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Lymphatic Endothelial Cells Express Viral Entry Receptors and Restriction Factors

Bowen, Christopher David (2011) Lymphatic Endothelial Cells Express Viral Entry Receptors and Restriction Factors. Master's Thesis, University of Pittsburgh.

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    Abstract

    Lymphatic endothelial cells (LECs) line lymphatic vessels and are present at mucosal portals of entry for many pathogens, including simian immunodeficiency virus (SIV) and human immunodeficiency virus type-1 (HIV-1). Recent studies have shown that LECs express pattern recognition receptors (PRRs), such as Toll-like receptors (TLRs), capable of recognizing pathogen-associated molecular patterns (PAMPs). PAMPs are structurally similar molecules expressed by groups of pathogens. LECs have also been shown to express chemokines, a group of small molecules secreted by cells that induce chemotaxis in responsive cells, such as CCL21, which is used by CCR7+ mature antigen presenting dendritic cells (DCs) to migrate to draining lymph nodes (LNs). These previous findings indicate that LECs might play an integral role in innate immune responses to a wide variety of microbes. In this study, I set out to characterize the expression of antiviral restriction factors as well as possible viral entry receptors for SIV/HIV-1 within three populations of human LECs. Real-time RT-PCR and immunofluorescent staining techniques were used to determine the relative expression of the restriction factors BST-2/Tetherin, APOBEC3G, and TRIM5-á. All of these factors have been shown to inhibit the replicative cycle of HIV-1 and have othologs present in nonhuman primates (NHPs). Expression of the viral entry receptors CD4, CXCR4, CCR5, DEC-205/CD205, D6/CCBP2, and CD209 as well as the LEC-specific markers podoplanin and LYVE-1 was also investigated. In addition, LEC populations were exposed to SIV, HIV-1, and markers internalization to determine to what extent LECs interact with virus in vitro. Data from populations exposed to HIV-1 as well as other substrates for internalization of extracellular materials illustrate the ability of LECs to actively monitor the extracellular milieu. LECs exposed to SIV showed multi-spliced viral transcripts possibly due to de novo transcription. Taken together, this study provides evidence that LECs are equipped with tools not only to bind and internalize pathogens, but may also serve as a low-level replicative cellular substrate for virus. Further studies to characterize LECs are of great public health relevance, particularly at mucosa sites of microbial exposure, due to their potential roles during transmission/infection.


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    Item Type: University of Pittsburgh ETD
    ETD Committee:
    ETD Committee TypeCommittee MemberEmailORCID
    Committee ChairReinhart, Todd A.reinhar@pitt.edu
    Committee MemberFerrell, Robert E.rferrell@pitt.edu
    Committee MemberWang, Tianyitywang@pitt.edu
    Title: Lymphatic Endothelial Cells Express Viral Entry Receptors and Restriction Factors
    Status: Unpublished
    Abstract: Lymphatic endothelial cells (LECs) line lymphatic vessels and are present at mucosal portals of entry for many pathogens, including simian immunodeficiency virus (SIV) and human immunodeficiency virus type-1 (HIV-1). Recent studies have shown that LECs express pattern recognition receptors (PRRs), such as Toll-like receptors (TLRs), capable of recognizing pathogen-associated molecular patterns (PAMPs). PAMPs are structurally similar molecules expressed by groups of pathogens. LECs have also been shown to express chemokines, a group of small molecules secreted by cells that induce chemotaxis in responsive cells, such as CCL21, which is used by CCR7+ mature antigen presenting dendritic cells (DCs) to migrate to draining lymph nodes (LNs). These previous findings indicate that LECs might play an integral role in innate immune responses to a wide variety of microbes. In this study, I set out to characterize the expression of antiviral restriction factors as well as possible viral entry receptors for SIV/HIV-1 within three populations of human LECs. Real-time RT-PCR and immunofluorescent staining techniques were used to determine the relative expression of the restriction factors BST-2/Tetherin, APOBEC3G, and TRIM5-á. All of these factors have been shown to inhibit the replicative cycle of HIV-1 and have othologs present in nonhuman primates (NHPs). Expression of the viral entry receptors CD4, CXCR4, CCR5, DEC-205/CD205, D6/CCBP2, and CD209 as well as the LEC-specific markers podoplanin and LYVE-1 was also investigated. In addition, LEC populations were exposed to SIV, HIV-1, and markers internalization to determine to what extent LECs interact with virus in vitro. Data from populations exposed to HIV-1 as well as other substrates for internalization of extracellular materials illustrate the ability of LECs to actively monitor the extracellular milieu. LECs exposed to SIV showed multi-spliced viral transcripts possibly due to de novo transcription. Taken together, this study provides evidence that LECs are equipped with tools not only to bind and internalize pathogens, but may also serve as a low-level replicative cellular substrate for virus. Further studies to characterize LECs are of great public health relevance, particularly at mucosa sites of microbial exposure, due to their potential roles during transmission/infection.
    Date: 28 September 2011
    Date Type: Completion
    Defense Date: 14 July 2011
    Approval Date: 28 September 2011
    Submission Date: 05 July 2011
    Access Restriction: 5 year -- Restrict access to University of Pittsburgh for a period of 5 years.
    Patent pending: No
    Institution: University of Pittsburgh
    Thesis Type: Master's Thesis
    Refereed: Yes
    Degree: MS - Master of Science
    URN: etd-07052011-145351
    Uncontrolled Keywords: HIV-1; Lymphatic Endothelial Cells; SIV
    Schools and Programs: Graduate School of Public Health > Infectious Diseases and Microbiology
    Date Deposited: 10 Nov 2011 14:50
    Last Modified: 11 Jan 2012 14:05
    Other ID: http://etd.library.pitt.edu/ETD/available/etd-07052011-145351/, etd-07052011-145351

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