Evaluation of Serum Netrin-1 Levels in Psoriasis Patients

Introduction

Psoriasis is immune-mediated chronic and recurrent, inflammatory dermatosis skin disease, which is closely associated with oxidative stres [1]. This disease is recurrent apparition of erythematous plaques often covered by silvery scales. The etiologic and genetic factors which affecting psoriasis are not clear. Trauma, drugs, smoking, emotional stress, alcohol and skin infection are among the factors that affect the clinical development of psoriasis [1, 2]. According to current understanding of the pathomechanisms integral to the psoriatic process, the skin disorder is characterized by decreased apoptotic cell death of keratinocytes, increased resistance of intralesional keratinocytes to apoptosis, and angiogenesis [1, 2, 3].

Complex interaction between cells in the skin and the immune system leads to chronic inflammation of the skin (3,4). The formation and maintenance of psoriatic plaques is due to an imbalance between anti-inflammatory agents that support leukocyte infiltration and proliferation of keratinocytes. The linkages between genetic susceptibility and inflammatory activity in this disease are thought to be effective in many molecules that concern T cell activation or differentiation [2]. In addition, vascular changes observed in lesions are thought to be an important feature for the disease. In fact, they are a key component not only as a result of the disease, but also because it can promote skin inflammation through recruitment of leukocytes. Increased angiogenesis resulting from psoriatic skin cells, activation of endothelial cells through pro-inflammatory cytokines, thus demonstrates the relationship between the altered epidermis and the immunological component of the disease.

Recently, angiogenesis has become very important in psoriasis. The vascular network found in these lesions has changed considerably, especially in papillary dermis infiltrated by numerous curved and dilated capillaries. Endothelial cells of the vessels constituting vascularization are activated and promote the expression of several adhesion molecules such as ICAM-1, VCAM-1, Thy-1, E- and β-selectin for leukocyte uptake. Therefore, this pathological angiogenesis is not only a consequence of the disease, but also it causes leukocyte deposition, inflammation and hence the formation of skin lesions. Psoriatic skin cells promote the expansion of the vascular network, particularly by secretion of pro-angiogenic factors such as keratinocytes, VEGF and angiopoietins. Indeed, angiogenesis supported by VEGF secreting keratinocytes leads to skin lesions mimicking inflammation and psoriasis [1, 2, 3, 4]. The inflammatory events associated with psoriatic diseases are not limited to the skin, but also are responsible for numerous comorbid conditions including obesity, hypertension, dyslipidemia and diabetes, cardiometabolic disease, stroke, metabolic syndrome, chronic kidney disease, malignancy [5, 6, 7].

Netrin-1 is a laminin-related molecule that was discovered as a diffusible molecule produced by the ventral structure in the initially developing spinal cord. Netrin-1 is a member of the family of homologous molecules such as netrin-3, netrin-G1, netrin-G2 and netrin-4 / b-netrin [8, 9]. In recent data, netrin-1 and its receptors are also expressed in nonneural tissues such as pancreas [9, 10], mammary gland [11], or lung [12] that has suggested their role in the morphogenesis of ‘branched’ organs. Netrin-1 was shown to act as a chemoattractive or chemorepulsive cue for many migrating axons and neurons. This effect is believed to occur through the binding to two main families of type 1 transmembrane receptors: DCC (for deleted in colorectal cancer) and its homolog neogenin and the UNC5H [9, 11]. Furthermore, the interaction of netrin-1/receptors and, more specifically, netrin-1/UNC5H2, was recently shown to be involved in the morphogenesis of endothelial vessels. Indeed, although it is clearly shown that UNC5H2 expressed by the tip cells is a key player in developmental angiogenesis [12], the present data published either suggest an antiangiogenic role or a proangiogenic role for netrin-1 [13, 14, 15]. Some recent studies have shown that netrin-1 plays an active role in autoimmune diseases and inflammatory reactions associated with these diseases [16, 17, 18, 19].

There have been no previous studies of the potential roles of netrin-1 in psoriasis. Based on the aforementioned literature on the roles of inflammation in the pathophysiology of psoriasis, this study suggests that netrin-1 levels might be linked to psoriasis. We tried to to find out if there is any change in serum netrin-1 levels in patients with psoriasis, and to determine the relationship between serum netrin-1 with demographic, clinical and laboratory characteristic.

Materials and Methods

This cross-sectional study was performed in accordance with the guidelines of Helsinki Declaration and it was approved by the local ethical committee.

