Psoriasis is a common chronic autoimmune and inflammatory skin disease that affects 2.7% of the world’s population (Naldi, 2004). Although it is a disease whose etiology and pathogenesis remain unclear, many factors are involved (Krueger, 2002). It is characterized by an immune-related pathogenesis, a genetic background that may be triggered by several environmental factors, and T-cell-mediated cytokine production (Mercuri and Naldi, 2010; Park and Lee, 2010).
Psoriasis is unique because it represents excessive keratinocyte proliferation with incomplete differentiation. This aberrant epidermal growth results in the loss of normal epidermal barrier function. However, the underlying mechanism of increased keratinocyte proliferation remains controversial (Zhou et al., 2003). Keratinocytes interact with each other through intercellular junctions to regulate cellular shape, proliferation, and the passage of ions and molecules through the paracellular pathway. Their proliferation outside the basal layer suggests an alteration in cell–cell interactions in active psoriasis (Li et al., 2008a, 2008b).
Adherens junctions (AJs) are required for the establishment and maintenance of epithelial layers, mediating intercellular adhesion, sensing the presence of neighboring cells, and anchoring the actin cytoskeleton to protein complexes and the membrane in this region of the cell (Perez-Moreno et al., 2003). E-cadherin is one of the major classical AJ expressed in the epidermis of skin. It is a cell adhesion transmembrane glycoprotein that mediates Ca2-dependent intercellular cellular adhesion (Young et al., 2003). It is important for epidermal keratinocytes’ intercellular adherence and skin differentiation (Tinkle et al., 2004). The aim of this study was to evaluate the immunohistochemical expression of E-cadherin in Egyptian psoriasis patients to clarify its possible role in the pathogenesis of this disease.
Patients and methods
Patients and controls
This study was carried out on 25 patients presenting with psoriasis. The diagnosis was made on clinical and histopathologic bases and all patients had received neither topical nor surgical treatment for at least 2 weeks before skin biopsy. For comparison purposes, 10 specimens from age-matched and sex-matched apparently normal volunteers during plastic surgery were included in the study as controls. Both patients and control participants were selected randomly from the Outpatient Dermatology Clinic, Menoufia University Hospital, during the period between June 2011 and January 2013.
Complete assessment of history
All patients were subjected to a complete assessment of history, with a special focus on the duration of the disease, dermatological examination, and registration of Psoriasis Area and Severity Index (Schmitt and Wozel, 2005).
Skin biopsies were taken local anesthesia from all cases (lesions) and controls after obtaining a written informed consent. All biopsies were processed in the Pathology Department, Faculty of Medicine, Menoufia University, where they were fixed in 10% neutral-buffered formalin, dehydrated in ascending grades of ethanol, followed by immersion in xylene, and then embedded in paraffin. Three 5-mm-thick sections from each block were taken. One was stained with hematoxylin and eosin for routine histopathological examination. The other two sections were mounted on Superfrost Plus slides and stored at room temperature: one to be stained immunohistochemically for E-cadherin and the other as a negative control.
Examination of the hematoxylin and eosin-stained sections
Histologic examination of the lesional skin was performed for identification and histopathological confirmation of the clinical diagnosis of psoriasis. This was done by evaluating epidermal and dermal changes:
- Evaluation of the epidermis in terms of acanthosis, parakeratosis, granular cell layer, Munro’s microabscess, and suprapapillary epidermis.
- Evaluation of the dermis in terms of the degree of angiogenesis and perivascular inflammatory infiltrate.
Immunohistochemical staining for E-cadherin
Mouse monoclonal antibody raised against E-cadherin was used. It was received as 7 ml ready to use (Cat. #MS-9470-R7; Lab Vision, Fremont, California, USA). The detection kit was the ultravision detection system antipolyvalent horse radish peroxidase/diaminobenzidine (ready to use, Cat. #TP-015-HD; Lab Vision). A chromogenic reaction was carried out with diaminobenzidine substrate and the sections were counterstained with Mayer’s hematoxylin. Human colonic carcinoma was used as a positive control. Negative controls obtained by substitution of primary antibodies with rat nonimmune serum were included in the staining procedure.
