M. Peter Marinkovich

NF-kappaB blockade and oncogenic Ras trigger invasive human epidermal neoplasia.

The nuclear factor NF-κB and oncogenic Ras can alter proliferation in epidermis, the most common site of human cancer. These proteins are implicated in epidermal squamous cell carcinoma in mice, however, the potential effects of altering their function are uncertain. Whereas inhibition of NF-κB enhances apoptosis in certain tumours, blockade of NF-κB predisposes murine skin to squamous cell carcinoma. Because therapeutics inhibiting Ras and NF-κB pathways are being developed to treat human cancer, it is essential to assess the effects of altering these regulators. The medical relevance of murine studies is limited, however, by differences between mouse and human skin, and by the greater ease of transforming murine cells. Here we show that in normal human epidermal cells both NF-κB and oncogenic Ras trigger cell-cycle arrest. Growth arrest triggered by oncogenic Ras can be bypassed by IκBα-mediated blockade of NF-κB, generating malignant human epidermal tissue resembling squamous cell carcinoma. Human cell tumorigenesis is dependent on laminin 5 and α6β4 integrin. Thus, IκBα circumvents restraints on growth promotion induced by oncogenic Ras and can act with Ras to induce invasive human tissue neoplasia.

Laminin 332 in squamous-cell carcinoma

Basement membranes can be a barrier to tumour growth, but basement membrane molecules, including laminins, are also important autocrine factors produced by cancers to promote tumorigenesis. Many studies have shown the importance of laminin 332 (previously known as laminin 5) in this process, especially in squamous cell carcinoma. Through interactions with several cell-surface receptors (including α6β4 and α3β1 integrins, epidermal growth factor receptor and syndecan 1) and other basement membrane components (including type VII collagen), laminin 332 drives tumorigenesis through phosphatidylinositol-3 kinase (PI3K) and RAC1 activation, promoting tumour invasion and cell survival. The extracellular interactions of laminin 332 appear amenable to antibody-mediated therapies.

Autocrine laminin-5 ligates α6β4 integrin and activates RAC and NFκB to mediate anchorage-independent survival of mammary tumors

Invasive carcinomas survive and evade apoptosis despite the absence of an exogenous basement membrane. How epithelial tumors acquire anchorage independence for survival remains poorly defined. Epithelial tumors often secrete abundant amounts of the extracellular matrix protein laminin 5 (LM-5) and frequently express α6β4 integrin. Here, we show that autocrine LM-5 mediates anchorage-independent survival in breast tumors through ligation of a wild-type, but not a cytoplasmic tail–truncated α6β4 integrin. α6β4 integrin does not mediate tumor survival through activation of ERK or AKT. Instead, the cytoplasmic tail of β4 integrin is necessary for basal and epidermal growth factor–induced RAC activity, and RAC mediates tumor survival. Indeed, a constitutively active RAC sustains the viability of mammary tumors lacking functional β1 and β4 integrin through activation of NFκB, and overexpression of NFκB p65 mediates anchorage-independent survival of nonmalignant mammary epithelial cells. Therefore, epithelial tumors could survive in the absence of exogenous basement membrane through autocrine LM-5–α6β4 integrin–RAC–NFκB signaling.

Injection of genetically engineered fibroblasts corrects regenerated human epidermolysis bullosa skin tissue.

Current therapeutic strategies for genetic skin disorders rely on the complex process of grafting genetically engineered tissue to recipient wound beds. Because fibroblasts synthesize and secrete extracellular matrix, we explored their utility in recessive dystrophic epidermolysis bullosa (RDEB), a blistering disease due to defective extracellular type VII collagen. Intradermal injection of RDEB fibroblasts overexpressing type VII collagen into intact RDEB skin stably restored correctly localized type VII collagen expression in vivo and normalized hallmark RDEB disease features, including subepidermal blistering and anchoring fibril defects. This article was published online in advance of the print edition. The date of publication is available from the JCI website, http://www.jci.org.

