Journal: bioRxiv

Article Title: IL-1-conferred gene expression pattern in ERα + BCa and AR + PCa cells is intrinsic to ERα − BCa and AR − PCa cells and promotes cell survival

doi: 10.1101/773978

Figure Lengend Snippet: RT-qPCR was performed for select genes for MCF7 and LNCaP cell lines treated with vehicle control or 25 ng/ml IL-1α or IL-1β for 5 days. MDA-MB-231, BT549, PC3 and DU145 cell lines were treated with vehicle control only. (A) RT-qPCR shows that CCL20 , CD68 , IL8 , p62 , SOX9 and Zeb1 are induced by IL-1 in MCF7 and/or LNCaP cell lines and are basally high in MDA-MB-231, BT549, PC3 and/or DU145 cell lines. (B) RT-qPCR shows that CXCR7 and MMP16 , but not CDK2 or PLK1 , are downregulated by IL-1 in MCF7 and/or LNCaP cell lines and are basally high in MDA-MB-231, BT549, PC3 and/or DU145 cell lines. N = 3 biological replicates; error bars, +/−STDEV; p-value, * ≤ 0.05, ** ≤ 0.005, ***≤ 0.0005. mRNA fold change is normalized to MCF7 vehicle control for the BCa cell lines and to LNCaP vehicle control for the PCa cell lines.

Article Snippet: Human recombinant IL-1α (GoldBio, St Louis MO; 1110-01A-100) and IL-1β (GoldBio, St Louis MO; 1110-01B-100) were resuspended in 0.1% bovine serum albumin (BSA, Thermo Fisher Scientific; BP 1600-1) in 1X phosphate buffered saline (PBS).

Techniques: Quantitative RT-PCR

Journal: Cell Death & Disease

Article Title: CHIP-mediated ubiquitin degradation of BCAT1 regulates glioma cell proliferation and temozolomide sensitivity

doi: 10.1038/s41419-024-06938-6

Figure Lengend Snippet: A Predicted ubiquitin ligase for BCAT1 on http://ubibrowser.bio-it.cn/ubibrowser/ . B , C U251 cells were transfected with HA-BCAT1 with or without Flag-CHIP plasmid. The interaction between BCAT1 and CHIP was detected by immunoprecipitation and western blot. D Schematic diagram of wile-type CHIP and its truncated mutants. E U251 cells were transfected with HA-BCAT1 plasmid with or without Flag-tagged wild-type CHIP or CHIP mutant plasmid, and immunoprecipitation and western blot was performed. F U251 cells were transfected with Flag-tagged CHIP plasmid and HA-tagged wild-type BCAT1 or BCAT1 mutant plasmid, and immunoprecipitation and western blot was performed. G Subcellular location of BCAT1 and CHIP was detected by immunofluorescence. Scale bar = 50 μm. H In vitro binding assay of GST-BCAT1 and CHIP proteins.

Article Snippet: For the GST-pull down assay, recombinant CHIP protein (#HY-P71340, MedChemExpress) was incubated with GSTSep Glutathione Magbeads (#20562ES03, Yeasen Biotechnology) and GST (#HY-P70270, MedChemExpress) or GST-BCAT1 (#Ag4574, Proteintech) protein at 4 °C for 2 h. Subsequently, the beads were washed thrice with PBS and boiled at 100 °C for 10 min with 1×loading buffer.

Techniques: Transfection, Plasmid Preparation, Immunoprecipitation, Western Blot, Mutagenesis, Immunofluorescence, In Vitro, Binding Assay

Journal: The Journal of Infectious Diseases

Article Title: Interleukin 1α (IL-1α) Promotes Pathogenic Immature Myeloid Cells and IL-1β Favors Protective Mature Myeloid Cells During Acute Lung Infection

