Amyloid P components in normal human skin and skin with lesions

Amyloid P component (AP) is a glycoprotein which is found in tissue deposits of all types of amyloid and is identical to and derived from serum amyloid P component (SAP). SAP binds in a calcium-dependent fashion to various ligands, such as agarose, desoxyribonucleic acid, fibronectin, C4-binding protein, glycosaminoglycans and isolated amyloid fibrils. Tissue AP (TAP) is also a constituent of the normal human renal glomerular basement membrane and is, in adult humans, invariably associated with elastic fiber microfibrils in connective tissue throughout the body, including that of blood vessels. In normal human skin anti-AP antibody binding was localized to the microfibrils of oxytalan fibers in the papillary dermis and to the peripheral microfibrillar mantle of elaunin and mature elastic fibers in the reticular dermis. Since SAP binds to fibronectin and glycosaminoglycans, which in turn bind to collagen fibers, TAP on elastic fiber microfibrils may play an important role in the maintenance of the normal dermal architecture and in dermo-epidermal adhesion. Under pathological conditions, AP is found in all forms of cutaneous amyloidosis, including primary localized cutaneous amyloid (PLCA); it is also detectable on keratin bodies, which represent precursor structures for PLCA. The association of AP with elastic fiber microfibrils and amyloid fibrils and their close anatomical relationship in vivo may reflect the significance of AP in the deposition of cutaneous amyloid.     

Amyloid P component binds to keratin bodies in human skin and to isolated keratin filament aggregates in vitro

Dermal keratin bodies, consisting mainly of keratin intermediate filament aggregates (KIFA) coated with IgM anti-KIF autoantibodies, are present in normal human skin and occur in increased quantities in certain skin diseases. Keratin bodies are normally rapidly removed, but in primary localized cutaneous amyloidosis (PLCA) they are converted by an unknown mechanism to amyloid. Amyloid P component (AP), a glycoprotein identical to, and derived from, the normal plasma protein serum amyloid P component (SAP), is present in all forms of amyloid including PLCA. We investigated the interaction between SAP, keratin bodies, and KIFA. Immunofluorescence staining of normal skin using fluoresceinated anti-SAP and rhodamine-conjugated anti-IgM, or AE-1/AE-3 anti-keratin antibodies followed by Texas Red-conjugated anti-mouse immunoglobulin, showed that 52% +/- 4 (mean +/- sem, n = 6) of keratin bodies bound anti-SAP. Similar findings were present in a biopsy from a patient with lichen planus. Isolated KIFA, prepared by 8M urea extraction of normal human epidermis or cultured keratinocytes, were preincubated with normal human serum as a source of SAP and then stained with fluoresceinated anti-SAP. Bright fluorescence seen when the incubation medium contained Ca++ was absent in the presence of ethylenediamine tetraacetic acid. Specific Ca++-dependent binding of SAP to KIFA was confirmed using immunoblotting. Binding of SAP to KIFA did not prevent their degradation following exposure to trypsin or alpha-chymotrypsin. Similarly, partial enzymatic digestion of KIFA did not abrogate their ability to bind SAP. Our findings, that SAP is associated with keratin bodies in skin and exhibits Ca++-dependent binding to KIFA in vitro, identify keratin filaments as a newly recognized ligand for SAP.     

Amyloidogenesis in organ-limited cutaneous amyloidosis: an antigenic identity between epidermal keratin and skin amyloid

Epidermal keratin was extracted and antibody against this protein was produced in rabbits. Various forms of organ-limited cutaneous amyloidosis (lichenoid, macular, and nodular amyloidosis, and basal cell epithelioma) and primary systemic amyloidosis were immunohistochemically examined to test the identity between epidermal keratin and skin amyloid. Amyloids in lichenoid and macular amyloidoses, and in basal cell epithelioma had an identical antigenicity with epidermal keratin, whereas amyloids in nodular amyloidosis and systemic amyloidosis did not have this identity. In addition, amyloid in lichen amyloidosis contained disulfide bonds as in keratin. Connective tissue components including filaments of fibroblasts and vascular endothelial cells did not react with this antikeratin antibody. It was concluded that at least some of the amyloid substance in organ-limited cutaneous amyloidosis is derived from degenerated epidermal keratinocytes through filamentous degeneration or apoptosis.     

