Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2020 Dec 8:9:e59258.
doi: 10.7554/eLife.59258.

Neutrophil infiltration regulates clock-gene expression to organize daily hepatic metabolism

María Crespo #  1 , Barbara Gonzalez-Teran #  1 , Ivana Nikolic  1 , Alfonso Mora  1 , Cintia Folgueira  1 , Elena Rodríguez  1 , Luis Leiva-Vega  1 , Aránzazu Pintor-Chocano  1 , Macarena Fernández-Chacón  1 , Irene Ruiz-Garrido  1 , Beatriz Cicuéndez  1 , Antonia Tomás-Loba  1 , Noelia A-Gonzalez  1 , Ainoa Caballero-Molano  1 , Daniel Beiroa  2   3 , Lourdes Hernández-Cosido  4 , Jorge L Torres  5 , Norman J Kennedy  6 , Roger J Davis  6 , Rui Benedito  1 , Miguel Marcos  5 , Ruben Nogueiras  2   3 , Andrés Hidalgo  1 , Nuria Matesanz  1 , Magdalena Leiva #  1 , Guadalupe Sabio  1
Affiliations

Neutrophil infiltration regulates clock-gene expression to organize daily hepatic metabolism

María Crespo et al. Elife. .

Abstract

Liver metabolism follows diurnal fluctuations through the modulation of molecular clock genes. Disruption of this molecular clock can result in metabolic disease but its potential regulation by immune cells remains unexplored. Here, we demonstrated that in steady state, neutrophils infiltrated the mouse liver following a circadian pattern and regulated hepatocyte clock-genes by neutrophil elastase (NE) secretion. NE signals through c-Jun NH2-terminal kinase (JNK) inhibiting fibroblast growth factor 21 (FGF21) and activating Bmal1 expression in the hepatocyte. Interestingly, mice with neutropenia, defective neutrophil infiltration or lacking elastase were protected against steatosis correlating with lower JNK activation, reduced Bmal1 and increased FGF21 expression, together with decreased lipogenesis in the liver. Lastly, using a cohort of human samples we found a direct correlation between JNK activation, NE levels and Bmal1 expression in the liver. This study demonstrates that neutrophils contribute to the maintenance of daily hepatic homeostasis through the regulation of the NE/JNK/Bmal1 axis.

Keywords: JNK; cell biology; circadian rhythm; immunology; inflammation; mouse; neutrophil elastase; steatosis.

Plain language summary

Every day, the body's biological processes work to an internal clock known as the circadian rhythm. This rhythm is controlled by ‘clock genes’ that are switched on or off by daily physical and environmental cues, such as changes in light levels. These daily rhythms are very finely tuned, and disturbances can lead to serious health problems, such as diabetes or high blood pressure. The ability of the body to cycle through the circadian rhythm each day is heavily influenced by the clock of one key organ: the liver. This organ plays a critical role in converting food and drink into energy. There is evidence that neutrophils – white blood cells that protect the body by being the first response to inflammation – can influence how the liver performs its role in obese people, by for example, releasing a protein called elastase. Additionally, the levels of neutrophils circulating in the blood change following a daily pattern. Crespo, González-Terán et al. wondered whether neutrophils enter the liver at specific times of the day to control liver’s daily rhythm. Crespo, González-Terán et al. revealed that neutrophils visit the liver in a pattern that peaks when it gets light and dips when it gets dark by counting the number of neutrophils in the livers of mice at different times of the day. During these visits, neutrophils secreted elastase, which activated a protein called JNK in the cells of the mice’s liver. This subsequently blocked the activity of another protein, FGF21, which led to the activation of the genes that allow cells to make fat molecules for storage. JNK activation also switched on the clock gene, Bmal1, ultimately causing fat to build up in the mice’s liver. Crespo, González-Terán et al. also found that, in samples from human livers, the levels of elastase, the activity of JNK, and whether the Bmal1 gene was switched on were tightly linked. This suggests that neutrophils may be controlling the liver’s rhythm in humans the same way they do in mice. Overall, this research shows that neutrophils can control and reset the liver's daily rhythm using a precisely co-ordinated series of molecular changes. These insights into the liver's molecular clock suggest that elastase, JNK and BmaI1 may represent new therapeutic targets for drugs or smart medicines to treat metabolic diseases such as diabetes or high blood pressure.

