The role of cytokines in the pathomechanism of cutaneous disorders.

Cytokines are produced transiently by cutaneous cells in response to immune stimuli. They are non-immunoglobulin proteins and glycoproteins. Cytokines, which are signaling molecules, act by sending downstream signals to various cells through a number of signal transduction pathways and by binding to cytokine receptors on the surface of cells. Cytokines regulate immunity and inflammation and thus play an important role in the pathogenesis of various cutaneous disorders.

Introduction

Cytokines are small molecules produced by a wide variety of cells including B and T lymphocytes, macrophages, endothelial cells, fibroblasts, mast cells and stromal cells of spleen, thymus and bone marrow de novo in response to any immune stimulus. They are signaling water-soluble non-immunoglobulin proteins and glycoproteins with a mass of 8 to 30 kd. They act by regulating immunity and inflammation as well as cell activation, migration, proliferation, and apoptosis. Hence, they are critical to the functioning of both innate and adaptive immune responses and play a vital role in the development of the immune system. They send downstream signals through signaling pathways such as the JAK/STAT and the NF-kB pathways. Their function may be autocrine, paracrine, or endocrine but differ from hormones which are produced by endocrine glands and are not always proteins. Although released transiently in the tissue microenvironment as diverse emergency molecules, they can produce chronic disease when released persistently.[1]

Functional Classification

The cytokines can be classified into different groups according to activities and mechanisms[2]:

Mediators of Innate Immunity: IL-1; IL-6; IL-12; IL-16; TNF-α, IFN-α, IFN-γ

Mediators of Hematopoiesis: GM-CSF; IL-3; IL-7; Precursor cell stimulating factor; M-CSF; Erythropoietin (Epo)

Mediators of Chemotaxis: Chemokines

Mediators in the activation, proliferation, differentiation, and death of T and B lymphocytes: IL-2; TNF-α; TNF-β; TNF-γ; IFN-γ

Cytokines of Cutaneous Cells

Keratinocytes

Keratinocytes are the major sources of cytokines in the epidermis. Normally, keratinocytes do not actively secrete cytokines; however, a number of agents mediate keratinocyte cytokine production, including cytokines themselves. IL-1α, IL-6, IL-8, and GM CSF play an important role in the normal regulation of the epidermis.[3-5]

The major cytokines secreted or produced by keratinocytes can be divided into:

Cytokines of undisturbed keratinocytes: IL-1, IL-7, and TGF-β.[6]

Mediators of inflammatory and immune response: IL-1, IL-6, IL-7, IL-10, IL-12, IL-15, IL-7, IL-18, IL-20, TNF-α, GM-CSF, M-CSF, SCF, CXC, and CC chemokines. When confronted by a foreign antigen, IL-1, IL-6, TNF-α, and chemokines initiate the inflammation; IL-1, GM-CSF, TNF-α, IL-10, and IL-15 activate and modulate the function of Langerhans cells; IL-15 and IL-18 activate the T cells; IL-10 and TGF-β cause T-cell inhibition and IL-12 modulates the lymphocyte activity.[7]

Regulators of growth of epithelial and/or mesenchymal cells: TGF-α, TGF-β, PDGF, FGF, Keratinocyte growth factor, Nerve growth factor, Vascular endothelial cell growth factor 1.[8]

The cytokine receptors present on keratinocytes are IL-1R Type I and Type II; TNF-αR type I; IL-4Rα; IL-6R; IL-10R1; IL-13Rα1; IL-18R; IL-20R1/Il-20R2; IL-22R1/IL-20R2; IFN-γ receptor; CXCR1; CXCR2; CCR3; CCR611.[9]

Langerhans cells

The Langerhans cells (LC) are dendritic cells (DC) which are the professional antigen-presenting cells (APC) in the epidermis. They produce IL-1α, IL-1β, IL-6,IL-7, IL-12, IL-15, IL-18, TNF, TGF-β, GM-CSF and the chrmokines MIP1α, MIP1β, MIP2, CCR1, CCR5, CCR7. TGF-β1 plays a very important role in the development of LC/DC.[10] LC precursors express CCR6 which plays an important role in the skin homing of LC/DC and their precursors. IL-10 and TGF-β cause CCR6 induction which is suppressed by IL-4 and IFN-γ. Recent studies suggest that cytokines are able to modulate LC/DC migration. Proinflammatory cytokines including IL-1β and TNF-α as well as IL-1α, IL-6, IL-12, IL-16, and IL-18 promote LC migration from the skin, whereas the anti-inflammatory cytokine IL-10 suppresses the process. Chemokines like CCR7 generated from lymphatic endothelial cells and lymph node cells play a role in the directional migration of LC/DC into lymph nodes.[11]

