Interleukin-6 mRNA and protein increase in vivo following induction of acute thrombocytopenia in mice

Abstract

Abstract Induction of experimental thrombocytopenia in rodents results in the enhancement of megakaryocytic growth and differentiation. Interleukin-3 (IL-3) and IL-6, cytokines with a broad spectrum of biologic activities, stimulate megakaryocytopoiesis in vitro. To determine if expression of these factors might increase in response to experimental thrombocytopenia, we measured steady-state levels of IL-3 and IL-6 mRNA following rabbit antiplatelet serum (APS) injection. Groups of mice were injected intravenously with 0.2 mL APS while control animals received rabbit antilymphocyte serum (ALS), normal rabbit serum (NRS), or phosphate-buffered saline (PBS). At various times up to 72 hours after injection mice were exsanguinated and splenectomized. Platelet counts in the experimental animals were less than 12% of controls. Splenic RNA was hybridized in solution to 32P-UTP-labeled cRNA probes for IL-3 and IL-6. RNase-resistant hybrids were resolved on denaturing gels and visualized autoradiographically. IL-3 hybrids were undetectable at all time points tested, irrespective of the film exposure time or specific activity of the probe. Conversely, IL-6 hybrids were easily visualized and showed peak expression at 1.5 to 2.0 hours. By 3 hours, IL-6 mRNA had returned almost to the level of the controls. Similar results were observed in the bone marrow, although maximal IL-6 mRNA in that tissue was observed 4 hours following APS administration. To determine if this mRNA increment was associated with a concomitant increase in bioactive protein, serum was tested for its ability to stimulate IL-6-dependent B9 cells. At 1.75 hours following injection, experimental animals showed a small but significant increment in IL-6 activity compared with controls (200 +/- 30 U/mL IL-6 compared with 129 +/- 17 U/mL in ALS-injected controls, 106 +/- 17 U/mL in NRS-injected controls and 84 +/- 17 U/mL in PBS-injected controls). The data show that IL-6 mRNA and bioactive protein increase in response to acute immunothrombocytopenia, while no increment in IL-3 is detectable. These results suggest that IL-6 may play a role in the physiologic response to acute immunothrombocytopenia.