Kinin-B1 receptors in ischaemia-induced pancreatitis: Functional importance and cellular localisation

In this study we compare the role of kininB1 and B2 receptors during ischaemia/reperfusion of rat pancreas. Our investigations were prompted by the observation that infusion of a kininB2 receptor antagonist produced significant improvement in acute experimental pancreatitis. In an acute model with two hours of ischaemia/two hours of reperfusion, application of the kininB1 receptor antagonist (CP-0298) alone, or in combination with kininB2 receptor antagonist (CP-0597), significantly reduced the number of adherent leukocytes in postcapillary venules. In a chronic model with five days of reperfusion, the continuous application of kininB1 receptor antagonist or a combination of kininB1 and B2 receptor antagonists markedly reduced the survival rate. In kininreceptor binding studies kininB1 receptor showed a 22-fold increase in expression during the time of ischaemia/ reperfusion. Carboxypeptidase M activity was upregulated 10-fold following two hours of ischaemia and two hours of reperfusion, provided the appropriate specific ligand, desArg10-kallidin and/or desArg9-bradykinin, was used. The occurrence of kininB1 receptor binding sites on acinar cell membranes was demonstrated by microautoradiography. With a specific antibody, the localisation of kininB1 receptor protein was confirmed at the same sites. In conclusion, we have demonstrated the upregulation of the pancreatic acinar cell kininB1 receptors during ischaemia/reperfusion. The novel functional finding was that antagonism of the kininB1 receptors decreased the survival rate in an experimental model of pancreatitis

Guanosine nucleotides regulate B2 kinin receptor affinity of agonists but not of antagonists: Discussion of a model proposing receptor precoupling to G protein

The effect of nucleotides on binding of the B2 kinin (BK) receptor agonist {[}H-3]BK and the antagonist {[}H-3]NPC17731 to particulate fractions of human foreskin fibroblasts was studied. At 0 degrees C, particulate fractions exhibited a single class of binding sites with a Kd of 2.3 nM for {[}H-3]BK and a K-d Of 3.8 nM for the antagonist {[}H-3]NPC17731. Incubation with radioligands at 37 degrees C for 5 min gave a reduction of agonist, as well as antagonist, binding that was between 0-40% depending on the preparation, even in the absence of guanosine nucleotides. As shown by Scatchard analysis, this reduction in specific binding was due to a shift in the affinity of at least a fraction of the receptors. The presence at 37 degrees C of the guanine nucleotides GTP, GDP and their poorly hydrolyzable analogs left {[}H-3]-NPC17731 binding unaffected, but reduced the receptor affinity for {[}H-3]BK to a K-d Of about 15 nM. The maximal number of receptors, however, was unchanged. This affinity change was strongly dependent on the presence of bivalent cations, in particular Mg2+. It was reversed by incubation at 0 degrees C, The rank order of the guanosine nucleotides for {[}H-3]BK binding reduction was GTP{[}gamma S] = Gpp{[}NH]p > GTP = GDP > GDP{[}beta S]. GMP, ATP, ADP and AMP showed no influence on agonist binding. A model for the interaction of the B2 kinin receptor with G proteins is discussed

The effects of a plant proteinase inhibitor from Enterolobium contortisiliquum on human tumor cell lines

Supplementary to the efficient inhibition of trypsin, chymotrypsin, plasma kallikrein, and plasmin already described by the EcTI inhibitor from Enterolobium contortisiliquum, it also blocks human neutrophil elastase (K(iapp)=4.3 nM) and prevents phorbol ester (PMA)-stimulated activation of matrix metalloproteinase (MMP)-2 probably via interference with membrane-type 1 (MT1)-MMP. Moreover, plasminogen-induced activation of proMMP-9 and processing of active MMP-2 was also inhibited. Furthermore, the effect of EcTI on the human cancer cell lines HCT116 and HT29 (colorectal), SkBr-3 and MCF-7 (breast), K562 and THP-1 (leukemia), as well as on human primary fibroblasts and human mesenchymal stem cells (hMSCs) was studied. EcTI inhibited in a concentration range of 1.0-2.5 mu M rather specifically tumor cell viability without targeting primary fibroblasts and hMSCs. Taken together, our data indicate that the polyspecific proteinase inhibitor EcTI prevents proMMP activation and is cytotoxic against tumor cells without affecting normal tissue remodeling fibroblasts or regenerative hMSCs being an important tool in the studies of tumor cell development and dissemination