Journal: BMC Cancer

Article Title: Bladder cancer-derived interleukin-1 converts the vascular endothelium into a pro-inflammatory and pro-coagulatory surface

doi: 10.1186/s12885-020-07548-z

Figure Lengend Snippet: Pro-inflammatory activation of endothelial cells after stimulation with tumour cell SN. a After incubation with T24 cell SN, CXCL-1, IL-6, IL-8, GM-CSF and PAI-1 were significantly released from HUVECs (HUVECs + T24 cell SN). The SN of UROtsa was not able to stimulate HUVECs (HUVECs + UROtsa SN). HUVECs: baseline cytokine levels produced by HUVECs; T24 and UROTsa cell SN: baseline levels of cytokine produced by the tumour cells; ( n = 8–12) ** P ≤ 0.01. b Incubation of HUVECs for 6 h with T24 cell SN promoted the trans-localization of NF-kB (red) into the nucleus (blue) of the endothelial cells. Treatment of HUVECs with the SN of RT112 and UROtsa cells had no effect. RT4 cell SN induced a weak trans-localization of NF-kB. Addition of IL-1ra blocked NF-kB translocation upon treatment with T24 cell SN. The bar diagram shows the fluorescence intensity of nuclear NF-kB. Scale bar corresponds to 50 μm; ( n = 10 images) ** P ≤ 0.01. c IL-1ra inhibited the T24 cell SN induced release of the indicated pro-inflammatory cytokines from HUVECs. Cytokine levels were shown in pg/ml; ( n = 3) * P ≤ 0.05

Article Snippet: Slides were incubated overnight at 4 °C with a rabbit polyclonal anti-human IL-1ra antibody (Sigma-Aldrich, St. Louis, MO, USA) 1:200 in DAKO Real antibody diluent (DAKO, Glostrup, Denmark).

Techniques: Activation Assay, Incubation, Produced, Translocation Assay, Fluorescence

Journal: BMC Cancer

Article Title: Bladder cancer-derived interleukin-1 converts the vascular endothelium into a pro-inflammatory and pro-coagulatory surface

doi: 10.1186/s12885-020-07548-z

Figure Lengend Snippet: Impact of UBC cell SN on the integrity of the endothelial cell layer. a HUVECs grown on gelatine coated ECIS electrodes were treated with starvation medium (control) or UBC cell SN. The impedance was measured continuously up to 12 h after treatment. T24 cell SN led to marked breakdown of the endothelial impedance. RT4 cell SN had a lower impact whereas RT112 cell SN, UROtsa cell SN or starvation medium (control) did not cause significant alterations after 12 h; n = 5; ** P ≤ 0.01. b HUVEC cells were incubated for 12 h with T24 cell SN (T24) or starvation medium (control) and then analysed by immunofluorescence staining of β-Actin (red) and CD31/PECAM-1 (green). In comparison to the control (left), treatment with T24 cell SN (right) induced the disintegration of adherent junctions (CD31/PECAM-1) indicating a weakening of the endothelial barrier. c Inhibition of cytokine signalling partially conserved endothelial barrier function. Antibodies against IL-6, CXCL-1 slightly mitigated impedance breakdown. In contrast, IL-1ra strongly attenuated tumour mediated endothelial dysfunction, whereas IL-8 neutralization had no impact; n = 4; ** P ≤ 0.01

Article Snippet: Slides were incubated overnight at 4 °C with a rabbit polyclonal anti-human IL-1ra antibody (Sigma-Aldrich, St. Louis, MO, USA) 1:200 in DAKO Real antibody diluent (DAKO, Glostrup, Denmark).

