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Investigating the role of Nef in HIV infection-induced dysregulation of innate immune cell function

Senthilkumar, Shreyaa (2023) Investigating the role of Nef in HIV infection-induced dysregulation of innate immune cell function. Master's Thesis, University of Pittsburgh. (Unpublished)

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Nef is an accessory protein of HIV that is associated with promoting HIV pathogenesis through increasing replication and immune evasion. This has been proven through delta Nef mutation studies which caused low viral load and no development of AIDS. However, Nef isn't an easy target for creating a therapeutic drug due to its lack of enzymatic activity. Here we examined HIV expression in its wildtype and delta Nef mutated forms in humanized mice. The BLTS model, which has been a cornerstone in aiding HIV studies, was re-examined here for functionality. This was done using immunohistochemistry (IHC) staining of healthy BLTS spleen tissues to analyze characterize the major human immune cells, especially T and B cells. The model was then infected with WT and delta Nef HIV types to analyze the differences in infectivity. This was detected through IHC which detects proteins and RNAScope which detects RNA genome. Wildtype HIV was found to have a higher viral infectivity and tissue damage within 2 weeks post infection (wpi) whereas delta Nef mutated HIV took nearly two months post infection (mpi) to reach lesser infectivity, viral genome load and almost no structural damage. Here, we also introduce the HSC-humanized mice model which has different transplant origins from the BLTS mice. This model was phenotyped using flow cytometry and assessed for its functionality through T cell stimulation assays, which gave a positive functionality indicating that this model, once further optimized, can be used for HIV studies. Finally, we also determine the tissue resident presence of natural killer (NK) cells and dendritic cells (DCs) in the BLTS spleen and thymus and their localization within the tissue. This will allow us to further study the effect of Nef on the localization of these crucial innate immune cells and determine how innate immunity combined with defective Nef could potentially be used to suppress the viral reservoirs of HIV.


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Item Type: University of Pittsburgh ETD
Status: Unpublished
CreatorsEmailPitt UsernameORCID
Senthilkumar, Shreyaashs259@pitt.edushs2590009-0000-8823-5207
ETD Committee:
TitleMemberEmail AddressPitt UsernameORCID
Thesis AdvisorBility, Dr. Moses T.mtbility@pitt.edumtbility0000-0001-5153-2718
Committee MemberMailliard, Dr. Robbie B.rbm19@pitt.edurbm190000-0001-5501-503X
Committee MemberMattila, Dr. Joshua Tjmattila@pitt.edujmattila
Date: 28 June 2023
Date Type: Publication
Defense Date: 23 June 2023
Approval Date: 28 June 2023
Submission Date: 23 June 2023
Access Restriction: 1 year -- Restrict access to University of Pittsburgh for a period of 1 year.
Number of Pages: 58
Institution: University of Pittsburgh
Schools and Programs: School of Public Health > Infectious Diseases and Microbiology
Degree: MS - Master of Science
Thesis Type: Master's Thesis
Refereed: Yes
Uncontrolled Keywords: HIV, Nef, Humanized mouse model, BLTS, NK-DC Crosstalk
Date Deposited: 28 Jun 2023 17:54
Last Modified: 28 Jun 2023 17:54


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