Journal: Translational Andrology and Urology

Article Title: Functional characterization of the immunomodulatory properties of human urine-derived stem cells

doi: 10.21037/tau-21-506

Figure Lengend Snippet: USCs inhibited the proliferation of PBMNCs in MLR. USC-TA+, USC-TA−, BMSCs, and SMCs were used as stimulator cells and human PBMNCs from two healthy donors were used as responder cells. One-way MLR with single donor PBMNCs (donor A or B) as responder cells and two-way MLR with aliquot half dose of two different donors’ PBMNCs (donor A and B) as responder cells were detected in the same multi-well plate. Cultures were incubated at 37 °C for 5 days in 5% CO2 and 100% humidity, the cells were checked with inverted microscope on the 5th day before further experiment. There were different cell densities of PBMNCs be noticed in different wells for 5 days mix-culture, Scale bar =50 µm (A). The proliferation of PBMNCs was assessed with the BrdU cell proliferation colorimetric ELISA. Newly synthesized BrdU-DNA was quantified using a scanning multi-well spectrophotometer (B). *, P<0.05, post-hoc paired-comparisons. MLR, mixed lymphocyte reaction; BMSC, human bone marrow mesenchymal stem cells; SMC, human smooth muscle cells; USC TA+, human urine derived stem cells with high telomerase activity; USC TA−, human urine derived stem cells with low telomerase activity; PBMNC, human peripheral blood mononuclear cells; SMC + PBMNC A, one-way mixed lymphocyte reaction as human smooth muscle cells mixed with human peripheral blood mononuclear cells from donor A; SMC + PBMNC B, one-way mixed lymphocyte reaction as human smooth muscle cells mixed with human peripheral blood mononuclear cells from donor B; SMC + PBMNC A + B, two-way mixed lymphocyte reaction as human smooth muscle cells mixed with aliquot half dose of two different donors’ human peripheral blood mononuclear cells from donor A and donor B; BMSC + PBMNC A, one-way mixed lymphocyte reaction as human bone marrow mesenchymal stem cells mixed with human peripheral blood mononuclear cells from donor A; BMSC + PBMNC B, one-way mixed lymphocyte reaction as human bone marrow mesenchymal stem cells mixed with human peripheral blood mononuclear cells from donor B; BMSC + PBMNC A + B, two-way mixed lymphocyte reaction as human bone marrow mesenchymal stem cells mixed with aliquot half dose of two different donors’ human peripheral blood mononuclear cells from donor A and donor B; USC + PBMNC A, one-way mixed lymphocyte reaction as human urine derived stem cells mixed with human peripheral blood mononuclear cells from donor A; USC + PBMNC B, one-way mixed lymphocyte reaction as human urine derived stem cells mixed with human peripheral blood mononuclear cells from donor B; USC + PBMNC A + B, two-way mixed lymphocyte reaction as human urine derived stem cells mixed with aliquot half dose of two different donors’ human peripheral blood mononuclear cells from donor A and donor B.

Article Snippet: Human peripheral blood mononuclear cells (PBMNCs, normal human, ATCC PCS-800-011) from two different donors were purchased from ATCC (American Type Culture Collection, MD, USA).

Techniques: Incubation, Inverted Microscopy, Enzyme-linked Immunosorbent Assay, Synthesized, Spectrophotometry, Derivative Assay, Activity Assay

Journal: Translational Andrology and Urology

Article Title: Functional characterization of the immunomodulatory properties of human urine-derived stem cells

doi: 10.21037/tau-21-506

Figure Lengend Snippet: USC showed a characteristic cytokine release profile. The supernatants from USC-TA+, USC-TA−, and BMSCs cultured alone or co-cultured with PBMNCs (direct mixed culture with PBMNCs, or indirect mixed culture with PBMNCs in transwell insert for 48 h) (A) were assessed for their relative levels of cytokines and chemokines by using the human cytokine array panel A, as described in “Methods” (B). A: human cytokine array panel template; B: PBMNCs background; C: USC-TA+ culture alone; D: USC-TA− culture alone; E: BMSCs culture alone; F: USC-TA+ direct mix culture with PBMNCs; G: USC-TA− direct mix culture with PBMNCs; H: BMSCs direct mix culture with PBMNCs; I: USC-TA+ in-direct mix culture with PBMNCs in transwell insert; J: USC-TA− in-direct mix culture with PBMNCs in transwell insert; K: BMSCs in-direct mix culture with PBMNCs in transwell insert: The immunoblot of cytokine expression levels were visualized and quantifed (C). *, P<0.05, comparing with BMSCs alone; †, P<0.05, comparing direct and indirect mixed culture with PBMNCs. BMSC, human bone marrow mesenchymal stem cells; SMC, human smooth muscle cells; USC TA+, human urine derived stem cells with high telomerase activity; USC TA−, human urine derived stem cells with low telomerase activity; PBMNC, human peripheral blood mononuclear cells.

Article Snippet: Human peripheral blood mononuclear cells (PBMNCs, normal human, ATCC PCS-800-011) from two different donors were purchased from ATCC (American Type Culture Collection, MD, USA).

Techniques: Cell Culture, Western Blot, Expressing, Derivative Assay, Activity Assay

Journal: STAR Protocols

Article Title: Mouse splenocyte enrichment strategies via negative selection for broadened single-cell transcriptomics

doi: 10.1016/j.xpro.2022.101402

Figure Lengend Snippet: Flow cytometry enriched antibody cocktail (22 μL) panel for analyzing rarer granulocyte and myeloid cells in mouse spleen for downstream scRNA-seq

Article Snippet: CD11b Antibody, anti-mouse, VioBlue®, REAfinityTM Clone REA592 , Miltenyi Biotec , Cat# 130-113-810; RRID: AB_2726327.

Techniques: Flow Cytometry, Marker

Journal: STAR Protocols

Article Title: Mouse splenocyte enrichment strategies via negative selection for broadened single-cell transcriptomics

doi: 10.1016/j.xpro.2022.101402

Figure Lengend Snippet:

Article Snippet: CD11b Antibody, anti-mouse, VioBlue®, REAfinityTM Clone REA592 , Miltenyi Biotec , Cat# 130-113-810; RRID: AB_2726327.

Techniques: Recombinant, Sterility, Saline, Extraction, Blocking Assay, Software, Solvent, Sequencing, RNA Sequencing, Transferring, Aerosol, Irradiation, Inverted Microscopy, Flow Cytometry, Fluorescence