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Bronchial smooth muscle hypoplasia in mouse embryonic lungs exposed to a laminin beta1 chain antisense
1Department of Pathology, Wayne State University School of Medicine, Detroit 48201, USA.
Abstract:
We used an antisense oligonucleotide (ODN) to inhibit laminin (LM) beta1 chain synthesis in mouse embryonic lung explants and cell cultures. The ODN spanned 17 bases located 13 bases downstream the initiation codon and contained phosphorothioate and C-5 propynyl pyrimidine modifications. Penetration of the ODN into the lung explants was confirmed by fluorescein isothiocyanate (FITC) tagging. 50 microM of antisense ODN decreased LM beta1 chain synthesis by 82+/-6.9% with no significant changes in the synthesis of other LM chains. The same antisense probe but without C-5 propynyl pyrimidine modification, another 17-mer ODN complementary to the LM beta1 initiation codon, and a 17-mer ODN complementary to the LM alpha1 initiation codon had no antisense activity. Lung explants exposed to the active LM beta1 antisense ODN showed decreased LM-1 and collagen type IV deposition at the epithelial-mesenchymal interface and an arrest in bronchial smooth muscle (SM) development. Histological examination and cell motility assays suggested that this arrest was due to impaired spreading and migration of SM cell precursors over the defective basement membrane (BM). Our studies indicate that beta1-chain containing LMs play a role in bronchial myogenesis.
Insights
Antisense inhibition of laminin beta1 chain synthesis disrupted bronchial smooth muscle development in mouse lungs. This suggests beta1-containing laminins are crucial for bronchial myogenesis and basement membrane formation.
Area of Science:
- Developmental Biology
- Molecular Biology
- Cell Biology
Background:
- Laminins (LM) are critical extracellular matrix proteins involved in cell adhesion, differentiation, and tissue development.
- The specific role of beta1-chain containing laminins in bronchial smooth muscle (SM) development is not fully understood.
Purpose of the Study:
- To investigate the function of laminin beta1 chain in mouse embryonic lung development.
- To determine the effect of inhibiting laminin beta1 chain synthesis on bronchial smooth muscle formation and function.
Main Methods:
- Utilized a modified antisense oligonucleotide (ODN) targeting laminin beta1 chain synthesis in mouse embryonic lung explants and cell cultures.
- Confirmed ODN penetration using fluorescein isothiocyanate (FITC) tagging.
- Assessed laminin chain synthesis, extracellular matrix deposition (LM-1, collagen type IV), and smooth muscle development through histological examination and cell motility assays.
Main Results:
- Antisense ODN significantly reduced laminin beta1 chain synthesis by 82% without affecting other LM chains.
- Exposure to the active antisense ODN led to decreased LM-1 and collagen type IV deposition.
- Observed an arrest in bronchial smooth muscle development, attributed to impaired migration of smooth muscle cell precursors.
Conclusions:
- Beta1-chain containing laminins are essential for proper bronchial smooth muscle development.
- Defective basement membrane formation due to reduced laminin beta1 impacts smooth muscle cell precursor migration and spreading.
- These findings highlight the critical role of laminin beta1 in bronchial myogenesis.