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NK Cell-Mediated Regulation of Protective Memory Responses against Intracellular Ehrlichial Pathogens.

Habib S, El Andaloussi A, Hisham A, Ismail N - PLoS ONE (2016)

Bottom Line: Ehrlichiae are gram-negative obligate intracellular bacteria that cause potentially fatal human monocytic ehrlichiosis.However, the contribution of NK cells to the memory response against Ehrlichia remains elusive.Together, these data suggest that E. muris-induced memory-like NK cells, which contribute to the protective, recall response against Ehrlichia.

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, Medical College of Georgia, Augusta University, Augusta, Georgia, United States of America.

ABSTRACT
Ehrlichiae are gram-negative obligate intracellular bacteria that cause potentially fatal human monocytic ehrlichiosis. We previously showed that natural killer (NK) cells play a critical role in host defense against Ehrlichia during primary infection. However, the contribution of NK cells to the memory response against Ehrlichia remains elusive. Primary infection of C57BL/6 mice with Ehrlichia muris provides long-term protection against a second challenge with the highly virulent Ixodes ovatus Ehrlichia (IOE), which ordinarily causes fatal disease in naïve mice. Here, we show that the depletion of NK cells in E. muris-primed mice abrogates the protective memory response against IOE. Approximately, 80% of NK cell-depleted E. muris-primed mice succumbed to lethal IOE infection on days 8-10 after IOE infection, similar to naïve mice infected with the same dose of IOE. The lack of a recall response in NK cell-depleted mice correlated with an increased bacterial burden, extensive liver injury, decreased frequency of Ehrlichia-specific IFN-γ-producing memory CD4+ and CD8+ T-cells, and a low titer of Ehrlichia-specific antibodies. Intraperitoneal infection of mice with E. muris resulted in the production of IL-15, IL-12, and IFN-γ as well as an expansion of activated NKG2D+ NK cells. The adoptive transfer of purified E. muris-primed hepatic and splenic NK cells into Rag2-/-Il2rg-/- recipient mice provided protective immunity against challenge with E. muris. Together, these data suggest that E. muris-induced memory-like NK cells, which contribute to the protective, recall response against Ehrlichia.

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NK cells expand and persist in the liver of E. muris- but not IOE-primed mice.Liver mononuclear cells were isolated from indicated mice groups on day 21 post-infection, and the frequency and activation of NK cells were analyzed. (A) shows gating strategy on NK cells and activation marker NKG2D. (B) The percentage and (C) absolute number of NK cells in the livers of indicated mice groups. as determined by flow cytometry. (D) The percentage of activated NK cells expressing NKG2D. The results demonstrate a higher frequency of activated NKG2D+ NK cells in the livers of E. muris-primed mice as compared to other groups (* P <0.05 and ** P <0.01). The data shown are the means ± SD from three mice per group and are representative of three independent experiments.
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pone.0153223.g001: NK cells expand and persist in the liver of E. muris- but not IOE-primed mice.Liver mononuclear cells were isolated from indicated mice groups on day 21 post-infection, and the frequency and activation of NK cells were analyzed. (A) shows gating strategy on NK cells and activation marker NKG2D. (B) The percentage and (C) absolute number of NK cells in the livers of indicated mice groups. as determined by flow cytometry. (D) The percentage of activated NK cells expressing NKG2D. The results demonstrate a higher frequency of activated NKG2D+ NK cells in the livers of E. muris-primed mice as compared to other groups (* P <0.05 and ** P <0.01). The data shown are the means ± SD from three mice per group and are representative of three independent experiments.

Mentions: We previously showed that a primary non-lethal infection of wild type (WT)-B6 mice with E. muris (EM), but not a sublethal IOE infection, provides long-term protection of primed mice against an ordinarily lethal secondary IOE infection [23,25]. E. muris, but not IOE, causes persistent infection, which is critical for the induction and maintenance of memory CD4+ T cells. However, the adoptive transfer of memory CD4+ T cells alone provides partial protection to naïve mice against lethal IOE infection [6]. Thus, we hypothesized that the protective recall response in E. muris-primed mice could be mediated by cells other than memory CD4+ T cells. Because the liver, which is enriched with NK and NKT cells, serves as the main site of infection in ehrlichiosis, we examined whether cross-protection in E. muris-primed mice was due to differences in NK cell responses. To this end, we analyzed the activation status, and frequency of NK cells in the liver and spleen of mice infected with E. muris or IOE via the IP route. Our data demonstrated that NK cells expand and persist in the liver Fig 1A–1C of E. muris-, but not IOE-, primed mice 21 DPI [the time point that corresponds to the contraction of effector T cells and the generation of central and effector memory T cells [23,25]. Our data further showed that approximately 70% of liver NK cells in E. muris-primed mice expressed NK cell-activating receptor (NKG2D) Fig 1D, suggesting that these cells were activated in vivo. Similar to hepatic NK cells, the frequency of splenic NK cells was higher in the spleen of E. muris-primed mice as compared to IOE-primed mice and uninfected controls Fig 2A–2C. Approximately 40% of E. muris-primed splenic NK cells expressed NKG2D, while ~ 20% of IOE-primed splenic NK cells expressed NKG2D Fig 2D. Since the total number of splenic NK cells was significantly higher in E. muris-infected mice than that detected in IOE-infected mice, these data suggest that E. muris also induces significant expansion of activated NKG2D+NK cells in the spleen compared to IOE-primed mice. These data suggest that E. muris infection promotes the expansion of activated NK cells that persist in the primed host, in the liver and spleen.


NK Cell-Mediated Regulation of Protective Memory Responses against Intracellular Ehrlichial Pathogens.

