SPINK5 and Netherton Syndrome: Novel Mutations, Demonstration of Missing LEKTI, and Differential Expression of Transglutaminases

Abstract

Netherton syndrome (NTS) is an autosomal recessive congenital ichthyosis featuring chronic inflammation of the skin, hair anomalies, epidermal hyperplasia with an impaired epidermal barrier function, failure to thrive and atopic manifestations. The disease is caused by mutations in the SPINK5 gene encoding the serine proteinase inhibitor lympho-epithelial Kazal-type inhibitor (LEKTI). Sequence analyses of SPINK5 in seven NTS patients from five different families allowed us to identify two known and three novel mutations all creating premature termination codons. We developed a monoclonal antibody giving a strong signal for LEKTI in the stratum granulosum of normal skin and demonstrated absence of the protein in NTS epidermis. Immunoblot analysis revealed presence of full length LEKTI and of LEKTI cleavage fragments in normal hair roots, whereas in NTS hair roots LEKTI and its cleavage products were completely missing. Transglutaminase1 activity was present throughout almost the entire suprabasal epidermis in NTS, whereas in normal skin it is restricted to the stratum granulosum. In contrast, immunostaining for transglutaminase3 was absent or faint. Moreover, comparable with the altered pattern in psoriatic skin the epidermis in NTS strongly expressed the serine proteinase inhibitor SKALP/elafin and the anti-microbial protein human beta-defensin 2. These studies demonstrate LEKTI deficiency in the epidermis and in hair roots at the protein level and an aberrant expression of other proteins, especially transglutaminase1 and 3, which may account for the impaired epidermal barrier in NTS.

Elastase 2 is expressed in human and mouse epidermis and impairs skin barrier function in Netherton syndrome through filaggrin and lipid misprocessing

“…SLPI and SKALP are thus potential physiological inhibitors of ELA2 in the skin, which is in agreement with their specific capacity to inhibit members of the elastase family (31,32). High expression of SKALP has been reported in NS patient skin in response to skin inflammation and recurrent infections (14,33). However, SKALP overexpression is not sufficient to counteract ELA2 hyperactivity, as revealed by in situ zymography on patient skin sections.…”

“…We have previ-ously identified serine protease inhibitor kazal-type 5 (SPINK5) as the defective gene in NS (7). The vast majority of causative mutations reported to date lead to the complete absence of the encoded protein, LEKTI (7,(12)(13)(14). LEKTI is expressed in the SG of the epidermis, and we showed that bioactive LEKTI fragments inhibit 3 major proteases involved in SC desquamation, kallikreins (KLK) 5, 7, and 14 (15).…”

Inherited ichthyoses/generalized Mendelian disorders of cornification

“…42,43 Immunohistochemical staining confirms FLG deficiency in IV (Figures 1l and m) 44 and reveals absent or reduced expression of lympho-epithelial kazal-type-related inhibitor and SNAP29 in NS and CEDNIK, respectively. [45][46][47] In addition, light microscopy of hair shafts demonstrates 'bamboo hairs' (trichorrhexis invaginata) in NS (Figure 1j) 48 and polarization microscopy of TTD patient hairs reveals a tiger-tail pattern that corresponds to the diagnostic low sulfur protein content (Figure 1i). 49,50 Electron microscopy.…”

Revised nomenclature and classification of inherited ichthyoses: Results of the First Ichthyosis Consensus Conference in Sorèze 2009

“…219,220 Special immunohistochemical procedures can be combined, eg, to confirm filaggrin deficiency in IV, 202,221 or demonstrate absent or reduced expression of LEKTI that supports the diagnosis of NS. [222][223][224] To screen for TGase-1 deficiency in ARCI unfixed cryostat sections are used for the enzyme activity assay. 225,226 Alternatively, superficial SC material can be subjected to a SDS heating test that visualizes absent cross-linked envelopes in TGase-1 deficiency.…”