Research

Laminin-332 alters connexin profile, dye coupling and intercellular Ca²⁺ waves in ciliated tracheal epithelial cells

Brant E Isakson^1,2* , Colin E Olsen^3* and Scott Boitano^3,4

¹  Department of Molecular Physiology and Biological Physics, University of Virginia School of Medicine, University of Virginia Charlottesville, Virginia 22908, USA

²  Robert M. Berne Cardiovascular Research Center, University of Virginia School of Medicine, University of Virginia, Charlottesville, Virginia 22908, USA

³  Arizona Respiratory Center, Arizona Health Sciences Center, University of Arizona, Tucson, Arizona 85724, USA

⁴  Department of Physiology, Arizona Health Sciences Center, University of Arizona, Tucson, Arizona 85724, USA

author email corresponding author email* Contributed equally

Respiratory Research 2006, 7:105doi:10.1186/1465-9921-7-105

Published:	2 August 2006

Abstract

Background

Tracheal epithelial cells are anchored to a dynamic basement membrane that contains a variety of extracellular matrix proteins including collagens and laminins. During development, wound repair and disease of the airway epithelium, significant changes in extracellular matrix proteins may directly affect cell migration, differentiation and events mediated by intercellular communication. We hypothesized that alterations in cell matrix, specifically type I collagen and laminin α3β3γ2 (LM-332) proteins within the matrix, directly affect intercellular communication in ciliated rabbit tracheal epithelial cells (RTEC).

Methods

Functional coupling of RTEC was monitored by microinjection of the negatively charged fluorescent dyes, Lucifer Yellow and Alexa 350, into ciliated RTEC grown on either a LM-332/collagen or collagen matrix. Coupling of physiologically significant molecules was evaluated by the mechanism and extent of propagated intercellular Ca²⁺ waves. Expression of connexin (Cx) mRNA and proteins were assayed by reverse transcriptase – polymerase chain reaction and immunocytochemistry, respectively.

Results

When compared to RTEC grown on collagen alone, RTEC grown on LM-332/collagen displayed a significant increase in dye transfer. Although mechanical stimulation of RTEC grown on either LM-332/collagen or collagen alone resulted in intercellular Ca²⁺ waves, the mechanism of transfer was dependent on matrix: RTEC grown on LM-332/collagen propagated Ca²⁺waves via extracellular purinergic signaling whereas RTEC grown on collagen used gap junctions. Comparison of RTEC grown on collagen or LM-332/collagen matrices revealed a reorganization of Cx26, Cx43 and Cx46 proteins.

Conclusion

Alterations in airway basement membrane proteins such as LM-332 can induce connexin reorganizations and result in altered cellular communication mechanisms that could contribute to airway tissue function.