TRAIL: A New Therapeutic Drug Target In Psoriasis

Psoriasis is a T-lymphocyte-mediated and multifactorial autoimmune disease of the skin, including heredity and circumstance. It has a bad effect on people,s physical and mental health and normal life. Recently, TRAIL has been a lot research issue in skin pharmacology ,which it has done a lot research on disease associated with skin proliferation, inflammation ,immunity and so forth. Skin is prone to diseases associated with hypersensitiveness and inflammation. The pathogenesis is complicated. These reactions are to involve in keratinocytes and fibroblasts. TRAIL has a close relation with skin proliferation and differentiation, which retains homeostasis. It plays a important role in regulating skin inflammation. The importance of TRAIL in psoriasis pathogenesis and the studies with respect to a new therapeutic drug target in psoriasis would provide some theoretical evidences. Those are of important significance in increasing the effective and low- recurrence drugs. What is more, It would develop some new study fields and offer some original ideas. Immunohistochemical and Realtime PCR methods were established in this study to detect the expressions of TRAIL and to analyze the expression features of the lesions of psoriatic patient , healthy human skin tissue and fibroblasts. MTT , Annexin V-FITC and immunohistochemical methods were used to to study the inductive effect of sulfur mustard on human epidermal fibroblasts and explore its mechanism of treating psoriasis. Constructing Guinea pigs to produce the psoriasis-like models were used to dective the improvement in psoriasis-like skin pathology and its mechanism of treating psoriasis. Using genes of microarray We constructed the gene adjacency matrix A which has the gene information. The matrix reflects correlations among genes. Through computing them, we would get the gene adjacency matrix. We selected genes of the significant differential genes between psoriasis and normal samples for constructing co-expression network. The network edges were specified to be more than 0.9 of coefficient correlation for obtaining strong relationships of gene co-expression, which aim to find out key gene in psoriasis. Simultaneously, we may do some deep research wheather TRAIL is a new therapeutic drug target in psoriasis and the relations between TRAIL and other genes in the gene network.Part I The expressions of TRAIL in the lesions of psoriatic patient , healthy human skin tissue and fibroblasts.At first, we collect lesions of psoriatic patient and healthy human skin tissue. HE staining, immunohistochemical, realtime PCR methods to confirm the different expression between lesions of psoriatic patient and healthy human skin tissue. The results illustrated that:1. We confirm the cases of psoriasis by HE staining.The results show that skin is characteristed with lhyperkeratosis, parakeratosis and Munro microabscess. granular layer goes thinner or disapear dramaticly. spinous layer is thicker. The epidermis extend to dermis layer. The top of dermal papilla pose drumstick and the upper spinous layer is thinner. The blood capillary is extension and around them we may see lymphocyte and neutrophilic granulocyte.2. These positive cell fractions were calculated for both epidermal and dermal cells in psoriatic lesions and healthy control. There was no TRAIL positive expression in the epidermis of psoriatic skin lesions. On the contrast, there was significantly positive expression seen in healthy skin. The results of this study were indicative of decreased apoptosis in psoriatic epidermis with development. These findings support the idea that decreased apoptosis was seen in psoriatic epidermal cells with development.3. There are brown positive expressions in fibroblasts.4. TRAIL can all be detected in the lesions of psoriatic patient and healthy human skin tissue,while the expression levels of TRAIL have a significant difference.Part II Study on apoptosis of human fibroblasts and release of IL-10,TRAIL induced by sulfur mustardThe part of this study is aim to the feasibility that TRAIL is a new parameter in the pathogenesis of psoriatic.The inhibitive effect of sulfur mustard on proliferation of HB was detected by MTT method,its effect on apoptotic index of HB was studied by Annexin V-FITC method,its effect on the expression of tumor necrosis factor (TNF)-related apoptosis-inducing ligand receptor one (TRAILR1) in human Fibroblasts (FB) was exmamined by immunohistochemical method.Results and the release of IL-10,TRAIL was detected by ELISA method. The results suggest that :1. Different concentrations of SM had significant