Reaction of antithrombin with proteases. Nature of the reaction with trypsin

Wong, Raymond F.; Chang, Tien; Feinman, Richard D.

doi:10.1021/bi00530a002

Cited by 26 publications

(11 citation statements)

References 28 publications

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“…However, lepirudin, a direct thrombin inhibitor, did not affect C5a production. Apart from thrombin, antithrombin can also inactivate other proteases involved in blood coagulation, as well as serine proteases that are not involved in the coagulation cascade such as trypsin (38). It has also been shown that trypsin is able to cleave C5 into biologically active C5a-like fragments (39).…”

Section: Discussionmentioning

confidence: 99%

Anaphylatoxin C5a Creates a Favorable Microenvironment for Lung Cancer Progression

Corrales

Ajona

Rafail

et al. 2012

The Journal of Immunology

234

291

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The complement system contributes to various immune and inflammatory diseases, including cancer. In this study we investigated the capacity of lung cancer cells to activate complement, and characterized the consequences of complement activation on tumor progression. We focused our study on the production and role of the anaphylatoxin C5a, a potent immune mediator generated after complement activation. We first measured the capacity of lung cancer cell lines to deposit C5 and release C5a. C5 deposition, after incubation with normal human serum, was higher in lung cancer cell lines than in non-malignant bronchial epithelial cells. Interestingly, lung malignant cells produced complement C5a even in the absence of serum. We also found a significant increase of C5a in plasma from patients with non-small cell lung cancer, suggesting that the local production of C5a is followed by its systemic diffusion. The contribution of C5a to lung cancer growth in vivo was evaluated in the Lewis lung cancer model. Syngeneic tumors of 3LL cells grew slower in mice treated with an antagonist of the C5a receptor. C5a did not modify 3LL cell proliferation in vitro but induced endothelial cell chemotaxis and blood-vessels formation. C5a also contributed to the immunosuppressive microenvironment required for tumor growth. In particular, blockade of C5a receptor significantly reduced myeloid-derived suppressor cells and immunomodulators ARG1, CTLA-4, IL6, IL10, LAG3 and PDL1 (B7H1). In conclusion, lung cancer cells have the capacity to generate C5a, a molecule that creates a favorable tumor microenvironment for lung cancer progression.

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Section: Discussionmentioning

confidence: 99%

Anaphylatoxin C5a Creates a Favorable Microenvironment for Lung Cancer Progression

Corrales

Ajona

Rafail

et al. 2012

The Journal of Immunology

234

291

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“…Antithrombin I11 is an effective inhibitor of trypsin, and virtually all coagulation serine proteinases except factor VII [22,24,25].The removal from the circulation of ATIII-thrombin complexes is accomplished by a recently described receptor-mediated process located on hepatocytes [ 10,111. Previous studies from this laboratory have demonstrated that proteinase inhibitor-proteinase complexes clear in vivo and bind to cells in vitro independently of the proteinase [3,10,Il,[26][27][28].…”

Section: Discussionmentioning

confidence: 99%

Hepatocyte receptors for antithrombin III‐proteinase complexes

Fuchs¹,

Shifman²,

Michalopoulos³

et al. 1984

J of Cellular Biochemistry

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The in vivo clearance of antithrombin III-proteinase complexes occurs via a specific and saturable pathway located on hepatocytes. We now report studies of the catabolism of antithrombin III-proteinase complexes in vitro using rat hepatocytes in primary culture. Antithrombin III-thrombin and trypsin complexes were prepared and purified to homogeneity. Ligand uptake by hepatocytes was concentration, temperature, and time dependent. Initial rate studies were performed to characterize the maximum rate of uptake, V, and apparent Michaelis constant Kapp. These studies yielded a V of 12.8 fmol/mg cell protein/min and a Kapp of 144 nM for antithrombin-trypsin complexes. Competition experiments with antithrombin III, antithrombin III-proteinase complexes, alpha 2-macroglobulin-methylamine, asialoorosomucoid and the neoglycoproteins, fucosyl-bovine serum albumin (BSA), N-acetylglucosaminyl-BSA, and mannosyl-BSA indicated that only antithrombin III-proteinase complexes were recognized by the hepatocyte receptor. Uptake studies were performed at 37 degrees C with 125I-antithrombin III-trypsin and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in conjunction with autoradiography. These studies demonstrate time-dependent uptake and degradation of the ligand to low molecular weight peptides. In addition, there was a time-dependent accumulation of a high molecular weight complex of ligand and a cellular protein. This complex disappeared when gels were performed under reducing conditions.

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“…The active concentrations of stock solutions of thrombin were determined by active-site titration with pNPGB according to the method of Chase & Shaw (1969). Measurements of thrombin activity by the clotting test were as described earlier (Wong et al, 1982). Antithrombin active concentrations were assayed by the inhibition of thrombin activity as measured by pNPGB.…”

Section: Methodsmentioning

confidence: 99%