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Natural Killer Cells-Produced IFN-γ Improves Bone Marrow-Derived Hepatocytes Regeneration in Murine Liver Failure Model.

Li L, Zeng Z, Qi Z, Wang X, Gao X, Wei H, Sun R, Tian Z - Sci Rep (2015)

Bottom Line: Hepatic NK cells became activated during BMDH generation and were the major IFN-γ producers.Indeed, both NK cells and IFN-γ were required for BMDH generation since WT, but not NK-, IFN-γ-, or IFN-γR1-deficient BM transplantation successfully generated BMDHs and rescued survival in Fah(-/-) hosts.BM-derived myelomonocytes were determined to be the IFN-γ-responding cells.

View Article: PubMed Central - PubMed

Affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, China.

ABSTRACT
Bone-marrow transplantation (BMT) can repopulate the liver through BM-derived hepatocyte (BMDH) generation, although the underlying mechanism remains unclear. Using fumarylacetoacetate hydrolase-deficient (Fah(-/-)) mice as a liver-failure model, we confirmed that BMDHs were generated by fusion of BM-derived CD11b(+)F4/80(+)myelomonocytes with resident Fah(-/-) hepatocytes. Hepatic NK cells became activated during BMDH generation and were the major IFN-γ producers. Indeed, both NK cells and IFN-γ were required for BMDH generation since WT, but not NK-, IFN-γ-, or IFN-γR1-deficient BM transplantation successfully generated BMDHs and rescued survival in Fah(-/-) hosts. BM-derived myelomonocytes were determined to be the IFN-γ-responding cells. The IFN-γ-IFN-γR interaction contributed to the myelomonocyte-hepatocyte fusion process, as most of the CD11b(+) BMDHs in mixed BM chimeric Fah(-/-) hosts transplanted with a 1:1 ratio of CD45.1(+) WT and CD45.2(+) Ifngr1(-/-) BM cells were of CD45.1(+) WT origin. Confirming these findings in vitro, IFN-γ dose-dependently promoted the fusion of GFP(+) myelomonocytes with Fah(-/-) hepatocytes due to a direct effect on myelomonocytes; similar results were observed using activated NK cells. In conclusion, BMDH generation requires NK cells to facilitate myelomonocyte-hepatocyte fusion in an IFN-γ-dependent manner, providing new insights for treating severe liver failure.

No MeSH data available.


Related in: MedlinePlus

NK cells are essential for liver reconstitution.(a) Hepatic NK cell (CD3−NK1.1+) percentage, absolute number, and CD69 expression in Fah−/− mice 16 weeks after WT BMT and 12 weeks after NTBC withdrawal (NTBC off); mice maintained on NTBC were used as controls (NTBC on). Fah−/− mice transplanted with WT BM were treated with anti-ASGM1 mAb (n = 9), anti-NK1.1 mAb (PK136) (n = 9), or PBS control (n = 10) throughout the period of NTBC withdrawal. (b) Survival rate is shown. (c) Liver tissues were collected from mice in the PBS-or anti-NK1.1–treated groups 14 weeks after NTBC withdrawal, and immunohistochemical staining of Fah was performed (scale bar, 500 μm). (d) Survival rate of Fah−/− mice transplanted with Nfil3−/− (n = 9), CD1d−/− (n = 14), or WT (n = 12) BM. (e) Immunohistochemical staining of Fah (scale bar, 200 μm) in liver tissues from Fah−/− mice that received BMT from the indicated mouse strain 16 weeks after NTBC withdrawal. Representative data from 2 or 3 independent experiments are shown as the mean ± SEM.
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f2: NK cells are essential for liver reconstitution.(a) Hepatic NK cell (CD3−NK1.1+) percentage, absolute number, and CD69 expression in Fah−/− mice 16 weeks after WT BMT and 12 weeks after NTBC withdrawal (NTBC off); mice maintained on NTBC were used as controls (NTBC on). Fah−/− mice transplanted with WT BM were treated with anti-ASGM1 mAb (n = 9), anti-NK1.1 mAb (PK136) (n = 9), or PBS control (n = 10) throughout the period of NTBC withdrawal. (b) Survival rate is shown. (c) Liver tissues were collected from mice in the PBS-or anti-NK1.1–treated groups 14 weeks after NTBC withdrawal, and immunohistochemical staining of Fah was performed (scale bar, 500 μm). (d) Survival rate of Fah−/− mice transplanted with Nfil3−/− (n = 9), CD1d−/− (n = 14), or WT (n = 12) BM. (e) Immunohistochemical staining of Fah (scale bar, 200 μm) in liver tissues from Fah−/− mice that received BMT from the indicated mouse strain 16 weeks after NTBC withdrawal. Representative data from 2 or 3 independent experiments are shown as the mean ± SEM.

