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Fasting enhances TRAIL-mediated liver natural killer cell activity via HSP70 upregulation.

Dang VT, Tanabe K, Tanaka Y, Tokumoto N, Misumi T, Saeki Y, Fujikuni N, Ohdan H - PLoS ONE (2014)

Bottom Line: Liver natural killer cells also showed high TRAIL-mediated antitumor function in response to 3-day fasting.Treatment of liver lymphocytes with 50 µg/mL of recombinant heat shock protein 70 led to the upregulation of both TRAIL and CD69 in liver natural killer cells (n = 6, p <0.05).These findings indicate that acute fasting enhances TRAIL-mediated liver natural killer cell activity against neoplastic cells through upregulation of heat shock protein 70.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterological and Transplant Surgery, Applied Life Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

ABSTRACT
Acute starvation, which is frequently observed in clinical practice, sometimes augments the cytolytic activity of natural killer cells against neoplastic cells. In this study, we investigated the molecular mechanisms underlying the enhancement of natural killer cell function by fasting in mice. The total number of liver resident natural killer cells in a unit weight of liver tissue obtained from C57BL/6J mice did not change after a 3-day fast, while the proportions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)+ and CD69+ natural killer cells were significantly elevated (n = 7, p <0.01), as determined by flow cytometric analysis. Furthermore, we found that TRAIL- natural killer cells that were adoptively transferred into Rag-2-/- γ chain-/- mice could convert into TRAIL+ natural killer cells in fasted mice at a higher proportion than in fed mice. Liver natural killer cells also showed high TRAIL-mediated antitumor function in response to 3-day fasting. Since these fasted mice highly expressed heat shock protein 70 (n = 7, p <0.05) in liver tissues, as determined by western blot, the role of this protein in natural killer cell activation was investigated. Treatment of liver lymphocytes with 50 µg/mL of recombinant heat shock protein 70 led to the upregulation of both TRAIL and CD69 in liver natural killer cells (n = 6, p <0.05). In addition, HSP70 neutralization by intraperitoneally injecting an anti- heat shock protein 70 monoclonal antibody into mice prior to fasting led to the downregulation of TRAIL expression (n = 6, p <0.05). These findings indicate that acute fasting enhances TRAIL-mediated liver natural killer cell activity against neoplastic cells through upregulation of heat shock protein 70.

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Adoptive transfer assay for TRAIL and CD69 expression on liver natural killer cells in response to starvation.(A) Isolated TRAIL− natural killer (NK) cells were separated from liver lymphocytes of wild type B6 mice. Proportion of lymphocytes expressing TCRβ, NK1.1, and TRAIL in whole liver lymphocytes, isolated NK cells, and isolated TRAIL− NK cells are presented in dot plots. The isolated TRAIL− NK cells were adoptively transferred into Rag-2−/− γ chain−/− mice (0.5×106 cells/mouse), which were then fed or fasted for 3 days before determining their NK phenotype. (B) Dot plots show the gated TCRβ− NK1.1+ NK cells and their percentage in the liver, spleen, and bone marrow of non-transferred (control), fed-transferred, and fasted-transferred mice. (C) Bar graph presents the mean percentage plus standard deviation of NK cells in the liver of fed and fasted-transferred mice. (D) Expression of TRAIL and CD69 (solid lines) on the liver NK cells of representative fed- and fasted-transferred mice with their percentages of positive cells are presented in histograms; dotted lines showed the negative control. (E) The proportion of liver TRAIL+ and CD69+ NK cells in fed and fasted-transferred mice are shown in bar graph as mean plus standard deviation; *p <0.05 as analyzed by the independent samples T test.
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pone-0110748-g004: Adoptive transfer assay for TRAIL and CD69 expression on liver natural killer cells in response to starvation.(A) Isolated TRAIL− natural killer (NK) cells were separated from liver lymphocytes of wild type B6 mice. Proportion of lymphocytes expressing TCRβ, NK1.1, and TRAIL in whole liver lymphocytes, isolated NK cells, and isolated TRAIL− NK cells are presented in dot plots. The isolated TRAIL− NK cells were adoptively transferred into Rag-2−/− γ chain−/− mice (0.5×106 cells/mouse), which were then fed or fasted for 3 days before determining their NK phenotype. (B) Dot plots show the gated TCRβ− NK1.1+ NK cells and their percentage in the liver, spleen, and bone marrow of non-transferred (control), fed-transferred, and fasted-transferred mice. (C) Bar graph presents the mean percentage plus standard deviation of NK cells in the liver of fed and fasted-transferred mice. (D) Expression of TRAIL and CD69 (solid lines) on the liver NK cells of representative fed- and fasted-transferred mice with their percentages of positive cells are presented in histograms; dotted lines showed the negative control. (E) The proportion of liver TRAIL+ and CD69+ NK cells in fed and fasted-transferred mice are shown in bar graph as mean plus standard deviation; *p <0.05 as analyzed by the independent samples T test.

