: Publication
Defense Date: 8 July 2013
Approval Date: 27 September 2013
Submission Date: 22 July 2013
Access Restriction: No restriction; Release the ETD for access worldwide immediately.
Number of Pages: 109
Institution: University of Pittsburgh
Schools and Programs: Graduate School of Public Health > Infectious Diseases and Microbiology
Degree: MS - Master of Science
Thesis Type: Master's Thesis
Refereed: Yes
Uncontrolled Keywords: gp130, single nucleotide polymorphism, G148R
Date Deposited: 27 Sep 2013 14:35
Last Modified: 15 Nov 2016 14:14
URI: http://d-scholarship.pitt.edu/id/eprint/19409

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