Dependence of innate lymphoid cell 1 development on NKp46

<div><p>NKp46, a natural killer (NK) cell–activating receptor, is involved in NK cell cytotoxicity against virus-infected cells or tumor cells. However, the role of NKp46 in other NKp46⁺ non-NK innate lymphoid cell (ILC) populations has not yet been characterized. Here, an NKp46 deficiency model of natural cytotoxicity receptor 1 (<i>Ncr1</i>)^gfp/gfp and <i>Ncr1</i>^gfp/+ mice, i.e., homozygous and heterozygous knockout (KO), was used to explore the role of NKp46 in regulating the development of the NKp46⁺ ILCs. Surprisingly, our studies demonstrated that homozygous NKp46 deficiency resulted in a nearly complete depletion of the ILC1 subset (ILC1) of group 1 ILCs, and heterozygote KO decreased the number of cells in the ILC1 subset. Moreover, transplantation studies confirmed that ILC1 development depends on NKp46 and that the dependency is cell intrinsic. Interestingly, however, the cell depletion specifically occurred in the ILC1 subset but not in the other ILCs, including ILC2s, ILC3s, and NK cells. Thus, our studies reveal that NKp46 selectively participates in the regulation of ILC1 development.</p></div

NKp46 is required for ILC1 development.

<p>(A) Gating strategy for ILC1s. ILC1s were gated on lymphocytes and then were further defined as Lin⁻NK1.1⁺NKp46⁺CD49b^─CD49a⁺, with the exception that GFP⁺ was used to replace NKp46⁺ when <i>Ncr1</i> ^gfp/+ or <i>Ncr1</i>^gfp/gfp mice were used. (B) Percentages or quantities of ILC1s were determined by flow cytometric analysis in the liver of <i>Ncr1</i>^gfp/gfp, <i>Ncr1</i>^gfp/+, and <i>Ncr1</i>^+/+ mice. (C) NK cells were gated on Lin^─NK1.1⁺NKp46⁺(or GFP⁺ for <i>Ncr1</i>^gfp/gfp mice)CD49b⁺CD49a^─ among lymphocytes. ILC1s were gated on Lin^─NK1.1⁺NKp46⁺(or GFP⁺ for <i>Ncr1</i>^gfp/gfp mice)CD49b^─CD49a⁺ among lymphocytes. (D) Quantification of ILC1s in different organs or tissues in <i>Ncr1</i>^gfp/gfp mice and <i>Ncr1</i>^+/+ littermates, i.e., summary data for (C). Each line demonstrates percentages of ILC1s for a pair of <i>Ncr1</i>^gfp/gfp and <i>Ncr1</i>^+/+ littermates. (E) Quantities of NK cells or ILC1s in different organs of <i>Ncr1</i>^gfp/gfp mice and their <i>Ncr1</i>^+/+ littermates (<i>n</i> = 4). Error bars, standard deviations; ***, <i>p</i> < 0.001; **, <i>p</i> < 0.01; *, <i>p</i> < 0.05. The numerical data for panels B, D and E can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2004867#pbio.2004867.s007" target="_blank">S1 Data</a>. Lin^─, CD3^─CD19^─; BM, bone marrow; FSC-A, forward scatter area; FSC-H, FSC height; FSC-W, FSC width; GFP, green fluorescent protein; ILC1, innate lymphoid cell 1; NK, natural killer; <i>Ncr1</i>, natural cytotoxicity receptor 1; SI, small intestine; SSC-A, side scatter area; SSC-H, SSC height; SSC-W, SSC width.</p

NKp46 plays a cell-intrinsic role in regulating ILC1 development.

<p>(A) Scheme of BM transplantation using BM cells of CD45.2 <i>Ncr1</i>^gfp/gfp mice or <i>Ncr1</i>^+/+ littermate controls as donor cells to inject into CD45.1 recipients via tail vein. Development of ILC subsets was analyzed 2 weeks after transplantation. (B) Percentages of CD45.2⁺ NK cells or CD45.2⁺ ILC1s were analyzed by flow cytometric analysis in the liver of CD45.1 recipients, which were engrafted with BM cells of <i>Ncr1</i>^gfp/gfp mice (<i>n</i> = 5) or <i>Ncr1</i>^+/+ littermates (<i>n</i> = 4). (C) Percentages of CD45.2⁺ NK cells or CD45.2⁺ ILC1s were analyzed in the spleen or BM of CD45.1 recipient mice, which were engrafted with BM cells of <i>Ncr1</i>^gfp/gfp mice (<i>n</i> = 5) or their <i>Ncr1</i>^+/+ littermates (<i>n</i> = 4). (D and E) Data shown are representative dot plots of flow cytometric analysis (left panel) and summary data (right panel) of CD45.2⁺ILC2 (D) or CD45.2⁺ ILC3 (E) in SI in CD45.1 recipients, which were engrafted with BM cells of <i>Ncr1</i>^gfp/gfp mice (<i>n</i> = 4) or their <i>Ncr1</i>^+/+ littermates (<i>n</i> = 4). Error bars, standard deviations; ***, <i>p</i> < 0.001; **, <i>p</i> < 0.01; *, <i>p</i> < 0.05. The numerical data for panels B, C, D and E can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2004867#pbio.2004867.s007" target="_blank">S1 Data</a>. Lin^─, CD3^─CD19^─; BM, bone marrow; ILC, innate lymphoid cells; <i>Ncr1</i>, natural cytotoxicity receptor 1; NK, natural killer; RORγt, retinoic acid receptor (RAR) related orphan receptor gamma t; SI, small intestine.</p

