title: Endometriotic mesenchymal stem cells significantly promote fibrogenesis in ovarian endometrioma through the Wnt/β-catenin pathway by paracrine production of TGF-β1 and Wnt1.
abstract: Are endometriotic mesenchymal stem cells (Ecto-MSCs) involved in the fibrosis of ovarian endometrioma?
  
  
  
  Ecto-MSCs enhanced the fibrotic behavior of stromal cells in ovarian endometrioma through the Wnt/β-catenin pathway by paracrine production of transforming growth factor-β1 (TGF-β1) and Wnt1.
  
  
  
  Endometriosis is characterized by ectopic outgrowth of endometrial stroma and glands surrounded by dense fibrous tissues. The pathogenesis of endometriosis, especially ovarian endometrioma-associated fibrosis, is still unknown.
  
  
  
  We analyzed endometrial samples from 15 patients of reproductive age with ovarian endometrioma and normal menstrual cycles. A total of 54 nude mice received a single injection of proliferative endometrial fragments from 14 individuals without endometriosis.
  
  
  
  Conditioned medium (CM) was collected from endometrial mesenchymal stem cells (Euto-MSCs) and Ecto-MSCs. The effects of CM on cell proliferation, migration, invasion and collagen gel contraction of endometrial and endometriotic stromal cells (Euto- and Ecto-ESCs) in ovarian endometrioma were evaluated by cell counting kit-8, transwell and collagen gel contraction assays. Effects of CM on fibrotic markers' expression [including α-smooth muscle actin, Type I collagen, connective tissue growth factor and fibronectin (FN)] in Euto- and Ecto-ESCs were determined by real-time reverse-transcription-polymerase chain reaction and western blotting. Additionally, fibrogenic effects of Ecto-MSC CM treatment on endometriotic implants were analyzed using a xenograft model of endometriosis in immunodeficient nude mice.
  
  
  
  Our results demonstrated that Ecto-MSC CM significantly promoted cell proliferation, migration, invasion and collagen gel contraction of Euto- and Ecto-ESCs from patients with ovarian endometrioma compared with control and Euto-MSC CM. Expression levels of fibrotic markers in Euto- and Ecto-ESCs were dramatically elevated after treatment with Ecto-MSC CM. Ecto-MSCs secreted higher levels of TGF-β1 and Wnt1 compared with Euto-MSCs. Furthermore, both TGF-β1 and Wnt1 significantly increased expression of fibrotic markers in Euto- and Ecto-ESCs, which was reversed by an anti-TGF-β1 antibody or Wnt1 negative regulator, Dickkopf-related protein 1 (Dkk1). Mechanistic studies demonstrated that Wnt/β-catenin signaling pathways in stromal cells were activated by Ecto-MSC CM. Animal experiments showed that TGF-β1 and Wnt1 as well as Ecto-MSC CM markedly increased the expression of FN and collagen I, which enhanced the progression of fibrosis in endometriosis.
  
  
  
  To our knowledge, this is the first study to identify the role of Ecto-MSCs in the pathogenesis of fibrosis in ovarian endometrioma. However, numerous other growth factors and cell types may also be involved in the pathogenesis. Therefore, further studies are required to elucidate the paracrine effects of cells in ovarian endometrioma.
  
  
  
  Ecto-MSCs may be involved in the pathogenesis of fibrosis in ovarian endometrioma.
  
  
  
  This study was supported in part by the National Natural Science Foundation of China (81471505 and 81270657). No competing interests are declared.
authors.affiliation: Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, No. 3 Qingchun East Road, Jianggan District, Hangzhou 310016, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Li
authors.affiliation: Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, No. 3 Qingchun East Road, Jianggan District, Hangzhou 310016, China.
authors.forename: Yongdong
authors.initials: Y
authors.lastname: Dai
authors.affiliation: Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, No. 3 Qingchun East Road, Jianggan District, Hangzhou 310016, China.
authors.forename: Haiyan
authors.initials: H
authors.lastname: Zhu
authors.affiliation: Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, No. 3 Qingchun East Road, Jianggan District, Hangzhou 310016, China.
authors.forename: Yinshen
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, No. 3 Qingchun East Road, Jianggan District, Hangzhou 310016, China zhangsongying@126.com.
authors.forename: Songying
authors.initials: S
authors.lastname: Zhang
mesh_terms: D000818:Animals; D002465:Cell Movement; D049109:Cell Proliferation; D003094:Collagen / Q000096:biosynthesis; D017077:Culture Media, Conditioned; D004715:Endometriosis / Q000378:metabolism / Q000473:pathology*; D005260:Female; D005353:Fibronectins / Q000096:biosynthesis; D064593:Heterografts; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D019899:Paracrine Communication; D053773:Transforming Growth Factor beta1 / Q000096:biosynthesis; D060449:Wnt Signaling Pathway; D051155:Wnt1 Protein / Q000096:biosynthesis; D051176:beta Catenin / Q000378:metabolism*
chemical_list: D017077:Culture Media, Conditioned; D005353:Fibronectins; D053773:Transforming Growth Factor beta1; D051155:Wnt1 Protein; D051176:beta Catenin; D003094:Collagen
keywords: TGF-β1; Wnt1; fibrosis; mesenchymal stem cells; ovarian endometrioma