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  Table of Contents    
ORIGINAL ARTICLE  
Year : 2014  |  Volume : 57  |  Issue : 2  |  Page : 205-208
Expression of tumor necrosis factor-α and nuclear factor-kappaB/RelA and the pathogenesis of psoriasis


1 Department of Pathology, Armed Forces Medical College, Pune, Maharashtra, India
2 Department of Dermatology, Armed Forces Medical College, Pune, Maharashtra, India

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Date of Web Publication19-Jun-2014
 

   Abstract 

Context: Tumor necrosis factor-α (TNF-α) is an important mediator in the pathogenesis of psoriasis. Nuclear factor-kappaB (NF-κB) is a transcription factor that regulates the activity of the proinflammatory genes. Psoriasis is an inflammatory disease and the role of TNF-α and NF-κB, should be considerable. Aims: We studied the role of TNF-α and NF-κB in psoriasis. Materials and Methods: A total of 61 cases of psoriatic skin biopsies were studies and the grade of TNF-α and NF-κB, staining was correlated with the histopathological indices of severity. Statistical Analysis Used: Pearson's correlation coefficient and Chi-square test. Statistical Package for Social Sciences version 13 was used. Results: The TNF-α immunostain in the cytoplasm of the epidermal cells and basal cells showed a strong inverse correlation with the grade of epidermal hyperplasia (P -0.019 and P -0.009, respectively). The epidermal cytoplasmic positivity and lymphocyte positivity for TNF-α did not correlate with the grade of NF-κB immunostaining in the epidermal cell nuclei, basal cells or lymphocytes. The basal cell cytoplasmic positivity for TNF-α correlated with the grade of NF-κB immunostaining in the nucleus of basal cells at a P - 0.005. There was a strong correlation between the epidermal cytoplasmic TNF-α immunostaining with the lymphocyte immunostaining (P -0.08); however, there was no correlation between the TNF-α expression in the other two locations. Conclusions: The study outlines the relationship between NF-κB and TNF-α and their combined role in the development of the characteristic histopathological changes in psoriasis. We hypothesize that NF-κB is involved in stimulating the release of TNF-α which would account for the characteristic histopathological changes of psoriasis. However, it is likely that NF-κB can act independently of TNF-α also in the pathogenesis of psoriasis.

Keywords: Nuclear factor-kappaB, psoriasis, tumor necrosis factor-α

How to cite this article:
Moorchung N, Vasudevan B, Mani NS, Verma R. Expression of tumor necrosis factor-α and nuclear factor-kappaB/RelA and the pathogenesis of psoriasis. Indian J Pathol Microbiol 2014;57:205-8

How to cite this URL:
Moorchung N, Vasudevan B, Mani NS, Verma R. Expression of tumor necrosis factor-α and nuclear factor-kappaB/RelA and the pathogenesis of psoriasis. Indian J Pathol Microbiol [serial online] 2014 [cited 2022 Jan 18];57:205-8. Available from: https://www.ijpmonline.org/text.asp?2014/57/2/205/134664



   Introduction Top


Psoriasis is a chronic inflammatory skin condition characterized by an inflammatory dermal infiltrate and hyperproliferative keratinocytes. The pathogenesis of this disease is complex and is believed to be mediated by a dysregulation of the innate immunity and cytokine production. The cytokines are regulatory proteins, which are known to have an important role in several inflammatory diseases. Of the cytokines, tumor necrosis factor (TNF) alpha is considered to be an important mediator in the pathogenesis of psoriasis. [1] Increased levels and activity of this cytokine have been observed in blood and skin of patients with psoriasis. [2] The role of TNF-α is of great therapeutic significance since TNF-α inhibitors play an important role in the treatment of psoriasis. [3]

The rel/nuclear factor-kappaB (NF-κB) proteins belong to a family of related transcription factors, which over the last 25 years have been characterized in most cell types and lineages. [4] NF-κB is one of the most important regulators of proinflammatory gene expression. TNF-α is an important cytokine, which is inducible by NF-κB. [5] Since NF-κB regulates cytokine gene expression and in particular TNF-α expression, it is likely that NF-κB is a second key factor involved in the pathogenesis of the disease.

