Effect of endothelin-1 on croton oil-induced granulation tissue in the rat. A pharmacologic and immunohistochemical study.
BACKGROUND
To date no attempts have been made to determine the role of the endothelial cell derived product, endothelin-1 (ET-1) in granulation tissue development. This study investigates the cellular immunolocalization of ET-1 and its pharmacologic effect on myofibroblast-mediated rat croton oil-induced granulation tissue contraction.


METHODS
The distribution, cellular localization and temporal production of ET-1 in the tissues was determined by immunohistochemistry at days 7, 14, 21, and 28. The contractile response of the granulation tissue to ET-1 was tested over the same time period, and it effects modified by use of calcium antagonists. The pharmacologic profile was correlated to the ultrastructural development of contractile fibroblast-like cells within the tissue.


RESULTS
Endothelin-1 caused reversible concentration-dependent contraction of the granulation tissue. The 21-day granulation tissue was the most responsive, with a maximum increase in tension of 458.9 +/- 41.1 mg; this response could be inhibited by use of calcium antagonists. Of the calcium antagonists tested, verapamil (1 x 10(-4) M) was the most potent inhibitor, giving a 43% reduction in maximum amplitude of the response. It is suggested that entry of extracellular calcium via the L-type potential operated calcium channel, is involved in ET-1 induced responses in contractile fibroblast-like cells or myofibroblasts. Ultrastructural analysis showed a correlation between the pharmacologic sensitivity of the tissue and the development of contractile fibroblast-like cells. The number of cells expressing the phenotypic characteristics of a myofibroblast increased with time, and were first observed at day 7. Immunohistochemistry revealed the presence of increasing numbers of ET-1 labeled cells throughout the time course of study. The ET-1 positive cells were localized to the capillaries. Immunolabeling of serial sections with the rodent endothelial cell specific lectin, Bandeiraea simplicifolia isolectin B4 and factor VIII-related antigen, confirmed the specific localization of ET-1 to endothelial cells.


CONCLUSIONS
We present evidence that ET-1 may be an endogenous modulator of myofibroblast-mediated granulation tissue contraction and that the use of calcium antagonists could afford a possible therapeutic control in the treatment of fibrocontractive diseases.
Appleton
I
I
Department of Experimental Pathology, William Harvey Research Institute, St. Bartholomew's Hospital Medical College, London, United Kingdom.
Tomlinson
A
A
Chander
C L
CL
Willoughby
D A
DA
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002454:Cell Differentiation; D003286:Contracture / Q000139:chemically induced / Q000378:metabolism / Q000473:pathology*; D003436:Croton Oil; D016232:Endothelins / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000494:pharmacology* / Q000502:physiology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006097:Granulation Tissue / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D007124:Immunoenzyme Techniques; D066298:In Vitro Techniques; D008297:Male; D051381:Rats; D017208:Rats, Wistar
D002121:Calcium Channel Blockers; D016232:Endothelins; D003436:Croton Oil