PREFERENTIAL BINDING OF HEPARIN TO GRANULOCYTES OF VARIOUS SPECIES

Objectives-To address the problem of heparin binding to leukocytes of various animal species. Design-Leukocytes of the various species were incubated with fluorescent-labeled, low molecular mass heparin (LMMH). Fluorescence intensity on granulocytes, lymphocytes, and monocytes wa s analyzed by flow cytometry analysis. Sample Population-leukocytes we re prepared from EDTA-anticoagulated blood of human subjects, rats, ra bbits, dogs, pigs, and sheep 3 times. Procedure-The leukocyte populati ons were identified by their light scatter properties. In addition, ph ycoerythrin-labeled CD4, CD13, and CD14 antibodies were used to identi fy human lymphocytes, granulocytes, and monocytes; CD4, GR-1, and CD11 b antibodies were used for mouse, and CD45, RP 3, and ED 9 antibodies were used for identification of rat leukocyte subpopulations. Results- Granulocytes, monocytes, and lymphocytes of ail species bound LMMH in dose-dependent manner. Binding of LMMH-tyramine (tyr)-fluorescein-5-is othiocyanate (FITC) to granulocytes was higher in human subjects, rats , rabbits, dogs, and pigs, compared with binding to monocytes and lymp hocytes, Mouse and sheep granulocytes did not bind more heparin than m onocytes or lymphocytes. Binding of LMMH-tyr-FITC was reversible in th e presence of unlabeled heparin or LMMH. More than 99% of human, rat, rabbit, dog, and sheep granulocyte populations were distinguished from monocytes and lymphocytes by means of their fluorescence intensity ow ing to LMMH-tyr-FITC, This separation was not obtained for mouse and p ig granulocytes, Conclusion-Evidence of specific heparin binding to gr anulocytes of many species indicates the relevance of fluorescent-labe led LMMH for biological investigations. Clinical Relevance-Binding of heparin and LMMH to granulocytes, lymphocytes, and monocytes may have a substantial role in atherosclerosis, inflammation, malignancy, and i mmunologic diseases.