PubMed Disclaimer

Conflict of interest statement

MC, BG, IN, AM, CF, ER, LL, AP, MF, IR, BC, AT, NA, AC, DB, LH, JT, NK, RD, RB, MM, RN, AH, NM, ML, GS No competing interests declared

Figures

Figure 1.
Figure 1.. Neutrophil infiltration into the liver controls hepatic clock-gene expression.
(A) Flow cytometry analysis of the CD11b+Ly6G+ liver myeloid subset, isolated from C57BL6J mice at the indicated ZTs. Left, CD11b+Ly6G+ liver myeloid subset analyzed at 6 hr intervals and normalized by the tissue weight. Right, percentage of CD11b+Ly6G+ population analyzed at 4 hr intervals and normalized to ZT2 (n = 5). (B) Representative 3-D image of liver section showing the distribution on infiltrated neutrophils. Livers were stained with anti-S100A9 (Mrp14) (red) and vessels were stained with anti-CD31 and anti-endomucin (grey). Sizes of the liver sections are 510 x 510 x 28 µm and 160 x 160 x 28 µm, respectively. (C) qRT-PCR analysis of circadian clock-gene and nuclear-receptor mRNA expression in livers from C57BL6J mice at the indicated ZTs (n = 5). (D) Liver triglycerides and oil-red-stained liver sections prepared from C57BL6J mice at ZT2 and ZT14. Scale bar, 50 μm (n = 5). (E) qRT-PCR analysis of clock-gene mRNA in hepatocyte cultures exposed to freshly isolated FMLP-activated neutrophils (n = 4-6 wells of 3 independent experiments). (F) qRT-PCR analysis of clock-gene mRNA in hepatocyte cultures treated with 5 nM elastase (n = 3-4 wells of 3 independent experiments). (G) qRT-PCR analysis of clock-gene and nuclear-receptor mRNA expression in livers from control mice (Mrp8-Cre) and neutropenic mice (MCL1Mrp8-KO) sacrificed at ZT2 (n = 5). (H) Hepatic triglycerides detected in livers from control mice (Mrp8-Cre) and neutropenic mice (MCL1Mrp8-KO) at ZT2 (n = 5). Data are means ± SEM from at least 2 independent experiments. *p<0.05; **p<0.01; ***p<0.005 (A, left panel) One-way ANOVA with Tukey’s post hoc test. (A, right panel) Kruskal-Wallis test with Dunn’s post hoc test. (C) One-way ANOVA with Tukey’s post hoc test or Kruskal-Wallis test with Dunn’s post hoc test. (D to H) t-test or Welch’s test. ZT2 point is double plotted to facilitate viewing.
Figure 1—figure supplement 1.
Figure 1—figure supplement 1.. Neutrophils follow a circadian rhythm.
(A) Left, circulating neutrophils quantified at 4 hr intervals in whole blood of C57BL6J mice. Right, flow cytometry analysis at 6 hr intervals of the CD11b+Ly6G+ myeloid subset in bone marrow from C57BL6J mice. ZT2 point is double plotted to facilitate viewing (n = 5). (B) Representative 3-D image of liver section showing the distribution of Kupffer cells. Livers were stained with anti-Clec4F (green) and vessels were stained with anti-CD31 and anti-endomucin (grey). Sizes of the liver sections are 510 x 510 x 28 µm and 160 x 160 x 28 µm, respectively (n = 5-7). (C) qRT-PCR of Ccl3, Cxcl2, Cxcl12 and Cxcl1 chemokines mRNA expression at ZT2 and ZT14 and qRT-PCR of Cxcl1 mRNA expression at 6 hr intervals in livers from C57BL6J mice (n = 5). (D) qRT-PCR of Bmal1 mRNA expression in hepatocyte cultures exposed to freshly isolated T-lymphocytes, B-lymphocytes or bone-marrow derived macrophages (BMDM) and 1 µM FMLP; Bmal1 mRNA expression in hepatocyte cultures treated with 0.5 mg/mL collagenase (n = 3 wells of 2 to 3 independent experiments) (E) Left, flow cytometry analysis of number of liver Kupffer cells (KCs) in control Lyzs-Cre and MCL1Lyzs-KO mice and in Mrp8-Cre and MCL1Mrp8-KO mice normalized by tissue weight. Right, representative dot plots showing F4/80+Clec4F+ population gated on total intrahepatic CD45+CD11b+ leukocyte population (n = 4-6). (F) Flow cytometry analysis of the CD11b+ Gr-1high liver myeloid subset isolated from control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice. The bar chart shows the CD11b+ Gr-1high population as a percentage of the total intrahepatic CD11b+ leukocyte population (n = 7-10). Data are means ± SEM. *p<0.05; **p<0.01; ***p<0.005 (A, left) Kruskal-Wallis with Dunn’s post-hoc test. (A, right) One-way ANOVA with Tukey’s pots hoc test. (C, left) t-test. (C, right) Kruskal-Wallis with Dunn’s post-hoc test. (D) t-test. (E) One-way ANOVA with Tukey’s pots-hoc test. (F) t-test.
