Journal: PLoS Pathogens

Article Title: Mast Cells Expedite Control of Pulmonary Murine Cytomegalovirus Infection by Enhancing the Recruitment of Protective CD8 T Cells to the Lungs

doi: 10.1371/journal.ppat.1004100

Figure Lengend Snippet: (A) Infected cells in alveolar epithelium and lung-infiltrating T cells co-localize in NIF. Lung tissue sections were taken on day 6 after intravenous infection of WT C57BL/6 mice. 2-color immunohistological staining identifies infected cells by black staining of viral intranuclear IE1 protein and T cells by red staining of CD3ε. (a) pulmonary alveoli; (b) pulmonary bronchioles lined with bronchiolar epithelium; (s) alveolar septum. The bar marker represents 10 µm. (B) Cytofluorometric phenotyping of pulmonary leucocyte subpopulations revealing a high proportion of CD3 + CD8 + T cells and virtual absence of CD335 + NK cells. The analysis was performed for a pool of 6 mice on day 6, corresponding to the immunohistological localization study. Shown are color-coded density plots with red and blue representing highest and lowest density, respectively. Percentages of main interest are indicated for gated areas and quadrants. (SSC) side scatter in the live gate; (FL) fluorescence channel and fluorescence intensity. (C) Infection of lungs and spleen is controlled by CD8 T cells. C57BL/6 mice were depleted of CD8 T cells on the day before infection. Virus titers in homogenates of the indicated organs were determined on day 6 post-infection. Dots represent virus titers in individual mice revealing the sample sizes and ranges, with the median values indicated. The dotted lines indicate the detection limits.

Article Snippet: In a second round, CD3 + T cells were labeled with a rat monoclonal antibody directed against CD3ε (clone CD3-12; catalog. no. SM1754P, acris antibodies, Hiddenhausen, Germany), followed by the biotin-conjugated antibody and the peroxidase-coupled avidin biotin complex as above.

Techniques: Infection, Staining, Marker, Fluorescence, Virus

Journal: PLoS Pathogens

Article Title: Mast Cells Expedite Control of Pulmonary Murine Cytomegalovirus Infection by Enhancing the Recruitment of Protective CD8 T Cells to the Lungs

doi: 10.1371/journal.ppat.1004100

Figure Lengend Snippet: (A) Localization of CD3 + T cells in intravascular (in) and extravascular (ex) compartments of the lungs. Shown are 2-color immunohistological images of lung tissue sections taken on day 6 after infection of C57BL/6 mice, with endothelial cells being identified by black staining of CD31 and T cells by turquoise-green staining of CD3ε. (v) small vessel; (a) pulmonary alveoli; (b) pulmonary bronchiole lined with bronchiolar epithelium; (s) alveolar septum; (c) capillary in a thickened alveolar septum. The bar marker represents 25 µm. (B) Frequencies of viral epitope-specific, IFNγ-secreting CD8 T cells among CD45 + leucocytes in different compartments. CD8 T cells recovered by BAL are those that are only loosely attached to the epithelial lining of the alveoli or bronchioles and are thus definitely located outside of the lung vasculature. Shown are data from ELISpot assays using stimulator cells exogenously-loaded with saturating concentrations of antigenic peptides representing the epitopes indicated. Bars represent most probable numbers determined by intercept-free linear regression analysis and error bars represent the 95% confidence intervals.

Article Snippet: In a second round, CD3 + T cells were labeled with a rat monoclonal antibody directed against CD3ε (clone CD3-12; catalog. no. SM1754P, acris antibodies, Hiddenhausen, Germany), followed by the biotin-conjugated antibody and the peroxidase-coupled avidin biotin complex as above.

Techniques: Infection, Staining, Marker, Enzyme-linked Immunospot

Journal: PLoS Pathogens

Article Title: Mast Cells Expedite Control of Pulmonary Murine Cytomegalovirus Infection by Enhancing the Recruitment of Protective CD8 T Cells to the Lungs

doi: 10.1371/journal.ppat.1004100

Figure Lengend Snippet: Pulmonary infiltrates were analyzed 6 days after intravenous infection of MC-sufficient WT C57BL/6 and MC-deficient Kit W-sh mice. (A) Representative immunohistological images show pulmonary T cell (with a negligible contribution of NKT cells, see ) infiltrates in situ by black staining of CD3ε, comparing lung tissue sections from WT C57BL/6 (left panel) and Kit W-sh (right panel) mice. Bar markers represent 50 µm. (B) Comparative analysis of absolute quantities of CD3ε + cells per lung, determined by counting of stained cells in tissue sections extrapolated to the whole organ. Symbols (closed circles: WT C57BL/6; open circles: Kit W-sh mutant) represent data from individual mice revealing sample sizes and ranges, with the median values indicated.

