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Abnormalities in fibrillin 1-containing microfibrils in dermal fibroblast cultures from patients with systemic
D D Wallis1, F K Tan, C M Kielty
1University of Texas-Houston Medical School, 77030, USA.
Objective:
To determine if there are abnormalities in fibrillin 1-containing microfibrils in the extracellular matrix (ECM) of primary dermal fibroblasts explanted from patients with systemic sclerosis (SSc).
Methods:
Explanted fibroblasts from unaffected skin of 12 SSc patients were used to examine fibrillin 1-containing microfibrils by immunofluorescence (IF) using a monoclonal antibody (mAb) to fibrillin 1. Metabolic labeling of the fibroblast cultures was used to study the synthesis, secretion, and processing of fibrillin 1, as well as to observe microfibril formation and stability. Microfibrils elaborated by the SSc cells were analyzed by electron microscopy for ultrastructural abnormalities, and the results were confirmed by immunoblotting.
Results:
Control and SSc fibroblasts displayed a prominent meshwork of fibrillin 1-containing microfibrils when visualized by IF using a fibrillin 1 mAb. Paradoxically, metabolic studies indicated a paucity of fibrillin 1 in the ECM in the majority of the SSc fibroblast strains. Subsequent rotary-shadowed electron microscopy revealed reduced amounts of and ultrastructural abnormalities in the microfibrils elaborated by all strains of SSc cells. Immunoblots confirmed the lack of the high molecular weight form of fibrillin 1 in the SSc fibroblasts of Choctaw American Indians. Finally, in vitro studies indicated that the amount of fibrillin 1 in the ECM of SSc cells diminished at a faster rate than the amount of fibrillin 1 in the ECM of control cells with time.
Conclusion:
Although SSc fibroblasts assemble microfibrils, these microfibrils are unstable, suggesting that an inherent defect of fibrillin 1-containing microfibrils may play a role in the pathogenesis of SSc.
Insights
Systemic sclerosis (SSc) fibroblasts produce unstable microfibrils due to inherent defects in fibrillin-1. This instability suggests a role for these extracellular matrix abnormalities in SSc pathogenesis.
Area of Science:
- Extracellular matrix research
- Connective tissue disorders
- Fibroblast biology
Background:
- Systemic sclerosis (SSc) is a complex autoimmune disease characterized by fibrosis.
- The extracellular matrix (ECM) plays a critical role in tissue structure and function.
- Fibrillin-1 is a major component of microfibrils in the ECM, essential for tissue elasticity.
Purpose of the Study:
- To investigate potential abnormalities in fibrillin-1-containing microfibrils within the ECM of dermal fibroblasts from SSc patients.
- To determine if SSc fibroblasts exhibit defects in the synthesis, secretion, or stability of fibrillin-1.
Main Methods:
- Immunofluorescence microscopy was used to visualize fibrillin-1 microfibrils in explanted SSc fibroblasts.
- Metabolic labeling assessed fibrillin-1 synthesis, secretion, and processing.
- Electron microscopy and immunoblotting analyzed microfibril ultrastructure and fibrillin-1 forms.
- In vitro degradation studies evaluated ECM fibrillin-1 stability over time.
Main Results:
- SSc fibroblasts showed reduced amounts and ultrastructural abnormalities in fibrillin-1 microfibrils compared to controls.
- Metabolic studies revealed a paucity of fibrillin-1 in the ECM of most SSc fibroblast strains.
- Immunoblots confirmed the absence of high molecular weight fibrillin-1 in some SSc fibroblasts.
- In vitro experiments demonstrated faster degradation of fibrillin-1 in the SSc fibroblast ECM.
Conclusions:
- SSc fibroblasts can assemble microfibrils, but these structures are inherently unstable.
- The observed instability of fibrillin-1-containing microfibrils suggests a potential role in the pathogenesis of systemic sclerosis.
- These findings highlight intrinsic defects in ECM components as contributors to SSc.