Figure 1—figure supplement 2.
Figure 1—figure supplement 2.. Neutrophil deficiency alters clock-gene expression.
(A) Representative dot plots showing the decrease in the CD11b+ Gr-1high population in blood, bone marrow, and spleen from neutropenic mice (MCL1Lyzs-KO) compared with control mice (Lyzs-Cre). Bar charts show the CD11b+ Gr-1high population as a percentage of the total CD11b+ leukocyte population. (B) Blood levels of monocytes and neutrophils in control and neutropenic mice. (C) Myeloid cell populations in bone marrow and liver determined by flow cytometry and representative dot plots (CD11b+ Gr-1neg as macrophages, CD11b+ Gr-1int as monocytes and CD11b+ Gr-1high as neutrophils). (D) qRT-PCR of clock genes in the livers from control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice. ZT2 point is double plotted to facilitate viewing (n = 5-7). (E) Left, flow cytometry analysis of the CD11b+ Ly6G+ lung myeloid subset of control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice at the indicated ZTs (n = 4). Right, qRT-PCR analysis of Bmal1 in lungs of control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice at the indicated ZTs (n = 4-6). Data are means ± SEM. *p<0.05; **p< 0.01; ***p<0.005. All tests are t-test or Welch’s test.
Figure 2.
Figure 2.. Increased hepatic neutrophil infiltration alters clock-genes expression and augments triglyceride content in the liver.
(A–D) Control (Lyzs-Cre) (A–B) and control and neutropenic (MCL1Lyzs-KO) mice (C–D) were housed for 3 weeks with a normal 12 hr: 12 hr light/dark cycle (Normal Cycle) or with the dark period extended by 12 hr every 5 days (JetLag). Samples were obtained at the indicated ZTs. (A) Left, flow cytometry analysis of the CD11b+Ly6G+ liver myeloid subset. Data represents the percentage CD11b+Ly6G+ normalized to Normal Cycle ZT2. Right, circulating neutrophils in whole blood. (n = 5-8). (B) Liver triglycerides and representative oil-red-stained liver sections at ZT14. Scale bar, 50 μm (n = 9-10). (C) Hepatic triglyceride content analyzed at 6 hr intervals, and representative oil-red-stained liver sections at ZT14. Scale bar, 50 μm (n = 4-6). (D) qRT-PCR analysis of Bmal1 mRNA in livers. (n = 5-8). (E) Flow cytometry analysis of the CD11b+Ly6G+ liver myeloid subset isolated at 6 hr intervals from C57BL6J mice fed a ND, a HFD (8 weeks) or a MCD (3 weeks). The chart shows the CD11b+Ly6G+ population as a percentage of the total intrahepatic CD11b+ leukocyte population normalized to ND group at ZT2 (n = 5 to 10). (F–I) Control mice (Lyzs-Cre) and neutropenic mice (MCL1Lyzs-KO) or p38γ/δLyzs-KO were fed a ND or the MCD diet for 3 weeks and sacrificed at ZT2. (F) Representative images of the infiltration of neutrophils in the liver stained with anti-Mrp14 (blue) and anti-NE (red); nuclei with Sytox Green. Scale bar, 50 μm (Top) and 25 μm (Bottom). (G) qRT-PCR analysis of clock-gene expression in livers (n = 6). (H) Liver triglycerides and representative oil-red-stained liver sections. Scale bar, 50 μm (n = 7-6). (I) qRT-PCR analysis of clock genes in livers at ZT2 (n = 9-17). Data are means ± SEM from at least two independent experiments. *p<0.05; **p<0.01; ***p<0.005 (A to D) t-test or Welch’s test. (E) Two-way ANOVA with Fisher’s post hoc test; p<0.05 ND vs HFD; p<0.0001 ND vs MCD. *p<0.05; ***p<0.005 (G to I) t-test or Welch’s test. ZT2 point is double plotted to facilitate viewing.