Anti-keratin filament autoantibodies

IgG anti-keratin-filament autoantibodies occur in all human sera and reach high titres in the sera of patients with various cutaneous and non-cutaneous diseases. At indirect immunofluorescence, these autoantibodies may, depending on their titre, appear as 'upper-cytoplasmic' antibodies or 'stratum-corneum' antibodies. In vitro, IgG anti-keratin-filament autoantibodies significantly enhance (as opsonins) the phagocytosis of keratin-filament aggregates by human monocytes and polymorphonuclear leucocytes. They may therefore, also in vivo, play a role in the removal of insoluble extracellular keratin-filament aggregates generated by necrotic or apoptotic cell death. Keratin (Civatte) bodies and deposits of localized cutaneous amyloid are examples of such keratin-filament aggregates that are found in various skin diseases but are also regularly present in large amounts in normal human skin. IgM anti-keratin-filament autoantibodies, which may, at indirect immunofluorescence, appear as 'general cytoplasmic' antibodies, also occur in all human sera and reach high titres in the sera of patients with various diseases. The regular presence of IgM in keratin bodies or deposits of localized cutaneous amyloid therefore probably represents the binding of IgM anti-keratin-filament autoantibodies to their respective antigens. To what extent these in vivo-bound autoantibodies prevent any further damaging reaction by the immune system in response to the liberation of keratin-filament autoantigen is as yet unclear.     

Ultrastructural localization of amyloid P component in primary localized cutaneous amyloidosis

Amyloid P component (AP) was specifically localized to dermal amyloid deposits in the papular and nodular variants of primary localized cutaneous amyloidosis by an immunoperoxidase technique using an antibody to serum amyloid P component (SAP). Specific staining with anti-SAP of elastic fibre microfibrils which has previously been observed in normal skin, was also present and was noted in close proximity 10 deposits of amyloid material. AP associated with normal elastic fibre microfibrils may be involved in the deposition of amyloid fibrils in vivo.     

Epidermal origin of the amyloid in localized cutaneous amyloidosis

A case of localized cutaneous amyloidosis which developed after a lichen planus-like skin reaction is reported. The amyloid consisted of amyloid fibrils enveloped by heparan sulphate granules. These amyloid fibrils reacted to anti-human keratin antibody, indicating an epidermal origin for the fibrils.     

The molecular skin pathology of familial primary localized cutaneous amyloidosis

Please cite this paper as: The molecular skin pathology of familial primary localized cutaneous amyloidosis. Experimental Dermatology 2010; 19: 416–423. Abstract: Familial primary localized cutaneous amyloidosis (FPLCA) is an autosomal dominant disorder associated with chronic itching and skin lichenification. In lesional skin, there are apoptotic basal keratinocytes and deposits of amyloid material on degenerate keratin filaments in the upper dermis. The genetic basis of FPLCA involves mutations in the OSMR and IL31RA genes but the disease pathophysiology is not fully understood. In this study, we identified new pathogenic heterozygous missense mutations in the OSMR gene (p.Val631Leu and p.Asp647Tyr) in two Dutch FPLCA families. We then compared gene expression profiles between FPLCA lesional skin (n = 4) and site‐matched control skin (n = 6). There was twofold or greater upregulation of 34 genes and downregulation of 43 genes. Most changes in gene expression (verified by quantitative RT‐PCR) reflected alterations in epidermal differentiation and proliferation consistent with lichenification, but we also noted a reduction in several interfollicular keratinocyte stem cell markers in FPLCA skin. Differences in gene expression were also noted for proteins involved in apoptosis and nerve conduction. Collectively, this study expands the molecular basis of FPLCA and provides new insight into the skin pathology of this condition.     