Mentions: To investigate the local immune response in the liver during BMDH generation, we evaluated hepatic lymphocyte composition and activation after BMT. NTBC withdrawal (NTBC-off condition), which promotes BMDH generation, resulted in dramatic upregulation of CD69 expression of hepatic NK cells (Fig. 2a and Supplementary Fig. S2a), but not T cells (Supplementary Fig. S3a, b), in the livers of BM-transplanted Fah−/− mice compared to control mice continuously receiving NTBC (NTBC-on condition), even though the number and percentage of NK cells did not change (Fig. 2a and Supplementary Fig. S2a). This result indicated the possible involvement of NK cells during BMDH generation. We also found that NK cell depletion by anti-ASGM1 antibody treatment led to decreased survival and BMDH generation in WT BM-transplanted Fah−/− mice, which was further confirmed by NK cell depletion with anti-NK1.1 antibody (Fig. 2b,c and Supplementary Fig. S4). Since anti-NK1.1 antibody also depleted NKT cells, we specifically tested whether NK or NKT played a role in this process by transplanting Fah−/− mice with either Nfil3−/− or CD1d−/− BM, which led to NK or NKT deficiency1920, respectively. Interestingly, Nfil3−/−, but not CD1d−/−, BM lost the ability to rescue Fah−/− mice from liver failure since Nfil3−/− BMT markedly decreased Fah−/− mouse survival and Fah+ BMDH generation after NTBC withdrawal (Fig. 2d,e). This result suggested that NK, but not NKT, cells were essential for BMDH generation. We additionally determined that CD4+ or CD8+ T cells were also not required for BMDH generation, as expected (Supplementary Fig. S3c, d).


Natural Killer Cells-Produced IFN-γ Improves Bone Marrow-Derived Hepatocytes Regeneration in Murine Liver Failure Model.

Li L, Zeng Z, Qi Z, Wang X, Gao X, Wei H, Sun R, Tian Z - Sci Rep (2015)

NK cells are essential for liver reconstitution.(a) Hepatic NK cell (CD3−NK1.1+) percentage, absolute number, and CD69 expression in Fah−/− mice 16 weeks after WT BMT and 12 weeks after NTBC withdrawal (NTBC off); mice maintained on NTBC were used as controls (NTBC on). Fah−/− mice transplanted with WT BM were treated with anti-ASGM1 mAb (n = 9), anti-NK1.1 mAb (PK136) (n = 9), or PBS control (n = 10) throughout the period of NTBC withdrawal. (b) Survival rate is shown. (c) Liver tissues were collected from mice in the PBS-or anti-NK1.1–treated groups 14 weeks after NTBC withdrawal, and immunohistochemical staining of Fah was performed (scale bar, 500 μm). (d) Survival rate of Fah−/− mice transplanted with Nfil3−/− (n = 9), CD1d−/− (n = 14), or WT (n = 12) BM. (e) Immunohistochemical staining of Fah (scale bar, 200 μm) in liver tissues from Fah−/− mice that received BMT from the indicated mouse strain 16 weeks after NTBC withdrawal. Representative data from 2 or 3 independent experiments are shown as the mean ± SEM.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4561890&req=5