Techniques: Incubation, Immunofluorescence, Staining, Inhibition, Neutralization

Journal: BMC Cancer

Article Title: Bladder cancer-derived interleukin-1 converts the vascular endothelium into a pro-inflammatory and pro-coagulatory surface

doi: 10.1186/s12885-020-07548-z

Figure Lengend Snippet: Impact of IL-1 on bladder cancer progression. a IL-1ra staining intensity in a cohort of 105 patients with UBC measured semiquantitatively by IHC. Controls (benign urothelium) showed a higher IL1-ra expression than in patients with cancer. No significant difference was found between low grade and high grade tumours. The staining intensity score was defined as follows: no staining = 1, low intensity = 2, moderate intensity = 3 and high intensity = 4. * P ≤ 0.05. n.s. = not significant. Representative IHC images are shown in b - c . Scale bar corresponds to 500 μm. b IHC very low staining intensity (score = 1). c Intermediate staining intensity (score = 3). d High staining intensity (score = 4). e High IL-1β mRNA levels were linked to adverse clinicopathological features high grade disease, muscle invasiveness and tumour progression. MIBC: muscle invasive bladder cancer, NMIBC: non-muscle invasive bladder cancer; * P ≤ 0.05

Article Snippet: Slides were incubated overnight at 4 °C with a rabbit polyclonal anti-human IL-1ra antibody (Sigma-Aldrich, St. Louis, MO, USA) 1:200 in DAKO Real antibody diluent (DAKO, Glostrup, Denmark).

Techniques: Staining, Expressing

Journal: Stem cells (Dayton, Ohio)

Article Title: Newly defined ABCB5 + dermal mesenchymal stem cells promote healing of chronic iron overload wounds via secretion of interleukin-1 receptor antagonist

doi: 10.1002/stem.3022

Figure Lengend Snippet: (A) ABCB5+-MSCs cultured alone (black bars) or with murine macrophages, secreted IL-1RA following IFN-γ/LPS stimulation, whereas ABCB5− HDFs did not. (B) Production of human IL-1RA+ (green) by β2M+ (red) human ABCB5+-derived MSCs was observed six hours after ABCB5+ MSCs injection around iron-overload wounds Scale bars = 50 μm. (C) A ~22 kDa band for lysates of three pooled ABCB5+-derived MSCs injected day three wounds, but not in ABCB5− HDF injected wounds, was detected by human IL-1RA specific immunoblotting. (D) IL-1RA silencing in ABCB5+-derived MSCs abrogated adaptive upregulation of IL-1RA (hatched bars) and significantly diminished TNFα sup-pression in activated murine (black bars) and human (white bars) macrophages in corresponding co-cultures. (E) Rescued wound healing and decreased wound areas following ABCB5+-derived MSC injection (blue squares) in iron overload mice was almost completely abrogated in IL-1RA siRNA (red diamonds) but not control siRNA transfected ABCB5+-derived MSCs (blue diamonds) from day five wounds onward. (F) Day five pro-inflammatory cytokine profiles of iron overload wounds remained unaltered when IL-1RA silenced ABCB5+-derived MSCs had been injected. (A, D, F) Bar graphs represent ELISA results. Significance level indications relate to control activated macrophages alone (A), respective siRNA-transfected groups (D) and PBS-treated group (F). * p < .05; ** p < .01; *** p < .001, t-test.

Article Snippet: A rabbit anti-IL-1RA IgG 1 antibody (Abcam #ab124962) which detects human and murine IL-1RA at a dilution of 1:1000 and a secondary HRP-coupled anti-rabbit IgG (H + L) antibody (Dianova) at a dilution of 1:10,000 was used.

Techniques: Cell Culture, Derivative Assay, Injection, Western Blot, Transfection, Enzyme-linked Immunosorbent Assay

Journal: Stem cells (Dayton, Ohio)