Mentions: We next examined the mechanism of TRAIL upregulation in fasted mice. To clarify whether TRAIL− NK cells convert into TRAIL+ NK cells in fasting mice, we transferred TRAIL− NK cells that were isolated from liver lymphocytes obtained from wild type B6 mice into Rag-2−/− γ chain−/− B6 mice. The NK cell purity and TRAIL expression rate on the isolated NK cells are shown in Figure 4A. It is noteworthy that these mice present macrophages, but not NK cells or other lymphocytes. The absence of NK cells in Rag-2−/− γ chain−/− mice was analyzed in Figure 4B (control mice). Three days after injection, the injected NK cells homed to the liver, but not to the spleen or the bone marrow (Figure 4B, C). Furthermore, fasted transferred mice showed significantly high expression of TRAIL and CD69 in liver NK cells in comparison with fed transferred mice (Figure 4D, E). These results indicate that TRAIL upregulation is induced in liver-resident NK cells by converting TRAIL− cells into TRAIL+ cells.


Fasting enhances TRAIL-mediated liver natural killer cell activity via HSP70 upregulation.

Dang VT, Tanabe K, Tanaka Y, Tokumoto N, Misumi T, Saeki Y, Fujikuni N, Ohdan H - PLoS ONE (2014)

Adoptive transfer assay for TRAIL and CD69 expression on liver natural killer cells in response to starvation.(A) Isolated TRAIL− natural killer (NK) cells were separated from liver lymphocytes of wild type B6 mice. Proportion of lymphocytes expressing TCRβ, NK1.1, and TRAIL in whole liver lymphocytes, isolated NK cells, and isolated TRAIL− NK cells are presented in dot plots. The isolated TRAIL− NK cells were adoptively transferred into Rag-2−/− γ chain−/− mice (0.5×106 cells/mouse), which were then fed or fasted for 3 days before determining their NK phenotype. (B) Dot plots show the gated TCRβ− NK1.1+ NK cells and their percentage in the liver, spleen, and bone marrow of non-transferred (control), fed-transferred, and fasted-transferred mice. (C) Bar graph presents the mean percentage plus standard deviation of NK cells in the liver of fed and fasted-transferred mice. (D) Expression of TRAIL and CD69 (solid lines) on the liver NK cells of representative fed- and fasted-transferred mice with their percentages of positive cells are presented in histograms; dotted lines showed the negative control. (E) The proportion of liver TRAIL+ and CD69+ NK cells in fed and fasted-transferred mice are shown in bar graph as mean plus standard deviation; *p <0.05 as analyzed by the independent samples T test.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4214715&req=5

