Regulation of thrombin-induced endothelial cell activation by bacterial toxins

C. E. Patterson, Joe GN Garcia

Research output: Contribution to journalArticle

23 Citations (Scopus)

Abstract

It has previously been shown that thrombin effects on endothelial cells can be mediated via G-proteins, which couple the thrombin receptor to several key physiological responses. As G-proteins are known targets of bacterial toxins, specific toxins were used to further characterize G-protein involvement in thrombin activation of bovine pulmonary arterial endothelial cells (BPAEC) and human umbilical vein endothelial cells (HUVEC). Homogenates were exposed to several bacterial toxins in the presence of 32P-NAD and ADP ribosylation of proteins determined by autoradiography of SDS-PAGE gels. Major substrates were a 40 kDa protein for pertussis toxin, 39, 45 and 52 kDa proteins (G(s)) for cholera toxin, a 21 kDa protein for botulinum toxin C, and a 43 kDa protein (actin) for botulinum toxin C(2a). The increase in either HUVEC or BPAEC PGI2 release induced by thrombin was not altered by pretreatment with any toxin. However, 1 h treatment of BPAEC monolayers with 1 μg/ml pertussis toxin resulted in dramatic barrier dysfunction, which was synergistic with the albumin permeability induced by 1 μM thrombin. In contrast, pretreatment with 1 μg/ml cholera toxin completely prevented the thrombin-induced barrier dysfunction. Moreover, contraction and gap formation due to thrombin challenge, observed by phase contrast microscopy, was greatly augmented by pertussis toxin and prevented by cholera toxin. Whereas 5 μg/ml botulinum toxin C did not affect either basal or thrombin-induced barrier dysfunction, botulinum toxin C(2a) increased basal BPAEC permeability over four-fold. Thus, bacterial toxins have specific and divergent effects on thrombin-induced endothelial cell responses. Botulinum toxin C(2a) appears to interact directly with actin to produce barrier dysfunction. In contrast, cholera toxin promotes barrier function via its known effects on G(s), stimulating adenylate cyclase and increasing cAMP. Because cholera toxin and pertussis toxin (via inhibition of G(i)) both increase cAMP, yet have opposing effects on barrier function, the present results suggest that pertussis toxin produces barrier dysfunction via ADP ribosylation of a novel G-protein other than G(i) or via a novel action of G(i).

Original languageEnglish (US)
Pages (from-to)63-72
Number of pages10
JournalBlood Coagulation and Fibrinolysis
Volume5
Issue number1
StatePublished - 1994
Externally publishedYes

Fingerprint

Bacterial Toxins
Thrombin
Endothelial Cells
Cholera Toxin
Pertussis Toxin
GTP-Binding Proteins
Lung
Human Umbilical Vein Endothelial Cells
Proteins
Adenosine Diphosphate
Actins
Permeability
Thrombin Receptors
Phase-Contrast Microscopy
Epoprostenol
Autoradiography
Adenylyl Cyclases
NAD
Polyacrylamide Gel Electrophoresis
Albumins

Keywords

  • botulinum toxin
  • cholera toxin
  • endothelium
  • G-proteins
  • pertussis toxin

ASJC Scopus subject areas

  • Hematology

Cite this

Regulation of thrombin-induced endothelial cell activation by bacterial toxins. / Patterson, C. E.; Garcia, Joe GN.

In: Blood Coagulation and Fibrinolysis, Vol. 5, No. 1, 1994, p. 63-72.

Research output: Contribution to journalArticle

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