Results

The patient group included 40 psoriasis patients (19 males and 21 females). The control group included 40 age- and sex- matched healthy individuals (22 females and 18 males). The characteristics of the study groups are summarized in Table 1. Psoriasis patients had significantly higher values for BMI (26.9 kg/m² vs. 23.4 kg/m², p<0.001) and WHR (0.8 kg/m vs. 0.7 kg/m, p=0.01, Table 1) compared to the controls. Psoriasis patients had significantly higher mean FPG (90 mg/dL vs. 82 mg/dL, p<0.001, Table 1), CRP (9.4 mg/dL vs. 3.3 mg/dL, p<0.001, Table 1), and triglyceride levels (111 mg/dL vs. 88.5, p<0.05, Table 1) compared to the control group. When compared to controls, netrin levels (442.7±245.8 pg/ml vs. 655.5±393.3 pg/ml, p<0.001, Table 1) were found to be lower in psoriasis patients According to disease severity, psoriasis patients were divided into two groups: 25 psoriasis patients with PASI≤10 (15 females and 10 males), 15 psoriasis patients with PASI>10 (6 females and 9 males). Table 2 summarizes the characteristics of the healthy controls, psoriasis patients with PASI≤10 and PASI>10. When compared to healthy controls, psoriasis patients with PASI≤10 and PASI>10 had significantly higher BMI, WHR, CRP and MCV (p’<0.05 for all, Table 2). The psoriasis patients with PASI≤10 and psoriasis patients with PASI >10 had significantly lower netrin-1 values than healthy controls (p’<0.05 for all, Table 2). The psoriasis patients with PASI>10 had significantly higher PASI, CRP and WBC values than psoriasis patients with PASI≤10 (p’<0.001 for all, Table 2). There were no statistically significant differences in netrin-1 levels between psoriasis patients with PASI>10 and with PASI≤10 (p> 0.05 for all, Table 2).

Table 3 shows the correlations among BMI, disease duration, PASI and CRP with netrin-1 parameters in psoriasis patients. We founded that psoriasis patients had netrin was negatively correlated with BMI, duration of disease, PASI and CRP (r-0.284; p<0.05, Table 3, r: -0.461, r:-0.441 and r:-0.424, p<0.001 for all, Table 3).

Table 1: Comparison of the clinical characteristics and biochemical parameters of the control and psoriasis patients. Data were presented as mean ± SD or median (min–max) N=number of volunteer; NS =nonsignificant; BMI= body mass index; WHR=waist to hip ratio; F/M females/males; PASI=Psoriasis area severity index; BP=blood pressure; FPG=Fasting plasma glucose; CRP=C-reactive protein; LDL=low- density lipoprotein; TC=Total cholesterol; HDL=high density lipoprotein. The p values< 0.05 were assessed as statistically significant.

Psoriasis patients

Parameters Control

with PASI≤10

(n=40) (n=27) P’-value F/M 22/18 15/10 6/9 NS Age (year) 32.5 (20-50) 33 (20-60) 32 (24-39) NS Weight (Kg) 63.3±7.7 73.8±13.2 69.1±14.2 <0.05† BMI (kg/m²) 23.4 (19.8-25.4) 27.3 (19.1-43.1) 25.3 (20.8-40.2) <0.05†‡ Psoriasis patients with PASI≤10 and Psoriasis patients with PASI>10 PASI>10 (n=13)

Table 2: Comparision of Biochemical Characteristics in Psoriasis Patients Whose PASI≤10 and PASI >10 with Healthy Controls. Data were presented as mean ± SD or median (min-max). N=number of volunteer; NS=nonsignificant; F/M=females/males; PASI= psoriasis area severity index; BMI=body mass index;WHR=waist to hip ratio; BP=blood pressure; FPG=Fasting plasma glucose; CRP=C-reactive protein; LDL=low density lipoprotein; TC=Total cholesterol; HDL=high density lipoprotein. The p’ values< 0.05 were assessed as statistically significant. *p<0.05 was accepted to be statistically significant. †There was statistical significance between healthy controls and psoriasis patients with PASI≤10 (p<0.05). ‡There was statistical significance between healthy controls and psoriasis patients with PASI >10 (p<0.05). •There was statistical significance between psoriasis patients with PASI≤10 and PASI>10 (p<0.05).