Positive E-cadherin expression was assigned when 10% or more of keratinocytes showed brown membranous staining. The staining intensity was evaluated using a four-tier grading system (0, negative; 1, weak; 2, moderate; and 3, strong staining intensity). For each intensity score, the percentage of cells with that score was estimated. H score=1×% of mildly stained cells+2×% of moderately stained cells+3×% of strongly stained cells (Bilalovic et al., 2004).
Data were collected, tabulated, and analyzed statistically using a personal computer with the statistical package for the social sciences (SPSS, version 16; SPSS Inc., Chicago, Illinois, USA). χ2-Test and Fisher’s exact tests were used for comparison between qualitative variables. Mann–Whitney U-test and Student’s t test were used for comparison between quantitative variables. P value 0.05 or less was considered statistically significant.
Clinical and histopathological data of selected psoriasis cases are presented in Table 1.
E-cadherin expression in control cases
All normal control cases showed diffuse positive E-cadherin expression in all layers of the epidermis (Fig. 1). The intensity of expression was mild in one (10%) case, moderate in five (50%) cases, and strong in four (40%) cases (Table 2).
E-cadherin expression in psoriasis cases
The diseased skin showed positive E-cadherin expression in 18/25 (72%) cases. Only 5/18 (27.8%) cases showed diffuse staining of all layers of the epidermis. The remaining 13/18 (72.2%) cases showed staining of the spinous layer, with absent expression in basal and granular layers (Fig. 2). The intensity of expression was mild in three (16.7%) cases (Fig. 3), moderate in 11 (61.1%) cases (Fig. 4), and strong in four (22.2%) cases (Fig. 5) (Table 2).
Comparison between cases and control in E-cadherin expression
Normal skin differed from lesional psoriasis in the degree and pattern of epidermal E-cadherin expression. There was a tendency of E-cadherin to be expressed in keratinocytes of normal skin than in psoriasis skin (P=0.08). Normal skin showed higher H scores of E-cadherin expression (P=0.01) in addition to its diffuse staining in all epidermal layers in normal skin compared with the focal expression in the spinous layer only in lesional skin (P<0.001) (Table 2).
Correlation between E-cadherin expression and the clinical and histopathological parameters of psoriasis cases
We did not find any significant relationship between positive and negative E-cadherin expression in keratinocytes and the parameters studied. However, lower H scores of E-cadherin expression were associated significantly with thinning of the suprapapillary epidermis (P=0.007) and marked dermal angiogenesis (P=0.05) (Tables 3 and 4).
Psoriasis is a chronic inflammatory disease with heterogeneous clinical manifestations ranging from limited to very extensive disease (Neimann et al., 2006; Park and Lee, 2010). It responds poorly to topical and/or systemic treatment (Lemini-López et al., 2006). The most characteristic change in psoriasis is the markedly increased, persistent, keratinocyte proliferation. Molecular alterations in the epidermal barrier function and mechanisms underlying the perturbed state of proliferation and differentiation in psoriatic epidermis remain poorly understood (Chung et al., 2005). AJs that are critical for normal epidermal barrier function as well as overall homeostasis have been shown to be altered in psoriasis (Pummi et al., 2001; Perez-Moreno et al., 2003; Zhou et al., 2003; Harhaj and Antonetti, 2004).
E-cadherin represents the main AJ responsible for intercellular adhesion of keratinocytes. It plays a critical role in keratinocytes adherence, epidermal thickness, and differentiation (Young et al., 2003; Tinkle et al., 2004). Loss of E-cadherin could therefore lead to progressive hyperproliferation in the epidermis (Christopher et al., 2004).
In the current work, normal skin tissues showed diffuse uniform E-cadherin expression within all layers of the epidermis. In agreement with our results, several studies reported that the stain of E-cadherin was detected uniformly within all layers of the normal epidermis at the sites of cell–cell junctions (Hampton et al., 2007; Li et al., 2008a, 2008b).