Article Title: Newly defined ABCB5 + dermal mesenchymal stem cells promote healing of chronic iron overload wounds via secretion of interleukin-1 receptor antagonist

doi: 10.1002/stem.3022

Figure Lengend Snippet: (A) Representative microphotographs of double immunostaining for macrophage F4/80 (green) and murine TNFα (red) in skin sections of day 5 wounds illustrate reduction of M1 macrophages following injection with ABCB5+-derived MSCs but not ABCB5− HDFs at the wound margins. This M1 macrophage suppressive effect was abrogated when IL-1RA silenced ABCB5+ MSCs were injected. (B) Double immunos-taining for F4/80 (green) and the M2 marker CD206 (red) displayed an IL-1RA-dependent increase in M2 macrophages in iron overload wounds, comparable to M2 macrophage numbers in physiological wound healing controls (Dextran/PBS) at day 5 post wounding. (A-B) Scale bars = 50 μm; inserts are three-fold enlarged. Nuclei are stained with DAPI (blue). (C) Multi-colour flow cytometry on singlet F4/80+ wound single cell preparations at day 5 post wounding confirm a wound macrophage immune-phenotype shift with downregulation of M1 markers like TNFα, IL-12/IL-23p40 and NOS2 and upregulation of the M2 markers CD206, Dectin-1 and ARG1 IL-1RA-dependently mediated by ABCB5+ MSCs but not by donor-matched ABCB5− HDFs. ns = not significant; * p < .05; ** p < .01; *** p < .001, t-test.

Article Snippet: A rabbit anti-IL-1RA IgG 1 antibody (Abcam #ab124962) which detects human and murine IL-1RA at a dilution of 1:1000 and a secondary HRP-coupled anti-rabbit IgG (H + L) antibody (Dianova) at a dilution of 1:10,000 was used.

Techniques: Double Immunostaining, Injection, Derivative Assay, Marker, Staining, Flow Cytometry

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Quantitative RT-PCR Primers.

Article Snippet: IL-1ra/CD11b 1 , Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL , , Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL.

Techniques: Quantitative RT-PCR

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Immunohistochemistry Antibodies

Article Snippet: IL-1ra/CD11b 1 , Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL , , Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL.

Techniques: Immunohistochemistry, Marker

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Chronic morphine treatment (25 mg s.c., ×2; 6 d) and LP-BM5 infection (8 wks) altered expression of pro-inflammatory (iNOS, IL-1β) and anti-inflammatory (Arg1, IL-1ra) enzymes and cytokines. Pro-/anti-inflammatory markers were analyzed by qRT-PCR (n=11-12) and normalized to GAPDH. Values represent percent change in relative expression from non-infected, placebo-treated mice. A-B, iNOS expression declined in all three areas with morphine treatment and LP-BM5 infection, while Arg1 was unaffected by either treatment. C-D, IL-1β and IL-1ra expression was reduced by LP-BM5 infection and chronic morphine treatment in hippocampus and frontal lobe, but not striatum. Legend: NI, non-infected; BM5, LP-BM5 infected; Plac, placebo; Mor, morphine. Symbols indicate significance by one-way ANOVA with a Fisher's PLSD post hoc test: *p ≤ 0.05 versus NI Plac; #p ≤ 0.05 versus NI Mor; ‡p ≤ 0.05 versus BM5 Plac

Article Snippet: IL-1ra/CD11b 1 , Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL , , Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL.

Techniques: Infection, Expressing, Quantitative RT-PCR

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Chronic morphine treatment (6 d) altered colocalization of pro-inflammatory (iNOS, IL-1β) and anti-inflammatory (Arg1, IL-1ra) markers with microglia (CD11b) and astrocytes (GFAP) in a cell- and region-specific manner. IHC colocalization of glial markers with pro-/anti-inflammatory cytokines and enzymes was assessed in the hippocampus, striatum, and frontal lobe (n=4). A-P, Representative confocal images (65×, scale = 50 μM) of glia positive for inflammatory markers (arrows). Q, Diagram of quantification workflow. Confocal z-stacks (15 slices, 5 μM depth) were obtained at 40× on a Leica SP5 system. Cells were counted in ImageJ on a maximum projection, with positive cell criterion being: (i) an adjacent nucleus (DAPI, blue) and (ii) clear colocalization of glial markers (red) with pro-/anti-inflammatory markers (green). Colocalization was confirmed on a single z-slice. AB-OP, Percentage of inflammatory marker-positive microglia (AB, CD, IJ, KL) and astrocytes (EF, GH, MN, OP) in hippocampus (left), striatum (center), and frontal lobe (right). Legend: NI, non-infected; BM5, LP-BM5 infected; Plac, placebo; Mor, morphine. Symbols indicate significance by one-way ANOVA with a Fisher's PLSD post hoc test: *p ≤ 0.05 versus NI Plac; #p ≤ 0.05 versus NI Mor; ‡p ≤ 0.05 versus BM5 Plac.

