Rajarshi Bhadra

Control of Toxoplasma reactivation by rescue of dysfunctional CD8+ T-cell response via PD-1–PDL-1 blockade

In this study, we document that Toxoplasma gondii differentiation and reactivation are mediated by systemic CD8 T-cell dysfunction during chronic infection. We demonstrate that CD8+ T-cell exhaustion occurs despite control of parasitemia during early-chronic toxoplasmosis. During later phases, these cells become exhausted, leading to parasite reactivation and mortality. Concomitant with increased CD8+ T-cell apoptosis and decreased effector response, this dysfunction is characterized by a graded elevation in expression of inhibitory receptor PD-1 on these cells in both lymphoid and nonlymphoid tissue. Blockade of the PD-1–PDL-1 pathway reinvigorates this suboptimal CD8+ T-cell response, resulting in control of parasite reactivation and prevention of mortality in chronically infected animals. To the best of our knowledge, this report is unique in showing that exposure to a persistent pathogen despite initial control of parasitemia can lead to CD8+ T-cell dysfunction and parasite reactivation.

Cutting Edge: CD40–CD40 Ligand Pathway Plays a Critical CD8-Intrinsic and -Extrinsic Role during Rescue of Exhausted CD8 T Cells

CD8 exhaustion mediated by an inhibitory programmed death-1–programmed death ligand-1 (PD-L1) pathway occurs in several chronic infections, including toxoplasmosis. Although blockade of the programmed death-1–PD-L1 pathway revives this response, the role of costimulatory receptors involved in this rescue has not been ascertained in any model of CD8 exhaustion. This report demonstrates that one such costimulatory pathway, CD40–CD40L, plays a critical role during rescue of exhausted CD8 T cells. Blockade of this pathway abrogates the ameliorative effects of anti–PD-L1 treatment on CD8 T cells. Additionally, we demonstrate in an infectious disease model that CD8-intrinsic CD40 signaling is important for optimal CD8 polyfunctionality, proliferation, T-bet upregulation, and IL-21 signaling, albeit in the context of CD8 rescue. The critical role of CD40 during the rescue of exhausted CD8 T cells may provide a rational basis for designing novel therapeutic vaccination approaches.

PD-1–Mediated Attrition of Polyfunctional Memory CD8+ T Cells in Chronic Toxoplasma Infection

We reported earlier that during chronic toxoplasmosis CD8(+) T cells become functionally exhausted with concomitant PD-1 upregulation, leading to eventual host mortality. However, how immune exhaustion specifically mediates attrition of CD8 polyfunctionality, a hallmark of potent T-cell response, during persistent infections has not been addressed. In this study, we demonstrate that PD-1 is preferentially expressed on polyfunctional memory CD8(+) T cells, which renders them susceptible to apoptosis. In vitro blockade of the PD-1-PD-L1 pathway dramatically reduces apoptosis of polyfunctional and interferon γ(+)/granzyme B(-) memory but not effector CD8(+) T cells. In summary, the present report underscores the critical role of the PD-1-PD-L1 pathway in mediating attrition of this important CD8(+) T-cell subset and addresses the mechanistic basis of how αPD-L1 therapy reinvigorates polyfunctional CD8 response during chronic infections. The conclusions of this study can have profound immunotherapeutic implications in combating recrudescent toxoplasmosis as well other chronic infections.

Absence of both IL-7 and IL-15 severely impairs the development of CD8 T cell response against Toxoplasma gondii.