Melanocytes

βFGF and TGF-α (both produced by keratinocytes), EGF, NGF, PDGF, and POMC promote development of melanocytes.[12] Keratinocytes also produce melanocyte growth inhibiting factors like IL-1 and IL-6. POMC is the precursor of α, β, γ and delta MSH. αMSH acts as an immunomodulator by inhibiting production of IL-1, IL-2, IL-6, IFN, inducing production of IL-10 and downregulating IL-1 and TNF-α induced activation of NFkB.[13]

Fibroblasts

Proliferation of fibroblasts during wound healing is influenced by Fibroblast Growth Factors FGF1 and FGF2.[14] Production of cytokines from fibroblasts is controlled by IL-1α and IL-1β. The cytokines produced by fibroblasts are IL-1α IL-1β, IL-6, IL-8, and KGF. Fibroblasts from hypertrophic scars are sensitive to TGF-β.[15]

Mast cells

Prior to activation, mast cells contain mRNA onlyfor TNF-α. However, after activation by IL-4 there is a substantial increase in the amounts of mRNA for IL-3, IL-4, IL-5, IL-6, IL-8, IL-10, IL-13, IL-16, TGF-β1, MIP-1α, MCP-1, and GM-CSF with a minimal increase in amounts of mRNA for IL-4. IL-4, under the influence of IL-5 increases LTC4 synthetase levels IL-5 also enhances release of some cytokines without affecting degranulation. IL-6 plays a role in the IgE-dependant degranulation of mast cells.[16]

Macrophages

Macrophages secrete the following cytokines under different conditions:[17] (a) TNF-α: It is produced by many other cells as well as by macrophages, which are major sources, especially after priming by IFN-γ. (b) IL-1β: It is secreted by macrophages activated by a number of stimuli including TNF-α, bacterial endotoxin and IL-1β itself.IL-1β exerts its effects on many different cell types locally (at the site of production) and systemically (at a distance). (c) IFN-α/β: Macrophages (and many other cells) produce these Type I interferons which act as immunomodulatory, as well as antiviral cytokines. (d) IL-6: This pro-inflammatory cytokine is produced in response to infection and tissue injury. It exerts its effects on multiple cell types and can act systemically. (e) IL-10: It downregulates proinflammatory cytokine production hence it is a potent modulator of monocyte/macrophage (f) IL-12 (g) IL-18. Important cytokines secreted by macrophages in response to bacterial products include IL-1, IL-6, IL-8, IL-12, and TNF-α.

T Lymphocytes

CD4+ (helper) T cells were originally classified into two subgroups: Th1 and Th2 cells. The Th1 cells secrete IL-2 and IFN-γ which induce cellular immune responses (by activating macrophages to induce phagocytosis) and inhibit Th2 cells. On the other hand Th2 cells produce IL-4, IL-5, and IL-13 (which help in activating B cells and augmenting the humoral response), TGF-β and IL 10 which inhibit Th1 cells. Th1 cytokines are proinflammatory and Th2 cytokines are anti-inflammatory. Each type of cytokine downregulates the others’ responses.[1819] Later another cytokine secreting subgroup of CD4+ cells emerged and were named “type 1 regulatory T (Tr1)” or “type 3 regulatory T (Tr3) cells,” They secrete the immunosuppressive cytokines IL-10, TGF-β, or both.[20] Another subset that has emerged is the Th17 subset which produces high levels of IL-17 along with IL17 B through F. TH-17 cells promote inflammation and play an important role in autoimmunity.[21] Activated T cells send signals to B lymphocytes through cytokines such as IL-2, IL-4, IL-5, and IL-6 resulting in their activation, proliferation, and differentiation.