In psoriasis, keratinocytic differentiation and proliferation could be modulated and regulated by inflammatory mediators released from chronic inflammatory cells which accompany this lesion. [6] Important inflammatory mediators are NF-κB and TNF-α whose role in the pathogenesis of psoriasis, i.e., keratinocyte proliferation and inflammation is not clear. Unraveling the complex relationship between the histopathology, NF-κB and TNF-α expression may provide a better understanding of the pathogenesis of this complex disease.

In order to accurately evaluate the role of NF-κB and TNF-α in psoriasis, the expression of the above mediators has been compared to the inflammatory infiltrate and grade of keratinocyte proliferation. The aim of the study was to evaluate the role of TNF-α in the pathology of psoriasis. We also evaluated the synergy between NF-κB and TNF-α in psoriasis. Knowing the critical role, which TNF-α inhibitors play in inflammatory diseases, the evaluation of the role of TNF-α is critical in understanding the pathogenesis of the disease.


   Materials and Methods Top


This study was designed as a prospective study and included skin specimens of 61 patients with psoriasis vulgaris diagnosed at a tertiary care hospital. The study was conducted in accordance with the principles of the Helsinki declaration. 63 paraffin blocks of skin tissues of patients with psoriasis vulgaris were studied. Psoriasis was diagnosed by clinical features and histology. The patients had not received any kind of treatment for at least 1 month before biopsy procedure. Biopsies were taken from the lesion to include the perilesional skin.

Histopathology

Biopsies were taken from the lesion to include a small portion of the perilesional skin. The biopsies were immediately fixed in formalin for histopathological examination. Sections were stained with the standard hematoxylin and eosin stain using standard histological laboratory methods. Histopathological analysis of the biopsies was done and included eight criteria (epithelial hyperplasia, parakeratosis, Munro's and Kogoj's microabscesses, suprapapillary thinning, and inflammatory infiltrate in the papillary dermis, widened rete ridges and capillary dilatation). Of the criteria, we included only epidermal hyperplasia and the inflammatory infiltrate for analysis in the present study. Grading was done using a visual analog scale and the biopsies were graded as 0-3 (nil to marked).

Immunohistochemistry

Serial 4 mm thick sections were made and mounted on poly L lysine coated slides. Paraffin sections were immersed in xylene for 5 min and hydrated using a gradient series of alcohol. Antigen retrieval was routinely performed by immersing the sections in the citric acid buffer (pH 6.0); in a microwave oven for 15 min. Endogenous peroxidase activity was blocked with 3% hydrogen peroxide for 10 min and then incubated with a primary antibody in a humidified chamber at 4°C overnight. Primary antibody was monoclonal mouse anti-NF-kB/p65 antibody (Santa Cruz) and monoclonal mouse ant TNF-α antibody (Santa Cruz) both at a 1:200 dilution. A total of 500 keratinocytes were enumerated in an area that was stained intensely on each three serial slides at ×400 magnification under the light microscopy and an average was taken. The results of the immunostaining were analyzed semi quantitatively as a percentage of positive cells. Localization of immunohistochemical staining was grouped and classified such as epidermal, basal cell, and lymphocyte staining. Basal cell staining was evaluated separately from epidermal staining because of the differences in the kinetics of the epidermal cells.

The sample size was calculated taking the alpha (type I) error as 5%, the confidence interval of 95% and the power of the study as 80%. The minimum sample size was calculated using the Epi Info 2002 software. The sample size was calculated was 61.

The results of study were statistically analyzed using IBM SPSS Statistics (IBM 2014) . The descriptive data were given as mean and standard deviation. The Pearson's correlation coefficient was used for comparison the differences between groups. P < 0.05 was considered to indicate statistical significance.


   Results Top


Demographic variables

A total of 61 cases were analyzed in the present study. The means age was 40.7 years with a range of 13-76 years. 36 males and 25 females comprised the study population.