In the present study, marked downregulation of E-cadherin expression was detected in the granular and basal layers of psoriasis lesional skin compared with normal skin (P<0.001). Furthermore, normal skin showed higher H scores of E-cadherin expression than lesional skin (P=0.01). These results are similar to those observed by Chung et al. (2005), and Li et al. (2008a, 2008b), who reported that there was a downregulation of E-cadherin in the granular layer, upper spinous, and basal layers, with sparing of the lower spinous layer of psoriasis lesional tissue. They hence assumed that the breakdown of AJs in the psoriatic epidermis is probably involved in the hyperproliferation of keratinocytes in psoriasis. Zhou et al. (2003) and Lemini-López et al. (2006) believed that the distribution of E-cadherin in psoriasis is the same as in normal skin, and the difference is that the protein shows an irregular and tortuous pattern in the epidermis of the lesional skin. They suggested that in these cases, alterations in the organization of AJ proteins could contribute toward modifying interactions between neighboring cells, leading to inadequate function of the epithelial layers. However, Tsuji et al. (2001) reported that the distribution of E-cadherin is the same as in normal skin and hence cannot play an important role in the pathogenesis of psoriasis.
In agreement with the assumption that defective expression of E-cadherin in basal and granular layers may simply reflect the role of this adhesion molecule in the pathogenesis of psoriasis, several studies found that loss of function of the epithelial barrier that accompanies alterations of epithelial junctional molecules such as E-cadherin is associated with disrupted keratinocyte differentiation, hyperproliferation, and cutaneous epithelial inflammatory processes typified by psoriasis (Pummi et al., 2001; Zhou et al., 2003). E-cadherin mutant mice showed acanthosis, hyperkeratosis, widening of basal cell intercellular spaces, and induction of K6 expression, findings that are characteristically observed in psoriatic epidermis (Stoler et al., 1988; Pittelkow, 1998; Bhawan et al., 2004). In addition, Chung et al. (2005) found that psoriatic epidermis may contain mitogenic molecules such as amphiregulin, which induces psoriatic inflammation and contributes toward the pathogenesis of psoriasis by reducing levels of functional E-cadherin in the epidermis, resulting in an impaired junctional barrier and allowing transmigration of neutrophils into psoriatic epidermis. Furthermore, soluble levels of E-cadherin have been quantified in serum and were found to be increased in psoriasis vulgaris, correlating with disease severity. This finding may reflect increased proteolysis of cell surface E-cadherin in the epidermis, therefore reducing functional E-cadherin at the cell surface (Chung et al., 2005).
In the current work, a significant negative relationship was found between H scores of E-cadherin and thinning of suprapapillary epidermis (P=0.007) as well as dermal angiogenesis (P=0.05). Psoriasis is characterized by hyperproliferation and abnormal differentiation of epidermal keratinocytes, lymphocyte infiltration consisting mostly of T lymphocytes and various endothelial vascular changes in the dermal layer such as angiogenesis, dilatation, and high endothelial venule formation (El Darouti and Abdel Hay, 2010). Angiogenesis and vascular hyperpermeability are the result of increased production of vascular endothelial growth factor/vascular permeability factor by keratinocytes that have been stimulated by TGFα produced by both T lymphocytes and keratinocytes. Keratinocytes, as members of the cutaneous immune system, actively participate in the inflammatory pathogenesis of psoriasis by producing cytokines such as IL-6, IL-8, and TGFs (Krueger and Ellis, 2005). These vascular changes may help nourish the hyperproliferating skin and may be important for T-lymphocyte extravasation and trafficking (Guenther and Ortonne, 2002).
This negative correlation mostly occurs because of the presence of a strong relation between the defective E-cadherin in psoriasis and its histopathologic picture. The more fully developed the lesion, the less it will be likely to express E-cadherin. Such a negative correlation goes hand in hand with the progression of the histologic lesion, indicating a relationship between the loss of such adherence protein and the worsening of the histological picture (Mahfouz et al., 2012). In addition, as mentioned previously, soluble levels of E-cadherin in serum are increased in psoriasis vulgaris, correlating with disease severity and reflecting reduced functional E-cadherin at the cell surface of the epidermis (Chung et al., 2005). Finally, therapies that target cell adhesion molecules could prove to be useful because these molecules are known to mediate cell recognition, growth, and signaling events (Bayliffe et al., 2004).
In conclusion, this study showed that there are alterations in the expression of E-cadherin, one of the most important AJ proteins, in psoriasis. These alterations could play a critical role in keratinocytes adherence, proliferation, and differentiation, leading to development of the disease and correlating with its severity. These findings may provide new targets for therapeutic intervention. However, further studies that include a larger number of patients will be necessary for a generalized conclusion.
Conflicts of interest
There are no conflicts of interest.
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