Psoriasis is a chronic, inflammatory and proliferative skin disease, frequently seen with remissions and relapses [1, 2]. It is a chronic inflammatory disease affecting 2-4 % of the world-population. It is considered that there is a hereditary predisposition to the disease and the onset of disease can be triggered by trauma, infection, medications and stress at any time in life [1, 2, 3, 4]. Netrin-1 is accepted to partipicate a play role in the development and progression of some disease related to inflammation [8, 9, 10, 11, 12, 13, 14, 15, 16]. This study aimed to investigate how a netrin-1 level is altered in psoriasis patients and whether netrin-1 levels correlate with the severity and duration of psoriasis. In this study, patients with psoriasis were shown to have significantly lower levels of netrin-1 than healthy controls. There is no significant difference The complex interaction between the immune system and skin cells causes chronic inflammation of the skin [1, 3, 4]. The formation and maintenance of psoriatic plaques results from an imbalance between anti- inflammatory molecules that promote leaking of leukocyte and proliferation of keratinocytes. The relationship between genetic predisposition and inflammatory activity in this disease become more pronounced when the susceptibility locus associated with activation or differentiation of T cell is identified [1, 2]. On the other hand, vascular changes observed in the lesions are thought to be an important feature of the disease. In fact, they are a key component not only as a result of the disease, but also it can promote skin inflammation through the proliferation of leukocytes. An increased angiogenesis caused by psoriatic skin cells and activation of endothelial cells through pro-inflammatory cytokines therefore has a significant relationship between the altered epidermis and the immunological component of the disease [1]. Pro-angiogenic factors stimulate endothelial cells, and I first began to release the proteolytic enzymes that break down the extracellular matrix, invasion, migration and proliferation of endothelial cells there, and finally to the differences in a new capillary tube [22]. The effect of netrin-1 on endothelial cell migration has been shown in different studies. Lu et al. They demonstrated the inhibitory effect of netrin-1 on endothelial cell migration using both the Transwell migration system and the wound healing model [13]. Treatment of growing vessels with netrin-1, especially in the end-cell leads to filopodial retraction, endothelial cell migration has been reported to include the role of netrin-1 antiangiogenic [13, 15]. However, other studies suggest that using similar models, netrin-1 has a positive effect on endothelial cell motions, and the authors suggest that unlike netrin-1, it will have pro- angiogenic activity [23, 24]. Since the same endothelial cell lines are cross-checked between different studies, these discrepancies can be caused by two reasons: 1. The use of different netrin-1 groups, but more likely cell treatment at different netrin concentrations. 2. Generally, the netrin-1 physiological concentration between assays is 50 ng to 150 ng / ml, since it was first measured in the chicken brain [9, 25].

In this way, Wilson and colleagues have performed migratory assays using gradient doses of netrin-1, and observed a maximal activity at 50 ng/ml. On the contrary, there was no significant increase in endothelial cell migration for netrin-1 concentration superior to 1 ng/ml. Interestingly, this is the minimal dose used in assays performed by Lu et al. meaning that at high doses netrin-1 could lose its ability to induce endothelial cells migration and even prevents endothelial cells movement [13]. Such biphasic “bellshaped” dose-reponse curve has already been observed for other angiogenic molecules such as endostatin, thrombospondin- 1 or angiopoietin and even for PDGF, FGF and VEGF on other cell types, such as mast cells for example [26, 27]. Thus, discrepancies observed upon the action of netrin-1 during angiogenesis likely results from the use of rightfully different doses of netrin- 1. It will be of key importance to determine in which range of concentration netrin-1 exerts its physiological activity in vivo and whether abnormally high levels of netrin-1 could occur under certain pathological conditions. In our study, we found that netrin-1 levels of the patients with psoriasis lower than healthy controls. Morever, netrin-1 correlated negatively with disease severity, disease duration, CRP and BMI in all psoriasis patients.. The decresed netrin possibly reflects the association between inflammation and angiogenesis in psoriasis. When the groups were divided according to the severity of the disease, the netrin levels was not different between mildly severe psoriasis patients and moderate- severe psoriasis patients. Such a contradictory finding of correlation according to other studies may be attributed to the differences in demographic and clinical characteristics of the reviewed patients, variations in biochemical measurement methods and heterogeneity in study populations.

In conclusion, we evaluated netrin-1 levels and their relationship with duration and severity of disease in psoriasis patients in this study. We founded that netrin-1 decreased in psoriasis patients. Cardiovascular risk factors such as higher BMI, dyslipidemia, FPG and CRP values were found to be more common in psoriasis patients. Additionally, psoriasis patients were demonstrated to have a significant negative correlation among netrin-1 levels, disease duration, PASI, CRP and BMI paramaters. Decreased netrin-1 levels may play a role in the pathogenesis of chronic inflammation and increased CVD risk in psoriasis patients. However, these findings should be interpreted carefully as the power of this study is limited by its relatively small cohort and lack of prospective data. Further studies are needed to determine whether netrin levels play a role in the inflammation of psoriasis and its role in metabolic diseases associated with this disease. Acknowledgements: We are grateful to participants in this study. Study limitations: This study is limited by its relatively small cohort.