Correlation of tumor necrosis factor-α immunostaining with histopathology

The TNF-α immunostain in the cytoplasm of the epidermal cells showed a strong inverse correlation with the grade of epidermal hyperplasia (P -0.019). The TNF-α immunostain in the cytoplasm of the basal cells also showed a strong inverse correlation with the grade of epidermal hyperplasia (P -0.009). The TNF-α immunostain in the lymphocytes did not show any correlation with any of the histopathological parameters of severity. The results are shown in [Table 1] and [Table 2].
Table 1: The correlation between the grade of epidermal hyperplasia and the grade of TNF-α immunostaining in the cytoplasm of the epidermal cells

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Table 2: The correlation between the grade of epidermal hyperplasia and the grade of TNF-α immunostaining in the cytoplasm of the basal cells

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Correlation of tumor necrosis factor-α epidermal cytoplasmic staining and nuclear factor-kappaB staining

The percentage of epidermal cells showing a cytoplasmic staining for TNF-α ranged from 0% to 100% with an average of 16.80%. The epidermal cytoplasmic positivity for TNF-α did not correlate with the grade of NF-κB immunostaining in the three locations evaluated i.e., nucleus of epidermal cells, nucleus of basal cells or in the nucleus of lymphocytes which comprised the inflammatory infiltrate.

Correlation of tumor necrosis factor-α basal cell staining and nuclear factor-kappaB staining

The percentage of basal cells showing a cytoplasmic staining for TNF-α ranged from 0% to 100% with an average of 37.95%. The basal cell cytoplasmic positivity for TNF-α correlated with the grade of NF-κB immunostaining in the nucleus of basal cells at a P - 0.005. There was however no correlation with the grade of NF-κB staining in the nucleus of lymphocytes which comprised the inflammatory infiltrates (P - 0.956).

Correlation of tumor necrosis factor-α lymphocyte staining and nuclear factor-kappaB staining

The percentage of lymphocytes showing a cytoplasmic staining for TNF-α ranged from 0% to 90% with an average of 9.59%. The lymphocyte cytoplasmic positivity for TNF-α did not correlate with the grade of NF-κB immunostaining in any of the three locations evaluated i.e., nucleus of epidermal cells, nucleus of basal cells or in the nucleus of lymphocytes which comprised the inflammatory infiltrate.

Correlation of epidermal cytoplasmic tumor necrosis factor-α and basal cell immunostaining

There was no correlation between the epidermal cytoplasmic TNF-α immunostaining with the basal cell immunostaining at a P - 0.05.

Correlation of epidermal cytoplasmic tumor necrosis factor-α and lymphocyte immunostaining

There was a strong correlation between the epidermal cytoplasmic TNF-α immunostaining with the lymphocyte immunostaining at a P - 0.08.

Correlation of basal cell cytoplasmic tumor necrosis factor-α and lymphocyte immunostaining

There was no correlation between the basal cell TNF-α and the lymphocyte nucleus staining at a P < 0.05 (P - 0.491).


   Discussion Top


There exist two major pathological steps in psoriasis: Epidermal hyperproliferation with abnormal differentiation and inflammatory infiltration of epidermis and dermis. [5],[6] Of the two, it is likely that the inflammatory infiltrate is of more importance in the pathogenesis of psoriasis. [7],[8] TNF-α is an important inflammatory mediator and its expression has been shown to be increased in psoriatic lesions. [9] TNF-α has been shown to be one of the most abundant early mediators in inflamed tissue. [10] TNF-α is considered to be a "master regulator" of proinflammatory cytokine production. [11] TNF-α is also believed to play a critical role in the development of many chronic inflammatory diseases. [12] The role of TNF-α in psoriatic lesions is known, but the underlying mechanisms for its action are poorly understood. In the present study, we found an inverse correlation between the grades of TNF-α staining in different skin compartments and the histopathological indicators of severity. This is probably because the action of TNF-α is mediated through other molecules like interleukin 8 (IL-8) and transforming growth factor-α (TGF-α). It is known that TNF-α stimulates the release of IL-8 by keratinocytes and fibroblasts. [13],[14] and TGF-α by keratinocytes. Both IL-8 and TGF-α may be involved in autocrine stimulation of keratinocytes proliferation in psoriatic lesions. [15] Therefore, it is likely that the TNF-α levels rise and cause a rise in the levels of other molecules like IL-8 and TGF-α. These secondary mediators then cause a hyperproliferation of keratinocytes. By the time, there is an increase in the thickness of the epidermis, the TNF-α levels have already decreased. In another study, other authors [16],[17] have shown that TNF-α does not increase keratinocyte proliferation in normal or psoriatic skin. Their finding is in consonance with our observations. These findings underline the fact that although TNF-α is a very important cytokine involved in the inflammatory response in psoriasis, it is not the cytokine which is responsible for the proliferation of keratinocytes.

