title: Corroborating evidence for platelet-induced epithelial-mesenchymal transition and fibroblast-to-myofibroblast transdifferentiation in the development of adenomyosis.
abstract: OBJECTIVE
  Do platelets play any role in the development of adenomyosis?
  
  
  CONCLUSIONS
  As in endometriosis, adenomyotic lesions show significantly increased platelet aggregation, increased expression of transforming growth factor (TGF)-β1, phosphorylated Smad3, markers of epithelial-mesenchymal transition (EMT) and fibroblast-to-myofibroblast transdifferentiation (FMT), and smooth muscle metaplasia (SMM), in conjunction with increased fibrosis as compared with normal endometrium.
  
  
  BACKGROUND
  Both EMT and FMT are known to play vital roles in fibrogenesis in general and in endometriosis in particular. EMT has been implicated in the development of adenomyosis. SMM is universally seen in endometriosis and also in adenomyosis, and is correlated positively with the extent of fibrosis. However, there has been no published study on the role of platelets in fibrogenesis in adenomyosis, even though adenomyotic lesions undergo repeated cycles of tissue injury and repair, which suggests the involvement of platelets and their possible roles in fibrogenesis.
  
  
  METHODS
  Cross-sectional studies of ectopic endometrial and control endometrial tissue samples from three sets of women with and without adenomyosis (n= 34 and 20, 12 and 10, and 8 and 8, respectively) were carried out from 2014 to 2015.
  
  
  METHODS
  Immunohistochemistry analysis of ectopic endometrial tissues from women with (n= 34) and without (n= 20) adenomyosis with respect to biomarkers of EMT, FMT and highly differentiated smooth muscle cells as well as TGF-β1, phosphorylated Smad3, markers of proliferation, angiogenesis and extracellular matrix (ECM) deposits. Masson trichrome staining, Van Gieson staining and Pico-Sirius staining were performed to evaluate and quantify the extent of fibrosis in lesions. Progesterone receptor isoform B (PR-B) staining also was performed. In addition, CD42b-positive platelets in ectopic (n= 12) and control (n= 10) endometrium were counted by confocal microscopy and compared. The protein expression levels of TGF-β1 and phosphorylated Smad3 in both ectopic (n= 8) and control (n= 8) endometrium were measured by western blot analysis. Immunofluorescent staining of both platelets and hepatocyte growth factor (HGF) was also performed for adenomyotic tissue samples (n= 10).
  
  
  RESULTS
  Adenomyotic lesions had a significantly higher extent of platelet aggregation and increased staining for TGF-β1 and phosphorylated Smad3 (both P-values <0.001 versus control). In addition, E-cadherin staining was decreased while vimentin staining in adenomyotic epithelial cells was increased, along with increased staining of proliferating cell nuclear antigen, vascular endothelial growth factor and CD31 (all P-values <0.001), markers of proliferation and angiogenesis. Staining for α-SMA, a marker for myofibroblast, desmin, smooth muscle myosin heavy chain and oxytocin receptor was significantly increased in adenomyotic lesions versus control, concomitant with increased staining of collagen I and lysyl oxidase (all P-values <0.001). Histochemistry analysis indicates that the extent of fibrosis is high in adenomyotic lesions (P < 0.001), and the extent appeared to correlate negatively with the microvessel density (P < 0.05). PR-B staining was significantly decreased in adenomyotic lesion as compared with control endometrium (P < 0.001). Platelets and HGF were co-localized mostly in the stromal component of adenomyotic lesions, near the glandular epithelium.
  
  
  CONCLUSIONS
  The results are limited by the cross-sectional nature of the study and the use of histochemistry and immunohistochemistry analyses only, but nonetheless is a validation of our previous finding in mouse experiments.
  
  
  CONCLUSIONS
  The data presented are consistent with the notion that platelet-induced activation of the TGF-β/Smad signaling pathway may be a driving force in EMT, FMT and SMM in the development of adenomyosis, leading to fibrosis. This study provides the first piece of evidence that adenomyotic lesions are wounds that undergo repeated injury and healing, and, as such, platelets play critical roles in the development of adenomyosis by promoting proliferation, angiogenesis, increasing ECM deposits, and SMM, resulting in fibrosis. Platelets may also be involved in uterine hyperactivity and myometrial hyperinnervation. Our results provide one explanation as to why adenomyosis is a challenge for medical treatment, and shed new light onto the pathophysiology of adenomyosis.
  
  
  BACKGROUND
  Support for data collection and analysis was provided by grants from the National Science Foundation of China. None of the authors has anything to disclose.
authors.affiliation: Shanghai OB/GYN Hospital, Fudan University, Shanghai 200011, China Shanghai Key Laboratory of Female Reproductive Endocrine-Related Diseases, Fudan University, Shanghai, China.
authors.forename: Xishi
authors.initials: X
authors.lastname: Liu
authors.affiliation: Shanghai OB/GYN Hospital, Fudan University, Shanghai 200011, China.
authors.forename: Minhong
authors.initials: M
authors.lastname: Shen
authors.affiliation: Shanghai OB/GYN Hospital, Fudan University, Shanghai 200011, China.
authors.forename: Qiuming
authors.initials: Q
authors.lastname: Qi
authors.affiliation: Department of Anatomy, Histology and Embryology, Shanghai Medical Collage, Fudan University, Shanghai, China.
authors.forename: Hongqi
authors.initials: H
authors.lastname: Zhang
authors.affiliation: Shanghai OB/GYN Hospital, Fudan University, Shanghai 200011, China Shanghai Key Laboratory of Female Reproductive Endocrine-Related Diseases, Fudan University, Shanghai, China hoxa10@outlook.com.
authors.forename: Sun-Wei
authors.initials: SW
authors.lastname: Guo
mesh_terms: D062788:Adenomyosis / Q000378:metabolism / Q000473:pathology*; D000328:Adult; D015415:Biomarkers / Q000378:metabolism; D001792:Blood Platelets / Q000378:metabolism / Q000473:pathology; D049109:Cell Proliferation; D054338:Cell Transdifferentiation; D002478:Cells, Cultured; D003430:Cross-Sectional Studies; D004717:Endometrium / Q000098:blood supply / Q000378:metabolism / Q000473:pathology*; D058750:Epithelial-Mesenchymal Transition; D005260:Female; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005355:Fibrosis; D006801:Humans; D055806:Microvessels / Q000378:metabolism / Q000473:pathology; D008875:Middle Aged; D058628:Myofibroblasts / Q000378:metabolism / Q000473:pathology*; D009215:Myometrium / Q000098:blood supply / Q000473:pathology*; D009389:Neovascularization, Pathologic / Q000378:metabolism / Q000473:pathology; D010974:Platelet Aggregation
chemical_list: D015415:Biomarkers
keywords: adenomyosis; epithelial-mesenchymal transition; fibroblast-to-myofibroblast transdifferentiation; fibrosis; platelet; smooth muscle metaplasia; transforming growth factor-β1