Article Snippet: In a second round, CD3 + T cells were labeled with a rat monoclonal antibody directed against CD3ε (clone CD3-12; catalog. no. SM1754P, acris antibodies, Hiddenhausen, Germany), followed by the biotin-conjugated antibody and the peroxidase-coupled avidin biotin complex as above.

Techniques: Infection, In Situ, Staining, Mutagenesis

Journal: PLoS Pathogens

Article Title: Mast Cells Expedite Control of Pulmonary Murine Cytomegalovirus Infection by Enhancing the Recruitment of Protective CD8 T Cells to the Lungs

doi: 10.1371/journal.ppat.1004100

Figure Lengend Snippet: (A) CD8 T-cell recruitment by infection is curtailed in MC-deficient mice and reversed by MC reconstitution. Shown are results from cytofluorometric analyses (recall for the gating strategy), compiled from two independent experiments, revealing the influence of MC on the proportions of CD8 T cells relative to all leucocytes in lymphoid (spleen) and non-lymphoid (lungs) compartments as well as in the circulation (peripheral blood) on day 6 after intravenous infection. Comparisons were performed in absence or presence of mCMV infection in MC-sufficient WT C57BL/6 mice, MC-deficient Kit W-sh mice, and (BM)MC-reconstituted Kit W-sh mice. Symbols represent data from individual mice revealing sample sizes and ranges, with the median values indicated. P values are given for comparisons of interest, unless the difference is obvious. (B) CD8 T-cell recruitment to the lungs suffers preferentially from MC deficiency. Shown are results from an independent experiment focusing on differences between the three compartments regarding the proportions of CD3 + T cells among CD45 + leucocytes, of CD8 + cells among all T cells, and the CD8/CD4 ratios. Note that it is important to calculate CD8/CD4 ratios for each individual separately, as these values are linked in that the individual with the highest percentage of CD8 T cells is necessarily also the one with the lowest percentage of CD4 T cells, and inverse.

Article Snippet: In a second round, CD3 + T cells were labeled with a rat monoclonal antibody directed against CD3ε (clone CD3-12; catalog. no. SM1754P, acris antibodies, Hiddenhausen, Germany), followed by the biotin-conjugated antibody and the peroxidase-coupled avidin biotin complex as above.

Techniques: Infection

Journal: Biomaterials

Article Title: In vitro and in vivo evaluation of biohybrid tissue-engineered vascular grafts with transformative 1 H/ 19 F MRI traceable scaffolds.

doi: 10.1016/j.biomaterials.2024.122669

Figure Lengend Snippet: Fig. 6. Ex vivo histological analyses evaluating the effects of 19F-TPU on ECM deposition and host reaction. A-B) Unlabeled TEVGs lacking both MR contrast agents used throughout this study (A) and TEVGs containing SPION-PLGA (TEVG_SPION, B) are used as negative controls. C) Vascular prostheses labeled with only the novel 19F-TPU fibers (TEVG_TPU) were used as a supplementary negative control to exclude any additive, synergistic, or antagonistic effect between SPIONs and 19F-TPU. D) Representative images of complete TEVGs labeled with both MR contrast agents, i.e., SPION-PLGA and 19F-TPU fibers, (TEVG_SPION-TPU). H&E staining was performed to get an overview of ECM (in pink) and cell nuclei (in purple), highlighting the presence and infiltration of immune cells. The Masson’s trichrome staining complementary shows total collagen in light blue, while nuclei are stained in purple. Fluorescence images in the bottom rows show collagen III and IV, F4/80, and CD3 in red, respectively, αSMA in green, and nuclei (DAPI) in blue. For all stainings no significant differences are observed, indicating that SPIONs incorporation into PLGA and 19F-TPU does not change the body reactions after TEVGs implantation. Scale bar: 50 μm. The stars indicate TEVGs’ lumen. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The primary antibodies were incubated at room temperature for 1 h (anti-rat and human α-smooth muscle actin (αSMA), anti-rat and human elastin, anti-rat collagen I, anti-rat and human collagen III, antirat and human collagen IV, anti-rat F4/80, and anti-rat CD3; Acris Antibodies GmbH, Herford, Germany; Abcam, Berlin, Germany; Thermofisher Scientific, Massachusetts, USA).

Techniques: Ex Vivo, Labeling, Negative Control, Staining, Fluorescence