There was no correlation between the grade of TNF-α staining in the epidermal cells and the grade of NF-κB immunostaining in the epidermal cells. In another study, we have shown that there is a correlation between the grade of NF-κB immunostaining in the epidermal cells and the degree of epidermal thickness. [18] We hypothesized that in a setting of a chronic inflammatory state such as psoriasis, there is an imbalance between the anti-apoptotic role and the cell cycle inhibitory role of NF-κB. The anti-apoptotic role appears to be dominant and that would account for the increased epidermal thickness correlating with the nuclear NF-κB positivity. We do know that NF-κB is one of the most important regulators of proinflammatory gene expression. However, NF-κB has multiple roles and regulation of apoptosis is quite distinct from regulation of proinflammatory gene expression. It is likely therefore that NF-κB does not mediate its effects through TNF-α in inducing epidermal hyperplasia.

The basal cell cytoplasmic positivity for TNF-α correlated with the grade of NF-κB immunostaining in the nucleus of basal cells. It is important to remember that the kinetics of the basal cell is different from the rest of the epidermis. Most of the mitotic activity in normal human epidermis occurs in the basal cell layer. An increase in the NF-κB immunostaining in the nucleus of basal cells would cause an increase in the TNF-α expression. Conversely, it is also known that TNF-α may also increase the action of NF-κB by increasing degradation of IκB, its inhibitory protein. [19] It can be concluded that TNF-α acts through the NF-κB pathway and results in an increased proliferation of the basal cells and increased epidermal thickness.

There was also a strong correlation between the epidermal cytoplasmic TNF-α immunostaining and the lymphocyte immunostaining. In a previous study, we have shown that lymphocyte staining of NF-κB showed a strong correlation with epidermal and basal cell staining pattern for NF-κB. As mentioned earlier, NF-κB is one of the most important regulators of proinflammatory gene expression. We believe that the NF-κB translocation is responsible for the correlation between the TNF-α immunostaining in the epidermal compartment and the lymphocytes.

This study outlines the key role of TNF-α in the pathogenesis of psoriasis. The study also outlines the relationship between NF-κB and TNF-α and their combined role in the development of the characteristic histopathological changes in psoriasis. TNF-α inhibitors are already being used in the treatment of psoriasis, in the future it is likely that NF-κB and TNF-α inhibitors will play an increasingly important role in the treatment of this enigmatic disease.

 
   References Top

1.Victor FC, Gottlieb AB. TNF-alpha and apoptosis: Implications for the pathogenesis and treatment of psoriasis. J Drugs Dermatol 2002;1:264-75.  Back to cited text no. 1
    
2.Bonifati C, Carducci M, Cordiali Fei P, Trento E, Sacerdoti G, Fazio M, et al. Correlated increases of tumour necrosis factor-alpha, interleukin-6 and granulocyte monocyte-colony stimulating factor levels in suction blister fluids and sera of psoriatic patients - Relationships with disease severity. Clin Exp Dermatol 1994;19:383-7.  Back to cited text no. 2
    
3.Caldarola G, De Simone C, Carbone A, Tulli A, Amerio P, Feliciani C. TNFalpha and its receptors in psoriatic skin, before and after treatment with etanercept. Int J Immunopathol Pharmacol 2009;22:961-6.  Back to cited text no. 3
    
