Journal: Biochimica et biophysica acta

Article Title: Regulation of the release and function of tumor cell-derived soluble CD44.

doi: 10.1016/j.bbamcr.2005.02.006

Figure Lengend Snippet: Fig. 2. Molecular characterization of soluble CD44 from PMA-treated HTB58 cells. Cells were labeled with [35S]methionine (A, B) or [35S]sulfate (C, D) in the absence or presence of indicated cytokines for 48 h, washed once with culture media and then treated with 5107 M PMA or diluent control DMSO for 90 min (A, C) or 4 h (B, D). CD44 from conditioned media was then immunoprecipitated with anti-CD44 mAb followed by protein A/G plus agarose. Immune complexes were resolved on SDS-8% PAGE under non-reducing conditions followed by fluorography. Molecular weight markers in kDa are shown. OT=OSM+TGF-h1.

Article Snippet: HTB58 human lung squamous carcinoma cell line was obtained from the ATCC (Rockville, MD).

Techniques: Labeling, Control, Immunoprecipitation, Molecular Weight

Journal: Biochimica et biophysica acta

Article Title: Regulation of the release and function of tumor cell-derived soluble CD44.

doi: 10.1016/j.bbamcr.2005.02.006

Figure Lengend Snippet: Fig. 1. HTB58 cells shed CD44 in response to PMA. Cells were incubated for 48 h in MEM supplemented with 50 ng/ml OSM, 10 ng/ml TGF-h1 or both cytokines together (OT). Media were changed and cells were treated for 90 min with 5107 M PMA or diluent control DMSO. CD44 membrane levels, expressed as mean fluorescence intensity (MFI), were determined by flow cytometry using PE-conjugated anti-CD44 mAb. Paired results represent the cell surface expression of CD44 in the absence or presence of PMA. Data show one representative experiment out of four (A). The levels of CD44 in conditioned media collected at 90 min were quantified by ELISA (B and C). The data are shown as the mean of triplicate wells from five experimentsFS.D. (B). To block metalloprotei- nase activity, the cells were first pre-treated with 50 AM TAPI for 15 min at 37 8C and then treated with 50 AM TAPI simultaneously with PMA. The data are shown as the mean of duplicate wells from one representative experiment out of three (C). The difference between cells incubated in the absence or presence of PMA depicted in panel B was statistically significant as determined by Student’s t test ( P b0.05).

Article Snippet: HTB58 human lung squamous carcinoma cell line was obtained from the ATCC (Rockville, MD).

Techniques: Incubation, Control, Membrane, Fluorescence, Flow Cytometry, Expressing, Enzyme-linked Immunosorbent Assay, Blocking Assay, Activity Assay