Absence of TRAIL⁺ ILC1s in NKp46-deficient mice.

<p>(A) Percentages of TRAIL⁺ ILC1s were analyzed by flow cytometric analysis in the liver of <i>Ncr1</i>^gfp/gfp mice and their <i>Ncr1</i>^+/+ littermates. ILC1s were gated on Lin^─NK1.1⁺NKp46⁺(or GFP⁺ for <i>Ncr1</i>^gfp/gfp mice) TRAIL⁺CD49b^─ among lymphocytes. (B) Quantification of TRAIL⁺ ILC1s in the liver of <i>Ncr1</i>^gfp/gfp mice and their <i>Ncr1</i>^+/+ littermates for (A) (<i>n</i> = 5). (C) Quantification of TRAIL⁺ ILC1s in other organs (spleen, <i>n</i> = 5; BM, <i>n</i> = 5; SI, <i>n</i> = 4) of <i>Ncr1</i>^gfp/gfp mice and their <i>Ncr1</i>^+/+ littermates. **, <i>p</i> < 0.01; *, <i>p</i> < 0.05. The numerical data for panels B and C can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.2004867#pbio.2004867.s007" target="_blank">S1 Data</a>. Lin^─, CD3^─CD19^─; BM, bone marrow; GFP, green fluorescent protein; ILC1, innate lymphoid cell 1; <i>Ncr1</i>, natural cytotoxicity receptor 1; NK, natural killer; SI, small intestine; TRAIL, tumor necrosis factor–related apoptosis-inducing ligand.</p

Molecular Tailoring Based on Forster Resonance Energy Transfer for Initiating Two-Photon Theranostics with Amplified Reactive Oxygen Species

The fabrication of multifunctional photosensitizers (PSs) with abundant Type I/II ROS for efficient theranostics in the “therapeutic window” (700–900 nm) is an appealing yet significantly challenging task. We herein report a molecular tailoring strategy based on intramolecular two-photon Forster Resonance Energy Transfer (TP-FRET) to obtain a novel theranostic agent (Lyso-FRET), featuring the amplified advantage of energy donor (NH) and acceptor (COOH), because of the reuse of fluorescence energy with high efficiency of FRET (∼83%). Importantly, under the excitation by the near-infrared (840 nm) window, Lyso-FRET can not only penetrate the deeper tissue with a higher resolution for fluorescence imaging due to the nonlinear optical (NLO) nature, but also generate more Type I (superoxide anion) and Type II (singlet oxygen) reactive oxygen species for hypoxic PDT. Moreover, Lyso-FRET targeting lysosomes further promotes the effect of treatment. The experiments in vitro and in vivo also verify that the developed TP-FRET PS is conducive to treating deep hypoxic tumors. This strategy provides new and significant insights into the design and fabrication of advanced multifunctional PSs

Crystal Structures and Human Leukemia Cell Apoptosis Inducible Activities of Parthenolide Analogues Isolated from <i>Piptocoma rufescens</i>

The molecular structures of three parthenolide analogues, (−)-goyazensolide (<b>1</b>), (−)-15-deoxygoyazensolide (<b>2</b>), and (−)-ereglomerulide (<b>3</b>), isolated from the leaves of <i>Piptocoma rufescens</i> in a previous study were determined by X-ray analysis, and the absolute configuration of (−)-goyazensolide (<b>1</b>) was confirmed crystallographically using Cu Kα radiation at low temperature. Compounds <b>1</b>–<b>3</b>, (+)-rufesolide A (<b>4</b>), and commercial parthenolide were found to be growth inhibitory toward MOLM-13 and EOL-1 human acute myeloid leukemia cells using PKC412 (midostaurin) as the positive control, with <b>1</b>–<b>3</b> being more active than parthenolide. Also, compounds <b>1</b>–<b>4</b> exhibited synergistic effects when tested with PKC412, but parthenolide did not show this type of activity. At a concentration lower than 2.0 μM, both <b>1</b> and <b>2</b> induced approximately 50% of the cells to become apoptotic at a late stage of the cell cycle, but no similar apoptotic effects were observed for <b>3</b>, <b>4</b>, or parthenolide. Leukemia cell apoptosis was induced by these compounds through the activation of caspase-3 and the inhibition of NF-κB, as indicated by immunoblotting analysis, and compounds <b>1</b> and <b>2</b> seem to be promising leads for development as potential antileukemic agents

