1. The Cutaneous Microfibrillar Apparatus Contains Latent Transforming Growth Factor-β Binding Protein-1 (LTBP-1) and is a Repository for Latent TGF-β1 
Michael Raghunath, Christine Unsöld, Leena Bruckner-Tuderman 
Journal of Investigative Dermatology 
Volume 111, Issue 4, Pages (October 1998)
DOI: /j x
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Immunoblot and radioimmunoprecipitation analyses exclude cross-reactivity of Ab39 against LTBP-1 with fibrillin. (a) Immunoblot from proteins from conditioned fibroblast culture medium separated on a 4% sodium dodecyl sulfate-polyacrylamide gel under nonreducing conditions and transferred onto nitrocellulose. Immunodetection of LTBP-1 with Ab39 results in a broad double band at ≈200kDa (lane 1), whereas MoAb69 recognizes only fibrillin migrating at ≈320kDa (lane 2). Fibrillin is not recognized by Ab39 (lane 1) nor is LTBP-1 recognized by Ab39. (b) Ab39 immunoprecipitates LTPB-1 as a 200kDa protein from culture medium of metabolically labeled fibroblasts. There is no coprecipitation of fibrillin. Lane 1, normal rabbit serum;lane 2, fibronectin;lane 3, LBTP-1;lane 4, crude medium
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
LTBP-1 is a constituent of the microfibrillar apparatus of the skin. Confocal laser scanning localization of fibrillin and LTBP-1 in normal and regenerating skin. Each micrograph represents an extended focus view calculated from 10 section planes (increment μm). The yellow mix color of superimposed images obtained for LTBP-1 (green immunofluorescence) and fibrillin (red immunofluorescence) suggests in normal skin extensive colocalization of fibrillin and LTBP-1 on the transpapillary (a) and the reticular portion of the microfibrillar apparatus (b). Regenerating skin: extensive colocalization of LTBP-1 with fibrillin throughout all developmental stages of the microfibrillar apparatus from day 7 after grafting (c), 1 mo after grafting (d), 17 mo after grafting (e), and 2 y after grafting (f). Scale bar: 10 μm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
Microfibrils of different tissues are a repository for latent TGF-β1. Confocal laser scanning study of fibrillin and TGF-β1 precursor on microfibrillar structures in human skin and cynomolgus monkey eye. (a–c) Tissues were double stained with MoAb69 against fibrillin (red signal) and anti-β-LAP (green signal). (a, b) The yellow mix color of superimposed images demonstrates association of TGF-β1 precursor with fibrillin-microfibrils. Note that the cytoplasm of basal and suprabasal keratinocytes contains latent TGF-β1. (c) Eye: the picture shows a rod-like ciliary zonule as part of the suspensory ligament of the eye lens surrounded by parts of the iris and the processus ciliaris. The yellow mix color demonstrates colocalization of fibrillin and latent TGF-β1 on the ciliary zonule fiber. (d) Human papillary dermis simultaneously immunostained for fibrillin (blue signal), LTBP-1 (green signal), and TGF-β1 precursor (red signal). The white mix colors indicates colocalization of all three antigens on individual microfibrillar bundles. Scale bars: 10 μm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Fibrillin but not LBTP-1 is present on zonule fibrils of the cynomolgus monkey eye. Conventional immunofluorescence detection of fibrillin (a) and LTBP-1 (b) in equatorial cryosection sections of cynomolgus monkey eye. L, lens; C, lens capsule; ZF, zonule fibril. Fibrillin is present on zonule fibrils, whereas LTBP-1 cannot be detected. Scale bars: 25 μm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
The developing microfibrillar apparatus is capable of latent TGF-β1 storage. Confocal laser scanning localization of fibrillin and TGF-β1 precursor in regenerating skin. Tissues were double stained with MoAb69 against fibrillin (red immunofluorescence) and anti-β-LAP (green immunofluorescence). The yellow mix color of superimposed images demonstrates association of latent TGF-β1 with fibrillin-microfibrils: (a) 7 d after keratinocyte grafting; (b) 1 mo after keratinocyte grafting; (c and d) 17 mo after keratinocyte grafting; (d) represents a high power view of the region designated by the square in (c). (e) Papillary dermis after 24 mo; (f) deep reticular dermis after 24 mo. There is discordant distribution of both antigens on blood vessels (e) and m. arrector pili which are positive for latent TGF-β1 only. Scale bar: 10 μm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

7. Figure 6
Plasmin removes β-LAP from cutaneous microfibrils. Cryosections of normal human skin were immunostained for TGF-β1 precursor. (a) Untreated epidermis and papillary dermis; (c) untreated reticular dermis and a m. arrector pili; (b) cryosections treated with 0.1 U plasmin per ml for 1h prior to immunostaining; (d) comparable region of the reticular dermis after plasmin treatment; (e, f) fibrillin staining confirms absence of conspicuous damage to the microfibrillar apparatus by plasmin. Scale bars: 50 μm.
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions

8. Figure 7
Schematic drawing of microfibril-associated latent TGF-β1. Small latent TGF-β is composed of TGF-β dimer bound noncovalently to its prodomain dimer (β-LAP). Large latent TGF-β is formed from small latent TGF-β and LTBP. LTBP associates with β-LAP by a disulfide bond, and to microfibrils via an unknown mechanism (Nunes et al. 1997). A proteinase-sensitive hinge in LTBP beween the contact region with the ECM and the TGF-β binding domains is highlighted. Cleavage at this protease-sensitive region results in the release of latent TGF-β from microfibrils, whereas the cleavage of β-LAP results in the generation of active TGF-β
(modified fromSaharinen et al. 1996).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 1998 The Society for Investigative Dermatology, Inc Terms and Conditions