doi: 10.1093/infdis/jiy049

Figure Lengend Snippet: Interleukin 1α (IL-1α) elicits an immature myeloid cell response, but IL-1β is required for mature myeloid cells. Wild-type (Wt) mice were treated with anti–IL-1α or anti–IL-1β and infected with Francisella tularensis. A, Tissue bacterial burden and total numbers of mature neutrophils (mPMNs), immature neutrophils (iPMNs), mature macrophages (mMΦ), and immature macrophages (iMΦ) in the lungs of IL-1α–neutralized mice at day 6. Data are median values from 2 independent experiments. *P < .05 and **P < .01, by the unpaired Student t test. B, Tissue bacterial burden and total numbers of mPMNs, iPMNs, mMΦ, and iMΦ in lungs of IL-1β–neutralized mice at day 5. Data are median values from 2 independent experiments. *P < .05 and **P < .01, by the unpaired Student t test. C, Levels of cytokines and chemokines measured by the Luminex assay in lung homogenates of IL-1α–neutralized mice at day 6. Data are median values from 2 independent experiments. *P < .05, by the nonparametric Mann-Whitney test. D, Levels of cytokines and chemokines measured by the Luminex assay in lung homogenates of IL-1β–neutralized mice at day 5. Data are median values from 2 independent experiments. *P < .05 and **P < .01, by the nonparametric Mann-Whitney test. Ab, antibody; G-CSF, granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor; IL-17, interleukin 17; IsoAb, isotype control antibody.

Article Snippet: Cytokine Neutralization F. tularensis –infected mice were injected intraperitoneally with 200 µg/mouse of anti–IL-1β (clone B122), anti–IL-1α (clone ALF-161), or isotype control antibody (BioXcell, West Lebanon, NH) at −1, 1, 3, 4, and 5 days after infection.

Techniques: Infection, Luminex, MANN-WHITNEY

Journal: The Journal of Infectious Diseases

Article Title: Interleukin 1α (IL-1α) Promotes Pathogenic Immature Myeloid Cells and IL-1β Favors Protective Mature Myeloid Cells During Acute Lung Infection

doi: 10.1093/infdis/jiy049

Figure Lengend Snippet: Interleukin 1β (IL-1β) favors myeloid cell maturation, differentiation, and phagocytosis. A and B, Ly6C+ cells from bone marrow (A) or blood (B) were or were not treated with recombinant IL-1α (rIL-1α) or rIL-1β. Left, Percentage of cells that phagocytosed the bioparticles. Data are median values with ranges, from 2 experiments. *P < .05, by the unpaired Student t test. Right, Intracellular bacterial burden. Data are median values from 2 experiments. *P < .05, by the nonparametric Mann-Whitney test. C and D, Ly6G+ cells from bone marrow (C) or blood (D) show the percentage of cells that phagocytosed the bioparticles. Data are median values with ranges, from 2 experiments. *P < .05, by the unpaired Student t test.

Article Snippet: Cytokine Neutralization F. tularensis –infected mice were injected intraperitoneally with 200 µg/mouse of anti–IL-1β (clone B122), anti–IL-1α (clone ALF-161), or isotype control antibody (BioXcell, West Lebanon, NH) at −1, 1, 3, 4, and 5 days after infection.

Techniques: Recombinant, MANN-WHITNEY

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: (A to D) Representative gross findings (A), histology (hematoxylin and eosin staining) (B), epidermal thickness (C), and numbers of total infiltrating cells and neutrophils in the dorsal skin (D) of wild-type (n =6 or 8) and Elovl6 -/- (n =6 or 8) mice before and on day 9 after the start of tape stripping. (E, F) Epidermal thickness (E) and numbers of total infiltrating cells and neutrophils in the skin (F) of Elovl6 fl/fl (n = 5) and Elovl6 fl/fl K14 -Cre (n = 4) on day 9 after the start of tape stripping. Black bars indicate scale (50 μm) (B). Error bars indicate 1 SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001. Data are representative of three (A-D) and two (E, F) independent experiments.

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Staining, Stripping Membranes

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: ( A ) The fetuses at E13.5 and E17.5 from WT and Elovl6 -/- mice were stained with 0.1% toluidine blue for 24 h and photographed. ( B ) The transepidermal water loss of 6-8 weeks old WT and Elovl6 -/- mice was measured before and after tape stripping (n = 14). Error bars indicate SD. NS, not significant. Data are representative of three independent experiments.

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Staining, Stripping Membranes

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: (A) Quantitative RT-PCR analysis of epidermis of wild-type and Elovl6 -/- mice isolated 6 h after tape stripping (n = 10 in each group). (B) Epidermis was isolated before, and 12 h after, tape stripping from wild-type and Elovl6 -/- mice and cultured for 24 h. The concentrations of IL-1β and CXCL-1 in the supernatants were measured by using cytometric bead array (n = 11 per group). (C) Quantitative RT-PCR analysis of ¡lift and Cxcl1 in the epidermis of wild-type, Myd88 -/- , and Ticami -/- mice before, and 6 h after, tape stripping (n = 7 to 11 per group). Error bars indicate 1 SD; *, P < 0.05; ***, P < 0.001. Data are representative of three independent experiments.

