Photographs and skin-biopsy specimens were taken immediately before treatment and at scheduled time points after transplantation. The clinical photographs show specific areas that were consistently blistered before treatment. More rapid wound healing and reepithelialization were noted after transplantation in all six patients, with variable reductions in skin blistering. In Patient 7, marked cutaneous toxicity developed (presumably as a result of the chemotherapy) and was most pronounced on day 17 after transplantation but showed marked improvement by day 25. Immunofluorescence staining for C7 was carried out with skin-biopsy specimens obtained from all six patients, with the use of anti-C7 antibodies provided by Drs. D.T. Woodley and M. Chen (Keck School of Medicine, University of Southern California) for Patient 1 and purchased from BD Biosciences for the other five patients. For Patient 1, immunofluorescence staining before transplantation revealed faint, stippled labeling of C7; continuous, bright, linear C7 labeling was observed at all time points after bone marrow infusion. For Patient 3, faint, stippled C7 labeling was also observed at the dermal–epidermal junction before transplantation and through day 60 after transplantation, with continuous, bright, linear C7 labeling noted on day 145. For Patient 4, a bright, continuous band was observed at the dermal–epidermal junction before transplantation, with the use of anti-C7 antibody from BD Biosciences, whereas a thicker, brighter band was noted on day 198 after transplantation; this observation was confirmed with the use of LH7.2 anti-C7 antibody from Sigma-Aldrich, with no labeling before transplantation but bright labeling on day 198. For Patient 5, faint, stippled C7 labeling was observed at the dermal–epidermal junction with loss of demonstrable C7 labeling early after transplantation (day 28); however, continuous, bright linear, C7 labeling was observed on day 180. For Patient 6, no immunofluorescence staining was detectable before transplantation or at any time after transplantation, through day 102. For Patient 7, immunofluorescence staining before transplantation revealed stippled C7 labeling and continuous, bright, linear C7 labeling at the dermal–epidermal junction as early as day 30 after transplantation. For Patients 3 through 7, keratinocytes (green) were visualized with the use of anti–cytokeratin 5 antibody.

Panel A shows the relative fluorescence intensity with the use of an anti-C7 antibody from BD Biosciences. Panel B shows the results with the use of additional anti-C7 antibodies, which in some cases failed to bind mutant C7 before transplantation but did bind C7 after transplantation. An anti-C7 antibody provided by Drs. D.T. Woodley and M. Chen (Keck School of Medicine, University of Southern California) was used for Patients 1, 3, 6, and 7), and an anti-C7 antibody (LH7.2) made by Sigma-Aldrich was used for Patients 4 and 5. (Each antibody was applied to tissue from Patients 3 through 7; the data presented are for samples showing the most pronounced changes in C7 expression.) Species-specific and isotype-specific control antibodies, which were used in assays performed on the same day as those carried out with anti-C7 antibodies, yielded negative results (data not shown). Note that 100% is the maximal measurable fluorescence intensity on the gray scale.

Photomicrographs show thin fibrillar structures beneath the lamina densa before transplantation and on day 60 after transplantation in Patient 4 (Panel A, arrowheads) and before transplantation and on day 100 in Patient 7 (Panel B, arrowheads), with an increase in the number of fibrils after transplantation. Although some of these fibrils appear somewhat thicker in Patient 4 than in Patient 7 (Panel A, arrow), none bear the ultrastructural hallmarks of normal anchoring fibrils. The bars represent 250 nm.