Habib S, El Andaloussi A, Hisham A, Ismail N - PLoS ONE (2016)

NK cells expand and persist in the liver of E. muris- but not IOE-primed mice.Liver mononuclear cells were isolated from indicated mice groups on day 21 post-infection, and the frequency and activation of NK cells were analyzed. (A) shows gating strategy on NK cells and activation marker NKG2D. (B) The percentage and (C) absolute number of NK cells in the livers of indicated mice groups. as determined by flow cytometry. (D) The percentage of activated NK cells expressing NKG2D. The results demonstrate a higher frequency of activated NKG2D+ NK cells in the livers of E. muris-primed mice as compared to other groups (* P <0.05 and ** P <0.01). The data shown are the means ± SD from three mice per group and are representative of three independent experiments.
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getmorefigures.php?uid=PMC4836677&req=5

pone.0153223.g001: NK cells expand and persist in the liver of E. muris- but not IOE-primed mice.Liver mononuclear cells were isolated from indicated mice groups on day 21 post-infection, and the frequency and activation of NK cells were analyzed. (A) shows gating strategy on NK cells and activation marker NKG2D. (B) The percentage and (C) absolute number of NK cells in the livers of indicated mice groups. as determined by flow cytometry. (D) The percentage of activated NK cells expressing NKG2D. The results demonstrate a higher frequency of activated NKG2D+ NK cells in the livers of E. muris-primed mice as compared to other groups (* P <0.05 and ** P <0.01). The data shown are the means ± SD from three mice per group and are representative of three independent experiments.
Mentions: We previously showed that a primary non-lethal infection of wild type (WT)-B6 mice with E. muris (EM), but not a sublethal IOE infection, provides long-term protection of primed mice against an ordinarily lethal secondary IOE infection [23,25]. E. muris, but not IOE, causes persistent infection, which is critical for the induction and maintenance of memory CD4+ T cells. However, the adoptive transfer of memory CD4+ T cells alone provides partial protection to naïve mice against lethal IOE infection [6]. Thus, we hypothesized that the protective recall response in E. muris-primed mice could be mediated by cells other than memory CD4+ T cells. Because the liver, which is enriched with NK and NKT cells, serves as the main site of infection in ehrlichiosis, we examined whether cross-protection in E. muris-primed mice was due to differences in NK cell responses. To this end, we analyzed the activation status, and frequency of NK cells in the liver and spleen of mice infected with E. muris or IOE via the IP route. Our data demonstrated that NK cells expand and persist in the liver Fig 1A–1C of E. muris-, but not IOE-, primed mice 21 DPI [the time point that corresponds to the contraction of effector T cells and the generation of central and effector memory T cells [23,25]. Our data further showed that approximately 70% of liver NK cells in E. muris-primed mice expressed NK cell-activating receptor (NKG2D) Fig 1D, suggesting that these cells were activated in vivo. Similar to hepatic NK cells, the frequency of splenic NK cells was higher in the spleen of E. muris-primed mice as compared to IOE-primed mice and uninfected controls Fig 2A–2C. Approximately 40% of E. muris-primed splenic NK cells expressed NKG2D, while ~ 20% of IOE-primed splenic NK cells expressed NKG2D Fig 2D. Since the total number of splenic NK cells was significantly higher in E. muris-infected mice than that detected in IOE-infected mice, these data suggest that E. muris also induces significant expansion of activated NKG2D+NK cells in the spleen compared to IOE-primed mice. These data suggest that E. muris infection promotes the expansion of activated NK cells that persist in the primed host, in the liver and spleen.

Bottom Line: Ehrlichiae are gram-negative obligate intracellular bacteria that cause potentially fatal human monocytic ehrlichiosis.However, the contribution of NK cells to the memory response against Ehrlichia remains elusive.Together, these data suggest that E. muris-induced memory-like NK cells, which contribute to the protective, recall response against Ehrlichia.

View Article: PubMed Central - PubMed

Affiliation: Department of Obstetrics and Gynecology, Medical College of Georgia, Augusta University, Augusta, Georgia, United States of America.

ABSTRACT
Ehrlichiae are gram-negative obligate intracellular bacteria that cause potentially fatal human monocytic ehrlichiosis. We previously showed that natural killer (NK) cells play a critical role in host defense against Ehrlichia during primary infection. However, the contribution of NK cells to the memory response against Ehrlichia remains elusive. Primary infection of C57BL/6 mice with Ehrlichia muris provides long-term protection against a second challenge with the highly virulent Ixodes ovatus Ehrlichia (IOE), which ordinarily causes fatal disease in naïve mice. Here, we show that the depletion of NK cells in E. muris-primed mice abrogates the protective memory response against IOE. Approximately, 80% of NK cell-depleted E. muris-primed mice succumbed to lethal IOE infection on days 8-10 after IOE infection, similar to naïve mice infected with the same dose of IOE. The lack of a recall response in NK cell-depleted mice correlated with an increased bacterial burden, extensive liver injury, decreased frequency of Ehrlichia-specific IFN-γ-producing memory CD4+ and CD8+ T-cells, and a low titer of Ehrlichia-specific antibodies. Intraperitoneal infection of mice with E. muris resulted in the production of IL-15, IL-12, and IFN-γ as well as an expansion of activated NKG2D+ NK cells. The adoptive transfer of purified E. muris-primed hepatic and splenic NK cells into Rag2-/-Il2rg-/- recipient mice provided protective immunity against challenge with E. muris. Together, these data suggest that E. muris-induced memory-like NK cells, which contribute to the protective, recall response against Ehrlichia.

Show MeSH
Related in: MedlinePlus