Journal: Blood Advances

Article Title: Gene-edited pseudogene resurrection corrects p47 phox -deficient chronic granulomatous disease

doi: 10.1182/bloodadvances.2016001214

Figure Lengend Snippet: Functional analysis of gene-targeting correction of p47-CGD patient HPCs. HPCs from p47-CGD subject 3 (homozygous exon 2 ΔGT NCF1) were cultured in Stemspan SFEMII supplemented with stem cell factor, Flt3 ligand, and thrombopoietin and gene targeted with ZFN mRNA and donor B or donor D rAAV6 at day 2 of culture and analyzed at day 10 of culture under conditions inducing myeloid differentiation with 50 ng/mL granulocyte colony-stimulating factor. Shown first on the left are uncorrected myeloid differentiating HPCs from the patient in whom no oxidase-positive cells were detected. Shown last on the right are myeloid differentiating HPCs from a healthy control, where 33% of cells fall into the oxidase-positive gate. Shown on the middle-left and on the middle-right panels, respectively, are myeloid differentiating minigene (donor B) or exon 2 replacement (donor D) corrected HPCs from the patient where significant numbers of oxidase-positive cells are detected with MFI approaching that of a healthy control subject.

Article Snippet: The harvested HPCs were further expanded in stage 3 media or stem cell media (StemSpan SFEMII medium, STEMCELL Technologies) containing 50 ng/mL stem cell factor, FLT-3L, thrombopoietin, 5 ng/mL interleukin 3 (IL-3), and 25 ng/mL IL-6.

Techniques: Functional Assay, Cell Culture

Journal: Frontiers in Immunology

Article Title: Icaritin Induces Anti-tumor Immune Responses in Hepatocellular Carcinoma by Inhibiting Splenic Myeloid-Derived Suppressor Cell Generation

doi: 10.3389/fimmu.2021.609295

Figure Lengend Snippet: Icaritin reduces the accumulation of HSPCs in the spleen of Hepa mice. Spleen weight and total cell number from orthotopic (A) and subcutaneous (B) Hepa mice. Numbers of splenic LSK and LK cells from orthotopic (C) and subcutaneous (D) Hepa mice. Differences between groups were examined for statistical significance by Student's t -test. Data were pooled from two experiments and n = 8 mice per group. * P < 0.05, ** P < 0.01, *** P < 0.001. HSPCs, hematopoietic stem and progenitor cells; Hepa, Hepa1-6 cells; ICT, icaritin; Veh, Vehicle; LSK, Lin lo/− Sca-1 + c-Kit hi ; LK, Lin lo/− Sca-1 − c-Kit hi .

Article Snippet: In brief, human CD34 + cells were isolated from the cord blood mononuclear cells using a direct CD34 progenitor cell isolation kit (Miltenyi Biotec), and cultured in hematopoietic stem cell expansion media (StemSpan SFEM; Stem Cell Technologies) supplemented with 100 ng/mL stem cell factor, 100 ng/mL Fms-like tyrosine kinase 3, 100 ng/mL thrombopoietin, and 20 ng/mL IL-3 (R&D Systems) for 9 days ( ).

Techniques:

Journal: Frontiers in Immunology

Article Title: Icaritin Induces Anti-tumor Immune Responses in Hepatocellular Carcinoma by Inhibiting Splenic Myeloid-Derived Suppressor Cell Generation

doi: 10.3389/fimmu.2021.609295

Figure Lengend Snippet: Icaritin inhibits the generation of human PMN-MDSCs in vitro . Freshly isolated human CD34 + cells from cord blood mononuclear cells were cultured in hematopoietic stem cell expansion media for 8–10 days. The expanded cells were cultured with combined IL-6 and G-CSF in complete medium with 0.1% DMSO (vehicle) or icaritin (2.5 μM) for 3 days. (A) Flow cytometric analysis of CD115 and HLA-DR expression on CD14 + or CD15 + cells. (B) Frequencies of PMN-MDSCs (CD15 + CD115 + ) and M-MDSCs (CD14 + HLA-DR − ). Differences between groups were analyzed using one-way ANOVA, and corrected by Dunnett's test. Data are representative of three experiments. ** P < 0.01. Veh, Vehicle; ICT, Icaritin.

Article Snippet: In brief, human CD34 + cells were isolated from the cord blood mononuclear cells using a direct CD34 progenitor cell isolation kit (Miltenyi Biotec), and cultured in hematopoietic stem cell expansion media (StemSpan SFEM; Stem Cell Technologies) supplemented with 100 ng/mL stem cell factor, 100 ng/mL Fms-like tyrosine kinase 3, 100 ng/mL thrombopoietin, and 20 ng/mL IL-3 (R&D Systems) for 9 days ( ).

Techniques: In Vitro, Isolation, Cell Culture, Expressing