Journal: Frontiers in Immunology

Article Title: Insight into immune profile associated with vitiligo onset and anti-tumoral response in melanoma patients receiving anti-PD-1 immunotherapy

doi: 10.3389/fimmu.2023.1197630

Figure Lengend Snippet: IL-17-expressing cells are differently modulated in the skin lesion and in the circulation of melanoma patients developing vitiligo during anti-PD-1 therapy. (A) Representative images of the immunohistochemistry (panels a to d) and immunofluorescence (panels e, f) analysis performed using antibodies directed against CD56 (panels a–c), IL-17 (panels d–f) and TCR alpha V 7.2 (panels g–i) on normal skin biopsies and on biopsies from patients affected by vitiligo without melanoma (vitiligo) and who developed vitiligo during immunotherapy (vitiligo DI). Magnification 200x. (B) Quantitative analyses of immunohistochemical staining. The mean value + standard error of the mean (SEM) of the cell count obtained for five different fields is shown, ***p<0.05 as assessed by Kruskal-Wallis test.

Article Snippet: For immunofluorescence, 4-µm sections were dewaxed, rehydrated, and after quenching endogenous peroxidase with 3% bovine serum albumin in 1x phosphate buffer, achieving antigen retrieval and blocking non-specific binding sites, sections were incubated with the monoclonal antibody anti-TCR V alpha 7.2 FITC conjugated (cod.130-123-929, Miltenyi Biotech; 1:50) and incubated for 1 hour at 37°C in a humid chamber.

Techniques: Expressing, Immunohistochemistry, Immunofluorescence, Staining, Cell Counting

Journal: Frontiers in Immunology

Article Title: Insight into immune profile associated with vitiligo onset and anti-tumoral response in melanoma patients receiving anti-PD-1 immunotherapy

doi: 10.3389/fimmu.2023.1197630

Figure Lengend Snippet: Expression of the immune cell subsets in the primary melanoma and metastases of patients developing or not vitiligo during anti-PD-1 therapy. (A) Representative images of immunohistochemistry (panels a to i) and immunofluorescence (l–n) obtained using anti-CD25 (a–c), anti-CD56 (d–f), anti-IL-17A (g–i), and anti-TCR alpha V 7.2 (j–l) in primary melanoma and metastasis from patients who developed (vitiligo) or not vitiligo (not vitiligo) during therapy. A–I 100x magnification, panel L-N 200X magnification. Magnification 200x. (B) Quantitative analyses of immunohistochemical staining. The mean value + standard error of the mean (SEM) of the cell count obtained for five different fields is shown, *p<0.05 as assessed by Kruskal-Wallis test.

Article Snippet: For immunofluorescence, 4-µm sections were dewaxed, rehydrated, and after quenching endogenous peroxidase with 3% bovine serum albumin in 1x phosphate buffer, achieving antigen retrieval and blocking non-specific binding sites, sections were incubated with the monoclonal antibody anti-TCR V alpha 7.2 FITC conjugated (cod.130-123-929, Miltenyi Biotech; 1:50) and incubated for 1 hour at 37°C in a humid chamber.

Techniques: Expressing, Immunohistochemistry, Immunofluorescence, Staining, Cell Counting

Journal: Nature

Article Title: 53BP1 cooperation with the REV7-Shieldin complex underpins DNA structure-specific NHEJ

doi: 10.1038/s41586-018-0362-1

Figure Lengend Snippet: (A) Schematic representation of the screen. Rev7 single and combinatorial point mutant alleles were stably expressed in Rev7-/- CH12-F3. IgM to IgA CSR was measured 40h post stimulation with anti-CD40 antibody, IL-4 and TGFβ-1 (CIT).

Article Snippet: B cells were stimulated with 5 μg/ml LPS (Sigma, L7770-1MG), 10 ng/ml mouse recombinant IL-4 (Peprotech, 214-14-20), and agonist anti-CD40 antibody (0.5 μg/ml; Miltenyi Biotec; FGK45.5).

