Limits...
Prolonged antigen presentation is required for optimal CD8+ T cell responses against malaria liver stage parasites.

Cockburn IA, Chen YC, Overstreet MG, Lees JR, van Rooijen N, Farber DL, Zavala F - PLoS Pathog. (2010)

Bottom Line: Firstly, reducing the time primed CD8+ T cells were exposed to antigen in vivo severely reduced the final size of the developing memory population.Secondly, fully developed memory cells expanded in previously immunized mice but not when transferred to naïve animals.Finally, persisting antigen was able to prime naïve cells, including recent thymic emigrants, to become functional effector cells capable of eliminating parasites in the liver.

View Article: PubMed Central - PubMed

Affiliation: Johns Hopkins Malaria Research Institute and Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Baltimore, Maryland, USA.

ABSTRACT
Immunization with irradiated sporozoites is currently the most effective vaccination strategy against liver stages of malaria parasites, yet the mechanisms underpinning the success of this approach are unknown. Here we show that the complete development of protective CD8+ T cell responses requires prolonged antigen presentation. Using TCR transgenic cells specific for the malaria circumsporozoite protein, a leading vaccine candidate, we found that sporozoite antigen persists for over 8 weeks after immunization--a remarkable finding since irradiated sporozoites are incapable of replication and do not differentiate beyond early liver stages. Persisting antigen was detected in lymphoid organs and depends on the presence of CD11c+ cells. Prolonged antigen presentation enhanced the magnitude of the CD8+ T cell response in a number of ways. Firstly, reducing the time primed CD8+ T cells were exposed to antigen in vivo severely reduced the final size of the developing memory population. Secondly, fully developed memory cells expanded in previously immunized mice but not when transferred to naïve animals. Finally, persisting antigen was able to prime naïve cells, including recent thymic emigrants, to become functional effector cells capable of eliminating parasites in the liver. Together these data show that the optimal development of protective CD8+ T cell immunity against malaria liver stages is dependent upon the prolonged presentation of sporozoite-derived antigen.

Show MeSH

Related in: MedlinePlus

Persisting antigen primes naïve thymic emigrants.A. Outline of the experiment: mice were immunized with 5×104 irradiated sporozoites i.v. On days 3 and 4 after immunization some of the mice were depleted of CD8+ cells such that the observed response consists largely of later thymic emigrants (gray bars). Control mice were treated with control IgG1 leaving the pre-existing response in tact (total response – black bars), while other mice were left un-immunized but treated with anti-CD8 (naïve – white bars). B. Histograms of the number of tetramer+ cells recovered in each group, bar charts show means ± SE (n = 6, * = P<0.05, data pooled from two similar experiments). C. Histograms of the percentage of activated cells (CD62Llo, CD44hi) among the tetramer+ population recovered in each experiment (n = 3, * = P<0.05, data from one of two similar experiments). D. Dot plots of the CD62L and CD44 expression of tetramer+ cells in each group, data show concatenated data from 3 animals per group and are from one of two similar experiments.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2865532&req=5

ppat-1000877-g007: Persisting antigen primes naïve thymic emigrants.A. Outline of the experiment: mice were immunized with 5×104 irradiated sporozoites i.v. On days 3 and 4 after immunization some of the mice were depleted of CD8+ cells such that the observed response consists largely of later thymic emigrants (gray bars). Control mice were treated with control IgG1 leaving the pre-existing response in tact (total response – black bars), while other mice were left un-immunized but treated with anti-CD8 (naïve – white bars). B. Histograms of the number of tetramer+ cells recovered in each group, bar charts show means ± SE (n = 6, * = P<0.05, data pooled from two similar experiments). C. Histograms of the percentage of activated cells (CD62Llo, CD44hi) among the tetramer+ population recovered in each experiment (n = 3, * = P<0.05, data from one of two similar experiments). D. Dot plots of the CD62L and CD44 expression of tetramer+ cells in each group, data show concatenated data from 3 animals per group and are from one of two similar experiments.

Mentions: The previous experiments demonstrate that a prolonged period of antigen presentation throughout the expansion phase of the CD8+ T cell response is required to maximize the CD8+ T cell response. However antigen continues to be presented for weeks after this. In addition to showing that this late antigen presentation could stimulate the proliferation of memory cells we also hypothesized that prolonged antigen presentation may continue to recruit naïve cells, such as naïve thymic emigrants to the immune response. To test this we immunized mice with sporozoites and 3 days later depleted CD8+ T cells by antibody treatment. Preliminary experiments showed that this treatment has no effect on the presence of persisting antigen (data not shown). Around 10 days after depletion, CD8+ T cells begin to reappear, these repopulating cells are, in all likeliness, mostly thymic emigrants (Figure 7A). Thirty-five days after immunization we isolated the endogenous tetramer+ cells by magnetic bead selection and stained for the surface markers CD44 and CD62L. We found antigen specific thymic emigrants from immune mice expanded ∼2.5-fold compared to cells in naïve mice - a difference that was significant (Figure 7B; P = 0.027 by two-tailed T test; t = 2.58; df = 10). More importantly, thymic emigrants had up-regulated CD44 and down-regulated CD62L, a phenotype similar to that seen in immunized mice which had received control IgG instead of anti-CD8 (Figure 7C and 7D) suggesting that thymic emigrants are indeed primed by persisting antigen.


