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. 2006 Aug;169(2):617–632. doi: 10.2353/ajpath.2006.050876

Figure 9-6913.

Figure 9-6913

Western blot analyses for cellular and secreted cath-L and cath-D from FTC-133-EGFP (A–C, lanes 1) and different FTC-133-RLN2 transfectants (A–C, lanes 2 to 4). B: Immunoreactive forms at 24, 31, and 42 kd resembling the heavy chain, single chain, and procath-L, respectively, were detected with the mAb 33/123,24 in the cellular and secreted protein fractions of all FTC-133 transfectants investigated. When compared with FTC-133-EGFP clones (A and B, lanes 1), FTC-133-RLN2 (A and B, lanes 2 to 4) displayed increased production and secretion of the cath-L proform (42 kd) as determined by the mAb 2D422 specific for procath-L (A) and the mAb 33/123,24 against an epitope present in all three cath-L forms (B). B: In addition, FTC-133-RLN2 clones (lanes 2 to 4) showed enhanced production and secretion of the 31-kd and 24-kd bioactive cath-L forms as compared to FTC-133-EGFP clones (lane 1). C: Two immunoreactive cath-D forms at 52 kd and 25 kd were detected in the cellular protein fractions of both FTC-133-EGFP (lane 1) and FTC-133-RLN2 (lanes 2 to 4) transfectants. FTC-133-RLN2 displayed significantly increased amounts of procath-D at 52 kd in cellular extracts (lanes 2 to 4) as compared to FTC-133-EGFP controls (lane 1). The amount of the 25-kd cath-D in the cellular extracts was slightly reduced in all FTC-133 clones studied (lanes 1 to 4). The secreted protein fraction exclusively contained procath-D (52 kd) at similar levels in FTC-133-EGFP (lane 1) and FTC-133-RLN2 (lanes 2 to 4) transfectants. Equal protein loading was checked by β-actin staining of stripped membranes. D: Representative Western blots are shown for cellular and secreted cath-L of FTC-238-EGFP (lane 1) and FTC-238-RLN2 clones (lane 2). Similar to FTC-133-RLN2 clones, corresponding FTC-238-RLN2 transfectants demonstrated an up-regulation in cath-L single chain (31 kd) and heavy chain (24 kd), whereas the amount of procath-L (42 kd) produced by the FTC-238 clones was negligible. The pattern of cellular and secreted cath-D in the FTC-238-RLN2 transfectant was similar to corresponding FTC-133 clones (data not shown).