5 results on '"Ouwendijk, Werner J.D."'
Search Results
2. Herpes simplex virus infection induces necroptosis of neurons and astrocytes in human fetal organotypic brain slice cultures
- Author
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Rashidi, Ahmad S., Tran, Diana N., Peelen, Caithlin R., van Gent, Michiel, Ouwendijk, Werner J.D., Verjans, Georges M.G.M., Rashidi, Ahmad S., Tran, Diana N., Peelen, Caithlin R., van Gent, Michiel, Ouwendijk, Werner J.D., and Verjans, Georges M.G.M.
- Abstract
Background: Herpes simplex virus (HSV) encephalitis (HSE) is a serious and potentially life-threatening disease, affecting both adults and newborns. Progress in understanding the virus and host factors involved in neonatal HSE has been hampered by the limitations of current brain models that do not fully recapitulate the tissue structure and cell composition of the developing human brain in health and disease. Here, we developed a human fetal organotypic brain slice culture (hfOBSC) model and determined its value in mimicking the HSE neuropathology in vitro. Methods: Cell viability and tissues integrity were determined by lactate dehydrogenase release in supernatant and immunohistological (IHC) analyses. Brain slices were infected with green fluorescent protein (GFP-) expressing HSV-1 and HSV-2. Virus replication and spread were determined by confocal microscopy, PCR and virus culture. Expression of pro-inflammatory cytokines and chemokines were detected by PCR. Cell tropism and HSV-induced neuropathology were determined by IHC analysis. Finally, the in situ data of HSV-infected hfOBSC were compared to the neuropathology detected in human HSE brain sections. Results: Slicing and serum-free culture conditions were optimized to maintain the viability and tissue architecture of ex vivo human fetal brain slices for at least 14 days at 37 °C in a CO2 incubator. The hfOBSC supported productive HSV-1 and HSV-2 infection, involving predominantly infection of neurons and astrocytes, leading to expression of pro-inflammatory cytokines and chemokines. Both viruses induced programmed cell death—especially necroptosis—in infected brain slices at later time points after infection. The virus spread, cell tropism and role of programmed cell death in HSV-induced cell death resembled the neuropathology of HSE. Conclusions: We developed a novel human brain culture model in which the viability of the major brain-resident cells—including neurons, microglia, astrocytes and
- Published
- 2024
3. Varicella-zoster virus proteome-wide T-cell screening demonstrates low prevalence of virus-specific CD8 T-cells in latently infected human trigeminal ganglia
- Author
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van Gent, Michiel, Ouwendijk, Werner J.D., Campbell, Victoria L., Laing, Kerry J., Verjans, Georges M.G.M., Koelle, David M., van Gent, Michiel, Ouwendijk, Werner J.D., Campbell, Victoria L., Laing, Kerry J., Verjans, Georges M.G.M., and Koelle, David M.
- Abstract
Background: Trigeminal ganglia (TG) neurons are an important site of lifelong latent varicella-zoster virus (VZV) infection. Although VZV-specific T-cells are considered pivotal to control virus reactivation, their protective role at the site of latency remains uncharacterized. Methods: Paired blood and TG specimens were obtained from ten latent VZV-infected adults, of which nine were co-infected with herpes simplex virus type 1 (HSV-1). Short-term TG-derived T-cell lines (TG-TCL), generated by mitogenic stimulation of TG-derived T-cells, were probed for HSV-1- and VZV-specific T-cells using flow cytometry. We also performed VZV proteome-wide screening of TG-TCL to determine the fine antigenic specificity of VZV reactive T-cells. Finally, the relationship between T-cells and latent HSV-1 and VZV infections in TG was analyzed by reverse transcription quantitative PCR (RT-qPCR) and in situ analysis for T-cell proteins and latent viral transcripts. Results: VZV proteome-wide analysis of ten TG-TCL identified two VZV antigens recognized by CD8 T-cells in two separate subjects. The first was an HSV-1/VZV cross-reactive CD8 T-cell epitope, whereas the second TG harbored CD8 T-cells reactive with VZV specifically and not the homologous peptide in HSV-1. In silico analysis showed that HSV-1/VZV cross reactivity of TG-derived CD8 T-cells reactive with ten previously identified HSV-1 epitopes was unlikely, suggesting that HSV-1/VZV cross-reactive T-cells are not a common feature in dually infected TG. Finally, no association was detected between T-cell infiltration and VZV latency transcript abundance in TG by RT-qPCR or in situ analyses. Conclusions: The low presence of VZV- compared to HSV-1-specific CD8 T-cells in human TG suggests that VZV reactive CD8 T-cells play a limited role in maintaining VZV latency.
