Journal: PLOS Pathogens

Article Title: The impact of fatty acid synthase on HSV-1 infection dynamics

doi: 10.1371/journal.ppat.1013068

Figure Lengend Snippet: (A) Western blot analysis of protein lysates from mock or infected cells at the indicated hpi using antibodies against ICP27, gD, or actin. A representative blot and the densitometric analyses are shown. Values were normalized to actin and plotted as fold induction relative to mock-infected cells (set at 1). ( n = 2; unpaired t-test). (B) At 24 hpi, the number of intracellular HSV-1 genomes was measured by qPCR ( n = 3; unpaired t-test). (C) At 48 hpi, the number of genome-containing particles released by cells was measured by qPCR ( n = 3; unpaired t-test). (D) The supernatants from panel C were used to determine the number of infectious units/mL by the viral yield test ( n = 3; unpaired t-test). (E) The genome-to-infectious unit ratio has been determined. Data are expressed as fold induction relative to shCTRL cells (set at 1) ( n = 3; unpaired t-test). (F) Surface CD36 protein expression in shCTRL and shFASN cells was assessed by flow cytometry. Results show the mean fluorescence intensity (MFI) ( n = 3; unpaired t-test). (G) SSO toxicity was determined at 48 h post-treatment (hpt), using the MTT method ( n = 3). (H) Schematic representation of FA uptake using a fluorescent analog of palmitate (BODIPY FL C16) (Created in BioRender. De Andrea, M. (2025) https://BioRender.com/j23i568 ). (I) shCTRL and shFASN cells were infected with HSV-1 in serum-free conditions and at 48 hpi treated with DMSO or SSO (200 µM) for 1 h. Then, cells were cultured in the presence of 2 μM BODIPY FL C16 for 30 minutes and then assessed by flow cytometry. Results show the MFI ( n = 3; one-way ANOVA followed by Bonferroni’s post-tests). (J) shCTRL and shFASN cells were treated with DMSO or SSO in the presence of serum and then infected with HSV-1 (MOI 1). At 48 hpi, the supernatants were used to determine the number of infectious units/mL by the viral yield test ( n = 3; unpaired t-test). Data are shown as the mean ± SEM, * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: CMS121 (HY-135981, MedChemExpress), C75 (HY-12364, MedChemExpress), SSO (11211, Cayman Chemical), palmitate (P0500; Sigma-Aldrich), and BODIPY FL C16 (D3821; Invitrogen) were used at a concentration of 5 μM, 5 μM, 200 μM, 200 μM, and 2 μM, respectively. pLKO.1 puro-humanU6-shRNA FASN was a gift from Elizabeth Stoll (Addgene plasmid #82327; http://n2t.net/addgene:82327 ; RRID: Addgene_82327). pLKO.1 Puro shRNA Scramble was a gift from Antonia Follenzi (University of Piemonte Orientale, Novara, Italy). pAcUW51-CgE was a gift from Pamela Bjorkman (Addgene plasmid #13762; http://n2t.net/addgene:13762 ; RRID: Addgene_13762) [ ].

Techniques: Western Blot, Infection, Expressing, Flow Cytometry, Fluorescence, Cell Culture

Journal: Journal of Lipid Research

Article Title: Inflammatory stress promotes the development of obesity-related chronic kidney disease via CD36 in mice [S]

doi: 10.1194/jlr.M076216

Figure Lengend Snippet: Effects of inflammatory stress on cellular uptake and intracellular accumulation of FFA. HMCs and HK2 cells were treated with serum-free medium containing PA alone or with TNFα or IL-6 for 24 h. A: Then, cells were incubated with BODIPY-C16 (20 μM) followed by a confocal microscope analysis at the time indicated. One of four representative experiments is shown. B: Quantitative analysis of intracellular FFA and TG contents in HMCs and HK2 cells. Data are expressed as mean ± SEM from four independent experiments. *P < 0.05 versus the PA only group.

Article Snippet: Measurement of FFA uptake To assess the uptake and accumulation of FFA, HMCs and HK2 cells were washed with PBS and loaded with fluorescent probe, BODIPY FL C16 (D3821; Invitrogen, Eugene, OR).