Natural killer T cells

A distinctive population of T cells is the natural killer T cells (NKT cells). When stimulated by contact with antigen, NKT cells develop heightened killer-cell activity and secrete large amounts of IL-4, IFN-γ, TGF-β, and IL-10, which activate cells that mediate inflammation, innate immunity, and Th2-type immunity.[22]

Skin appendages

Eccrine sweat glands and sebaceous glands contain receptors for IL13 – which plays an important role in Atopic dermatitis.[23] Other cytokine receptors identified in clear cells and duct epithelium of eccrine sweat glands are IL-1αR, IL-1βR, TNF-αR, and IL-6R. TGF-β2 is found in hair follicles, sebaceous glands, and upper eccrine sweat ducts.[24]

Role of Cytokines in Some Cutaneous Diseases

Psoriasis

As psoriasis is an inflammatory and hyperproliferative dermatosis, T cells and cytokines play a major role in its pathogenesis.[25] One of the initial events in psoriasis is the migration of activated T lymphocytes into lesional skin.[26] This process is facilitated by the expression of ICAM-1 and e-selectin on endothelial cells. IL-8 plays a major role in the transfer of activated T cells. While CD4+ helper T cells localize in the dermis, those that localize in the epidermis are mainly of the CD8+ cytotoxic type.[27] The subsequent activation of T cells by antigen-presenting cells (APCs) involves a cascade of pathways that ultimately leads to the production of a variety of cytokines. The overexpression of pro-inflammatory cytokines, particularly type-1 cytokines such as IL-2, IL-6, IL-8, IL-12, IFN-γ, and TNF-α by Th1 cells causes initiation, maintenance, and recurrence of skin lesions.[28] The cytokines control the cellular composition of the inflammatory infiltrate within the plaques as well as the keratinocyte hyperproliferation. For example, overexpression of IL-8 causes the accumulation of granulocytes, a characteristic finding in psoriatic lesions. It has been shown that IL-1Rα, IL-1β, IL-6, IL-8, Groα/β/γ, and epidermal growth factor receptor are found in psoriatic plaques though they are absent in normal skin.[29] IFN-γ can induce epidermal thickening when injected into the skin. IFN-γ induces macrophages to release high levels of other inflammatory cytokines like TNF-α which are present at higher levels in psoriatic plaques and in the synovial fluid of patients with psoriatic arthritis. The IL-2 receptor, CD25, is upregulated on T cells within psoriatic lesions. However, there is a relatively low level of expression of the anti-inflammatory cytokines IL-1RA and IL-10 by Th2 cells. Hence, the effect of pro-inflammatory cytokines in Psoriatic plaques is much more than that of anti-inflammatory cytokines.[30] A potential role for the chemokines CXCR3 and CCR4 ligands whereby they mediate the transendothelial migration of CLA+ T cells and a role for CXCR3 ligands in directing lymphocytes into the epidermis has also been suggested.[31] Although psoriasis has traditionally been considered to be mediated by Th1 cells, recent reports have highlighted the possibility of Th17 cells and the IL-23/Il-17A pathway playing an important role in the pathogenesis of psoriasis.[32]

Atopic dermatitis

Atopic dermatitis (AD) is a chronic or chronically relapsing inflammatory dermatosis where the lesional skin shows a dermal infiltrate of mainly CLA+ memory T cells as well as APC. Langerhans cells predominate in the initial phases of AD and prime naοve T cells to the Th2 type.[33] There is a dysregulated Th2 biased immune response to antigens, particularly environmental and bacterial stimuli. Although Th2 cells secreting IL-4 predominate in early lesions and unaffected skin, a mixed Th1-Th2 profile or dominance of IFN-γ producing Th1 cells is seen in late lesions.[34] Acute T-cell infiltration in AD is associated with a predominance of IL-4 and IL-13 expression while in the chronic lesions IL-5, GM-CSF, IL-12, and IFN-γ expression is increased.[35] Chronic AD is also associated with Th1 cytokines like IL-12 and IL-18 as well as other cytokines such as IL-11 and TGF-β1.[3637] IL-10, IL-31, CCL1, CCL2, CCL3, CCL4, CCL5, CCL13, CCL18, CCL20, CCL22, CCL26, and CCL27 have also been implicated in the pathogenesis of AD.[38] Keratinocytes of AD produce cytokines such as GM- CSF[39] and thymic stromal lymphopoietin (TSLP) which activate mast cells and DCs which then prime T cells to support the maintenance of Th2 cells.[40] Chemokines like RANTES are produced by keratinocytes following stimulation with TNF-α and IFN-γ leading to increased recruitment of eosinophils which play an important role in the pathogenesis of AD. Among the various chemokine receptors expressed by eosinophils, CCR3 and its ligands CCL5, CCL11, CCL13, and CCL26 have been implicated in the development of AD.[41]

