Journal: Stem Cells (Dayton, Ohio)

Article Title: TAFA2 Induces Skeletal (Stromal) Stem Cell Migration Through Activation of Rac1‐p38 Signaling

doi: 10.1002/stem.2955

Figure Lengend Snippet: TAFA2 levels are regulated during the process of fracture healing. ( A): Immunohistochemical analysis of TAFA2 protein expression in a fractured femur, 48 hours after induction of fracture. Scale bar: 1 mm. (B): Analysis of TAFA2 immunoreactivity at the fracture site (boxed area from A ) at different time points (2, 24, and 48 hours, 4 days, and 1 week) after induction of femoral fracture (upper panel) and nonfractured femur (lower panel). Scale bar: 100 μm. (C): Quantitative reverse transcription polymerase chain reaction (qRT‐PCR) analysis of TAFA2 mRNA expression in nonfractured and fractured mouse femur at different time points after fracture induction (2 hours, day 1, day 4, and day 14). (D): Flow cytometry analysis of TAFA2 protein expression in bone marrow cells obtained from fractured femur at different time points. Data represent MFI ± SEM from three independent experiments. **, p ≤ .01; two‐tailed unpaired Student t test. (E): qRT‐PCR analysis of IL‐1β mRNA expression in mouse femur before fracture and at different time points after fracture induction (2 hours, day 1, day 4, and day 14). (F): qRT‐PCR analysis of TAFA2 mRNA expression in human skeletal (mesenchymal, stromal) stem cell (hMSC) cultures in the presence of different doses of IL‐1β (1, 10, and 50 ng/ml) for 48 hours. Data represent three independent experiments ± SEM; **, p ≤ .01, two‐tailed paired Student t test. (G, H): Western blot analysis and quantification of TAFA2 protein levels in hMSC cultures in the presence of different doses of IL‐1β. Data represent mean ± SEM from four independent experiments. *, p ≤ .05, two‐tailed unpaired Student t test. (I, J): Flow cytometry analysis of TAFA2 protein expression and percentage of leukocytes (CD45 + ), monocytes (CD14 + ), T cells (CD3 + ), and B cells (CD19 + ) at 1 and 3 days postfracture. Data are obtained from pooled bone marrow cells isolated from fractured femurs of three mice. (K): Enzyme‐linked immunosorbent assay measurement of TAFA2 levels in consecutive serum samples from patients with hip fracture. **, p ≤ .01, two‐tailed paired Student t test. (L): Proposed model for role of TAFA2 in recruiting MSC to the site of fracture. Abbreviations: IL, interleukins; MFI, mean fluorescence intensity; MSC, skeletal (mesenchymal) stem cell.

Article Snippet: ELISA Measurement of TAFA2 Concentrations in Clinical SamplesELISA measurements of TAFA2 concentrations in human serum samples were performed using an established ELISA procedure (Mybiosource, San Diego, USA) according to the manufacturer's instructions.

Techniques: Immunohistochemistry, Expressing, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Flow Cytometry, Two Tailed Test, Western Blot, Isolation, Mouse Assay, Enzyme-linked Immunosorbent Assay, Fluorescence

Journal: PLoS ONE

Article Title: Stepwise Release of Biologically Active HMGB1 during HSV-2 Infection

doi: 10.1371/journal.pone.0016145

Figure Lengend Snippet: HMGB1 released by HSV-2-infected cells activates fibroblast migration and stimulates HIV-1 expression. MRC5 fibroblasts were subjected to migration assays in the presence of (A) human recombinant HMGB1 or (B) supernatants from mock (M) or HSV-2-infected HEC-1 cells. HMGB1 concentrations reached 270 ng/ml in the supernatants collected from infected cells, versus 50 ng/ml in mock-infected cell media. Control experiments used glycyrrhizin or neutralizing antibodies against HMGB1 or its receptors RAGE, TLR-4 and TLR-2. Data are expressed as -fold increases in cell migration compared to non treated control cells. ACH-2 cells, that contain a latent HIV-1 provirus, were grown in the presence of (C) human recombinant HMGB1 or (D) supernatants from mock (M) or HSV-2-infected HEC-1 cells. P24 antigen was measured by ELISA after 36 h. Data are expressed as the -fold increase in p24 antigen compared with non treated control cells. Virion-free supernatants were obtained by ultracentrifugation. Results are either representative of several experiments (A, C, D) or expressed as the mean and SD of 3 independent experiments (B).

Article Snippet: HIV production was evaluated by measuring p24 antigen with an ELISA method (Innogenetics, Les Ulis, France).

Techniques: Infection, Migration, Expressing, Recombinant, Enzyme-linked Immunosorbent Assay

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Cultivation and Immortalization of Human B-Cells Producing a Human Monoclonal IgM Antibody Binding to MDA-LDL: Further Evidence for Formation of Atherogenic MDA-LDL Adducts in Humans In Vivo

doi: 10.1155/2017/6047142

Figure Lengend Snippet: The effect of several antioxidants used as competitors in the monoclonal anti-MDA-LDL IgM ELISA. Aldehyde residues of MDA were reduced, presumably to alcohols, by antioxidants, thus precluding binding of the MCA.

Article Snippet: Identification of Productive Clones At the end of the HAT selection period, supernatants of each well were tested for antibodies binding to Cu++ -oxidized LDL and/or MDA-LDL with commercially available ELISA methods (oLAb-ELISA, Biomedica, Vienna, Austria; MDA-LDL IgG/IgM, LDN Ltd., Nordhorn, Germany) adapted for the purposes of this investigation [ ].

Techniques: Multiple Displacement Amplification, Enzyme-linked Immunosorbent Assay, Binding Assay