CD8+ T cells play an essential role in the protection against both acute as well as chronic Toxoplasma gondii infection. Although the role of IL-15 has been reported to be important for the development of long-term CD8+ T cell immunity against the pathogen, the simultaneous roles played by both IL-15 and related γ-chain family cytokine IL-7 in the generation of this response during acute phase of infection has not been described. We demonstrate that while lack of IL-7 or IL-15 alone has minimal impact on splenic CD8+ T cell maturation or effector function development during acute Toxoplasmosis, absence of both IL-7 and IL-15 only in the context of infection severely down-regulates the development of a potent CD8+ T cell response. This impairment is characterized by reduction in CD44 expression, IFN-γ production, proliferation and cytotoxicity. However, attenuated maturation and decreased effector functions in these mice are essentially downstream consequences of reduced number of antigen-specific CD8+ T cells. Interestingly, the absence of both cytokines did not impair initial CD8+ T cell generation but affected their survival and differentiation into memory phenotype IL-7Rαhi cells. Significantly lack of both cytokines severely affected expression of Bcl-2, an anti-apoptotic protein, but minimally affected proliferation. The overarching role played by these cytokines in eliciting a potent CD8+ T cell immunity against T. gondii infection is further evidenced by poor survival and high parasite burden in anti IL-7 treated IL-15−/− mice. These studies demonstrate that the two cytokines, IL-7 and IL-15, are exclusively important for the development of protective CD8+ T cell immune response against T. gondii. To the best of our knowledge this synergism between IL-7 and IL-15 in generating an optimal CD8+ T cell immunity against intracellular parasite or any other infectious disease model has not been previously reported.

Psychiatric Disorders in Toxoplasma Seropositive Patients—The CD8 Connection

Although the highest numbers of studies linking an infectious agent with schizophrenia has involved the parasite Toxoplasma gondii, the mechanistic underpinnings of this correlation has remained unaddressed. Incidentally, CD8 T cells, which play a pivotal role in mediating long-term immunity to Toxoplasma, are downregulated in schizophrenia patients. Recent studies have demonstrated that CD8 response is also impaired during chronic toxoplasmosis in murine models. In light of these new findings, in this article, we discuss the potential role of CD8 T cells in causing altered mental status in Toxoplasma seropositive schizophrenia patients.

The CD8 T-cell road to immunotherapy of toxoplasmosis

Toxoplasma gondii infection induces a robust CD8 T-cell immunity that is critical for keeping chronic infection under control. In studies using animal models, it has been demonstrated that the absence of this response can compromise the host ability to keep chronic infection under check. Therapeutic agents that facilitate the induction and maintenance of CD8 T-cell response against the pathogen need to be developed. In the last decade, major strides in understanding the development of effector and memory response, particularly in viral and tumor models, have been made. However, factors involved in the generation of effector or memory response against T. gondii infection have not been extensively investigated. This information will be invaluable in designing immunotherapeutic regimens needed for combating this intracellular pathogen that poses a severe risk for pregnant women and immunocompromised individuals.

CD8 T Cells and Toxoplasma gondii: A New Paradigm

CD8 T cells are essential for control of Toxoplasma gondii infection. Once activated they undergo differentiation into short-lived effector and memory precursor effector cells. As effector cells, CD8 T cells exert immune pressure on the parasite via production of inflammatory cytokines and through their cytolytic activity. Once immune control has been established, the parasite encysts and develops into chronic infection regulated by the memory CD8 T-cell population. Several signals are needed for this process to be initiated and for development of fully differentiated memory CD8 T cells. With newly developed tools including CD8 T-cell tetramers and TCR transgenic mice, dissecting the biology behind T. gondii-specific CD8 T-cell responses can now be more effectively addressed. In this paper, we discuss what is known about the signals required for effective T. gondii-specific CD8 T-cell development, their differentiation, and effector function.

Intrinsic TGF-β signaling promotes age-dependent CD8+ T cell polyfunctionality attrition

Advanced age is associated with immune system deficits that result in an increased susceptibility to infectious diseases; however, specific mediators of age-dependent immune dysfunction have not been fully elucidated. Here we demonstrated that aged mice exhibit poor effector CD8+ T cell polyfunctionality, primarily due to CD8+ T cell-extrinsic deficits, and that reduced CD8+ T cell polyfunctionality correlates with increased susceptibility to pathogenic diseases. In aged animals challenged with the parasite Encephalitozoon cuniculi, effector CD8+ T cell survival and polyfunctionality were suppressed by highly elevated TGF-β1. Furthermore, TGF-β depletion reduced effector CD8+ T cell apoptosis in both young and aged mice and enhanced effector CD8+ T cell polyfunctionality in aged mice. Surprisingly, intrinsic blockade of TGF-β signaling in CD8+ T cells was sufficient to rescue polyfunctionality in aged animals. Together, these data demonstrate that low levels of TGF-β1 promote apoptosis of CD8+ effector T cells and high TGF-β1 levels associated with age result in both CD8+ T cell apoptosis and an altered transcriptional profile, which correlates with loss of polyfunctionality. Furthermore, elevated TGF-β levels are observed in the elderly human population and in aged Drosophila, suggesting that TGF-β represents an evolutionarily conserved negative regulator of the immune response in aging organisms.

