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Functional impairment of central memory CD4 T cells is a potential early prognostic marker for changing viral load in SHIV-infected rhesus macaques.

He H, Nehete PN, Nehete B, Wieder E, Yang G, Buchl S, Sastry KJ - PLoS ONE (2011)

Bottom Line: Relative to long-term non-progressors with low/undetectable viral loads, those with chronic plasma viremia, but clinically healthy, exhibited significantly lower numbers and functional impairment of CD4(+) T cells, but not CD8(+) T cells, in terms of IL-2 production by central memory subset in response to PMA and ionomycine (PMA+I) stimulation.Highly viremic animals showed impaired cytokine-production by all T cells subsets.These results suggest that functional impairment of CD4(+) T cells in general, and of central memory subset in particular, may be a potential indicator/predictor of chronic infection with immune dysfunction, which could be assayed relatively easily using non-specific PMA+I stimulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas, United States of America.

ABSTRACT
In HIV infection there is a paucity of literature about the degree of immune dysfunction to potentially correlate and/or predict disease progression relative to CD4(+) T cells count or viral load. We assessed functional characteristics of memory T cells subsets as potential prognostic markers for changing viral loads and/or disease progression using the SHIV-infected rhesus macaque model. Relative to long-term non-progressors with low/undetectable viral loads, those with chronic plasma viremia, but clinically healthy, exhibited significantly lower numbers and functional impairment of CD4(+) T cells, but not CD8(+) T cells, in terms of IL-2 production by central memory subset in response to PMA and ionomycine (PMA+I) stimulation. Highly viremic animals showed impaired cytokine-production by all T cells subsets. These results suggest that functional impairment of CD4(+) T cells in general, and of central memory subset in particular, may be a potential indicator/predictor of chronic infection with immune dysfunction, which could be assayed relatively easily using non-specific PMA+I stimulation.

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Gating scheme for the analyses of the different T cell subsets in the                        peripheral blood mononuclear cells (PBMC) from a representative                        animal.The lymphocytes were first gated based on forward scatter (FCS) versus side                        scatter (SSC), and then live lymphocytes were identified based on SSC and                        aqua-negative population. The T cells were then positively identified by CD3                        expression followed by the detection of the CD4+                            CD8− (CD4+ T cells) and                            CD4− CD8+ (CD8+ T                        cells) populations within the CD3+ T cells. On the basis of                        CD28 and CD95 expression, the CD4+ and CD8+                        T cells were further differentiated into naïve (Tn                            CD28+ CD95−), central memory (Tcm                            CD28+ CD95+) and effector memory (Tem                            CD28− CD95+) subsets.
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pone-0019607-g001: Gating scheme for the analyses of the different T cell subsets in the peripheral blood mononuclear cells (PBMC) from a representative animal.The lymphocytes were first gated based on forward scatter (FCS) versus side scatter (SSC), and then live lymphocytes were identified based on SSC and aqua-negative population. The T cells were then positively identified by CD3 expression followed by the detection of the CD4+ CD8− (CD4+ T cells) and CD4− CD8+ (CD8+ T cells) populations within the CD3+ T cells. On the basis of CD28 and CD95 expression, the CD4+ and CD8+ T cells were further differentiated into naïve (Tn CD28+ CD95−), central memory (Tcm CD28+ CD95+) and effector memory (Tem CD28− CD95+) subsets.

Mentions: We used multicolor flow cytometry analyses to investigate the functional features paralleling the circulating total and memory subsets of CD4+ and CD8+ T cells in SHIV-infected rhesus macaques exhibiting varying levels of plasma viral loads along with or without clinical symptoms. Peripheral blood mononuclear cells (PBMC) collected from a total of 20 rhesus macaques at the necropsy and preserved at liquid nitrogen were used for the multicolor flow cytometry analyses. These 20 animals were used in the past for vaccine studies [12], [13] and changes in viral loads and CD4+ T cell counts were recorded for approximately one year post-infection with SHIV (Fig. S1). Based on the viral loads and presence or absence of clinical symptoms at necropsy the animals were grouped as long-term non-progressors (LTNP, n = 10), chronically infected but healthy (Chronic, n = 5), and symptomatic with high viral loads (Viremic, n = 5), as described in the Methods section. The gating scheme used for the eight-color flow cytometry analyses of the different T cells subsets from a representative animal is shown in Fig. 1. The lymphocytes were first gated based on forward scatter (FSC) versus side scatter (SSC) with the dead cells excluded using the violet live/dead stain. The live T cells were then positively identified based on CD3 expression followed by detecting CD4+ CD8− (CD4+ T cells) and CD4− CD8+ (CD8+ T cells) populations. On the basis of CD28 and CD95 expression, the CD4+ and CD8+ T cells were further differentiated into naïve (Tn CD28+ CD95−), central memory (Tcm CD28+ CD95+) and effector memory (Tem CD28− CD95+) subsets as described in the literature [15]. Next, the total and the different subsets of CD4+ and CD8+ T cells were assessed for functional capacity in terms of cytokine production (IFN-γ and/or IL-2) in response to stimulation with PMA and ionomycin (PMA+I). This analysis enabled identification of three functionally distinct populations of total as well as memory subsets of CD4+, CD8+ T cells, each producing IFN-γ and IL-2 alone or the two cytokines together (Fig. S2). Since these cytokines are important for sustaining memory (IL-2) and mediating effector function (IFN-γ), this analysis provides a snap shot of the quantity and quality of the inherent cell-mediated immunity in relation to viral load in the three different groups of monkeys studied.


