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Chronic-phase HIV and simian immunodeficiency virus (SIV) replication is reduced by as much as 10,000-fold in elite controllers (ECs) compared with typical progressors (TPs), but sufficient viral replication persists in EC tissues to allow viral sequence evolution and induce excess immune activation. Here we show that productive SIV infection in rhesus monkey ECs, but not TPs, is markedly restricted to CD4(+) follicular helper T (TFH) cells, suggesting that these EC monkeys' highly effective SIV-specific CD8(+) T cells can effectively clear productive SIV infection from extrafollicular sites, but their relative exclusion from B cell follicles prevents their elimination of productively infected TFH cells. CD8(+) lymphocyte depletion in EC monkeys resulted in a dramatic re-distribution of productive SIV infection to non-TFH cells, with restriction of productive infection to TFH cells resuming upon CD8(+) T cell recovery. Thus, B cell follicles constitute 'sanctuaries' for persistent SIV replication in the presence of potent anti-viral CD8(+) T cell responses, potentially complicating efforts to cure HIV infection with therapeutic vaccination or T cell immunotherapy.

Inflammation contributes to ischemic brain injury. However, translation of experimental findings from animal models into clinical trials is still ineffective, since the majority of human stroke studies mainly focus on acute neuroprotection, thereby neglecting inflammatory mechanisms and inflammation-associated co-morbidity factors such as hypercholesterolemia. Therefore, both wildtype and ApoE(-/-) mice that exhibit increased serum plasma cholesterol levels fed with normal or high cholesterol diet were exposed to transient middle cerebral artery occlusion. Analysis of peripheral immune responses revealed an ischemia-induced acute leukocytosis in the blood, which was accompanied by enhanced myeloid cell and specifically granulocyte cell counts in the spleen and blood of ApoE(-/-) mice fed with Western diet. These cellular immune changes were further associated with increased levels of pro-inflammatory cytokines like IL-6 and TNF-α. Moreover, endogenous stroke-induced endothelial activation as well as CXCL-1 and CXCL-2 expression were increased, thus resulting in accelerated leukocyte, particularly granulocyte accumulation, and enhanced ischemic tissue damage. The latter was revealed by larger infarct volumes and increased local DNA fragmentation in ischemic brains of ApoE(-/-) mice on Western diet. These effects were not observed in wildtype mice on normal or Western diet and in ApoE(-/-) mice on normal diet. Our data demonstrate that the combination of both ApoE knockout and a high cholesterol diet leads to increased ischemia-induced peripheral and cerebral immune responses, which go along with enhanced cerebral tissue injury. Thus, clinically predisposing conditions related to peripheral inflammation such as hypercholesterolemia should be included in up-coming preclinical stroke research., (© 2013.)

Prime-boost immunization with heterologous vaccines elicits potent cellular immunity. In this study, we assessed the influence of various TLR ligands on SIV Gag-specific T cell immunity and protection following prime-boost immunization. Rhesus macaques (RMs) were primed with SIV Gag protein emulsified in Montanide ISA51 with or without TLR3 (polyinosinic-polycytidylic acid [poly-IC]), TLR4 (monophosphoryl lipid A), TLR7/8 (3M-012), TLR9 (CpG), or TLR3 (poly-IC) combined with TLR7/8 ligands, then boosted with replication defective adenovirus 5 expressing SIV Gag (rAd5-Gag). After priming, RMs that received SIV Gag protein plus poly-IC developed significantly higher frequencies of SIV Gag-specific CD4(+) Th1 responses in blood and bronchoalveolar lavage (BAL) fluid lymphocytes compared with all other adjuvants, and low-level SIV Gag-specific CD8(+) T cell responses. After the rAd5-Gag boost, the magnitude and breadth of SIV Gag-specific CD8(+) T cell responses were significantly increased in RM primed with SIV Gag protein plus poly-IC, with or without the TLR7/8 ligand, or CpG. However, the anamnestic, SIV Gag-specific CD8(+) T cell response to SIVmac251 challenge was not significantly enhanced by SIV Gag protein priming with any of the adjuvants. In contrast, the anamnestic SIV Gag-specific CD4(+) T cell response in BAL was enhanced by SIV Gag protein priming with poly-IC or CpG, which correlated with partial control of early viral replication after SIVmac251 challenge. These results demonstrate that prime-boost vaccination with SIV Gag protein/poly-IC improves magnitude, breadth, and durability of CD4(+) T cell immune responses, which could have a role in the control of SIV viral replication.

Live attenuated simian immunodeficiency virus (SIV) vaccines (LAVs) remain the most efficacious of all vaccines in nonhuman primate models of HIV and AIDS, yet the basis of their robust protection remains poorly understood. Here we show that the degree of LAV-mediated protection against intravenous wild-type SIVmac239 challenge strongly correlates with the magnitude and function of SIV-specific, effector-differentiated T cells in the lymph node but not with the responses of such T cells in the blood or with other cellular, humoral and innate immune parameters. We found that maintenance of protective T cell responses is associated with persistent LAV replication in the lymph node, which occurs almost exclusively in follicular helper T cells. Thus, effective LAVs maintain lymphoid tissue-based, effector-differentiated, SIV-specific T cells that intercept and suppress early wild-type SIV amplification and, if present in sufficient frequencies, can completely control and perhaps clear infection, an observation that provides a rationale for the development of safe, persistent vectors that can elicit and maintain such responses.

