Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts
Aims: To evaluate the role of autophagy in primary knee fibroblasts undergoing myofibroblast differentiation as an in vitro model of arthrofibrosis, a complication after total knee arthroplasty characterized by aberrant intra-articular scar tissue formation and limited range of motion. Methods: We...
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The British Editorial Society of Bone & Joint Surgery
2025-04-01
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| Series: | Bone & Joint Research |
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| Online Access: | https://online.boneandjoint.org.uk/doi/epdf/10.1302/2046-3758.144.BJR-2024-0312.R1 |
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| author | Oliver B. Dilger Mason F. Carstens Cole E. Bothun Ashley N. Payne Daniel J. Berry Joaquin Sanchez-Sotelo Mark E. Morrey Roman Thaler Amel Dudakovic Matthew P. Abdel |
| author_facet | Oliver B. Dilger Mason F. Carstens Cole E. Bothun Ashley N. Payne Daniel J. Berry Joaquin Sanchez-Sotelo Mark E. Morrey Roman Thaler Amel Dudakovic Matthew P. Abdel |
| author_sort | Oliver B. Dilger |
| collection | DOAJ |
| description | Aims: To evaluate the role of autophagy in primary knee fibroblasts undergoing myofibroblast differentiation as an in vitro model of arthrofibrosis, a complication after total knee arthroplasty characterized by aberrant intra-articular scar tissue formation and limited range of motion. Methods: We conducted a therapeutic screen of autophagic-modulating therapies in primary human knee fibroblasts undergoing transforming growth factor-beta 1 (TGF-β1)-mediated myofibroblast differentiation. Autophagy was induced pharmacologically with rapamycin or by amino acid deprivation. Picrosirius red staining was performed to quantify collagen deposition. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting were conducted to evaluate fibrotic gene expression levels. Results: Rapamycin, an mTOR complex 1 (mTORC1) inhibitor and autophagy inducer, reduced TGF-β1-mediated collagen deposition. Interestingly, we simultaneously report that myofibrogenic genes, including ACTA2, were highly upregulated following rapamycin-TGF-β1 treatment. When autophagy was induced through amino acid deprivation, we demonstrated suppressed extracellular matrix levels, fibrotic gene expression (e.g. ACTA2), and SMAD2 phosphorylation levels in TGF-β1-stimulated fibroblasts. Conclusion: Our findings demonstrate that the induction of cellular autophagy suppresses TGF-β1-induced collagen deposition in primary human knee fibroblasts. Taken together, these data suggest that cellular autophagy may be prophylactic against the pathogenesis of arthrofibrosis. Cite this article: Bone Joint Res 2025;14(4):328–337. |
| format | Article |
| id | doaj-art-de4ae838e2ae410baed47785ee2ca84e |
| institution | Kabale University |
| issn | 2046-3758 |
| language | English |
| publishDate | 2025-04-01 |
| publisher | The British Editorial Society of Bone & Joint Surgery |
| record_format | Article |
| series | Bone & Joint Research |
| spelling | doaj-art-de4ae838e2ae410baed47785ee2ca84e2025-08-20T03:53:23ZengThe British Editorial Society of Bone & Joint SurgeryBone & Joint Research2046-37582025-04-0114433134010.1302/2046-3758.144.BJR-2024-0312.R1Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblastsOliver B. Dilger0Mason F. Carstens1Cole E. Bothun2Ashley N. Payne3Daniel J. Berry4Joaquin Sanchez-Sotelo5Mark E. Morrey6Roman Thaler7Amel Dudakovic8Matthew P. Abdel9Department of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USADepartment of Orthopedic Surgery, Mayo Clinic, Rochester, Minnesota, USAAims: To evaluate the role of autophagy in primary knee fibroblasts undergoing myofibroblast differentiation as an in vitro model of arthrofibrosis, a complication after total knee arthroplasty characterized by aberrant intra-articular scar tissue formation and limited range of motion. Methods: We conducted a therapeutic screen of autophagic-modulating therapies in primary human knee fibroblasts undergoing transforming growth factor-beta 1 (TGF-β1)-mediated myofibroblast differentiation. Autophagy was induced pharmacologically with rapamycin or by amino acid deprivation. Picrosirius red staining was performed to quantify collagen deposition. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting were conducted to evaluate fibrotic gene expression levels. Results: Rapamycin, an mTOR complex 1 (mTORC1) inhibitor and autophagy inducer, reduced TGF-β1-mediated collagen deposition. Interestingly, we simultaneously report that myofibrogenic genes, including ACTA2, were highly upregulated following rapamycin-TGF-β1 treatment. When autophagy was induced through amino acid deprivation, we demonstrated suppressed extracellular matrix levels, fibrotic gene expression (e.g. ACTA2), and SMAD2 phosphorylation levels in TGF-β1-stimulated fibroblasts. Conclusion: Our findings demonstrate that the induction of cellular autophagy suppresses TGF-β1-induced collagen deposition in primary human knee fibroblasts. Taken together, these data suggest that cellular autophagy may be prophylactic against the pathogenesis of arthrofibrosis. Cite this article: Bone Joint Res 2025;14(4):328–337.https://online.boneandjoint.org.uk/doi/epdf/10.1302/2046-3758.144.BJR-2024-0312.R1arthrofibrosisautophagytotal knee arthroplasty (tka)myofibroblastextracellular matrixfibroblastsautophagytransforming growth factor-beta 1kneewestern blottingarthrofibrosisamino acidscollagensquantitative polymerase chain reaction |
| spellingShingle | Oliver B. Dilger Mason F. Carstens Cole E. Bothun Ashley N. Payne Daniel J. Berry Joaquin Sanchez-Sotelo Mark E. Morrey Roman Thaler Amel Dudakovic Matthew P. Abdel Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts Bone & Joint Research arthrofibrosis autophagy total knee arthroplasty (tka) myofibroblast extracellular matrix fibroblasts autophagy transforming growth factor-beta 1 knee western blotting arthrofibrosis amino acids collagens quantitative polymerase chain reaction |
| title | Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts |
| title_full | Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts |
| title_fullStr | Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts |
| title_full_unstemmed | Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts |
| title_short | Induction of cellular autophagy impairs TGF-β1-mediated extracellular matrix deposition in primary human knee fibroblasts |
| title_sort | induction of cellular autophagy impairs tgf β1 mediated extracellular matrix deposition in primary human knee fibroblasts |
| topic | arthrofibrosis autophagy total knee arthroplasty (tka) myofibroblast extracellular matrix fibroblasts autophagy transforming growth factor-beta 1 knee western blotting arthrofibrosis amino acids collagens quantitative polymerase chain reaction |
| url | https://online.boneandjoint.org.uk/doi/epdf/10.1302/2046-3758.144.BJR-2024-0312.R1 |
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