THE LIPID-PEROXIDATION PRODUCT 4-HYDROXYNONENAL IS FORMED BY - AND ISABLE TO ATTRACT - RAT NEUTROPHILS IN-VIVO

4-Hydroxynonenal (HNE), a major aldehydic product of lipid peroxidatio n, is a chemoattractant for neutrophilic polymorphonuclear granulocyte s in vitro. The question was studied, whether HNE is formed during the ingress of neutrophils in the Sephadex model of inflammation. The pol ydextrane Sephadex G-200, which causes an acute aseptic traumatic infl ammation, was injected subcutaneously into rats. The implants were exc ised 6-36 hours later, and the neutrophils separated from the exsudate by centrifugation. After extraction with dichloromethane HNE was iden tified in the exsudate by non-derivative reversed phase HPLC in combin ation with on-line uv-spectroscopy. The concentration of HNE in the in flammatory focus did not correlate with the number of neutrophils pres ent. While the peak of HNE coincided with the time point of the highes t turnover rate of neutrophils (0.13 mu M at 6 hrs after implantation) , the highest number of neutrophils (about 100 million cells) occurred not earlier than 18 hrs later (24 hrs after onset of inflammation). W hen neutrophils were isolated from the inflammatory focus and stimulat ed with Zymosan, they were able to produce HNE in vitro depending on t he time of isolation. The highest production of HNE (0.17 mu M) by pha gocyting neutrophils was observed at the shortest inflammation time st udied (3 hrs). In order to compare these results with the oxidative bu rst of neutrophils the formation of superoxide was also measured by th e cytochrome c reduction assay in vitro. The maximum of the production rate of superoxide anion was observed at the same inflammation time ( 6 hrs), when the HNE maximum occurred. Cells which ingressed earliest (at 3 hrs) showed the highest production rate of superoxide per cell ( 307 x 10(-18) moles per cell and 30 min). The ability of HNE to attrac t neutrophils in vivo was studied by adding synthetic HNE to the Sepha dex gel and measuring the ingression of neutrophils afterwards. The ap plication of 1 mu M HNE in the focus did not change the number of neut rophils but 10 mu M HNE increased the cell number by a factor of 3. Th e results indicate that HNE is not only a chemoattractant for rat neut rophils in vitro but also in vivo. It is suggested that HNE is produce d by selfdestruction of neutrophils during a traumatic inflammation an d its production seems to be tightly connected to the oxidative burst of neutrophils. The idea of HNE as part of an autocatalytic cycle is s upported whereby neutrophils which immigrate into an inflammatory focu s produce HNE which stimulates the ingress of new neutrophils.