Glycans of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) play pivotal roles in modulating virus-target cell interactions. We have previously reported that, whereas SIVmac239 is pathogenic, its deglycosylated essentially nonpathogenic mutant (Δ5G) serves as a live-attenuated vaccine, although both replicate similarly during primary infection. These findings prompted us to determine whether such a polarized clinical outcome was due to differences in the immune tissues targeted by these viruses, where functionally and phenotypically different memory CD4(+) T cells reside. The results showed that Δ5G replicates in secondary lymphoid tissue (SLT) at 1- to 2-log-lower levels than SIVmac239, whereas SIVmac239-infected but not Δ5G-infected animals deplete CXCR3(+) CCR5(+) transitional memory (TrM) CD4(+) T cells. An early robust Δ5G replication was localized to small intestinal tissue, especially the lamina propria (effector site) rather than isolated lymphoid follicles (inductive site) and was associated with the induction and depletion of CCR6(+) CXCR3(-) CCR5(+) effector memory CD4(+) T cells. These results suggest that differential glycosylation of Env dictates the type of tissue-resident CD4(+) T cells that are targeted, which leads to pathogenic infection of TrM-Th1 cells in SLT and nonpathogenic infection of Th17 cells in the small intestine, respectively.

Vascular endothelial growth factor (VEGF) has potent angiogenic and neuroprotective effects in the ischemic brain. Its effect on axonal plasticity and neurological recovery in the post-acute stroke phase was unknown. Using behavioral tests combined with anterograde tract tracing studies and with immunohistochemical and molecular biological experiments, we examined effects of a delayed i.c.v. delivery of recombinant human VEGF(165), starting 3 days after stroke, on functional neurological recovery, corticorubral plasticity and inflammatory brain responses in mice submitted to 30 min of middle cerebral artery occlusion. We herein show that the slowly progressive functional improvements of motor grip strength and coordination, which are induced by VEGF, are accompanied by enhanced sprouting of contralesional corticorubral fibres that branched off the pyramidal tract in order to cross the midline and innervate the ipsilesional parvocellular red nucleus. Infiltrates of CD45+ leukocytes were noticed in the ischemic striatum of vehicle-treated mice that closely corresponded to areas exhibiting Iba-1+ activated microglia. VEGF attenuated the CD45+ leukocyte infiltrates at 14 but not 30 days post ischemia and diminished the microglial activation. Notably, the VEGF-induced anti-inflammatory effect of VEGF was associated with a downregulation of a broad set of inflammatory cytokines and chemokines in both brain hemispheres. These data suggest a link between VEGF's immunosuppressive and plasticity-promoting actions that may be important for successful brain remodeling. Accordingly, growth factors with anti-inflammatory action may be promising therapeutics in the post-acute stroke phase., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Although CMV infection is largely benign in immunocompetent people, the specific T cell responses associated with control of this persistent virus are enormous and must be maintained for life. These responses may increase with advanced age and have been linked to an "immune risk profile" that is associated with poor immune responsiveness and increased mortality in aged individuals. Based on this association, it has been suggested that CMV-specific T cell responses might become dysfunctional with age and thereby contribute to the development of immune senescence by homeostatic disruption of other T cell populations, diminished control of CMV replication, and/or excess chronic inflammation. In this study, we use the rhesus macaque (RM) model of aging to ask whether the quantity and quality of CMV-specific T cell responses differ between healthy adult RMs and elderly RMs that manifest hallmarks of immune aging. We demonstrate that the size of the CD4(+) and CD8(+) CMV-specific T cell pools are similar in adult versus old RMs and show essentially identical phenotypic and functional characteristics, including a dominant effector memory phenotype, identical patterns of IFN-γ, TNF-α, and IL-2 production and cytotoxic degranulation, and comparable functional avidities of optimal epitope-specific CD8(+) T cells. Most importantly, the response to and protection against an in vivo CMV challenge were identical in adult and aged RMs. These data indicate that CMV-specific T cell immunity is well maintained in old RMs and argue against a primary role for progressive dysfunction of these responses in the development of immune senescence.

Primary simian immunodeficiency virus (SIV) infections of rhesus macaques result in the dramatic depletion of CD4(+) CCR5(+) effector-memory T (T(EM)) cells from extra-lymphoid effector sites, but in most infections, an increased rate of CD4(+) memory T cell proliferation appears to prevent collapse of effector site CD4(+) T(EM) cell populations and acute-phase AIDS. Eventually, persistent SIV replication results in chronic-phase AIDS, but the responsible mechanisms remain controversial. Here, we demonstrate that in the chronic phase of progressive SIV infection, effector site CD4(+) T(EM) cell populations manifest a slow, continuous decline, and that the degree of this depletion remains a highly significant correlate of late-onset AIDS. We further show that due to persistent immune activation, effector site CD4(+) T(EM) cells are predominantly short-lived, and that their homeostasis is strikingly dependent on the production of new CD4(+) T(EM) cells from central-memory T (T(CM)) cell precursors. The instability of effector site CD4(+) T(EM) cell populations over time was not explained by increasing destruction of these cells, but rather was attributable to progressive reduction in their production, secondary to decreasing numbers of CCR5(-) CD4(+) T(CM) cells. These data suggest that although CD4(+) T(EM) cell depletion is a proximate mechanism of immunodeficiency, the tempo of this depletion and the timing of disease onset are largely determined by destruction, failing production, and gradual decline of CD4(+) T(CM) cells.

