Journal: Journal of Cellular and Molecular Medicine

Article Title: Senescence determines the fate of activated rat pancreatic stellate cells

doi: 10.1111/j.1582-4934.2012.01573.x

Figure Lengend Snippet: Histological characteristics of DBTC-induced pancreatitis. Pancreatitis was induced by the injection of DBTC (8 mg/kg). All microphotographs correspond to pancreatic sections of a rat killed on day 7 after DBTC application, and are representative of at least 10 animals analysed. ( A ) H&E staining of paraffin-embedded tissue reveals typical histology of the disease, including acinar cell damage, ductal changes, periductal and interstitial fibrosis and inflammatory cell infiltrates. ( B–F ) Cryostat tissue sections were used to detect CD4, CD8 and CD161-positive lymphocytes as well as cells expressing the PSC markers α-SMA and desmin, employing specific antibodies and the APAAP technique. ( A , B , D , E ) original magnification ×100, ( C and F ): ×200. The lymphocyte stainings were performed on serial sections of the tissue.

Article Snippet: Rat NK cells were separated in an autoMACS Separator (Miltenyi Biotec GmbH, Bergisch Gladbach, Germany) by a negative selection for CD3 and positive selection for CD161, using R-phycoerythrin (PE)-labelled anti-CD3 and FITC-labelled anti-CD161 antibodies (Immunotools, Friesoythe, Germany).

Techniques: Injection, Staining, Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: Senescence determines the fate of activated rat pancreatic stellate cells

doi: 10.1111/j.1582-4934.2012.01573.x

Figure Lengend Snippet: Semi-quantitative and quantitative assessment of histopathological changes in rats with DBTC-induced pancreatitis. To induce mild or more severe chronic pancreatitis, DBTC was injected at 6 and 8 mg/kg, respectively. At the indicated time points, the rats were killed and pancreatic tissue subjected to histopathological evaluation. ( A ) Applying a semi-quantitative score from 0 to 3, an overall assessment of disease severity was conducted as described in the Materials and methods section, using H&E stained samples. ( B ) After H&E staining, extension of fibrosis was judged on a scale from 0 to 6 points (up to 3 points for most severe periductal and interstitial fibrosis respectively). ( C ) Extension of areas covered with SA β-Gal-positive cells was evaluated on a scale from 0 to 3. ( D–F ) After APAAP staining with specific antibodies, α-SMA expression was assessed using a scale from 0 to 3 (lower panel). CD4 and CD161-positive cells (upper and middle panel) were counted, employing an ocular grid, at a magnification of 400×. All results are averaged values (±S.E.M.) of at least five rats per experimental group.

Article Snippet: Rat NK cells were separated in an autoMACS Separator (Miltenyi Biotec GmbH, Bergisch Gladbach, Germany) by a negative selection for CD3 and positive selection for CD161, using R-phycoerythrin (PE)-labelled anti-CD3 and FITC-labelled anti-CD161 antibodies (Immunotools, Friesoythe, Germany).

Techniques: Injection, Staining, Expressing

Journal: Journal for Immunotherapy of Cancer

Article Title: STAT3 inhibitor Napabucasin abrogates MDSC immunosuppressive capacity and prolongs survival of melanoma-bearing mice

doi: 10.1136/jitc-2021-004384

Figure Lengend Snippet: STAT3 activation in MDSC from patients with melanoma and its inhibition by Napaucasin. PBMC from untreated patients with melanoma (stages I–IV) were analyzed by flow cytometry. (A) Representative dot plots for M-MDSC defined as CD33 high HLA-DR low/neg and PMN-MDSC defined as CD33 dim HLA-DR neg CD66b + LIN − are presented. (B) Cumulative data are shown as the percentage of pSTAT3 + cells among respective MDSC subsets (mean±SD, n=19). pSTAT3 + MDSC were gated according to isotype control. (C) PFS of patients with high (>58.2%, n=8) and low (<58.2%, n=7) pSTAT3 + M-MDSC frequencies is shown as a Kaplan–Meier curve. Follow-up period was 2 years. (D) Cumulative data are presented as the percentage of pSTAT3 + M-MDSC within total M-MDSC in patients with (n=11) or without (n=4) progression. (E) Human M-MDSCs were generated in vitro for 4 days with IL-6 and GM-CSF followed by coculture with activated CD3 + T cells labeled with cell proliferation dye eFluor 450 in the presence of 0.5 µM Napa or 0.005% DMSO. On 96 hours of incubation, T-cell proliferation was assessed by proliferation dye dilution measured by flow cytometry. Cumulative data for T-cell proliferation are presented as the percentage of divided T cells norm. to the respective control of stimulated T cells alone (mean±SD, n=4). Statistics were performed on not norm. data. *P<0.05, **P<0.01, ***P<0.001. gen., generated; GM-CSF, granulocyte–macrophage colony-stimulating factor; IL, interleukin; MDSC, myeloid-derived suppressor cell; M-MDSC, monocytic myeloid-derived suppressor cell; Napa, Napabucasin; norm., normalized; PFS, progression-free survival; PMN, polymorphonuclear; STAT3, signal transducer and activator of transcription 3.

Article Snippet: The proliferation of T cells was assessed after 4 days of coculture by measuring proliferation dye eFluor 450 dilution using the BD FACSLyric flow cytometer.

Techniques: Activation Assay, Inhibition, Flow Cytometry, Generated, In Vitro, Labeling, Incubation, Derivative Assay