Figure 2—figure supplement 1.
Figure 2—figure supplement 1.. Defective neutrophil migration to the liver alters hepatic clock- gene expression and triglyceride content.
(A) Schematic representation of JetLag protocol with stepwise increases in the dark period of 12 h12h every 5 days (B) Flow cytometry analysis of the CD11b+Ly6G+ liver myeloid subset isolated from control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice housed for 3 weeks under a 12 hr:12 hr light/dark cycle (Normal Cycle) or Jetlag. The bar chart shows the percentage of CD11b+Ly6G+ total intrahepatic CD11b+ leukocyte population analyzed at 6-h intervals and normalized to Normal Cycle ZT2 (n = 5-7). Dot plots show CD11b+ Ly6G+ population at ZT14. (C–D) After bone -marrow (BM) reconstitution of irradiated WT mice using Mrp8-Cre (Mrp8-Cre BM) or CXCR2Mrp8-KO (CXCR2Mrp8-KO) mice as BM donors, mice were housed for 3 weeks under JetLag (n = 6-8) (C) qRT-PCR analysis of Bmal1 mRNA in livers at ZT14. (D) Hepatic triglyceride content and representative oil-red-stained liver sections at ZT14. Scale bar, 50 µm. (E–G) Control (Lyzs-Cre) and p38γ/δLyzs-KO mice were housed for 3 weeks under JetLag (n = 6-7) (E) Flow cytometry analysis of the CD11b+ Ly6G+ liver myeloid subset analyzed at 6 hr intervals and normalized by the tissue weight. (F) qRT-PCR analysis of Bmal1 mRNA in livers at ZT14. (G) Hepatic triglyceride content and representative oil-red-stained liver sections at ZT14. Scale bar, 50 µm. Data are means ± SEM. *p<0.05; **p< 0.01; ***p<0.005 All tests are t-test or Welch’s test. ZT2 point is double plotted to facilitate viewing.
Figure 2—figure supplement 2.
Figure 2—figure supplement 2.. Neutrophil depletion alters hepatic clock-gene expression.
(A-C) Osmotic minipumps containing saline or Ly6G antibody were implanted subcutaneously in Lyzs-Cre mice. These animals were fed with a MCD diet for 3 weeks and sacrificed at ZT2. (A-B) Blood levels of neutrophils and monocytes in Lyzs-Cre after 3 weeks of MCD diet treated or not with Ly6G antibody. (C) qRT-PCR of clock genes in the liver (n = 7-9). Data are means ± SEM. *p<0.05; ***p<0.005. All tests are t-test or Welch’s test.
Figure 3.
Figure 3.. Diurnal regulation of liver metabolism involves neutrophil-mediated regulation of JNK and the hepatokine FGF21.
Immunoblot analysis of JNK content and activation at ZT2 in liver extracts prepared from control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice fed a MCD diet for 3 weeks (A) or Lyzs-Cre and p38γ/δLyzs-KO mice after 3 weeks of MCD diet (B). Immunoblot analysis of JNK content and activation (C) and Bmal1 RNA expression (D) in hepatocyte cultures exposed to NE for 2 hr (n = 14 wells of 3 independent experiments). Immunoblot quantification is shown in Figure 3—figure supplement 1D (E) qRT-PCR analysis of clock genes and Fgf21 in livers from Alb-Cre, and JNK1/2Alb-KO mice after 3 weeks of MCD diet at ZT2 (n = 9-12). (F) Immunoblot analysis of FGF21 content in liver extracts prepared from control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice, or from Lyzs-Cre, and p38γ/δLyzs-KO mice after 3 weeks of MCD diet sacrificed at ZT2. Immunoblot quantification is shown in Figure 3—figure supplement 1I,J. (G–I) Lyzs-Cre and p38γ/δLyzs-KO mice were injected with 2 shRNA independent clones targeting FGF21. Seven days after infection, mice were placed on the MCD diet and sacrificed after 3 weeks at ZT2. (G) Immunoblot analysis of FGF21 content in liver extracts prepared from Lyzs-Cre, p38γ/δLyzs-KO, and p38γ/δLyzs-KO mice infected with FGF21 shRNA. Immunoblot quantification is shown in Figure 3—figure supplement 1K. (H) Representative H&E-stained liver sections. Scale bar, 50 μm. (I) Hepatic triglyceride content at the end of the treatment period (n = 8-10). Data are means ± SEM from at least 2 independent experiments. *p<0.05; **p<0.01; ***p<0.005 (A, B, D and E) t-test or Welch’s test. (I) One-way ANOVA with Bonferroni post hoc test or t-test.