Vitronectin shows complement-independent binding to isolated keratin filament aggregates

Keratinocyte cell death, whether produced by skin disease or by physiologic apoptosis in normal skin, may result in formation of dermal keratin bodies, consisting mainly of keratin intermediate filament aggregates. Vitronectin, a multifunctional plasma and tissue glycoprotein, which inhibits the complement membrane attack complex and promotes cell attachment and spreading, is, like amyloid P component, associated with keratin bodies in vivo. To investigate a potential role for vitronectin in the removal of keratin bodies, we studied the interaction of vitronectin with keratin intermediate filaments in normal human skin and in Hep-2 cells, as well as with isolated keratin intermediate filament aggregates in vitro. Following pre-incubation of skin sections and Hep-2 cells with normal human serum (as a source of vitronectin), cytoplasmic staining of keratinocytes and of cytoskeletal filaments in Hep-2 cells was observed by immuno-fluorescence staining with polyclonal and monoclonal anti-vitronectin antibodies. Vitronectin binding to keratin intermediate filament aggregates extracted from normal human epidermis was demonstrated by immunofluorescence and by immunoblotting, and was not dependent on complement activation, because it occurred even when heat-inactivated human serum or C4-deficient serum was used as a source of vitronectin. Amyloid P component shows Ca++- dependent binding to keratin intermediate filament aggregates. does not involve amyloid P component because it occurred when binding of the latter protein was inhibited by EDTA buffer. Moreover, purified vitronectin also bound to keratin intermediate filament aggregates in immunofluorescence studies. Vitronectin binding to keratin intermediate filaments may play a role both in limiting complement-mediated tissue damage (because keratin bodies may activate complement) and in promoting removal of keratin bodies by fibroblasts and/or macrophages.     

Amyloid K

Electron-microscopic, histochemical and immunological studies have shown that the deposits of primary and secondary localized cutaneous amyloid (amyloid K) consist mainly of keratin filament material. The amyloid P component is the second most significant constituent. Pathogenetically, defective keratinocyte apoptosis is the most likely mechanism of generation for amyloid K. Apoptosis is a distinct mode of cell death according to which individual basal keratinocytes disintegrate in physiological and pathological conditions to form membrane-bound "apoptotic" bodies consisting mainly of keratin filament aggregates. These lose their protective membrane and drop into the upper dermis, where they present as keratin (Civatte, cytoid) bodies, which are regularly coated with immunoglobulins. Overproduction of keratin bodies and/or a defect in their degradation by enzymatic digestion or phagocytosis by macrophages and fibroblasts, followed by conversion into amyloid fibrils, may be responsible for the generation of deposits of amyloid K.     

Immunohistochemical studies on vitronectin in elastic tissue disorders, cutaneous amyloidosis, lichen ruber planus and porphyria

Vitronectin, identical with serum-spreading factor and S-protein of complement, is a glycoprotein present in both plasma and tissue. It stimulates cell adhesion and spreading and affects the complement and coagulation pathways. Vitronectin immunoreactivity was recently found in conjunction with dermal and renal elastic fibres, in renal amyloid deposits in cases of AL- and AA-amyloidosis, and in sclerotic glomerular lesions. Skin specimens from lesions of patients with selected skin diseases were investigated with an avidin-biotin peroxidase technique using both monoclonal and polyclonal anti-vitronectin antibodies and an alkaline phosphatase anti-alkaline phosphatase technique using monoclonal anti-vitronectin antibodies. Vitronectin immunoreactivity was found in association with the abnormal elastic tissue in solar elastosis and pseudoxanthoma elasticum. It was also found in conjunction with dermal amyloid deposits in primary localized cutaneous amyloidosis and in Civatte bodies in cases of lichen ruber planus. In cases of erythropoietic protoporphyria and porphyria cutanea tarda, hyaline perivascular deposits also demonstrated positive vitronectin immunoreactivity. The presence of vitronectin immunoreactivity not only in normal and degenerated elastic fibres but also in various pathological tissue deposits suggests that vitronectin occurs both in elastic fibres and in different types of abnormal protein deposits.     

