1. Pancreatitis Associated Protein I (PAP-I) Alters Adhesion and Motility of Human Melanocytes and Melanoma Cells 
Christine Valery, Jean-Jacques Grob, Patrick Verrando 
Journal of Investigative Dermatology 
Volume 116, Issue 3, Pages (March 2001)
DOI: /j x
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Infection of melanocytic cells with adenoviral shuttle vector Ad-PAP results in intracellular PAP-I expression. (A) Normal human melanocytes (NHM), SK-MEL-2, and KAL melanoma cell lines were infected with pδE1sp1B-CMV-PAP construct (Ad-PAP) or pδE1sp1B-CMV-CAT (control) construct (Ad-CAT) at 0, 40, and 100 multiplicity of infection per cell. Twenty-four hours after the removal of viruses, cells were allowed to react with a specific polyclonal antibody to human PAP-I (H8) in an indirect immunofluorescence assay and analyzed by cytofluorimetry. Histograms report the fluorescence intensities of Ad-PAP-infected cells, after subtraction of the background fluorescence intensity of the corresponding Ad-CAT-infected cells. Percentages of fluorescent cells that express PAP are given. (B) Indirect immunofluorescence assay detection with antibody H8 of PAP from Ad-PAP-infected normal human melanocytes at 40 multiplicity of infection per cell (left) compared with Ad-CAT-infected normal human melanocytes (right) (scale bar: 8.5 μm).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Adhesion of human melanocytic cells to extracellular matrix components can be modified by infection with Ad-PAP. Normal human melanocytes (NHM), SK-MEL-2, and KAL melanoma cell lines were infected with pδE1sp1B-CMV-PAP construct (Ad-PAP, black boxes) or pδE1sp1B-CMV-CAT control construct (Ad-CAT, gray boxes) and assayed 24 h after the removal of viruses. Cells were allowed to adhere for 1 h on immobilized laminin, fibronectin, and collagen I as described in Materials and Methods. Adhesion is expressed as the optical density (OD) of adherent cells stained by crystal violet (mean ±SEM, n = 3; Ad-PAP vs Ad-CAT: *p <0.1, NS = not significant).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Cell surface expression of integrin α chains in human melanocytic cells infected with Ad-PAP and Ad-CAT is almost unchanged. Normal human melanocytes (NHM), SK-MEL-2, and KAL melanoma cell lines were infected with pδE1sp1B-CMV-PAP construct (Ad-PAP, black boxes) or pδE1sp1B-CMV-CAT control construct (Ad-CAT, gray boxes). Twenty-four hours after the removal of viruses, they were allowed to react with specific integrin α-chain antibodies and the corresponding fluorophore-conjugated secondary antibody. Fluorescence was determined by flow cytometry.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Loss of integrin-dependent adhesion to some extracellular matrix components of human melanocytes infected with Ad-PAP. Normal human melanocytes (NHM) were infected with pδE1sp1B-CMV-PAP construct (Ad-PAP, black boxes) or pδE1sp1B-CMV-CAT control construct (Ad-CAT, gray boxes) and assayed 24 h after the removal of viruses as described in Materials and Methods. Adhesion rates were evaluated as indicated in Figure 1 (mean ±SEM, n = 3; *p <0.1, NS = not significant). Ab: integrin-function-blocking antibody.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Haptotactic migration of some melanocytic cells infected with Ad-PAP is enhanced on laminin-1, fibronectin, and collagen I. Normal human melanocytes (NHM), SK-MEL-2, and KAL melanoma cell lines were infected with pδE1sp1B-CMV-PAP construct (Ad-PAP, black boxes) or pδE1sp1B-CMV-CAT control construct (Ad-CAT, gray boxes) and assayed 24 h after the removal of viruses in the phagokinetic track assay (mean ±SEM, n = 8; *p <0.1, NS = not significant).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

7. Figure 6
Directed movement of some melanocytic cells infected with Ad-PAP is enhanced in transmigration assays. Normal human melanocytes (NHM), SK-MEL-2, and KAL melanoma cell lines were infected with pδE1sp1B-CMV-PAP construct (Ad-PAP, black boxes) or pδE1sp1B-CMV-CAT control construct (Ad-CAT, gray boxes) and assayed 24 h after the removal of viruses on uncoated (transmigration) or Matrigel-coated (invasion) Boyden chamber filters. Results are expressed as the number of melanocytic cells observed per microscopic field (mean ±SEM; n = 8; *p <0.01, NS = not significant).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

8. Figure 7
Stimulation of directed movement of human melanocytic cells, human keratinocytes, and human fibroblasts in the presence of purified PAP-I. Normal human melanocytes (NHM), SK-MEL-2, and KAL melanoma cell lines (A) as well as human epidermal keratinocytes and dermal fibroblasts (B) were assayed for directed movement in the presence of 0, 1, and 10 ng per ml purified rat PAP-I protein in uncoated Boyden chambers. Results are expressed as the stimulation factor of movement obtained by comparison with cells that migrated in the absence of exogenous PAP-I (mean ±SEM; n = 8). Statistical significance versus 0 ng per ml PAP-I (control): (A) normal human melanocytes, NS = not significant; SK-MEL-2, NS = not significant; KAL, *p <0.03, **p <0.03; (B) fibroblasts and keratinocytes, *p =0.02, **p <0.02.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions

9. Figure 8
Exogenous PAP-I enhances the spreading of KAL melanoma cells on fibronectin. Normal human melanocytes (NHM) (A, B), SK-MEL-2 (C, D), and KAL (E, F) melanoma cell lines, previously cultivated for 3 h in the presence (left) or absence (right) of 10 ng per ml PAP-I, were allowed to attach and to spread for 1 h on immobilized fibronectin (30 μg per ml). The figure shows the immunofluorescence staining of the cells after actin-binding of rhodamine-phalloidin (scale bar 8.5 μm).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2001 The Society for Investigative Dermatology, Inc Terms and Conditions