Functional impairment of central memory CD4 T cells is a potential early prognostic marker for changing viral load in SHIV-infected rhesus macaques.

He H, Nehete PN, Nehete B, Wieder E, Yang G, Buchl S, Sastry KJ - PLoS ONE (2011)

Gating scheme for the analyses of the different T cell subsets in the                        peripheral blood mononuclear cells (PBMC) from a representative                        animal.The lymphocytes were first gated based on forward scatter (FCS) versus side                        scatter (SSC), and then live lymphocytes were identified based on SSC and                        aqua-negative population. The T cells were then positively identified by CD3                        expression followed by the detection of the CD4+                            CD8− (CD4+ T cells) and                            CD4− CD8+ (CD8+ T                        cells) populations within the CD3+ T cells. On the basis of                        CD28 and CD95 expression, the CD4+ and CD8+                        T cells were further differentiated into naïve (Tn                            CD28+ CD95−), central memory (Tcm                            CD28+ CD95+) and effector memory (Tem                            CD28− CD95+) subsets.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3094340&req=5

pone-0019607-g001: Gating scheme for the analyses of the different T cell subsets in the peripheral blood mononuclear cells (PBMC) from a representative animal.The lymphocytes were first gated based on forward scatter (FCS) versus side scatter (SSC), and then live lymphocytes were identified based on SSC and aqua-negative population. The T cells were then positively identified by CD3 expression followed by the detection of the CD4+ CD8− (CD4+ T cells) and CD4− CD8+ (CD8+ T cells) populations within the CD3+ T cells. On the basis of CD28 and CD95 expression, the CD4+ and CD8+ T cells were further differentiated into naïve (Tn CD28+ CD95−), central memory (Tcm CD28+ CD95+) and effector memory (Tem CD28− CD95+) subsets.
Mentions: We used multicolor flow cytometry analyses to investigate the functional features paralleling the circulating total and memory subsets of CD4+ and CD8+ T cells in SHIV-infected rhesus macaques exhibiting varying levels of plasma viral loads along with or without clinical symptoms. Peripheral blood mononuclear cells (PBMC) collected from a total of 20 rhesus macaques at the necropsy and preserved at liquid nitrogen were used for the multicolor flow cytometry analyses. These 20 animals were used in the past for vaccine studies [12], [13] and changes in viral loads and CD4+ T cell counts were recorded for approximately one year post-infection with SHIV (Fig. S1). Based on the viral loads and presence or absence of clinical symptoms at necropsy the animals were grouped as long-term non-progressors (LTNP, n = 10), chronically infected but healthy (Chronic, n = 5), and symptomatic with high viral loads (Viremic, n = 5), as described in the Methods section. The gating scheme used for the eight-color flow cytometry analyses of the different T cells subsets from a representative animal is shown in Fig. 1. The lymphocytes were first gated based on forward scatter (FSC) versus side scatter (SSC) with the dead cells excluded using the violet live/dead stain. The live T cells were then positively identified based on CD3 expression followed by detecting CD4+ CD8− (CD4+ T cells) and CD4− CD8+ (CD8+ T cells) populations. On the basis of CD28 and CD95 expression, the CD4+ and CD8+ T cells were further differentiated into naïve (Tn CD28+ CD95−), central memory (Tcm CD28+ CD95+) and effector memory (Tem CD28− CD95+) subsets as described in the literature [15]. Next, the total and the different subsets of CD4+ and CD8+ T cells were assessed for functional capacity in terms of cytokine production (IFN-γ and/or IL-2) in response to stimulation with PMA and ionomycin (PMA+I). This analysis enabled identification of three functionally distinct populations of total as well as memory subsets of CD4+, CD8+ T cells, each producing IFN-γ and IL-2 alone or the two cytokines together (Fig. S2). Since these cytokines are important for sustaining memory (IL-2) and mediating effector function (IFN-γ), this analysis provides a snap shot of the quantity and quality of the inherent cell-mediated immunity in relation to viral load in the three different groups of monkeys studied.

Bottom Line: Relative to long-term non-progressors with low/undetectable viral loads, those with chronic plasma viremia, but clinically healthy, exhibited significantly lower numbers and functional impairment of CD4(+) T cells, but not CD8(+) T cells, in terms of IL-2 production by central memory subset in response to PMA and ionomycine (PMA+I) stimulation.Highly viremic animals showed impaired cytokine-production by all T cells subsets.These results suggest that functional impairment of CD4(+) T cells in general, and of central memory subset in particular, may be a potential indicator/predictor of chronic infection with immune dysfunction, which could be assayed relatively easily using non-specific PMA+I stimulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas, United States of America.

ABSTRACT
In HIV infection there is a paucity of literature about the degree of immune dysfunction to potentially correlate and/or predict disease progression relative to CD4(+) T cells count or viral load. We assessed functional characteristics of memory T cells subsets as potential prognostic markers for changing viral loads and/or disease progression using the SHIV-infected rhesus macaque model. Relative to long-term non-progressors with low/undetectable viral loads, those with chronic plasma viremia, but clinically healthy, exhibited significantly lower numbers and functional impairment of CD4(+) T cells, but not CD8(+) T cells, in terms of IL-2 production by central memory subset in response to PMA and ionomycine (PMA+I) stimulation. Highly viremic animals showed impaired cytokine-production by all T cells subsets. These results suggest that functional impairment of CD4(+) T cells in general, and of central memory subset in particular, may be a potential indicator/predictor of chronic infection with immune dysfunction, which could be assayed relatively easily using non-specific PMA+I stimulation.

Show MeSH
Related in: MedlinePlus