Article Snippet: IL-1ra/CD11b 1 , Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL , , Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL.

Techniques: Marker, Infection

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Correlation to Regional Viral Load

Article Snippet: IL-1ra/CD11b 1 , Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL , , Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL.

Techniques: RNA Expression

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Quantitative RT-PCR Primers.

Article Snippet: For Arg1-GFAP probed sections, slides were fixed for 10 minutes in 100% ethanol instead of 4% PFA. table ft1 table-wrap mode="anchored" t5 caption a7 Target Pair Cytokine Primary Cytokine Secondary Glial Marker Primary Glial Marker Secondary iNOS/CD11b 1 , 3 Rabbit anti-iNOS (Abcam #ab15323) Dilution: 0.2 μg/mL Donkey anti-Rabbit AlexaFluor 488 (Jackson #711-545-152) 3 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Donkey anti-Rat Cy3 (Jackson #712-165-153) 3 μg/mL iNOS/GFAP 2 , 3 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Arg1/CD11b 1 Goat anti-Arginase-1 (Novus #NB100-59740) 2 μg/mL Donkey anti-Goat AlexaFluor 488 (Jackson #705-545-147) 2.8 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Arg1/GFAP 2 , † Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1β/CD11b 1 Rabbit anti-IL-1β (Abcam #ab9722) 0.5 μg/mL Goat anti-Rabbit AlexaFluor 488 (Life Technologies # {"type":"entrez-nucleotide","attrs":{"text":"A11034","term_id":"489250","term_text":"A11034"}} A11034 ) 4 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Goat anti-Rat Dylight 549 (Jackson #112-505-143) 3 μg/mL IL-1β/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1ra/CD11b 1 Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL IL-1ra/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Open in a separate window 1 Slides were fixed for 10 min with 100% EtOH for optimal CD11b staining.

Techniques: Quantitative RT-PCR

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Immunohistochemistry Antibodies

Article Snippet: For Arg1-GFAP probed sections, slides were fixed for 10 minutes in 100% ethanol instead of 4% PFA. table ft1 table-wrap mode="anchored" t5 caption a7 Target Pair Cytokine Primary Cytokine Secondary Glial Marker Primary Glial Marker Secondary iNOS/CD11b 1 , 3 Rabbit anti-iNOS (Abcam #ab15323) Dilution: 0.2 μg/mL Donkey anti-Rabbit AlexaFluor 488 (Jackson #711-545-152) 3 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Donkey anti-Rat Cy3 (Jackson #712-165-153) 3 μg/mL iNOS/GFAP 2 , 3 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Arg1/CD11b 1 Goat anti-Arginase-1 (Novus #NB100-59740) 2 μg/mL Donkey anti-Goat AlexaFluor 488 (Jackson #705-545-147) 2.8 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Arg1/GFAP 2 , † Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1β/CD11b 1 Rabbit anti-IL-1β (Abcam #ab9722) 0.5 μg/mL Goat anti-Rabbit AlexaFluor 488 (Life Technologies # {"type":"entrez-nucleotide","attrs":{"text":"A11034","term_id":"489250","term_text":"A11034"}} A11034 ) 4 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Goat anti-Rat Dylight 549 (Jackson #112-505-143) 3 μg/mL IL-1β/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1ra/CD11b 1 Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL IL-1ra/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Open in a separate window 1 Slides were fixed for 10 min with 100% EtOH for optimal CD11b staining.