Techniques: Incubation, Microscopy

Journal: Immunity

Article Title: The white adipose tissue is a reservoir for memory T cells that promotes protective memory responses to infection

doi: 10.1016/j.immuni.2017.11.009

Figure Lengend Snippet: (A–D) Mice were either naïve, 6 days post-infection with Yptb (Yptb WT 6 days), >4 weeks post-infection with Yptb ΔyopM (Yptb ΔyopM), or >4 weeks post-infection with Yptb ΔyopM followed by challenge with Yptb for 6 days (Yptb ΔyopM + Yptb WT) before imaging or isolation of cells for flow cytometry. (A) Representative images of the entire mAT from actin-DsRed reporter mice are shown. White arrows indicate FALCs. (B) Number of FALCs per mAT represented as fold change over the number of FALCs per naïve mAT. (C) mATs from CD11c–YFP reporter mice were stained for CD8 and LYVE-1 and imaged by confocal microscopy. CD8, CD11c, and LYVE-1 staining (top) or CD8+ T cells alone (bottom) in areas of the mAT with (left) or without (right) FALCs are shown. (D) Numbers of CD44+YopE:Kb+CD8+ T cells from mAT. (E) Pooled mAT, scAT and gAT isolated from either naïve donors or donors >4 weeks post-infection with Yptb ΔyopM were subcutaneously transplanted into Rag1−/− mice. Rag1−/− mice receiving ATs from previously infected mice were either left untreated or injected with anti-CD4 and anti-CD8 depleting antibodies. 2 weeks post surgery, animals were challenged i.v. with 200 CFU of Yptb WT. Data are representative of at least 2 experiments with ≥5 mice per group. Error bars in all bar graphs represent standard deviation. Statistical comparisons in (B) and (D) were performed using one way Anova adjusted for multiple comparisons. Statistical comparisons in (E) were performed using Log-rank (Mantel Cox) test. ns not significant, *p<0.05, **p<0.01, ****p<0.0001. See Figure S4.

Article Snippet: N/A Toxoplasma gondii (ME-49 C1) Laboratory of Michael Grigg N/A Biological Samples Chemicals, Peptides, and Recombinant Proteins Bodipy FL C16 Life Technologies Cat# D-3821 BrdU Thermo Fisher or BD Biosciences Cat# {"type":"entrez-nucleotide","attrs":{"text":"B23151","term_id":"2508782"}} B23151 or 550891 Collagenase IV Sigma Cat# C5138-1G DNase I Sigma Cat# DN25-5G Liberase TL Roche Cat# 5401020001 Tgd_057 59–66 Genscript N/A TGME49_012300 605–619 Genscript N/A YopE 69–77 peptide Genscript N/A Critical Commercial Assays Mouse adiponectin/Acrp30 ELISA duoset R&D systems Cat# DY1119 Arcturus PicoPure RNA Isolation kit Thermo Fisher Cat# KIT0204 FITC BrdU Flow kit BD Biosciences Cat# 557891 Fixation/Permeabilization eBioscience Cat# 00-5523-00 Free Glycerol Determination kit Sigma Cat# FG0100 Infinity Cholesterol Liquid Stable Reagent Thermo Scientific Cat# TR13421 iQ SYBR Green Supermix Life Technologies Cat# 4368702 Live/Dead fixable stain Life Technologies Cat# {"type":"entrez-nucleotide","attrs":{"text":"L23105","term_id":"1185069"}} L23105 miRNeasy kit Qiagen Cat# 217084 MitoTracker Deep Red Thermo Fisher Cat# {"type":"entrez-nucleotide","attrs":{"text":"M22426","term_id":"197107"}} M22426 Omniscript RT kit Qiagen Cat# 205111 RNeasy mini kit Qiagen Cat# 74106 Vybrant DyeCycle Violet eBioscience Cat# V35003 Deposited Data Raw RNA-Seq data This paper BioProject ID: PRJNA414132 Raw microarray data This paper GEO: {"type":"entrez-geo","attrs":{"text":"GSE104955","term_id":"104955"}} GSE104955 Experimental Models: Cell Lines Experimental Models: Organisms/Strains Mouse: C57BL/6 Taconic Farms Mouse strain: B6 Mouse: Tg(CAG-DsRed*MST)1Nagy/J (DsRed reporter) Jackson Laboratory Mouse strain: Jax005441 Mouse: B6.SJL-Ptprc a /BoyAiTac (CD45.1) Taconic Farms – NIAID exchange Mouse strain: Tac8478 Mouse: C57BL/6-[Tg]CD11c( Itgax ):EYFP (CD11c–eYFP reporter) Taconic Farms – NIAID exchange Mouse strain: Tac307 B6.SJL-Cd45a(Ly5a)Nai-[KO]RAG1 (Rag KO) Taconic Farms – NIAID exchange Mouse strain: Tac165 Non-human primate: Macaca mulatta Laboratory of J.M.B.