Figure 3—figure supplement 1.
Figure 3—figure supplement 1.. Neutrophils regulate hepatic metabolism and clock genes through JNK and FGF21.
(A) qRT-PCR analysis of the metabolic gene Acaca in livers of control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice fed the MCD diet for 3 weeks (n = 4). (B) Immunoblot analysis of ACC content in livers from Lyzs-Cre and p38γ/δLyzs-KO mice at the end of the MCD diet. (C) Immunoblot analysis of ACC content and JNK content and activation in extracts prepared from hepatocyte cultures exposed to freshly isolated FMLP-activated for 1 h. Quantification is shown in the bottom panels. (D) Immunoblot analysis quantification of JNK content and activation in hepatocyte cultures treated with neutrophil elastase (NE) for 2 h. (E) qRT-PCR analysis of the metabolic gene Acaca mRNA expression from livers of Alb-Cre and JNK1/2Alb-KO mice fed a MCD for 3 weeks (n = 10-12). (F) qRT-PCR analysis of the clock genes Bmal1 and Clock and the metabolic gene Acaca mRNA expression from livers of control and neutropenic mice treated with the JNK inhibitor SP600125. Mice were sacrificed at ZT2 (n = 6-7). (G) Immunoblot of c-Jun activation at ZT2 in livers from control and neutropenic mice treated with the JNK inhibitor SP600125. (H) qRT-PCR analysis of Fgf21 mRNA expression in hepatocyte cultures exposed to freshly isolated FMLP-activated neutrophils 1 hr (n = 4 to 6 wells of 3 independent experiments). (I-K) Quantification of the immunoblot analysis of FGF21 content in extracts prepared from livers of control (Lyzs-Cre) and neutropenic (MCL1Lyzs-KO) mice fed the MCD diet for 3 weeks (I), Lyzs-Cre, and p38γ/δLyzs-KO mice fed the MCD diet for 3 weeks (JC), and Lyzs-Cre and p38γ/δLyzs-KO mice injected with 2 shRNA independent clones targeting FGF21 and fed the MCD diet for 3 weeks (K) (n = 3). Data are means ± SEM. *p< 0.05; **p< 0.01; ***p<0.005 (A–E) t-test. (F) One-way ANOVA with Tukey’s post hoc test, Kruskal-Wallis with Dunn’s post hoc test or t-test. (H to J) t-test or Welch’s test. (K) One-way ANOVA with Bonferroni post hoc test or t-test.
Figure 4.
Figure 4.. Elastase controls liver clock-gene expression modulating JNK activation.
(A) Extracellular NE levels in livers from WT mice at ZT2 and ZT14. (B) qRT-PCR analysis of clock-genes and nuclear-receptor mRNA expression in livers from WT and NE KO mice (NE-/-) at ZT2 (n = 5–6). (C) Respiratory exchange ratio of WT and NE-/- mice fed with ND. Results are from the lights-on period (n = 9). (D–H) WT and NE-/- mice were fed a MCD diet for 3 weeks and sacrificed at the indicated time. (D) Liver triglycerides at the end of the diet period. (E) Representative oil-red-stained liver sections. Scale bar, 50 μm (n = 10). (F) Immunoblot analysis and quantifications of JNK content and activation in liver extracts prepared from WT and NE-/-. (G) Immunoblot analysis and quantification of ACC content in liver extracts from WT and NE-/- mice. (H) qRT-PCR analysis of clock-genes and nuclear-receptor mRNA expression in livers from WT and NE-/- mice at ZT2 and ZT14 (n = 7–8). Data are means ± SEM from at least two independent experiments. *p<0.05; **p<0.01; ***p<0.005 (A to G) t-test or Welch’s test. (H) One-way ANOVA with to Tukey’s post hoc test, t-test or Welch’s test.
Figure 4—figure supplement 1.
Figure 4—figure supplement 1.. Neutrophil elastase regulates daily hepatic metabolism through JNK.
