Joseph A. Borkowski

Cloning and pharmacological characterization of a human bradykinin (BK-2) receptor

A human BK-2 bradykinin receptor was cloned from the lung fibroblast cell line CCD-16Lu. The cDNA clone encodes a 364 amino acid protein that has the characteristics of a seven transmembrane domain G-protein coupled receptor. The predicted amino acid sequence of the human BK-2 receptor is 81% identical to the smooth muscle rat BK-2 receptor (1). Transfection of the human BK-2 receptor cDNA into COS-7 cells results in the expression of high levels of specific BK binding sites. Saturation binding analysis indicates that the human BK-2 receptor expressed in COS-7 cells binds BK with a KD of 0.13 nM. Pharmacological characterization of the expressed BK receptor is consistent with the cDNA encoding a receptor of the BK-2 subtype. The BK-2 receptor antagonist Hoe 140 (2), D-Arg0[Hyp3, Thi5, D-Tic7, Oic8]BK has a high affinity (IC50 = 65 pM) for the cloned human receptor. The tissue distribution of the human BK-2 receptor was analyzed by competitive PCR with human tissue cDNA and is similar to that determined for the BK-2 receptor in the rat.

Effects of the bradykinin B1 receptor antagonist des-Arg9[Leu8]bradykinin and genetic disruption of the B2 receptor on nociception in rats and mice

Abstract The contributions of B1 and B2 bradykinin receptors to acute and chronic inflammatory hyperalgesia were examined using the peptide B1 receptor antagonist des‐Arg9[Leu8]bradykinin and transgenic Bk2r‐/‐ mice. In normal rats and mice, des‐Arg9[Leu8]bradykinin (30 nmol/kg i.v. or s.c.) inhibited carrageenan‐induced hyperalgesia and the late phase nociceptive response to formalin. The active dose range was narrow, suggesting partial agonist activity of this peptide. In rats with monoarthritis, des‐Arg9[Leu8]bradykinin (up to 30 nmol/kg i.v.) failed to reduce the number of vocalisations elicited by gentle flexion and extension of the inflamed limb; however, hyperalgesia was exacerbated by administration of the B1 receptor agonist des‐[Arg9]bradykinin (100 nmol/kg i.v.), consistent with other evidence for local induction of B1 receptors during adjuvant‐induced arthritis. The nociceptive response to intraplantar injection of bradykinin (10 nmol) and hyperalgesia induced by carrageenan (0.6 mg) were absent in Bk2r‐/‐ mice, indicating that stimulation of B2 receptors is an essential step in the initiation of some nociceptive and inflammatory reactions. However, the nociceptive response to formalin (2.5% intraplantar), including inhibition of the late phase by des‐Arg9[Leu8]bradykinin (0.3 nmol), and induction of thermal hyperalgesia by Freunds adjuvant (0.1%) appeared intact in Bk2r‐/‐ mice. These findings support other evidence for an involvement of B1 receptors in inflammatory hyperalgesia and suggest that B1 receptor antagonists may be clinically useful as anti‐inflammatory and analgesic drugs.

Presence of a simple tandem repeat in the ITS1 region of the Xylariales.

A Simple Tandem Repeat sequence of 11 nucleotides has been found in the ITS1 region of the rDNA of members of Order Xylariales. The number of repetitions detected ranged from one to six, and they could be found in pure tandem or interspersed. The same core sequences have also been found in DNA from other organisms, although usually not repeated in tandem. These repetitions could have been generated by slipped strand mispairing. The presence of this sequence increases the normal rate of divergence in the ITS1 of the Xylariales. The phylogenetic implications of the presence of this sequence in the molecular taxonomy of Xylariales are also discussed.