Investigating the Role of P311 in the Hypertrophic Scar Jianglin Tan1., Xu Peng1., Gaoxing Luo1,2., Bing Ma3*, Chuan Cao4, Weifeng He1,2, Shunzong Yuan1, Shirong Li4, John A. Wilkins5, Jun Wu1,2* 1 State Key Laboratory for Trauma, Burn and Combined Injury, Institute of Burn Research, Southwest Hospital, Third Military Medical University, Chongqing, China, 2 Chongqing Key Laboratory for Disease Proteomics, Chongqing, China, 3 Department of Plastic Surgery, First Affiliated Hospital of Chengdu Medical College, Chengdu, Sichuan Province, China, 4 Department of Plastic Surgery, Southwest Hospital, Third Military Medical University, Chongqing, China, 5 Manitoba Centre for Proteomics and Systems Biology, Department of Internal Medicine, University of Manitoba, Winnipeg, Canada Abstract The mechanisms of hypertrophic scar formation are not fully understood. We previously screened the differentially expressed genes of human hypertrophic scar tissue and identified P311 gene as upregulated. As the activities of P311 in human fibroblast function are unknown, we examined the distribution of it and the effects of forced expression or silencing of expression of P311. P311 expression was detected in fibroblast-like cells from the hypertrophic scar of burn injury patients but not in peripheral blood mononuclear cells, bone marrow mesenchymal stem cells, epidermal cells or normal skin dermal cells. Transfection of fibroblasts with P311 gene stimulated the expression of alpha-smooth muscle actin (a- SMA), TGF-b1 and a1(I) collagen (COL1A1), and enhanced the contraction of fibroblast populated collagen lattices (FPCL). In contrast, interference of fibroblast P311 gene expression decreased the TGF-b1 mRNA expression and reduced the contraction of fibroblasts in FPCL. These results suggest that P311 may be involved in the pathogenesis of hypertrophic scar via induction of a myofibroblastic phenotype and of functions such as TGF-b1 expression. P311 could be a novel target for the control of hypertrophic scar development. Citation: Tan J, Peng X, Luo G, Ma B, Cao C, et al. (2010) Investigating the Role of P311 in the Hypertrophic Scar. PLoS ONE 5(4): e9995. doi:10.1371/ journal.pone.0009995 Editor: Christophe Egles, Tufts University, United States of America Received January 27, 2010; Accepted March 12, 2010; Published April 9, 2010 Copyright: 2010 Tan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by NSFC (No: 30872695 and 30672174), and partly by ‘‘863’’ project in China (2006AA02A121), by State Key Lab Funding in China (SKLZZ200808 to J.W., SKLZZ200820 to G.L.). The relevant grant number is 5. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: dmabing@hotmail.com (BM); junwupro1@yahoo.com.cn (JW) . These authors contributed equally to this work. Introduction Hypertrophic scars are a common problem after an injury. Clinically they present as erythematous or elevated firm plaques that remain confined to the damaged area. However if the formation of hypertrophic scar occurs in organs such as the lungs, liver, kidneys, joints, and skin, it can interfere with organ function, and ultimately result in organ failure. Even with the increasing knowledge of wound healing, hypertrophic scars are still difficult to prevent and to treat [1]. The formation of hypertrophic scar represents a dysregulated response to cutaneous wound healing and is characterized by inflammation, excessive proliferation of fibroblasts, and abnormal deposition of extracellular matrix (ECM) proteins [2]. Despite the fact that intrinsic differences between normal skin fibroblasts and hypertrophic scar myofibroblasts have been demonstrated by many studies, the underlying mechanisms promoting scar formation remain to be elucidated. We previously screened for genes that were differentially expressed in hypertrophic scar and normal skin tissues of burn patients using gene microarrays and identified 97 differentially expressed genes. Among these P311 gene expression increased conspicuously in hypertrophic scar tissue [3]. The P311 gene is mapped to the long arm of chromosome 5 and the protein it encodes for does not belong to any known protein family [4]. The N-terminus of this 8 kDa protein contains a PEST domain that is highly conserved among human, murine and chicken proteins. This domain is a binding site of the ubiquitin/proteasome pathway which is involved in the regulation of expression of transcription factors, cytokines and signal factors [5,6]. Pan et. al. reported that P311 protein and mRNA could be only found in myofibroblasts present in human wounds. Moreover, in their study the expression of P311 was associated with downreg- ulation of TGF-b1 and collagen type I expression in the murine cell lines NIH 3T3 and C3H10T1/2 [7]. However, further study of the possible role(s) of P311 in human fibroblast function is required as hypertrophic scar is a unique feature of humans which is absent in other species including mice [8,9,10]. Therefore, in the present study, we examined the effects of forced and silenced P311 gene expression on human fibroblasts functions in vitro. Materials and Methods Patient Inclusion Criteria Hypertrophic scar patients were selected according to the Vancouver Scar Scale (VSS) ranging from 10 score to 13 score [11]. 1.Tissues were collected within 1 year of the burn injury; 2. clinical manifestations: scar exhibited obvious hyperemia, red appearance, and obvious hypertrophy; patient experienced pruritus, pain, and dysesthesia of scar; local growth of the scars PLoS ONE | www.plosone.org 1 April 2010 | Volume 5 | Issue 4 | e9995