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Quantitative RT-PCR, Isolation, Stripping Membranes, Cell Culture

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: (A) Representative histopathology of wild-type and Elovl6 -/- mice 4 h after tape stripping. Black arrows indicate degenerated keartinocytes with irregularly shaped nuclei. Scale bar, 50 pm. (B, C) Flow cytometry of epidermal cells isolated from the skin of wild-type and Elovl6 -/- mice at the indicated time points after tape stripping. Cells were stained with anti-CD45.2, anti-CD49f and propidium iodide (PI) and the proportion of PI+ cells in CD45.2-CD49f+ cells were shown (n = 6 to 10 per group). Error bars indicate 1 SD; *, P < 0.05. Data are representative of two independent experiments.

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Histopathology, Stripping Membranes, Flow Cytometry, Isolation, Staining

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: (A) Live cell number of primary peritoneal macrophages 16 h after stimulation with oleic acid (OA) or CVA (300 µM) (n = 3 in each group). (B, E) Primary mouse keratinocytes were cultured for 6 h in the presence or absence of 10 μM of triacsin C (B), or 1 mM necrostatin (Nec-1), 1 mM necrosulfonamide (NSA), 2 mM IM-54, or 1 mM cyclosporine (CyA) (E), followed by stimulation by adding 300 μM CVA; live cells were counted 16 h afterward (n = 3). (C) A representative dead primary keratinocyte induced by stimulation with 300 μM CVA for 10 h under a transmission electron microscope. (D) Immunofluorescence microscopic study of primary keratinocyte 10 h after stimulation with CVA or 6 h after ultraviolet irradiation. Cells were stained with anti-cleaved caspase 9, followed by Alexa Fluor 594-conjugated secondary antibody and DAPI. White bars indicate a scale (20 μm). Percentage of cleaved caspase 9-positive cells was calculated (n = 3). Error bars indicate SD. NS, not significant. *, P < 0.05; **, P < 0.01; ***, P < 0.001. Data are representative of more than two independent experiments.

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Cell Culture, Transmission Assay, Microscopy, Immunofluorescence, Irradiation, Staining

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: (A) Enzyme-linked immunosorbent assay of HMGB-1 (n = 4 per group) and cytokine bead array of IL-1α (n = 3 per group) in the supernatant of cultured primary keratinocytes 10 h after initiation of stimulation with 300 pM OA or CVA. (B) Quantitative RT-PCR analysis of Il1β and Cxcl1 in the epidermis of wild-type mice 6 h after topical application of ethanol (control) (n = 10) or 15mM of OA (n = 13) or CVA (n = 14) (B). (C, D) Wild-type and Elovl6 -/- mice were treated with PBS (n = 10 and 12, respectively), an IL-1 receptor antagonist (n = 9 and 8, respectively), or a CXCR-2 antagonist (n = 5 and 4, respectively) daily for 9 days, from the beginning on the day of tape stripping. Epidermal thickness (C) and the number of infiltrating neutrophils (D) were analyzed on day 9. (E) A proposed signal pathway from mechanical damage onto the skin to skin inflammation. Error bars indicate SD; *, P < 0.05; **, P < 0.01, ***, P < 0.001; NS, not significant. Data are representative of at least two independent experiments.

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Enzyme-linked Immunosorbent Assay, Cell Culture, Quantitative RT-PCR, Stripping Membranes

Journal: bioRxiv

Article Title: A long-chain fatty acid elongase Elovl6 regulates mechanical damage–induced keratinocyte death and skin inflammation

doi: 10.1101/264838

Figure Lengend Snippet: Quantitative RT-PCR analysis of Il1β and Cxcl1 in keratinocytes of wild-type and Elovl6 -/- mice after stimulation or not with HMGB-1 or IL-1α in vitro (n=10 per group) (A) and in the epidermis isolated 4 h after injection intradermally with PBS, HMGB-1, or IL-1α (n = 8 per each group) (B).

Article Snippet: Primary mouse keratinocytes were stimulated with bovine HMGB-1 (Chondrex, Redmond, Washington, USA) or mouse IL-1α (Miltenyi Biotec, Bergisch Gladbach, Germany).

Techniques: Quantitative RT-PCR, In Vitro, Isolation, Injection