NE-/- and control mice were fed a HFD for 8 weeks. (A) Liver triglycerides at the end of the diet period (n = 5). (B) Representative oil-red-stained liver sections. Scale bar, 50 μm. (C) Liver weight at the end of the treatment (n = 5). (D) Immunoblot analysis and quantifications of ACC content and JNK content and activation in liver extracts prepared from WT and NE-/- mice. (E) qRT-PCR analysis of clock-genes mRNA expression in livers from WT mice fed a ND (upper panels) and in WT and NE-/- mice fed a HFD (at ZT2 and ZT14 (bottom panels) at ZT12 and ZT14 (n = 5)). Data are means ± SEM from at least 2 independent experiments.*p<0.05; **p<0.01; ***p<0.005 (A and C) One-way ANOVA with Bonferroni post hoc test. (D and E) t-test or Welch’s test.
Figure 5.
Figure 5.. Neutrophil elastase reverses neutropenic mice phenotype through regulation of daily hepatic metabolism.
(A–D) Neutropenic (MCL1Lyzs-KO) mice were housed for 2 weeks with the dark period extended by 12 hr every 5 days (JetLag). Mice were infused with purified WT or NE-/- neutrophils. Samples were obtained at ZT14. (A) Picture describing the neutrophil infusion schedule during the JetLag protocol. (B) qRT-PCR analysis of Bmal1 mRNA in livers. (C) Liver triglycerides and (D) representative oil-red-stained liver sections. Scale bar, 50 µm (n = 6-7). Data are means ± SEM. *p<0.05; t-test. (E) Correlation between mRNA levels of BMAL1 and ELANE (r = 0.6141; p = 0.0052) or JUN and ELANE (r = 0.7362; p = 0.001105) in human livers. The mRNA levels of JUN, BMAL1 and ELANE were determined by qRT-PCR. Linear relationships between variables were tested using Pearson’s correlation coefficient (n = 23). (F) Circadian neutrophil infiltration regulates hepatic metabolism through elastase, JNK and FGF21. Data are means ± SEM. *p< 0.05; **p< 0.01; (B) One-way ANOVA with Tukey’s pots hoc test. (C) t-test or Welch’s test.
Author response image 1.
Author response image 1.. Representative images of MRP8-Cre and MCL-1MRP8-KO mice.
8 weeks-old Mrp8-Cre (size: 8cm, weight: 23.36g) and MCL1Mrp8-KO (size: 6.8cm, weight: 17.3g) male mice and their size are shown.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Adrover JM, Del Fresno C, Crainiciuc G, Cuartero MI, Casanova-Acebes M, Weiss LA, Huerga-Encabo H, Silvestre-Roig C, Rossaint J, Cossío I, Lechuga-Vieco AV, García-Prieto J, Gómez-Parrizas M, Quintana JA, Ballesteros I, Martin-Salamanca S, Aroca-Crevillen A, Chong SZ, Evrard M, Balabanian K, López J, Bidzhekov K, Bachelerie F, Abad-Santos F, Muñoz-Calleja C, Zarbock A, Soehnlein O, Weber C, Ng LG, Lopez-Rodriguez C, Sancho D, Moro MA, Ibáñez B, Hidalgo A. A neutrophil timer coordinates immune defense and vascular protection. Immunity. 2019;50:390–402. doi: 10.1016/j.immuni.2019.01.002. - DOI - PubMed
    1. Belaaouaj A, McCarthy R, Baumann M, Gao Z, Ley TJ, Abraham SN, Shapiro SD. Mice lacking neutrophil elastase reveal impaired host defense against gram negative bacterial Sepsis. Nature Medicine. 1998;4:615–618. doi: 10.1038/nm0598-615. - DOI - PubMed
    1. Casanova-Acebes M, Pitaval C, Weiss LA, Nombela-Arrieta C, Chèvre R, A-González N, Kunisaki Y, Zhang D, van Rooijen N, Silberstein LE, Weber C, Nagasawa T, Frenette PS, Castrillo A, Hidalgo A. Rhythmic modulation of the hematopoietic niche through neutrophil clearance. Cell. 2013;153:1025–1035. doi: 10.1016/j.cell.2013.04.040. - DOI - PMC - PubMed
    1. Casanova-Acebes M, Nicolás-Ávila JA, Li JL, García-Silva S, Balachander A, Rubio-Ponce A, Weiss LA, Adrover JM, Burrows K, A-González N, Ballesteros I, Devi S, Quintana JA, Crainiciuc G, Leiva M, Gunzer M, Weber C, Nagasawa T, Soehnlein O, Merad M, Mortha A, Ng LG, Peinado H, Hidalgo A. Neutrophils instruct homeostatic and pathological states in naive tissues. The Journal of Experimental Medicine. 2018;215:2778–2795. doi: 10.1084/jem.20181468. - DOI - PMC - PubMed
    1. Das M, Sabio G, Jiang F, Rincón M, Flavell RA, Davis RJ. Induction of hepatitis by JNK-mediated expression of TNF-alpha. Cell. 2009;136:249–260. doi: 10.1016/j.cell.2008.11.017. - DOI - PMC - PubMed

Publication types