f2: NK cells are essential for liver reconstitution.(a) Hepatic NK cell (CD3−NK1.1+) percentage, absolute number, and CD69 expression in Fah−/− mice 16 weeks after WT BMT and 12 weeks after NTBC withdrawal (NTBC off); mice maintained on NTBC were used as controls (NTBC on). Fah−/− mice transplanted with WT BM were treated with anti-ASGM1 mAb (n = 9), anti-NK1.1 mAb (PK136) (n = 9), or PBS control (n = 10) throughout the period of NTBC withdrawal. (b) Survival rate is shown. (c) Liver tissues were collected from mice in the PBS-or anti-NK1.1–treated groups 14 weeks after NTBC withdrawal, and immunohistochemical staining of Fah was performed (scale bar, 500 μm). (d) Survival rate of Fah−/− mice transplanted with Nfil3−/− (n = 9), CD1d−/− (n = 14), or WT (n = 12) BM. (e) Immunohistochemical staining of Fah (scale bar, 200 μm) in liver tissues from Fah−/− mice that received BMT from the indicated mouse strain 16 weeks after NTBC withdrawal. Representative data from 2 or 3 independent experiments are shown as the mean ± SEM.
Mentions: To investigate the local immune response in the liver during BMDH generation, we evaluated hepatic lymphocyte composition and activation after BMT. NTBC withdrawal (NTBC-off condition), which promotes BMDH generation, resulted in dramatic upregulation of CD69 expression of hepatic NK cells (Fig. 2a and Supplementary Fig. S2a), but not T cells (Supplementary Fig. S3a, b), in the livers of BM-transplanted Fah−/− mice compared to control mice continuously receiving NTBC (NTBC-on condition), even though the number and percentage of NK cells did not change (Fig. 2a and Supplementary Fig. S2a). This result indicated the possible involvement of NK cells during BMDH generation. We also found that NK cell depletion by anti-ASGM1 antibody treatment led to decreased survival and BMDH generation in WT BM-transplanted Fah−/− mice, which was further confirmed by NK cell depletion with anti-NK1.1 antibody (Fig. 2b,c and Supplementary Fig. S4). Since anti-NK1.1 antibody also depleted NKT cells, we specifically tested whether NK or NKT played a role in this process by transplanting Fah−/− mice with either Nfil3−/− or CD1d−/− BM, which led to NK or NKT deficiency1920, respectively. Interestingly, Nfil3−/−, but not CD1d−/−, BM lost the ability to rescue Fah−/− mice from liver failure since Nfil3−/− BMT markedly decreased Fah−/− mouse survival and Fah+ BMDH generation after NTBC withdrawal (Fig. 2d,e). This result suggested that NK, but not NKT, cells were essential for BMDH generation. We additionally determined that CD4+ or CD8+ T cells were also not required for BMDH generation, as expected (Supplementary Fig. S3c, d).

Bottom Line: Hepatic NK cells became activated during BMDH generation and were the major IFN-γ producers.Indeed, both NK cells and IFN-γ were required for BMDH generation since WT, but not NK-, IFN-γ-, or IFN-γR1-deficient BM transplantation successfully generated BMDHs and rescued survival in Fah(-/-) hosts.BM-derived myelomonocytes were determined to be the IFN-γ-responding cells.

View Article: PubMed Central - PubMed

Affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027, China.

ABSTRACT
Bone-marrow transplantation (BMT) can repopulate the liver through BM-derived hepatocyte (BMDH) generation, although the underlying mechanism remains unclear. Using fumarylacetoacetate hydrolase-deficient (Fah(-/-)) mice as a liver-failure model, we confirmed that BMDHs were generated by fusion of BM-derived CD11b(+)F4/80(+)myelomonocytes with resident Fah(-/-) hepatocytes. Hepatic NK cells became activated during BMDH generation and were the major IFN-γ producers. Indeed, both NK cells and IFN-γ were required for BMDH generation since WT, but not NK-, IFN-γ-, or IFN-γR1-deficient BM transplantation successfully generated BMDHs and rescued survival in Fah(-/-) hosts. BM-derived myelomonocytes were determined to be the IFN-γ-responding cells. The IFN-γ-IFN-γR interaction contributed to the myelomonocyte-hepatocyte fusion process, as most of the CD11b(+) BMDHs in mixed BM chimeric Fah(-/-) hosts transplanted with a 1:1 ratio of CD45.1(+) WT and CD45.2(+) Ifngr1(-/-) BM cells were of CD45.1(+) WT origin. Confirming these findings in vitro, IFN-γ dose-dependently promoted the fusion of GFP(+) myelomonocytes with Fah(-/-) hepatocytes due to a direct effect on myelomonocytes; similar results were observed using activated NK cells. In conclusion, BMDH generation requires NK cells to facilitate myelomonocyte-hepatocyte fusion in an IFN-γ-dependent manner, providing new insights for treating severe liver failure.

No MeSH data available.


Related in: MedlinePlus