Journal: Frontiers in Cell and Developmental Biology

Article Title: lncRNA Gm16410 Mediates PM 2 . 5 -Induced Macrophage Activation via PI3K/AKT Pathway

doi: 10.3389/fcell.2021.618045

Figure Lengend Snippet: Macrophage activation caused inflammation of lung tissue in response to PM 2 . 5 . (A) Immunohistochemistry detection of the macrophage marker F4/80 in mouse lung tissues. Scale bar = 10 μm. (B) Pathological changes in the lung tissues. Scale bar = 10 μm. (C , D) Western blot analysis of TNF-α and IL-1β levels in the lung tissues. The plot gives a quantitative gray-scale analysis of the blot ( n = 3). (E) ELISA analysis of IL-6 in serum ( n = 5). (F) Real-time quantitative PCR analysis of IL-6, TNF-α, and IL-1β in the lung ( n = 8). (G , H) Western blot analysis of the P-PI3K, P-AKT, and NF-κB in mouse lung tissues. The plot gives a quantitative gray-scale analysis of the blot ( n = 3). (I) Immunohistochemistry detection of P-AKT and NF-κB in mouse lung tissues. Scale bar = 10 μm. Data are the means ± SEMs from at least three independent experiments, each performed in triplicate. * P < 0.5 and ** P < 0.01—levels of significance that are different from the control group at the levels, respectively.

Article Snippet: Mouse IL-6 ELISA kit was purchased from Boster (Wuhan, China).

Techniques: Activation Assay, Immunohistochemistry, Marker, Western Blot, Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction, Control

Journal: Molecular Therapy Oncolytics

Article Title: Therapeutic effects of mesenchymal stem cells loaded with oncolytic adenovirus carrying decorin on a breast cancer lung metastatic mouse model

doi: 10.1016/j.omto.2022.01.007

Figure Lengend Snippet: MSCs.Null and rAd.Null enhanced antitumor responses in peripheral blood (A) The CD4 + CD44 High CD62L + memory T cells were analyzed by flow cytometry on days 14 and 19. (B) The statistical results of flow cytometry detection. (C) The protein concentrations of mouse interleukin-6 (IL-6), TNF-α, and IFN-γ were measured using the CBA Mouse Flex Set for IL-6, IFN-γ, and TNF-α (BD Biosciences) and analyzed by flow cytometry on days 10 and 17. Data are the mean ± SEM. ∗p < 0.05 and ∗∗p < 0.01 versus the corresponding group.

Article Snippet: The concentrations of IL-6, IFN-γ, and TNF-α were measured using a CBA Mouse Flex Set for IL-6, IFN-γ, and TNF-α (BD Biosciences).

Techniques: Flow Cytometry

Journal: Nature microbiology

Article Title: Listeria monocytogenes upregulates mitochondrial calcium signaling to inhibit LC3-associated phagocytosis as a survival strategy