Immunohistochemical demonstration of amyloid P component in skin of normal subjects and patients with cutaneous amyloidosis

The presence of amyloid P component (AP) in dermal deposits of cutaneous atnyloidosis was demonstrated by a direct immunofluorescence technique using an antibody to serum amyloid P component (SAP). AP was also shown, for the first time, to be a constituent of normal human skin. It was present at the periphery of dermal elastic tissue fibres, in basement membranes of dermal blood vessels and surrounding eccrine sweat glands but was absent from the dermo-epidermal basement membrane. The staining pattern in cutaneous amyloidosis was morphologically distinctive and readily distinguishable from staining of thickened vascular basement membranes in porphyria. Itnmunofluorescence with anti-SAP is simple and specific and may become the procedure of choice in the differential diagnosis of amyloidosis.     

Elastic fibres in normal and sun-damaged skin: an immunohistochemical study

Sun-exposed and sun-protected skin obtained at post mortem from the nape of the neck in 14 subjects was immunostained using antisera to elastin, lysozyme, amyloid P component, and the plasma protease inhibitors alpha-I antitrypsin, alpha-I antichymotrypsin and alpha-2 macroglobulin. Both the normal elastic fibres in sun-protected skin, and elastosis in sun-exposed skin were positively immunostained for elastin, lysozyme and amyloid P component. Collagen fibres were unstained. No immunostaining of normal elastic fibres or elastosis in the skin was obtained with antisera to alpha-I antitrypsin, alpha-I antichymotrypsin or alpha-2 macroglobulin. It was concluded that the elastosis in sun-exposed skin does contain elastic fibres. The absence of immunostaining for plasma protease inhibitors probably indicates that the elastic material is mature, and not newly-formed.     

Deposition of complement CIq in primary localized cutaneous amyloidosis

Deposition of immunoglobulins and complement components was studied in twelve cases of primary localized cutaneous amyloidosis by direct immunofiuorescence microscopy. CIq was detected in amyloid islands in ten cases but immunoglobulins and C3 were found in only a few cases. It is suggested that CIq may be one of the nonepidermal components of cutaneous amyloid.     

Complement in skin diseases

Complement is one of the most important mechanisms of natural resistance preventing infections in humans and animals. It is actively involved in the pathogenesis of several diseases, including skin diseases, characterized by the presence of autoantibodies, foreign microorganisms, altered tissue cells, and the presence of mannan. Complement is intended to kill invading microorganisms but it can also destroy the organism's own damaged or altered cells. It is characterized by vigorous activity and is also potentially harmful for the host if triggered in its own body. This review discusses the significance of complement activation for emerging skin diseases and highlights the importance of serological laboratory tests for the detection of complement system activity alterations in skin diseases such as pemphigus vulgaris, bullous pemphigoid, herpes gestationis, dermatitis herpetiformis, porphyria, urticaria, angioedema, cutaneous vasculitis, systemic lupus erythematosus, partial lipodystrophy, lichen planus, xeroderma pigmentosum, psoriasis, and recurrent cutaneous infections. Finally, we draw attention to the current potential for treating these diseases with complement inhibitors.     

Immunohistochemical studies of amyloid P component distribution in normal human skin

The distribution of amyloid P component (AP) in normal human skin was investigated by a light and electron microscopic immunoperoxidase technique, using antibodies to serum amyloid P component (SAP). AP, or an immunologically cross-reactive protein, was found to be specifically localized to the microfibrils of papillary oxytalan fibers and to the peripheral microfibrillar mantle surrounding the elastin core of mature elastic fibers in the reticular dermis; collagen fibers were not stained with anti-SAP. AP was not detected in the dermal-epidermal basement membrane or in the basement membranes surrounding dermal papillary blood vessels and eccrine structures. These findings, which establish the detailed distribution of normal tissue AP in the skin, provide a basis for further studies of the function and behavior of this protein in health and disease.     

Immunofluorescence and histochemical studies of localized cutaneous amyloidosis

Lichen amyloidosus (LA) and macular amyloidosis (MA) are two forms of localized cutaneous amyloidosis in which the amyloid occurs as larger and smaller deposits respectively in the papillary dermis. The histogenesis of the amyloid of these conditions is unknown. By using an indirect immunofluarescence technique we showed that LA and MA do not react with antibodies against different previously characterized amyloid fibril proteins. These results indicate that the amyloid of LA and MA is different from other known types of amyloid. Protein AP, which was demonstrated in amyloid of MA and LA, is known to be present in all forms of amyloid and is of unknown significance. Antiserum against keratin did not react with the larger homogeneous amyloid bodies, but showed a weak reaction with some small deposits. Histochemical staining failed to show keratin in any of the tissues containing LA or MA.     