Urticaria

In some cases of Acute Urticaria not responding to antihistamines, levels of circulating IL-6 are increased but levels of IL-8 and TNF-α are not increased.[42] In the late phase reaction (LPR), besides the mast cell-derived cytokines (IL-4, Il-5, IL-6, and IL-8),[43] cytokines are also secreted by eosinophils (IL-1, IL-4, IL-5, IL-6, IL-8, RANTES, and MIP-α)[43] basophils (IL-4, IL-8, IL-13, and MIP-1α)[44] neutrophils (IL-8) and Th2 lymphocytes (IL-1, IL-4, IL-5, IL-6, IL-9, IL-10, and IL-13).[43] IL-1 and IL-6 activate lymphocytes. IL-4 and IL-5 with the help of IL-9 and IL-13 cause increased IgE production, help in the recruitment of eosinophils, and support a Th2 environment for LPR.[45] Chemokines which play an important role in LPR are RANTES, eotaxin, eotaxin-2, MIP-1α, MCP-3, and MCP-4.[46]

Contact dermatitis

In irritant contact dermatitiskeratinocytes release IL-1, TNFα, IL-6, IL-7, IL-8, IL- 10, IL-12, IL-15, IL-18, GM-CSF, and TGF-α in response to environmental stimuli such as ultraviolet light and chemical agents.[4748] TNF-α and IL-8 also play an important role in allergic contact dermatitis since they regulate accessory adhesion molecules, such as ICAM-1.[49]

Infections and infestations

In bacterial infections, superantigens such as Staphylococcal exotoxins, staphylococcal enterotoxins A-E (SEA-SEE), and toxic shock syndrome toxin-1 (TSST- 1) produced by Staphylococcus aureus and Streptococcus pyogenes activate APC and T cells causing a massive release of proinflammatory cytokines.[50] Cytokine levels have also been found to be raised in Leprosy patients. Among the non reactional Leprosy patients, the pure neuritic group have increased levels of IFN-γ which is directly proportional to the extent of nerve involvement. Lepromatous leprosy patients have high levels of IL6. Type I and Type II reactional patients have higher levels of IFN-γ and IL-6 respectively as compared to non-reactional patients.[51] Viral infections induce production of IL-12 which causes NK cells to produce IFN-γ which in turn contributes to an antiviral state. IL-12 expression can be regulated by IFN. Endogenous IFN-β is responsible for the lack of IL-12 during viral infections which are unable to produce IFN-β.[52] In patients with HIV infection, there is a shift from Th1 cytokines (IL-2, IL-12, TNF-α, and IFN-γ) to Th 2 type cytokines (IL-4, IL-5, IL-6, and IL10). This change is more pronounced in HIV infected patients with evidence of opportunistic infection than those without them.[53] The Th1 cytokines stimulate the in vitro replication of HIV, whereas Th2 cytokines (mainly interferons) inhibit it.[54] Keratinocytes play an important role in the defense against fungal infections by secreting cytokines such as IL-1, IL-8, and TNF-α. Dermatophytes directly induce production of IL-8 and GRO-α by keratinocytes.[55] Th1 cytokines provide protection against Candida albicans infection, while Th2 responses are associated with disease exacerbation and pathology. Production of some cytokines such as TNF-α and IL-6 is more important in the early part of the infection. Later, production of IL-4, IL-10, and IL-12 is necessary for maintenance of Th1 reactivity.[56] M. furfur suppresses inflammatory response by downregulating IL-1α, inhibiting IL-6 and TNF-α, and upregulating IL-10 and TGF-β1. This modulation of proinflammatory cytokines and immunomodulatory cytokine synthesis may play a role in production of chronic disease.[57] Patients with uncomplicated scabies exhibit increased expression of IFN-γ in peripheral blood mononuclear cells while in crusted scabies, the expression of IL-4, IL-5, and TNF-α is increased. Sarcoptes scabiei induces a marked increase in secretion of IL-6 and VEGF and a slight increase in production of G-CSF by keratinocytes and downregulates keratinocyte secretion of IL-1ra. The liberation of IL-6, IL-8, G-CSF and VEGF by fibroblasts is increased.[58]