Techniques: Immunohistochemistry, Marker

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Chronic morphine treatment (25 mg s.c., ×2; 6 d) and LP-BM5 infection (8 wks) altered expression of pro-inflammatory (iNOS, IL-1β) and anti-inflammatory (Arg1, IL-1ra) enzymes and cytokines. Pro-/anti-inflammatory markers were analyzed by qRT-PCR (n=11-12) and normalized to GAPDH. Values represent percent change in relative expression from non-infected, placebo-treated mice. A-B, iNOS expression declined in all three areas with morphine treatment and LP-BM5 infection, while Arg1 was unaffected by either treatment. C-D, IL-1β and IL-1ra expression was reduced by LP-BM5 infection and chronic morphine treatment in hippocampus and frontal lobe, but not striatum. Legend: NI, non-infected; BM5, LP-BM5 infected; Plac, placebo; Mor, morphine. Symbols indicate significance by one-way ANOVA with a Fisher's PLSD post hoc test: *p ≤ 0.05 versus NI Plac; #p ≤ 0.05 versus NI Mor; ‡p ≤ 0.05 versus BM5 Plac

Article Snippet: For Arg1-GFAP probed sections, slides were fixed for 10 minutes in 100% ethanol instead of 4% PFA. table ft1 table-wrap mode="anchored" t5 caption a7 Target Pair Cytokine Primary Cytokine Secondary Glial Marker Primary Glial Marker Secondary iNOS/CD11b 1 , 3 Rabbit anti-iNOS (Abcam #ab15323) Dilution: 0.2 μg/mL Donkey anti-Rabbit AlexaFluor 488 (Jackson #711-545-152) 3 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Donkey anti-Rat Cy3 (Jackson #712-165-153) 3 μg/mL iNOS/GFAP 2 , 3 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Arg1/CD11b 1 Goat anti-Arginase-1 (Novus #NB100-59740) 2 μg/mL Donkey anti-Goat AlexaFluor 488 (Jackson #705-545-147) 2.8 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Arg1/GFAP 2 , † Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1β/CD11b 1 Rabbit anti-IL-1β (Abcam #ab9722) 0.5 μg/mL Goat anti-Rabbit AlexaFluor 488 (Life Technologies # {"type":"entrez-nucleotide","attrs":{"text":"A11034","term_id":"489250","term_text":"A11034"}} A11034 ) 4 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Goat anti-Rat Dylight 549 (Jackson #112-505-143) 3 μg/mL IL-1β/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1ra/CD11b 1 Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL IL-1ra/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Open in a separate window 1 Slides were fixed for 10 min with 100% EtOH for optimal CD11b staining.

Techniques: Infection, Expressing, Quantitative RT-PCR

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Chronic morphine treatment (6 d) altered colocalization of pro-inflammatory (iNOS, IL-1β) and anti-inflammatory (Arg1, IL-1ra) markers with microglia (CD11b) and astrocytes (GFAP) in a cell- and region-specific manner. IHC colocalization of glial markers with pro-/anti-inflammatory cytokines and enzymes was assessed in the hippocampus, striatum, and frontal lobe (n=4). A-P, Representative confocal images (65×, scale = 50 μM) of glia positive for inflammatory markers (arrows). Q, Diagram of quantification workflow. Confocal z-stacks (15 slices, 5 μM depth) were obtained at 40× on a Leica SP5 system. Cells were counted in ImageJ on a maximum projection, with positive cell criterion being: (i) an adjacent nucleus (DAPI, blue) and (ii) clear colocalization of glial markers (red) with pro-/anti-inflammatory markers (green). Colocalization was confirmed on a single z-slice. AB-OP, Percentage of inflammatory marker-positive microglia (AB, CD, IJ, KL) and astrocytes (EF, GH, MN, OP) in hippocampus (left), striatum (center), and frontal lobe (right). Legend: NI, non-infected; BM5, LP-BM5 infected; Plac, placebo; Mor, morphine. Symbols indicate significance by one-way ANOVA with a Fisher's PLSD post hoc test: *p ≤ 0.05 versus NI Plac; #p ≤ 0.05 versus NI Mor; ‡p ≤ 0.05 versus BM5 Plac.