Image_5_Host species of freshwater snails within the same freshwater ecosystem shapes the intestinal microbiome.tif

BackgroundFreshwater snails are not only intermediate hosts for parasites but also an important part of the food chain as they convert plant biomass and humus into animal biomass. However, being widely distributed in freshwater environments, snails are highly affected by human activities, which makes their adaptation to altering environments challenging. The gut microbiome helps animals in their digestion, immune system, growth and adapting to changing environments. The effect of host species on intestinal microbial community has been poorly studied in snails.MethodsIn this study, single-molecule real-time sequencing technology (SMRT) was used to obtain full-length 16S rRNA genes to determine the intestinal microbiomes of three species of freshwater snails (SQ: Sinotaia quadrata, BU: Boreoelona ussuriensis, RP: Radix plicatula) with similar feeding habits in a same water environment.ResultsUnifrac PCoA (PConcludesLelliottia, Romboutsia, Clostridium_sensu_stricto_1, and Pirellula play an important role in the intestinal microbiota phenotype of the host snails. In general, the host species affects the structure of the gut microbial community, which in turn helps gastropods improve their environmental adaptability, but further study is still needed.</p

Image_4_Host species of freshwater snails within the same freshwater ecosystem shapes the intestinal microbiome.tif

BackgroundFreshwater snails are not only intermediate hosts for parasites but also an important part of the food chain as they convert plant biomass and humus into animal biomass. However, being widely distributed in freshwater environments, snails are highly affected by human activities, which makes their adaptation to altering environments challenging. The gut microbiome helps animals in their digestion, immune system, growth and adapting to changing environments. The effect of host species on intestinal microbial community has been poorly studied in snails.MethodsIn this study, single-molecule real-time sequencing technology (SMRT) was used to obtain full-length 16S rRNA genes to determine the intestinal microbiomes of three species of freshwater snails (SQ: Sinotaia quadrata, BU: Boreoelona ussuriensis, RP: Radix plicatula) with similar feeding habits in a same water environment.ResultsUnifrac PCoA (PConcludesLelliottia, Romboutsia, Clostridium_sensu_stricto_1, and Pirellula play an important role in the intestinal microbiota phenotype of the host snails. In general, the host species affects the structure of the gut microbial community, which in turn helps gastropods improve their environmental adaptability, but further study is still needed.</p

Table_1_Host species of freshwater snails within the same freshwater ecosystem shapes the intestinal microbiome.xlsx

BackgroundFreshwater snails are not only intermediate hosts for parasites but also an important part of the food chain as they convert plant biomass and humus into animal biomass. However, being widely distributed in freshwater environments, snails are highly affected by human activities, which makes their adaptation to altering environments challenging. The gut microbiome helps animals in their digestion, immune system, growth and adapting to changing environments. The effect of host species on intestinal microbial community has been poorly studied in snails.MethodsIn this study, single-molecule real-time sequencing technology (SMRT) was used to obtain full-length 16S rRNA genes to determine the intestinal microbiomes of three species of freshwater snails (SQ: Sinotaia quadrata, BU: Boreoelona ussuriensis, RP: Radix plicatula) with similar feeding habits in a same water environment.ResultsUnifrac PCoA (PConcludesLelliottia, Romboutsia, Clostridium_sensu_stricto_1, and Pirellula play an important role in the intestinal microbiota phenotype of the host snails. In general, the host species affects the structure of the gut microbial community, which in turn helps gastropods improve their environmental adaptability, but further study is still needed.</p

Image_3_Host species of freshwater snails within the same freshwater ecosystem shapes the intestinal microbiome.tif

BackgroundFreshwater snails are not only intermediate hosts for parasites but also an important part of the food chain as they convert plant biomass and humus into animal biomass. However, being widely distributed in freshwater environments, snails are highly affected by human activities, which makes their adaptation to altering environments challenging. The gut microbiome helps animals in their digestion, immune system, growth and adapting to changing environments. The effect of host species on intestinal microbial community has been poorly studied in snails.MethodsIn this study, single-molecule real-time sequencing technology (SMRT) was used to obtain full-length 16S rRNA genes to determine the intestinal microbiomes of three species of freshwater snails (SQ: Sinotaia quadrata, BU: Boreoelona ussuriensis, RP: Radix plicatula) with similar feeding habits in a same water environment.ResultsUnifrac PCoA (PConcludesLelliottia, Romboutsia, Clostridium_sensu_stricto_1, and Pirellula play an important role in the intestinal microbiota phenotype of the host snails. In general, the host species affects the structure of the gut microbial community, which in turn helps gastropods improve their environmental adaptability, but further study is still needed.</p