pone-0110748-g004: Adoptive transfer assay for TRAIL and CD69 expression on liver natural killer cells in response to starvation.(A) Isolated TRAIL− natural killer (NK) cells were separated from liver lymphocytes of wild type B6 mice. Proportion of lymphocytes expressing TCRβ, NK1.1, and TRAIL in whole liver lymphocytes, isolated NK cells, and isolated TRAIL− NK cells are presented in dot plots. The isolated TRAIL− NK cells were adoptively transferred into Rag-2−/− γ chain−/− mice (0.5×106 cells/mouse), which were then fed or fasted for 3 days before determining their NK phenotype. (B) Dot plots show the gated TCRβ− NK1.1+ NK cells and their percentage in the liver, spleen, and bone marrow of non-transferred (control), fed-transferred, and fasted-transferred mice. (C) Bar graph presents the mean percentage plus standard deviation of NK cells in the liver of fed and fasted-transferred mice. (D) Expression of TRAIL and CD69 (solid lines) on the liver NK cells of representative fed- and fasted-transferred mice with their percentages of positive cells are presented in histograms; dotted lines showed the negative control. (E) The proportion of liver TRAIL+ and CD69+ NK cells in fed and fasted-transferred mice are shown in bar graph as mean plus standard deviation; *p <0.05 as analyzed by the independent samples T test.
Mentions: We next examined the mechanism of TRAIL upregulation in fasted mice. To clarify whether TRAIL− NK cells convert into TRAIL+ NK cells in fasting mice, we transferred TRAIL− NK cells that were isolated from liver lymphocytes obtained from wild type B6 mice into Rag-2−/− γ chain−/− B6 mice. The NK cell purity and TRAIL expression rate on the isolated NK cells are shown in Figure 4A. It is noteworthy that these mice present macrophages, but not NK cells or other lymphocytes. The absence of NK cells in Rag-2−/− γ chain−/− mice was analyzed in Figure 4B (control mice). Three days after injection, the injected NK cells homed to the liver, but not to the spleen or the bone marrow (Figure 4B, C). Furthermore, fasted transferred mice showed significantly high expression of TRAIL and CD69 in liver NK cells in comparison with fed transferred mice (Figure 4D, E). These results indicate that TRAIL upregulation is induced in liver-resident NK cells by converting TRAIL− cells into TRAIL+ cells.

Bottom Line: Liver natural killer cells also showed high TRAIL-mediated antitumor function in response to 3-day fasting.Treatment of liver lymphocytes with 50 µg/mL of recombinant heat shock protein 70 led to the upregulation of both TRAIL and CD69 in liver natural killer cells (n = 6, p <0.05).These findings indicate that acute fasting enhances TRAIL-mediated liver natural killer cell activity against neoplastic cells through upregulation of heat shock protein 70.

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterological and Transplant Surgery, Applied Life Sciences, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.

ABSTRACT
Acute starvation, which is frequently observed in clinical practice, sometimes augments the cytolytic activity of natural killer cells against neoplastic cells. In this study, we investigated the molecular mechanisms underlying the enhancement of natural killer cell function by fasting in mice. The total number of liver resident natural killer cells in a unit weight of liver tissue obtained from C57BL/6J mice did not change after a 3-day fast, while the proportions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)+ and CD69+ natural killer cells were significantly elevated (n = 7, p <0.01), as determined by flow cytometric analysis. Furthermore, we found that TRAIL- natural killer cells that were adoptively transferred into Rag-2-/- γ chain-/- mice could convert into TRAIL+ natural killer cells in fasted mice at a higher proportion than in fed mice. Liver natural killer cells also showed high TRAIL-mediated antitumor function in response to 3-day fasting. Since these fasted mice highly expressed heat shock protein 70 (n = 7, p <0.05) in liver tissues, as determined by western blot, the role of this protein in natural killer cell activation was investigated. Treatment of liver lymphocytes with 50 µg/mL of recombinant heat shock protein 70 led to the upregulation of both TRAIL and CD69 in liver natural killer cells (n = 6, p <0.05). In addition, HSP70 neutralization by intraperitoneally injecting an anti- heat shock protein 70 monoclonal antibody into mice prior to fasting led to the downregulation of TRAIL expression (n = 6, p <0.05). These findings indicate that acute fasting enhances TRAIL-mediated liver natural killer cell activity against neoplastic cells through upregulation of heat shock protein 70.

Show MeSH
Related in: MedlinePlus