HIV infection selectively targets CD4+ effector memory T (T EM) cells, resulting in dramatic depletion of CD4+ T cells in mucosal effector sites in early infection. Regeneration of the T EM cell compartment is slow and incomplete, even when viral replication is controlled by antiretroviral therapy (ART). Here, we demonstrate that IL-15 dramatically increases in vivo proliferation of rhesus macaque (RM) CD4+ and CD8+ T EM cells with little effect on the naive or central memory T (T CM) cell subsets, a response pattern that is quite distinct from that of either IL-2 or IL-7. T EM cells produced in response to IL-15 did not accumulate in blood. Rather, 5-bromo-2'-deoxyuridine (BrdU) labeling studies suggest that many of these cells rapidly disperse to extralymphoid effector sites, where they manifest (slow) decay kinetics indistinguishable from that of untreated controls. In RMs with uncontrolled SIV infection and highly activated immune systems, IL-15 did not significantly increase CD4+ T EM cell proliferation, but with virologic control and concomitant reduction in immune activation by ART, IL-15 responsiveness was again observed. These data suggest that therapeutic use of IL-15 in the setting of ART might facilitate specific restoration of the CD4 + T cell compartment that is the primary target of HIV with less risk of exhausting precursor T cell compartments or generating potentially deleterious regulatory subsets.

The mechanisms linking human immunodeficiency virus replication to the progressive immunodeficiency of acquired immune deficiency syndrome are controversial, particularly the relative contribution of CD4+ T cell destruction. Here, we used the simian immunodeficiency virus (SIV) model to investigate the relationship between systemic CD4+ T cell dynamics and rapid disease progression. Of 18 rhesus macaques (RMs) infected with CCR5-tropic SIVmac239 (n=14) or CXCR4-tropic SIVmac155T3 (n=4), 4 of the former group manifested end-stage SIV disease by 200 d after infection. In SIVmac155T3 infections, naive CD4+ T cells were dramatically depleted, but this population was spared by SIVmac239, even in rapid progressors. In contrast, all SIVmac239-infected RMs demonstrated substantial systemic depletion of CD4+ memory T cells by day 28 after infection. Surprisingly, the extent of CD4+ memory T cell depletion was not, by itself, a strong predictor of rapid progression. However, in all RMs destined for stable infection, this depletion was countered by a striking increase in production of short-lived CD4+ memory T cells, many of which rapidly migrated to tissue. In all rapid progressors (P <0.0001), production of these cells initiated but failed by day 42 of infection, and tissue delivery of new CD4+ memory T cells ceased. Thus, although profound depletion of tissue CD4+ memory T cells appeared to be a prerequisite for early pathogenesis, it was the inability to respond to this depletion with sustained production of tissue-homing CD4+ memory T cells that best distinguished rapid progressors, suggesting that mechanisms of the CD4+ memory T cell generation play a crucial role in maintaining immune homeostasis in stable SIV infection.

The rhesus macaque (RM) is a critical animal model for studies of viral pathogenesis and immunity, yet fundamental aspects of their cellular immune response remain poorly defined. One such deficiency is the lack of validated phenotypic signatures for their naive and memory T cell subsets, and the resultant unavailability of accurate information on their memory T cell development, homeostasis, and function. In this study, we report a phenotypic paradigm allowing definitive characterization of these subsets and their comprehensive functional analysis. Naive T cells are optimally delineated by their homogeneous CD95(low)CD28(high)beta(7) integrin(int) (CD4+) or CD95(low)CD28(int)CD11a(low) (CD8+) phenotypes. This subset 1) was present in blood and secondary lymph tissues, but not effector sites; 2) vastly predominated in the fetal/neonatal immune system, but rapidly diminished with postnatal age; 3) lacked IFN-gamma production capability, and specific responses to RM CMV; and 4) demonstrated low in vivo proliferative activity. CD4+ and CD8+ memory subsets were CD95(high), but otherwise phenotypically heterogeneous and included all IFN-gamma production, RM CMV-specific responses, effector site T cells, and demonstrated high in vivo proliferative activity ( approximately 10 times the naive subset). These analyses also revealed the RM "effector memory" subset within the overall memory population. This population, best defined by lack of CD28 expression, contained the majority of RM CMV-specific cells, was highly enriched in extralymphoid effector sites, and comprised an increasing proportion of total memory cells with age. The effector memory subset demonstrated similar in vivo proliferative activity and survival as CD28+ "central memory" T cells, consistent with independent homeostatic regulation.