title: IL-4 administration exerts preventive effects via suppression of underlying inflammation and TNF-α-induced apoptosis in steroid-induced osteonecrosis.
abstract: Macrophages play an important role during the development of steroid-induced osteonecrosis. Interleukin (IL)-4 administration helped reduce the infiltration of M1 phenotypic macrophages and maintain the activation of M2 phenotypic macrophages, resulting in restriction of inflammation and decrease in osteocyte apoptosis. The results indicated the therapeutic potential of IL-4 in prevention of steroid-induced osteonecrosis.
  
  
  BACKGROUND
  Steroid-induced osteonecrosis (ON) is a debilitating disease characterized by the activation and infiltration of macrophages into the necrotic site. This study aimed to investigate the effects of IL-4 administration on macrophage polarization and the involved signaling pathways.
  
  
  METHODS
  Fifty-six BALB/c mice were randomly divided into two groups, group M (model group) and group MI (treatment group), each containing 28 mice. ON model was induced by the injection of methylprednisolone (MPS). The mice in group MI received intra-abdominal injections of 2 μg/100 g/day of rIL-4 for five consecutive days, following the administration of MPS. Osteonecrosis was verified by histopathological staining. The expression of tumor necrosis factor-alpha (TNF-α) was analyzed by ELISA and immunohistochemistry. The infiltration of M1/M2 macrophages was examined by the expression of specific makers of F4/80, CD11c, and CD206 protein. Cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and the apoptotic signal molecules such as STAT1 and caspase-3 were examined.
  
  
  RESULTS
  Histopathological observations indicated that IL-4 administration reduced the incidence of ON and the accumulation of osteoclasts. IL-4 administration inhibited the expression of TNF-α and reduced the infiltration of M1 phenotypic macrophages and maintained relatively high level of M2 phenotypic macrophages. Additionally, TUNEL assay suggested that IL-4 intervention could reduce the number of apoptotic cells in the necrotic zone. The anti-apoptotic mechanisms were related to STAT1 phosphorylation and the activation of caspase-3.
  
  
  CONCLUSIONS
  Il-4 administration could alleviate steroid associated ON in mice by inhibiting the inflammatory response, the infiltration of M1 phenotypic macrophages, and suppressing TNF-a-induced osteocytic apoptosis by inhibiting the STAT1-caspase-3 signal pathway.
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: X
authors.initials: X
authors.lastname: Wu
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: X
authors.initials: X
authors.lastname: Feng
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Y
authors.initials: Y
authors.lastname: He
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Y
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: S
authors.initials: S
authors.lastname: Yang
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: Z
authors.initials: Z
authors.lastname: Shao
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China.
authors.forename: C
authors.initials: C
authors.lastname: Yang
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China. wanghwh@126.com.
authors.forename: H
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China. 56138779@qq.com.
authors.forename: Z
authors.initials: Z
authors.lastname: Ye
mesh_terms: D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology / Q000627:therapeutic use*; D053148:Caspase 3 / Q000378:metabolism; D002478:Cells, Cultured; D004195:Disease Models, Animal; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005938:Glucocorticoids; D007249:Inflammation / Q000188:drug therapy*; D015847:Interleukin-4 / Q000494:pharmacology / Q000627:therapeutic use*; D008264:Macrophages / Q000187:drug effects; D008775:Methylprednisolone; D008807:Mice, Inbred BALB C; D010010:Osteoclasts / Q000187:drug effects / Q000473:pathology; D010020:Osteonecrosis / Q000139:chemically induced / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D011994:Recombinant Proteins / Q000494:pharmacology / Q000627:therapeutic use; D050794:STAT1 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D000071681:Tartrate-Resistant Acid Phosphatase / Q000097:blood; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors* / Q000378:metabolism / Q000494:pharmacology
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D005938:Glucocorticoids; D011994:Recombinant Proteins; D050794:STAT1 Transcription Factor; C494080:Stat1 protein, mouse; D014409:Tumor Necrosis Factor-alpha; D015847:Interleukin-4; D000071681:Tartrate-Resistant Acid Phosphatase; C505409:Casp3 protein, mouse; D053148:Caspase 3; D008775:Methylprednisolone
keywords: Apoptosis; IL-4; Macrophages; Osteonecrosis; TNF-α