Techniques: Mutagenesis, Stable Transfection

Journal: Biomolecules

Article Title: Supplementing L-Citrulline Can Extend Lifespan in C. elegans and Attenuate the Development of Aging-Related Impairments of Glucose Tolerance and Intestinal Barrier in Mice.

doi: 10.3390/biom13111579

Figure Lengend Snippet: Figure 3. Effect of enriching drinking water with L-Cit or hydrolyzed soy protein (=Iso-N-control) on markers of senescence in 4-, 16-, and 20-month-old C57BL/6J mice. (a) p16 protein concentration in the western blot of plasma; (b) total PAI-1 concentration in plasma; (c) IL6 concentration in the western blot of plasma; (d) CRP concentration in the western blot of plasma; and (e) representative images of p16, CRP, and IL6 and corresponding Ponceau images. Representative images of full- size membranes of western blots are shown in Supplemental Figures S1–S5. Data are presented as means ± SEM, * p ≤0.05. CRP, C-reactive protein; IL6, interleukin 6; Iso-N-control, hydrolyzed soy protein; L-Cit, L-Citrulline; PAI-1, plasminogen activator inhibitor-1.

Article Snippet: Membranes were incubated with primary antibodies for p16 (Biorbyt Ltd., Cambridge, UK), intestinal fatty acid binding protein (IFABP, Abcam, Cambridge, UK), IL6 (Santa Cruz Biotechnology, Inc., Heidelberg, Germany), and CRP (Santa Cruz Biotechnology, Inc., Heidelberg, Germany), and corresponding HRPlinked secondary antibodies.

Techniques: Control, Protein Concentration, Western Blot, Clinical Proteomics, Concentration Assay

Journal: Nature Communications

Article Title: Non-AUG HIV-1 uORF translation elicits specific T cell immune response and regulates viral transcript expression

doi: 10.1038/s41467-025-56772-3

Figure Lengend Snippet: ( A to D ), Peptides potentially encoded by uORFs (see peptides “A02; A03; A06; A07; A08” in panel (A) of Fig. ) were synthesized and used to screen for T cell responses in PBMCs of HIV-infected individuals. PBMCs from treated (ART) and untreated but aviremic (so called elite controllers, EC) individuals were stimulated with a pool of uORF-derived peptides (POOL) and cultured in the presence of IL-2 and IL-7 cytokines in order to expand peptide-specific memory T cells. On day 7 (not shown) and day 12, T cell responses against the POOL were assessed using IFNγ-Elispot. In addition, on day 12, except for donor EC-3, individual peptides of the pool (A02; A03; A06; A07; A08) were assessed using IFNγ-Elispot. As positive control for T cell expansion and activation, a pool of immunogenic peptides from HIV Env and Gag proteins was also used. A Number of uORF-derived peptides recognized by each HIV-infected individuals. The color code indicates from which uORF the peptides are derived (A02= Blue; A03= Orange; A07&A08= Red). B, C and D left panels, detailed IFNγ-Elispot data from the 3 individuals presenting T cell responses, expressed as spot forming units (SFU) per million PBMCs; right panels pictures of the raw data from the Elispot plates in technical triplicate ( n = 3 technical replicates) ( B and D ), or duplicate ( n = 2 technical replicates) ( C ), POOL (−) and POOL (+): PBMCs expanded with the pool of uORF-derived peptides but restimulated on the day of the Elispot with medium or the POOL, respectively. A02; A03; A06; A07 and A08 name of individual uORF-derived peptides used for re-stimulation. HIV (-) and HIV ( + ): PBMC expanded with the pool of Env- and Gag-derived peptides but re-stimulated for the Elispot assay with medium or the same pool of HIV peptides, respectively. Responses were considered positive when IFNγ production was superior to 20 spots/106 PBMCs and at least twofold higher than background production from cells re-stimulated with medium (dotted lines). SAT: saturated signal, where counts cannot be estimated due to overwhelm IFNγ secretion by activated T cells. Source data are provided as a Source Data file. Error bars correspond to the standard deviation of the mean.

Article Snippet: From 10-25 × 10 5 cells/well were seeded Elispot plates (MSIPN4550, Millipore) in IMDM supplemented with 10 % SAB, Pen/Strep, nonessential amino acids and sodium pyruvate, loaded with either 50, 10 or 2 μg/ml of uORF-derived peptides, HIV and CEF peptides, respectively and incubated at 37 °C for 16 h. Elispot plates were pre-coated with anti-IFNγ primary antibody and after cell-incubation IFNγ revealed using anti-IFNγ secondary antibody conjugated with biotin (both from Mabtech) as described .

Techniques: Synthesized, Infection, Derivative Assay, Cell Culture, Enzyme-linked Immunospot, Positive Control, Activation Assay, Standard Deviation