Connective tissue disorders

In Systemic Lupus Erythematosus, the immune dysregulation is due to lymphocyte hyperactivity due to autocrine and paracrine effects of the abnormally high IL-10 production. This results in an increased production of autoantibodies and impaired cell-mediated immunity (due to abnormalities of T lymphocyte and APC). The abnormality in the T-cell response results in an imbalance in the production of Th1 and Th2 cytokines.[59] In Scleroderma the serum contains increased amounts of IL-4, IL-6, and Il-2. IL-4 causes adult dermal fibroblasts to proliferate and produce IL-6. IL-6 has been shown to stimulate fibroblast synthesis of collagen and glycosaminoglycans. Fibroblasts, when exposed to TGF-β, overexpress the alpha-type receptor for platelet-derived growth factor.[60] In juvenile Dermatomyositis, the initial immune response is an IFN-α and IFN-β-induced cascade with secondary stimulation of IFN-γ. TNF-α synthesis is also increased. In adults with polymyositis and dermatomyositis IL-1α, TGF-β and IL-15 is produced early during the course of the disease.[61]

Vesicobullous disorders

The levels of TNF-α and IL-6 are high in the sera of patients with Pemphigus Vulgaris and decrease after one month of corticosteroid therapy. The skin biopsies show a mixed Th1/Th2 pattern of cytokine expression, including the presence of IL-2, IFN-γ, and IL-4 and the absence of IL-5 and IL-13.[62] IL-6 levels have been found to be increased in Paraneoplastic Pemphigus which may also be produced by treatment with cytokines such as IFN-α.[63] Perilesional skin biopsies from patients with Bullous Pemphigoid show deposition of IL-4, IL-13 and IL-5. While IL-4 and IL-13 localize in mononuclear cells within the dermal infiltrate, IL-5 is mainly expressed at the dermal-epidermal junction.[64] In Dermatitis Herpetiformis, the lesional skin shows increased levels of IL-8 in the basal cell layer and increased expression of endothelial leucocyte adhesion molecules (ELAM) in the deep dermis. Dendritic cells producing GM-CSF are also seen at the dermo-epidermal junction.[65]

Lichen planus

Activated CD8+ cytotoxic T-cells in the lymphocytic infiltrate of Lichen Planus secrete IL-2, IL-4, IL-10 IFN-γ, TGF-β1 and TNF-α after antigen recognition. These cytokines attract lymphocytes to the site of the lesion and regulate their activities. Also IFN-γ secretion by Th1 CD4+ T-cells stimulates TNF-α secretion by CD8+ T-cells resulting in keratinocyte apoptosis. Hence there is a mixed production of Th1/Th2 cytokines and the balance between them determines the course of the disease. Keratinocytes also produce IL-1β, IL-4, IL-6, GMCSF, and TNF-α which activate tissue macrophages and peripheral blood mononuclear cells and upregulate expression of keratin genes and cell adhesion molecules.[66]

Vitiligo

Cytokines play an important role in autoimmunity. Hence, their possible role in the autoimmune pathogenesis of Vitiligo has been investigated by various workers. Expression of GM-CSF, βFGF, and SCF (Stem cell factor)is decreased and that of IL-6 and TNF-a is increased in lesional skin of Vitiligo as compared with perilesional, non-lesional and healthy skin.[67] In non-segmental vitiligo, the lesional skin contains increased levels of IL-1α and TNF-α.[68] The serum of such patients has been shown to contain increased levels of IL-6 and GM-CSF.[69] Increased levels of IL-17 found in serum and lesional skin suggest an important role for this cytokine in the pathogenesis of vitiligo.[70]