A case of secondary cutaneous amyloidosis: epidermal keratinocytes produce amyloid in the cytoplasm

A case of secondary localized cutaneous amyloidosis associated with a seborrheic keratosis is reported. Amyloid was observed both in the stroma and in the tumor. Light and electron microscopy revealed amyloid within the cytoplasm of the tumor cells. This intracytoplasmic amyloid was seen in basaloid cells or in the border areas between basaloid cells and squamous cells, but it was not seen in squamous cells. The amyloid was positive for anti-keratin antibody and contained disulfide bonds. It is suggested that either abnormal keratinization or the degeneration of basaloid cells produced abnormal keratin proteins that formed this amyloid.     

Nylon cloth macular amyloidosis

In primary localized cutaneous amyloid, deposition of amyloid is confined to the skin without the involvement of any internal organs. Amyloid deposition in the skin is often scanty, and electron microscopy may be needed to confirm the presence of the typical amyloid fibrils. There have been several case reports of cutaneous amyloidosis associated with friction or rubbing of the skin. We report a case of primary localized cutaneous amyloid associated with the habitual use of a nylon cloth.     

Exacerbated and prolonged allergic and non-allergic inflammatory cutaneous reaction in mice with targeted interleukin-18 expression in the skin

Interleukin 18 induces both T helper 1 and T helper 2 cytokines, proinflammatory cytokines, chemokines, and IgE and IgG1 production. A role of interleukin 18 in inflammatory cutaneous reactions is still unclear, however. Here we generated keratin 5/interleukin 18 transgenic mice overexpressing mature murine interleukin 18 in the skin using a human keratin 5 promoter. In the contact hypersensitivity model, trinitrochlorobenzene elicited a stronger ear swelling in keratin 5/interleukin 18 transgenic mice compared with control littermate wild-type or immunoglobulin/interleukin 18 transgenic mice in which mature interleukin 18 was expressed by B and T cells under the control of the immunoglobulin promoter. Application of an irritant, croton oil, induced stronger and more sustained ear swelling in keratin 5/interleukin 18 transgenic mice than in immunoglobulin/interleukin 18 transgenic or wild-type mice. Repetitive topical application (weekly for six consecutive weeks) of trinitrochlorobenzene to their ears also elicited a stronger cutaneous inflammation in keratin 5/interleukin 18 transgenic mice than seen in immunoglobulin/interleukin 18 transgenic or wild-type mice. After these six trinitrochlorobenzene applications, the expression of interferon-gamma, interleukin-4, and CCL20 mRNA in the ear tissue was increased and dermal changes, such as acanthosis and eosinophilic, neutrophilic, and mast cell infiltration, were greater in keratin 5/interleukin 18 transgenic mice than in wild-type mice. Furthermore, the repetitive application elicited a significant increase in serum IgE levels and the number of B cells in the draining lymph node in keratin 5/interleukin 18 transgenic mice. These results suggest that overexpression of interleukin 18 in the skin aggravates allergic and nonallergic cutaneous inflammation, which is accompanied by high expression of T helper 1 and T helper 2 cytokines and chemokines in the skin.     

Primary Localized Cutaneous Nodular Amyloidosis Following Local Trauma

Primary localized cutaneous nodular amyloidosis (nodular amyloidosis) is a rare and distinct type of amyloidosis, in which amyloid L deposition is limited to the skin and typically manifested as a tumefactive nodule on the acral sites. However, the definite cause of nodular amyloidosis is still unknown. Although it is relatively well known that the amyloid deposits in nodular amyloidosis originate from immunoglobulin light chains secreted by local plasma cells, traumatic injury to the skin has rarely been recognized as a triggering factor of nodular amyloidosis. Herein, we present a case of a 50-year-old male patient with primary localized cutaneous nodular amyloidosis, which occurred after local trauma, and discuss the relationship between traumatic damage and dermal amyloid L deposition.