- Published
- 2023
4. T-cells in human trigeminal ganglia express canonical tissue-resident memory T-cell markers
- Author
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Unger, Peter Paul A., Oja, Anna E., Khemai-Mehraban, Tamana, Ouwendijk, Werner J.D., Hombrink, Pleun, Verjans, Georges M.G.M., Landsteiner Laboratory, Neurology, and Virology
- Subjects
General Neuroscience ,Immunology ,Programmed Cell Death 1 Receptor ,Herpes Simplex ,Herpesviridae Infections ,Herpesvirus 1, Human ,CD8-Positive T-Lymphocytes ,Normal-appearing white matter ,Cellular and Molecular Neuroscience ,Memory T Cells ,Ki-67 Antigen ,Neurology ,SDG 3 - Good Health and Well-being ,Trigeminal Ganglion ,Leukocytes, Mononuclear ,Humans ,Leukocyte Common Antigens ,Tissue-resident memory T-cells and herpes simplex virus ,Human - Abstract
Background Trigeminal ganglia (TG) neurons are the main site of lifelong latent herpes simplex virus type 1 (HSV-1) infection. T-cells in ganglia contribute to long-term control of latent HSV-1 infection, but it is unclear whether these cells are bona fide tissue-resident memory T-cells (TRM). We optimized the processing of human post-mortem nervous tissue to accurately phenotype T-cells in human TG ex vivo and in situ. Methods Peripheral blood mononuclear cells (PBMC; 5 blood donors) were incubated with several commercial tissue digestion enzyme preparations to determine off-target effect on simultaneous detection of 15 specific T-cell subset markers by flow cytometry. Next, optimized enzymatic digestion was applied to ex vivo phenotype T-cells in paired PBMC, normal appearing white matter (NAWM) and TG of 8 deceased brain donors obtained Results Collagenase IV digestion of human nervous tissue was most optimal to obtain high numbers of viable T-cells without disrupting marker surface expression. Compared to blood, majority T-cells in paired NAWM and TG were effector memory T-cells expressing the canonical TRM markers CD69, CXCR6 and the immune checkpoint marker PD1, and about half co-expressed CD103. A trend of relatively higher TRM frequencies were detected in TG of latently HSV-1-infected compared to HSV-1 naïve individuals. Subsequent in situ analysis of latently HSV-1-infected TG showed the presence of cytotoxic T-cells (TIA-1+), which occasionally showed features of proliferation (KI-67+) and activation (CD137+), but without signs of degranulation (CD107a+) nor damage (TUNEL+) of TG cells. Whereas majority T-cells expressed PD-1, traits of T-cell senescence (p16INK4a+) were not detected. Conclusions The human TG represents an immunocompetent environment in which both CD4 and CD8 TRM are established and retained. Based on our study insights, we advocate for TRM-targeted vaccine strategies to bolster local HSV-1-specific T-cell immunity, not only at the site of recurrent infection but also at the site of HSV-1 latency.
- Published
- 2022
5. Selective retention of virus-specific tissue-resident T cells in healed skin after recovery from herpes zoster
- Author
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Laing, Kerry J., Ouwendijk, Werner J.D., Campbell, Victoria L., McClurkan, Christopher L., Mortazavi, Shahin, Elder Waters, Michael, Krist, Maxwell P., Tu, Richard, Nguyen, Nhi, Basu, Krithi, Miao, Congrong, Schmid, D. Scott, Johnston, Christine, Verjans, Georges M.G.M., Koelle, David M., Laing, Kerry J., Ouwendijk, Werner J.D., Campbell, Victoria L., McClurkan, Christopher L., Mortazavi, Shahin, Elder Waters, Michael, Krist, Maxwell P., Tu, Richard, Nguyen, Nhi, Basu, Krithi, Miao, Congrong, Schmid, D. Scott, Johnston, Christine, Verjans, Georges M.G.M., and Koelle, David M.
- Abstract
Herpes zoster is a localized skin infection caused by reactivation of latent varicella-zoster virus. Tissue-resident T cells likely control skin infections. Zoster provides a unique opportunity to determine if focal reinfection of human skin boosts local or disseminated antigen-specific tissue-resident T cells. Here, we show virus-specific T cells are retained over one year in serial samples of rash site and contralateral unaffected skin of individuals recovered from zoster. Consistent with zoster resolution, viral DNA is largely undetectable on skin from day 90 and virus-specific B and T cells decline in blood. In skin, there is selective infiltration and long-term persistence of varicella-zoster virus-specific T cells in the rash site relative to the contralateral site. The skin T cell infiltrates express the canonical tissue-resident T cell markers CD69 and CD103. These findings show that zoster promotes spatially-restricted long-term retention of antigen-specific tissue-resident T cells in previously infected skin.
- Published
- 2022
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