doi: 10.1038/s41564-020-00843-2

Figure Lengend Snippet: (a) Gentamicin protection assay in Mcufl/fl and McuΔmye BMMs infected with L. monocytogenes (MOI, 10) for 1 h. At five minutes after changing to gentamicin-containing medium, the incubation time was considered as 0 h. (b) Flow cytometry analysis of GFP fluorescence signal in Mcufl/fl and McuΔmye BMMs infected with GFP-L. monocytogenes (MOI, 10) for indicated periods. (c–f) Immunoblotting assay (c and d) and microscopy analysis (e and f) of GFP fluorescence signal to measure bacteria killing ability in Mcufl/fl and McuΔmye BMMs. Scale bar, 5 μm. (g and h) Transmission electron microscopy (TEM) images of Mcufl/fl and McuΔmye BMMs challenged with L. monocytogenes with indicated periods (g). Quantification of intracellular L. monocytogenes was performed by counting bacteria in 50 cells (h). Scale bar, 2 μm. (i-k) Gentamicin protection assay in Mcufl/fl and McuΔmye peritoneal macrophages (i), or WT and MCU-KO THP-1 cells (k) infected with L. monocytogenes, or Mcufl/fl and McuΔmye BMMs challenged with F. novicida. (l and m) Survival (l) and body weight change (m) of Mcufl/fl and McuΔmye (for both groups, n = 20, 10 male and 10 female) mice after intraperitoneal injection with L. monocytogenes (0.1×106 c.f.u for female mice, 0.5×106 c.f.u for male mice). (n–p) Bacterial loads in the spleen (n) or liver (o) and IL-6 and TNF-α in serum (p) from Mcufl/fl and McuΔmye mice challenged with L. monocytogenes for 24 h (n=7 female per group). *P < 0.05, ** P < 0.01, ****P < 0.001 versus controls (two-tailed Student’s t-test (a, b, d, f, h-o). Data are from one experiment representative of five experiments (a, b, i and j; mean ± SEM of six biological replicates). The results presented are representative of three independent experiments (c). The image presented are representative of three independent experiments (e and g). Data are shown as mean ± SEM of all analyzed cells from three independent experiments (f and h).

Article Snippet: ELISA Cytokines in supernatant from in vitro cultured cells or cytokines in the peritoneal lavage fluids, serum or lung homogenates from animal experiments were quantified using the ELISA set for mouse IL-6 and TNF-α (BD Biosciences) according to the manufacturer’s protocol.

Techniques: Infection, Incubation, Flow Cytometry, Fluorescence, Western Blot, Microscopy, Transmission Assay, Electron Microscopy, Injection, Two Tailed Test

Journal: Nature microbiology

Article Title: Listeria monocytogenes upregulates mitochondrial calcium signaling to inhibit LC3-associated phagocytosis as a survival strategy

doi: 10.1038/s41564-020-00843-2

Figure Lengend Snippet: (a) Colony-forming units in BMMs generated from Mcufl/fl and McuΔmye mice challenged with L. monocytogenes (MOI, 10) for 1 hour followed by gentamicin treatment for indicated time before cell lysis. Intracellular bacteria were plated on brain-heart-infusion plates. (b) GFP-L. monocytogenes containing cells in BMMs generated from Mcufl/fl and McuΔmye mice challenged with L. monocytogenes (MOI, 10) for indicated periods were measured by FACS analysis. (c-f) BMMs (c-d) or peritoneal macrophages (e-f) from Mcufl/fl and McuΔmye mice were left untreated or challenged with L. monocytogenes (MOI, 10) for indicated periods. Gene transcripts of Il6 and Tnfa in the cells (c and e), IL-6 and TNF-α proteins in the supernatants (d and f) were measured with RT-PCR and ELISA, respectively. (g) Gene transcripts of IL6 and TNFA in THP-1 cells left untreated or stimulated with L. monocytogenes (MOI, 10) for indicated periods were measured with RT-PCR. (h-i) NF-κB signaling molecules including IKKα, p65 and IκBα, and MAPK signaling molecules including ERK1/2, JNK1/2 and p38, in Mcufl/fl and McuΔmye BMMs (h) or peritoneal macrophages (i) left untreated or stimulated with L. monocytogenes (MOI, 10) for indicated periods were analyzed by immunoblotting. (j) Immunoblotting of NF-κB signaling molecules and MAPK signaling molecules in THP-1 cells left untreated or stimulated with L. monocytogenes (MOI, 10) for indicated periods. The averages of n = 3 biologically independent samples are shown. The error bars represent the SEM. Statistical significance was determined using t test (and nonparametric tests).

Article Snippet: ELISA Cytokines in supernatant from in vitro cultured cells or cytokines in the peritoneal lavage fluids, serum or lung homogenates from animal experiments were quantified using the ELISA set for mouse IL-6 and TNF-α (BD Biosciences) according to the manufacturer’s protocol.

Techniques: Generated, Lysis, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Western Blot