α6β4 integrin regulates keratinocyte chemotaxis through differential GTPase activation and antagonism of α3β1 integrin

Growth factor-induced cell migration and proliferation are essential for epithelial wound repair. Cell migration during wound repair also depends upon expression of laminin-5, a ligand for α6β4 integrin. We investigated the role of α6β4 integrin in laminin-5-dependent keratinocyte migration by re-expressing normal or attachment-defective β4 integrin in β4 integrin null keratinocytes. We found that expression ofβ 4 integrin in either a ligand bound or ligand unbound state was necessary and sufficient for EGF-induced cell migration. In a ligand bound state, β4 integrin supported EGF-induced cell migration though sustained activation of Rac1. In the absence of α6β4 integrin ligation, Rac1 activation became tempered and EGF chemotaxis proceeded through an alternate mechanism that depended upon α3β1 integrin and was characterized by cell scattering. α3β1 integrin also relocalated from cell-cell contacts to sites of basal clustering where it displayed increased conformational activation. The aberrant distribution and activation ofα 3β1 integrin in attachment-defective β4 cells could be reversed by the activation of Rac1. Conversely, in WT β4 cells the normal cell-cell localization of α3β1 integrin became aberrant after the inhibition of Rac1. These studies indicate that the extracellular domain ofβ 4 integrin, through its ability to bind ligand, functions to integrate the divergent effects of growth factors on the cytoskeleton and adhesion receptors so that coordinated keratinocyte migration can be achieved.

Keratinocyte-Secreted Laminin 5 Can Function as a Transient Receptor for Human Papillomaviruses by Binding Virions and Transferring Them to Adjacent Cells

ABSTRACT Human papillomaviruses (HPVs) replicate only in the terminally differentiating epithelium of the skin and mucosa. While infection of basal keratinocytes is considered a requirement for permissive infection, it remains unclear whether virions can specifically target basal cells for adsorption and uptake following epithelial wounding. We present evidence that HPV binds specifically to laminin 5 (LN5), a component of the extracellular matrix (ECM) secreted by migrating and basal keratinocytes. HPV type 11 capsids colocalized with LN5 in the ECM secreted by vaginal keratinocytes. Binding of both virions and virus-like particles to purified LN5 and to the LN5-rich ECM secreted by cultured keratinocytes was effectively blocked by pretreatment with anti-LN5 antibodies. HPV capsid binding to human cervical mucosa sections included the basement membrane which contains LN5. Cultured keratinocytes expressing α6 integrin, a transmembrane protein known to bind LN5, were readily infected by virions preadsorbed to LN5-containing substrates, whereas mutant keratinocytes lacking α6 integrin were relatively resistant to infection via this route. These findings suggest a model of natural HPV infection in which proliferating keratinocytes expressing α6 integrin at the site of epithelial wounding might be targeted by virions adsorbed transiently to LN5 secreted by migrating keratinocytes.

Laminin-10 is crucial for hair morphogenesis

The role of the extracellular matrix in cutaneous morphogenesis is poorly understood. Here, we describe the essential role of laminin‐10 (α5β1γ1) in hair follicle development. Laminin‐10 was present in the basement membrane of elongating hair germs, when other laminins were downregulated, suggesting a role for laminin‐10 in hair development. Treatment of human scalp xenografts with antibodies to laminin‐10, or its receptor β1 integrin, produced alopecia. E16.5 Lama5 −/− mouse skin, lacking laminin‐10, contained fewer hair germs compared with controls, and after transplantation, Lama5 −/− skin showed a failure of hair germ elongation followed by complete hair follicle regression. Lama5 −/− skin showed defective basement membrane assembly, without measurable increases in anoikis. Instead, Lama5 −/− skin showed decreased expression of early hair markers including sonic hedgehog and Gli1, implicating laminin‐10 in developmental signaling. Intriguingly, treatment of Lama5 −/− skin with purified laminin‐10 corrected basement membrane defects and restored hair follicle development. We conclude that laminin‐10 is required for hair follicle development and report the first use of exogenous protein to correct a cutaneous developmental defect.