4.Baldwin AS Jr. The NF-kappa B and I kappa B proteins: New discoveries and insights. Annu Rev Immunol 1996;14:649-83.  Back to cited text no. 4
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5.Das RP, Jain AK, Ramesh V. Current concepts in the pathogenesis of psoriasis. Indian J Dermatol 2009;54:7-12.  Back to cited text no. 5
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6.Kawashima K, Doi H, Ito Y, Shibata MA, Yoshinaka R, Otsuki Y. Evaluation of cell death and proliferation in psoriatic epidermis. J Dermatol Sci 2004;35:207-14.  Back to cited text no. 6
    
7.Boruah D, Moorchung N, Vasudevan B, Malik A, Chatterjee M. Morphometric study of microvessels, epidermal characteristics and inflammation in psoriasis vulgaris with their correlations. Indian J Dermatol Venereol Leprol 2013;79:216-23.  Back to cited text no. 7
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8.Moorchung N, Khullar J, Mani N, Chatterjee M, Vasudevan B, Tripathi T. A study of various histopathological features and their relevance in pathogenesis of psoriasis. Indian J Dermatol 2013;58:294-8.  Back to cited text no. 8
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9.Ettehadi P, Greaves MW, Wallach D, Aderka D, Camp RD. Elevated tumour necrosis factor-alpha (TNF-alpha) biological activity in psoriatic skin lesions. Clin Exp Immunol 1994;96:146-51.  Back to cited text no. 9
    
10.Feldmann M, Brennan FM, Elliott M, Katsikis P, Maini RN. TNF alpha as a therapeutic target in rheumatoid arthritis. Circ Shock 1994;43:179-84.  Back to cited text no. 10
    
11.Maini RN, Elliott MJ, Brennan FM, Feldmann M. Beneficial effects of tumour necrosis factor-alpha (TNF-alpha) blockade in rheumatoid arthritis (RA) Clin Exp Immunol 1995;101:207-12.  Back to cited text no. 11
    
12.Clark IA. How TNF was recognized as a key mechanism of disease. Cytokine Growth Factor Rev 2007;18:335-43.  Back to cited text no. 12
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13.Larsen CG, Anderson AO, Oppenheim JJ, Matsushima K. Production of interleukin-8 by human dermal fibroblasts and keratinocytes in response to interleukin-1 or tumour necrosis factor. Immunology 1989;68:31-6.  Back to cited text no. 13
    
14.Barker JN, Sarma V, Mitra RS, Dixit VM, Nickoloff BJ. Marked synergism between tumor necrosis factor-alpha and interferon-gamma in regulation of keratinocyte-derived adhesion molecules and chemotactic factors. J Clin Invest 1990;85:605-8.  Back to cited text no. 14
    
15.Barker JN, Mitra RS, Griffiths CE, Dixit VM, Nickoloff BJ. Keratinocytes as initiators of inflammation. Lancet 1991;337:211-4.  Back to cited text no. 15
    
16.Takahashi H, Tsuji H, Hashimoto Y, Ishida-Yamamoto A, Iizuka H. Cell proliferation and cytokine induction by TNF-alpha of psoriatic keratinocytes are not different from normal keratinocytes in vitro. Indian J Dermatol 2009;54:237-9.  Back to cited text no. 16
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17.Fransson J. Tumour necrosis factor-alpha does not influence proliferation and differentiation of healthy and psoriatic keratinocytes in a skin-equivalent model. Acta Derm Venereol 2000;80:416-20.  Back to cited text no. 17
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18.Moorchung N, Kulaar JS, Chatterjee M, Vasudevan B, Tripathi T, Dutta V. Role of NF-κB in the pathogenesis of psoriasis elucidated by its staining in skin biopsy specimens. Int J Dermatol. 2013 Apr 28. doi: 10.1111/ijd.12050. [Epub ahead of print].  Back to cited text no. 18
    
19.Brotas AM, Cunha JM, Lago EH, Machado CC, Carneiro SC. Tumor necrosis factor-alpha and the cytokine network in psoriasis. An Bras Dermatol 2012;87:673-81.  Back to cited text no. 19
    

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Correspondence Address:
Nikhil Moorchung
Department of Pathology, Armed Forces Medical College, Sholapur Road, Pune - 411 040, Maharashtra
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0377-4929.134664

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    Tables

  [Table 1], [Table 2]

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