title: Increased CCL19 and CCL21 levels promote fibroblast ossification in ankylosing spondylitis hip ligament tissue.
abstract: BACKGROUND
  It is well-documented that both chemokine (C-C motif) ligand 19 (CCL19) and 21 (CCL21) mediate cell migration and angiogenesis in many diseases. However, these ligands' precise pathological role in ankylosing spondylitis (AS) has not been elucidated. The objective of this study was to examine the expression of CCL19 and CCL21 (CCL19/CCL21) in AS hip ligament tissue (LT) and determine their pathological functions.
  
  
  METHODS
  The expression levels of CCL19, CCL21 and their receptor CCR7 in AS (n = 31) and osteoarthritis (OA, n = 21) LT were analyzed via real-time polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC). The expression of CCL19, CCL21 and CCR7 in AS ligament fibroblasts was also detected. The proliferation of ligament fibroblasts was measured via a cell counting kit-8 (CCK8) assay after exogenous CCL19/CCL21 treatment. Additionally, the role of CCL19/CCL21 in osteogenesis was evaluated via RT-PCR and enzyme-linked immunosorbent assay (ELISA) in individual AS fibroblast cultures. Furthermore, the expression of the bone markers alkaline phosphatase (ALP), osteocalcin (OCN), collagenase I (COL1), integrin-binding sialoprotein (IBSP) and the key regulators runt-related transcription factor-2 (Runx-2) and osterix were investigated. Moreover, the CCL19/CCL21 levels in serum and LT were measured via ELISA.
  
  
  RESULTS
  The mRNA levels of CCL19/CCL21 in AS hip LT were significantly higher than that in OA LT, and IHC analysis revealed a similar result. Exogenous CCL19/CCL21 treatment did not affect the proliferation of ligament fibroblasts but significantly up-regulated the expression of bone markers, including ALP and OCN, and the key regulators Runx-2 and osterix. In addition, the serum levels of CCL19/CCL21 were apparently elevated in AS patients compared to healthy controls (HC), and the expression of the two chemokines correlated significantly in AS patients.
  
  
  CONCLUSIONS
  CCL19 and CCL21, two chemokines displaying significantly associated expression in serum, indicating a synergistic effect on AS pathogenesis, may function as promoters of ligament ossification in AS patients.
authors.forename: Yang
authors.initials: Y
authors.lastname: Qin
authors.forename: Li Da
authors.initials: LD
authors.lastname: He
authors.forename: Zhou Jian
authors.initials: ZJ
authors.lastname: Sheng
authors.forename: Miao Ming
authors.initials: MM
authors.lastname: Yong
authors.forename: Yang Sheng
authors.initials: YS
authors.lastname: Sheng
authors.affiliation: Department of Orthopedics, Changhai Hospital Affiliated to the Second Military Medical University, Changhai Road 168, Shanghai 200433, Yangpu district, P, R, China. xuwdshanghai@126.com.
authors.forename: Xu
authors.initials: X
authors.lastname: Wei Dong
authors.forename: Tong
authors.initials: T
authors.lastname: Wen Wen
authors.forename: Zou Yu
authors.initials: ZY
authors.lastname: Ming
mesh_terms: D000328:Adult; D015415:Biomarkers / Q000097:blood / Q000378:metabolism; D002478:Cells, Cultured; D054415:Chemokine CCL19 / Q000096:biosynthesis / Q000097:blood*; D054421:Chemokine CCL21 / Q000096:biosynthesis / Q000097:blood*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D006615:Hip / Q000473:pathology; D006801:Humans; D008022:Ligaments / Q000378:metabolism* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009999:Ossification, Heterotopic / Q000097:blood* / Q000175:diagnosis; D015207:Osteoarthritis, Hip / Q000097:blood / Q000175:diagnosis; D012333:RNA, Messenger / Q000096:biosynthesis / Q000097:blood; D013167:Spondylitis, Ankylosing / Q000097:blood* / Q000175:diagnosis; D055815:Young Adult
chemical_list: D015415:Biomarkers; C515460:CCL19 protein, human; C515478:CCL21 protein, human; D054415:Chemokine CCL19; D054421:Chemokine CCL21; D012333:RNA, Messenger