Integrin β4 Regulates Migratory Behavior of Keratinocytes by Determining Laminin-332 Organization

Whether α6β4 integrin regulates migration remains controversial. β4 integrin-deficient (JEB) keratinocytes display aberrant migration in that they move in circles, a behavior that mirrors the circular arrays of laminin (LM)-332 in their matrix. In contrast, wild-type keratinocytes and JEB keratinocytes, induced to express β4 integrin, assemble laminin-332 in linear tracks over which they migrate. Moreover, laminin-332-dependent migration of JEB keratinocytes along linear tracks is restored when cells are plated on wild-type keratinocyte matrix, whereas wild-type keratinocytes show rotation over circular arrays of laminn-332 in JEB keratinocyte matrix. The activities of Rac1 and the actin cytoskeleton-severing protein cofilin are low in JEB keratinocytes compared with wild-type cells but are rescued following expression of wild-type β4 integrin in JEB cells. Additionally, in wild-type keratinocytes Rac1 is complexed with α6β4 integrin. Moreover, Rac1 or cofilin inactivation induces wild-type keratinocytes to move in circles over rings of laminin-332 in their matrix. Together these data indicate that laminin-332 matrix organization is determined by the α6β4 integrin/actin cytoskeleton via Rac1/cofilin signaling. Furthermore, our results imply that the organizational state of laminin-332 is a key determinant of the motility behavior of keratinocytes, an essential element of skin wound healing and the successful invasion of epidermal-derived tumor cells.

Linear IgA bullous dermatosis

Linear immunoglobulin A (IgA) disease, also known as linear IgA bullous dermatosis (LAD), is a rare acquired subepidermal blistering disease characterized by continuous linear IgA deposits in the basement membrane zone (BMZ) when visualized on direct immunofluorescence microscopy (DIF). Linear IgA bullous dermatosis was first described in 1901 by Bowen, who saw 15 children with nonpruritic blistering eruptions mainly involving the genitalia; however, it was not recognized as a entity separate from dermatitis herpetiformes (DH) until 1979. In addition, a druginduced variant of LAD has been recognized in a minority of cases as well as several associations with various infections or chronic illnesses. In LAD, autoantibodies recognize and bind multiple antigens associated with the BMZ, and thus it constitutes a heterogeneous disease with respect to ultrastructural localization of target antigens.

Necrolytic migratory erythema without glucagonoma in patients with liver disease

Abstract Background: Necrolytic migratory erythema (NME) is an uncommon inflammatory dermatosis with a distinct clinical and histologic appearance. NME is usually associated with glucagonoma. Only a few cases of NME in the absence of glucagonoma have been previously reported. Objective: We sought to understand further the pathogenesis of NME by analyzing data from three patients. Methods: Three patients were examined both clinically and histopathologically. Results: Each patient had an extensive erythematous scaling eruption in intertriginous, perioral, and acral areas, and a markedly red, smooth tongue. Skin biopsy specimens showed confluent parakeratosis, epidermal pallor, papillary edema, and a lymphohistiocytic infiltrate. Two patients had alcoholic liver disease and one had liver dysfunction as a result of hemochromatosis. Serum albumin level was depressed, and liver enzyme values were increased in all three patients. Glucagonoma was undetectable in these patients. Conclusion: In the absence of glucagonoma, hepatocellular dysfunction and hypoalbuminemia appear to be the most common factors associated with NME.BACKGROUND Necrolytic migratory erythema (NME) is an uncommon inflammatory dermatosis with a distinct clinical and histologic appearance. NME is usually associated with glucagonoma. Only a few cases of NME in the absence of glucagonoma have been previously reported. OBJECTIVE We sought to understand further the pathogenesis of NME by analyzing data from three patients. METHODS Three patients were examined both clinically and histopathologically. RESULTS Each patient had an extensive erythematous scaling eruption in intertriginous, perioral, and acral areas, and a markedly red, smooth tongue. Skin biopsy specimens showed confluent parakeratosis, epidermal pallor, papillary edema, and a lymphohistiocytic infiltrate. Two patients had alcoholic liver disease and one had liver dysfunction as a result of hemochromatosis. Serum albumin level was depressed, and liver enzyme values were increased in all three patients. Glucagonoma was undetectable in these patients. CONCLUSION In the absence of glucagonoma, hepatocellular dysfunction and hypoalbuminemia appear to be the most common factors associated with NME.