Prolonged antigen presentation is required for optimal CD8+ T cell responses against malaria liver stage parasites.

Cockburn IA, Chen YC, Overstreet MG, Lees JR, van Rooijen N, Farber DL, Zavala F - PLoS Pathog. (2010)

Persisting antigen primes naïve thymic emigrants.A. Outline of the experiment: mice were immunized with 5×104 irradiated sporozoites i.v. On days 3 and 4 after immunization some of the mice were depleted of CD8+ cells such that the observed response consists largely of later thymic emigrants (gray bars). Control mice were treated with control IgG1 leaving the pre-existing response in tact (total response – black bars), while other mice were left un-immunized but treated with anti-CD8 (naïve – white bars). B. Histograms of the number of tetramer+ cells recovered in each group, bar charts show means ± SE (n = 6, * = P<0.05, data pooled from two similar experiments). C. Histograms of the percentage of activated cells (CD62Llo, CD44hi) among the tetramer+ population recovered in each experiment (n = 3, * = P<0.05, data from one of two similar experiments). D. Dot plots of the CD62L and CD44 expression of tetramer+ cells in each group, data show concatenated data from 3 animals per group and are from one of two similar experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2865532&req=5

ppat-1000877-g007: Persisting antigen primes naïve thymic emigrants.A. Outline of the experiment: mice were immunized with 5×104 irradiated sporozoites i.v. On days 3 and 4 after immunization some of the mice were depleted of CD8+ cells such that the observed response consists largely of later thymic emigrants (gray bars). Control mice were treated with control IgG1 leaving the pre-existing response in tact (total response – black bars), while other mice were left un-immunized but treated with anti-CD8 (naïve – white bars). B. Histograms of the number of tetramer+ cells recovered in each group, bar charts show means ± SE (n = 6, * = P<0.05, data pooled from two similar experiments). C. Histograms of the percentage of activated cells (CD62Llo, CD44hi) among the tetramer+ population recovered in each experiment (n = 3, * = P<0.05, data from one of two similar experiments). D. Dot plots of the CD62L and CD44 expression of tetramer+ cells in each group, data show concatenated data from 3 animals per group and are from one of two similar experiments.
Mentions: The previous experiments demonstrate that a prolonged period of antigen presentation throughout the expansion phase of the CD8+ T cell response is required to maximize the CD8+ T cell response. However antigen continues to be presented for weeks after this. In addition to showing that this late antigen presentation could stimulate the proliferation of memory cells we also hypothesized that prolonged antigen presentation may continue to recruit naïve cells, such as naïve thymic emigrants to the immune response. To test this we immunized mice with sporozoites and 3 days later depleted CD8+ T cells by antibody treatment. Preliminary experiments showed that this treatment has no effect on the presence of persisting antigen (data not shown). Around 10 days after depletion, CD8+ T cells begin to reappear, these repopulating cells are, in all likeliness, mostly thymic emigrants (Figure 7A). Thirty-five days after immunization we isolated the endogenous tetramer+ cells by magnetic bead selection and stained for the surface markers CD44 and CD62L. We found antigen specific thymic emigrants from immune mice expanded ∼2.5-fold compared to cells in naïve mice - a difference that was significant (Figure 7B; P = 0.027 by two-tailed T test; t = 2.58; df = 10). More importantly, thymic emigrants had up-regulated CD44 and down-regulated CD62L, a phenotype similar to that seen in immunized mice which had received control IgG instead of anti-CD8 (Figure 7C and 7D) suggesting that thymic emigrants are indeed primed by persisting antigen.

Bottom Line: Firstly, reducing the time primed CD8+ T cells were exposed to antigen in vivo severely reduced the final size of the developing memory population.Secondly, fully developed memory cells expanded in previously immunized mice but not when transferred to naïve animals.Finally, persisting antigen was able to prime naïve cells, including recent thymic emigrants, to become functional effector cells capable of eliminating parasites in the liver.

View Article: PubMed Central - PubMed

Affiliation: Johns Hopkins Malaria Research Institute and Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Baltimore, Maryland, USA.

ABSTRACT
Immunization with irradiated sporozoites is currently the most effective vaccination strategy against liver stages of malaria parasites, yet the mechanisms underpinning the success of this approach are unknown. Here we show that the complete development of protective CD8+ T cell responses requires prolonged antigen presentation. Using TCR transgenic cells specific for the malaria circumsporozoite protein, a leading vaccine candidate, we found that sporozoite antigen persists for over 8 weeks after immunization--a remarkable finding since irradiated sporozoites are incapable of replication and do not differentiate beyond early liver stages. Persisting antigen was detected in lymphoid organs and depends on the presence of CD11c+ cells. Prolonged antigen presentation enhanced the magnitude of the CD8+ T cell response in a number of ways. Firstly, reducing the time primed CD8+ T cells were exposed to antigen in vivo severely reduced the final size of the developing memory population. Secondly, fully developed memory cells expanded in previously immunized mice but not when transferred to naïve animals. Finally, persisting antigen was able to prime naïve cells, including recent thymic emigrants, to become functional effector cells capable of eliminating parasites in the liver. Together these data show that the optimal development of protective CD8+ T cell immunity against malaria liver stages is dependent upon the prolonged presentation of sporozoite-derived antigen.

Show MeSH
Related in: MedlinePlus