Techniques: Infection, Imaging, Isolation, Flow Cytometry, Staining, Confocal Microscopy, Injection, Standard Deviation

Journal: Immunity

Article Title: The white adipose tissue is a reservoir for memory T cells that promotes protective memory responses to infection

doi: 10.1016/j.immuni.2017.11.009

Figure Lengend Snippet: DATA AND SOFTWARE AVAILABILITY

Article Snippet: N/A Toxoplasma gondii (ME-49 C1) Laboratory of Michael Grigg N/A Biological Samples Chemicals, Peptides, and Recombinant Proteins Bodipy FL C16 Life Technologies Cat# D-3821 BrdU Thermo Fisher or BD Biosciences Cat# {"type":"entrez-nucleotide","attrs":{"text":"B23151","term_id":"2508782"}} B23151 or 550891 Collagenase IV Sigma Cat# C5138-1G DNase I Sigma Cat# DN25-5G Liberase TL Roche Cat# 5401020001 Tgd_057 59–66 Genscript N/A TGME49_012300 605–619 Genscript N/A YopE 69–77 peptide Genscript N/A Critical Commercial Assays Mouse adiponectin/Acrp30 ELISA duoset R&D systems Cat# DY1119 Arcturus PicoPure RNA Isolation kit Thermo Fisher Cat# KIT0204 FITC BrdU Flow kit BD Biosciences Cat# 557891 Fixation/Permeabilization eBioscience Cat# 00-5523-00 Free Glycerol Determination kit Sigma Cat# FG0100 Infinity Cholesterol Liquid Stable Reagent Thermo Scientific Cat# TR13421 iQ SYBR Green Supermix Life Technologies Cat# 4368702 Live/Dead fixable stain Life Technologies Cat# {"type":"entrez-nucleotide","attrs":{"text":"L23105","term_id":"1185069"}} L23105 miRNeasy kit Qiagen Cat# 217084 MitoTracker Deep Red Thermo Fisher Cat# {"type":"entrez-nucleotide","attrs":{"text":"M22426","term_id":"197107"}} M22426 Omniscript RT kit Qiagen Cat# 205111 RNeasy mini kit Qiagen Cat# 74106 Vybrant DyeCycle Violet eBioscience Cat# V35003 Deposited Data Raw RNA-Seq data This paper BioProject ID: PRJNA414132 Raw microarray data This paper GEO: {"type":"entrez-geo","attrs":{"text":"GSE104955","term_id":"104955"}} GSE104955 Experimental Models: Cell Lines Experimental Models: Organisms/Strains Mouse: C57BL/6 Taconic Farms Mouse strain: B6 Mouse: Tg(CAG-DsRed*MST)1Nagy/J (DsRed reporter) Jackson Laboratory Mouse strain: Jax005441 Mouse: B6.SJL-Ptprc a /BoyAiTac (CD45.1) Taconic Farms – NIAID exchange Mouse strain: Tac8478 Mouse: C57BL/6-[Tg]CD11c( Itgax ):EYFP (CD11c–eYFP reporter) Taconic Farms – NIAID exchange Mouse strain: Tac307 B6.SJL-Cd45a(Ly5a)Nai-[KO]RAG1 (Rag KO) Taconic Farms – NIAID exchange Mouse strain: Tac165 Non-human primate: Macaca mulatta Laboratory of J.M.B.

Techniques: Software, Recombinant, Enzyme-linked Immunosorbent Assay, Isolation, SYBR Green Assay, Staining, Microarray