Acne vulgaris

Propioni-bacterium acnes (P. acnes) is a commensal microorganism found in sebum-rich skin and plays a role in acne inflammation by stimulating production of proinflammatory cytokines by keratinocytes.[71] Viable P. acnes in the stationary phase of growth stimulate keratinocytes to produce significantly higher amounts of IL-1α, TNF-α, and GM-CSF. P. acnes induces TNF-alpha mRNA/protein expression in fibroblasts through the NF-kB pathway resulting in an increase of both proMMP-2 mRNA and protein expression.[72]

Alopecia areata

Serum IFN-γ, RANTES, IL-8, and eotaxin levels are increased in patients with alopecia areata. Levels of IFN-γ, RANTES, and IL-8 are also increased. Elevated serum IFN-γ and RANTES levels significantly correlate with the disease activity suggesting that MIG and RANTES play an important role in the development of Alopecia Areata and are useful as markers of disease activity and as therapeutic targets.[73] It has been postulated that in alopecia areata aberrant expression of IFN-γ as a result of antigen-dependent immune response plays an important role in the pathogenesis.[74]

Neutrophilic dermatoses

Overexpression of cytokines such as IL-8 is implicated in the pathogenesis of Pyoderma Gangrenosum.[75] TNF-α also enhances neutrophil activation, upregulates the expression of adhesion molecules, and induces the release of chemokines and cytokines from fibroblasts. Other cytokines involved are IL- 1, IL-3, IL-6, G-CSF, GM-CSF and IFN-γ.[76] TNF-α, IL-8, IL-17, VEGF, and Il-8 (cytokine chemotactic for neutrophils) are significantly elevated both in Pyoderma Gangrenosum and Sweet's syndrome.[77] G-CSF, GMCSF, IFN-γ, IL-1, IL-5, IL-6, and IL-8 have all been implicated in its pathogenesis.[78] TNFα, IL-2, IL-2r, IL-6, IL-8, and VEGF have been implicated in the pathogenesis of Behcet's syndrome.[79] Although both active and in remission patients show increased serum levels of IFN-γ, IL-4, IL-6, IL-10, IL-12, IL-17, and IL-18, the increase is more in patients with active Behcet's syndrome than in patients in remission.[80]

Melanoma

Cytokines facilitate autonomous melanoma growth and tumor progression. Cultured melanoma cell lines and melanoma cells derived from primary melanoma and metastases produce basic fibroblast growth factor bFGF, IL-1α, IL-1β, IL-6, VEGF, PDGF, and IL-8 which promote tumor cell growth and increase the capacity of tumor cells to survive. They also stimulate endothelial cell proliferation, migration, and angiogenesis that are important for melanoma growth and metastasis. Increased production of proangiogenic and growth factors by melanoma and stromal cells in vivo might result in elevation of these factors in the circulation. Serum IL-8 and IL-6 levels are raised in melanoma patients. In addition, levels of IL-1 and serum epidermal growth factor (EGF) receptor are also elevated. While increased levels of IL-10 have been associated with advanced stage III and IV melanoma, elevated levels of IL-6 indicate a poor prognosis in patients with stage IV melanoma.[81]

Cutaneous T cell lymphoma

Cutaneous T cell lymphoma (CTCL) is a malignancy of skin-homing Th2 T cells. Hence, clonal T cells and CTCL skin lesions typically express Th2 cytokines like IL-4, IL-5, IL-6, and IL-10 but fail to produce Th1 cytokines including IFN-γ. This profile indicates a direct contribution of the malignant cells in the Th1/Th2 imbalance observed frequently in CTCL patients and suggests their potential role in depressed cell-mediated immunity.[82] However, the reason for the Th2 bias seen in CTCL is unknown. It has been proposed that the high levels of IL-18 expression in lesional CTCL skin contribute to increased plasma levels of IL-18 and this may contribute to the Th2 bias seen in this disease.[83] Malignant T cells in CTCL lesions express IL-17 through the Jak3/Stat3 pathway.[84]

Conclusion

Cytokines play an important role in cutaneous tissues. Newer insights into their role in various cutaneous disorders have provided researchers with important clues to the etiological factors implicated not only in the pathomechanism but also into the responses to therapies in these diseases and has led to the development of a whole new genre of drugs known as biological which has revolutionized the therapy of these disorders.[1]