Article Snippet: For Arg1-GFAP probed sections, slides were fixed for 10 minutes in 100% ethanol instead of 4% PFA. table ft1 table-wrap mode="anchored" t5 caption a7 Target Pair Cytokine Primary Cytokine Secondary Glial Marker Primary Glial Marker Secondary iNOS/CD11b 1 , 3 Rabbit anti-iNOS (Abcam #ab15323) Dilution: 0.2 μg/mL Donkey anti-Rabbit AlexaFluor 488 (Jackson #711-545-152) 3 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Donkey anti-Rat Cy3 (Jackson #712-165-153) 3 μg/mL iNOS/GFAP 2 , 3 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Arg1/CD11b 1 Goat anti-Arginase-1 (Novus #NB100-59740) 2 μg/mL Donkey anti-Goat AlexaFluor 488 (Jackson #705-545-147) 2.8 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Arg1/GFAP 2 , † Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1β/CD11b 1 Rabbit anti-IL-1β (Abcam #ab9722) 0.5 μg/mL Goat anti-Rabbit AlexaFluor 488 (Life Technologies # {"type":"entrez-nucleotide","attrs":{"text":"A11034","term_id":"489250","term_text":"A11034"}} A11034 ) 4 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Goat anti-Rat Dylight 549 (Jackson #112-505-143) 3 μg/mL IL-1β/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1ra/CD11b 1 Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL IL-1ra/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Open in a separate window 1 Slides were fixed for 10 min with 100% EtOH for optimal CD11b staining.

Techniques: Marker, Infection

Journal: Journal of neuroimmunology

Article Title: Morphine-potentiated cognitive deficits correlate to suppressed hippocampal iNOS RNA expression and an absent type 1 interferon response in LP-BM5 murine AIDS

doi: 10.1016/j.jneuroim.2018.02.017

Figure Lengend Snippet: Correlation to Regional Viral Load

Article Snippet: For Arg1-GFAP probed sections, slides were fixed for 10 minutes in 100% ethanol instead of 4% PFA. table ft1 table-wrap mode="anchored" t5 caption a7 Target Pair Cytokine Primary Cytokine Secondary Glial Marker Primary Glial Marker Secondary iNOS/CD11b 1 , 3 Rabbit anti-iNOS (Abcam #ab15323) Dilution: 0.2 μg/mL Donkey anti-Rabbit AlexaFluor 488 (Jackson #711-545-152) 3 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Donkey anti-Rat Cy3 (Jackson #712-165-153) 3 μg/mL iNOS/GFAP 2 , 3 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Arg1/CD11b 1 Goat anti-Arginase-1 (Novus #NB100-59740) 2 μg/mL Donkey anti-Goat AlexaFluor 488 (Jackson #705-545-147) 2.8 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Arg1/GFAP 2 , † Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1β/CD11b 1 Rabbit anti-IL-1β (Abcam #ab9722) 0.5 μg/mL Goat anti-Rabbit AlexaFluor 488 (Life Technologies # {"type":"entrez-nucleotide","attrs":{"text":"A11034","term_id":"489250","term_text":"A11034"}} A11034 ) 4 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL Goat anti-Rat Dylight 549 (Jackson #112-505-143) 3 μg/mL IL-1β/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL IL-1ra/CD11b 1 Rabbit anti-IL-1ra (Abcam #ab124962, clone EPR6483) 0.1 μg/mL Rat anti-CD11b (eBioscience #14-0112-85, clone M1/70) 5 μg/mL IL-1ra/GFAP 2 Rat anti-GFAP (Life Technologies #130300, clone 2.2B10) 5 μg/mL Open in a separate window 1 Slides were fixed for 10 min with 100% EtOH for optimal CD11b staining.

Techniques: RNA Expression