Redefining chronic toxoplasmosis: A T cell exhaustion perspective

Toxoplasmosis caused by Toxoplasma gondii (T. gondii), an obligate intercellular protozoan, is considered to be a leading cause of foodborne mortality in the United States (www.cdc.gov). Even in the post-HAART era, fatal toxoplasmic encephalitis (TE) due to reactivation of chronic Toxoplasma infection remains a major problem in Toxoplasma-seropositive AIDS patients in developing countries [1]. In warm-blooded intermediate hosts (including humans), the parasite undergoes stage conversion between the rapidly proliferating tachyzoite, which is considered to be responsible for acute toxoplasmosis, and the relatively quiescent, slowly replicating, encysted bradyzoite that can persist for life. However, in the immunocompromised such as AIDS patients, the parasite converts from a bradyzoite to a tachyzoite stage, leading to TE [2]. Similarly, repeated reactivations can also occur in congenitally infected individuals [3]. Tachyzoite– bradyzoite interconversion is believed to play a central role not only in establishing the chronic infection but also in disease recrudescence [2]. However, factors responsible for the reactivation of chronic infection in vivo remain poorly understood [2,4]. Studies in murine models of chronic toxoplasmosis have demonstrated that CD8 T cells are pivotal for long-term protection [3]. Paradoxically, despite a robust CD8 T cell response during the acute phase of infection, long-term immunity against this pathogen is compromised in susceptible mouse strains, leading to reactivation and host mortality. Differential susceptibility to T. gondii reactivation in AIDS patients was also noted in a study conducted during the pre-HAART era, which reported that only 30% of AIDS patients with low CD4 T cell count and Toxoplasma seropositivity, who were not on effective prophylaxis, developed reactivated toxoplasmosis [5]. Why does a modest subset of this high-risk group develop TE? Considering that memory CD8 T cells can persist for a lifetime and can mediate protective recall responses upon antigen reencounter in other infectious diseases [6], it remains to be addressed whether this differential outcome is a consequence of potential attrition of T. gondii–specific memory CD8 T cells due to genetic polymorphisms or other microenvironment-associated factors. Recent studies from our group, which utilized a susceptible mouse model (C57BL/6), have demonstrated that CD8 T cells during the later phase of chronic toxoplasmosis exhibit progressive attrition of functionality, increased apoptosis, and poor recall response along with elevated expression of PD-1, an inhibitory receptor-a phenomenon referred to as CD8 exhaustion [7]. Concomitant with graded CD8 exhaustion, parasites undergo reactivation resulting in the mortality of the infected host (Figure 1). While the paradigm of CD8 exhaustion has been extensively explored in chronic viral models, it is just beginning to unfold in parasitic infections. Unlike chronic viral models of CD8 exhaustion, which are characterized by persistent high viremia, the T. gondii model represents a unique situation where, despite initial control of parasitemia, CD8 T cells eventually become exhausted [7,8]. Considering that current drugs against T. gondii are toxic and inefficacious against the encysted bradyzoite stage of the parasite [2,3], a thorough understanding of T cell exhaustion during chronic toxoplasmosis is critical for the development of improved immunotherapeutics against this pathogen. Significantly, our laboratory has demonstrated that a blockade of PD-1 interaction with its receptor PDL1, via anti-PD-L1 antibody treatment of chronically infected animals, not only reinvigorates CD8 response and controls parasite reactivation but also prevents host mortality [7].

Blimp-1–mediated CD4 T cell exhaustion causes CD8 T cell dysfunction during chronic toxoplasmosis

Khan et al. demonstrate that in chronic toxoplasmosis, CD4 T cell–intrinsic expression of Blimp-1 results in progressive exhaustion, which in turn contributes to CD8 T cell exhaustion and poor pathogen control.