Spinorphin

To characterize the inflammatory effect of spinorphin, an endogenous peptide purified from bovine spinal cord, its effects on chemotaxis, O2- generation, and exocytosis by N-formylmethionyl-leucyl-phenylalanine (FMLP)-stimulated human neutrophils (PMNs) in vitro were examined. At 10 microM, spinorphin significantly inhibited chemotaxis by FMLP-stimulated PMNs. Spinorphin at 100 microM also inhibited both O2- generation and exocytosis of beta-glucuronidase and collagenase by FMLP-stimulated PMNs. The mechanisms by which spinorphin inhibits these PMN functions were examined further. Spinorphin markedly suppressed the binding of FML[3H]P to its receptor on PMNs, as observed in a binding assay. However, other neuropeptides that were examined (angiotensin II and substance P) had no effect on FML[3H]P binding, suggesting the possibility that spinorphin plays a specific role in the inhibition of the binding between FMLP and its receptor. The suppression of FMLP binding also caused a decrease of the FMLP-induced intracellular calcium concentration [Ca2+]i, which acts as a second messenger leading to PMN functions. These results suggest that spinorphin may be a new endogenous inflammation-regulatory peptide that modulates the interaction of FMLP with its receptor.

We have found activity of dipeptidyl peptidase (DPP) III, one of the most important enkephalin-degrading enzymes in the central nervous system, in human neutrophils. HPLC analysis of the peptide fragments produced by treatment of leucine-enkephalin with isolated neutrophils in the presence of inhibitors of other enkephalin-degrading enzymes revealed that the enzyme in human neutrophils cleaved dipeptides from the NH(2) terminus of leucine-enkephalin, suggesting the presence of DPPIII activity in human neutrophils. Using a specific synthesized substrate and proteinase inhibitors, it was found that the neutrophils have 19.2 +/- 3.6 microM/h/5 x 10(6) cells of beta-naphthylamine for the enzyme. It was also confirmed that spinorphin and tynorphin, both reported to inhibit the activities of enkephalin-degrading enzymes, had potent inhibitory activities (IC(50): 4.0 and 0.029 microg/ml, respectively) against the enzyme. The presence of DPPIII activity in human neutrophils suggests that the biologically active peptides which are associated with enkephalin play a physiological role in regulating enkephalin or inflammatory mechanisms in peripheral tissues.

Spinorphin (LVVYPWT) has been isolated from the bovine spinal cord as an endogenous inhibitor of enkephalin-degrading enzymes. It has been reported that spinorphin has an antinociceptive effect, inhibitory effect on contraction of smooth muscle and anti-inflammatory effect. In the present study, the effects of leu-enkephalin and spinorphin on allodynia and mechanical and thermal nociceptions were examined in vivo using mice. Intrathecal (i.t.) administration of leu-enkephalin or spinorphin inhibited the allodynia induced by intrathecal nociceptin in a dose-dependent manner. Furthermore, spinorphin enhanced the inhibitory effect of enkephalin on allodynia induced by nociceptin. Naloxone antagonized both inhibitory effects of leu-enkephalin and spinorphin, suggesting that the endogenous opioidergic system can modulate allodynia. Intracerebroventricular (i.c.v.) administration of leu-enkephalin increased the nociceptive threshold of heat or mechanical stimulation to a mouse. Although i.c.v. administration of spinorphin had no effect on the threshold of heat or mechanical stimulation, spinorphin enhanced and prolonged the antinociceptive effect of leu-enkephalin. The enhancement of spinorphin on the antinociception produced by leu-enkephalin was reversed by pretreatment with naloxone.