and dose-dependent inhibitive effect on the growth of FB(P<0.01).The apoptotic index of FB increased as the concentration of SM was increased.2. TRAIL protein in human fibroblasts expressed positively correlated to the concentration of SM. It mainly expressed in cell membrane and cytoplasm.3. ELISA assays have elucidated the time-effect and does-effect relation in the secretion of IL-10,TRAIL in human fibroblasts after the treatment of SM.The results showed that the human fibroblasts can secret some IL-10,TRAIL in normal status.And after treatment with different does of SM for 0,24,48,72h, the concentration of IL-10,TRAIL in the supernatant fluids are elevated,and there is a more significant increasing trend for IL-10,TRAIL. Moreover,it had significant and dose-dependent and time-dependent trend.Part III Study on mechanism of action in Guinea pigs to produce the psoriasis-like models by sulfur mustardThe study in this part we produce the psoriasis-like models by 5% propranolol emulsion was applied on ear ,back of Guinea pigs . This model show that skin is characteristed with lhyperkeratosis, parakeratosis, spinous layer is thicker and so forth. From a global view we investigate the changes in HE pathology staining, the expression of TRAIL and the secretion of blood serum soluble TRAIL, which aims to potential mechanism of action on psoriasis by sulfur mustard.The results suggest that :1. We produce Guinea pigs to produce the psoriasis-like models successfully. This animal models fit for the requirement in this study, The evidences is provided from the results of HE staining and the thickness in Guinea pigs.2. To a large extent, the psoriatic pathology symptom have great improvement after the treatment of sulfur mustard and of TRAIL subcutaneous injection in ear ,back. Moreover,it inhibits the proliferation of skin cell and the infiltration of inflammatory cell. 3. In the spinous layer and granular layer, There are brown positive expressions of TRAIL after the treatment of sulfur mustard and sTRAIL subcutaneous injection in ear ,back. It mainly expressed in cell membrane and cytoplasm.Part IV Microarray: construction And Topological Analysis of Gene Co-Expression Network modeling in psoriasisThe study of this part we can look snapshots of protein-protein interaction, gene expression regulatory network and metabolism networks among different groups. To constructe two sorts of networks refered to psoriasis and normal group, we transform the normalized expression value of the Pearson correlation into measures of pair wise connection strengths. We selected genes of the significant differential genes between psoriasis and normal samples for constructing co-expression network. we identify differential genes between psoriasis and normal tissues , construct And Topologicaly Analyze of Gene Co-Expression Network ,which aim to find out key gene in psoriasis. We want to confirm the relationships between TRAIL and other genes in the gene network. The results illustrated that.1. According to normailzed signal of psoriasis and normal samples, we separately compared average normal signal of each genes within psoriasis or normal tissues groups and get signal ratio of genes between psoriasis vs normal tissue groups. 288 differetial genes between psoriasis and normal tissues were from filtering genes with ratio less than 2 or larger tha 0.5.2. According to degree distribution of genes in normal and psoriasis sample network, we identified the core regulatory genes with highest i-core value and degree.We find some important genes including MET,TRAIL,SULT1B1 and so on. Most of genes are attributed to protein folding, lipid metabolic process, steroid metabolic process, transport, oxidation reduction, sulfur metabolic process, multicellular organismal development, electron transport, carbohydrate metabolic process, signal transduction, regulation of transcription, inflammatory response, regulation of apoptosis by gene ontology hierarchical category. Simultaneously, some key regulation correlation were detected.3. To evaluate difference from psoriasis and normal tissues with insight on whole interaction networks, we compare 2 arguments of network structure including density, cluster coefficient between psoriasis and normal tissues .The network of normal tissues possessed much more interactions than psoriasis according to comparison of density. However, genes in two networks had similar capability of interactions independently on center regulatory genes based on comparison of cluster coefficient. As a result, we consider that genes in psoriasis tissues may more difficultly in connecting to archive normal process.