There is a pressing need to find countermeasures to treat RI and CI. At present, none of countermeasures yet evaluated to treat either condition has been approved by the U.S. FDA for the use in humans based upon evaluation in two suitable laboratory animal species. In the search for appropriate CI-treatment targets, it is necessary to understand the mechanisms underlying CI. Our research has been investigating many pathophysiological changes that occurred during the period of 30 days after CI, which identified increased DNA damage, stress-response gene expressions, serum cytokine and chemokine concentrations, ileal damage, bacterial translocation 
, and blood-cell depletion 
. Some of these changes were truly unique to CI, i.e.
, changes were great enough to distinguish CI from any other single injuries, and offered us multiple treatment targets or parameters to monitor prospective countermeasure development.
CIP is a second-generation fluoroquinolone (FQ) 
. The spectrum of antimicrobial activity of CIP is due to the specificity in inhibition directed to bacterial DNA gyrase and topoisomerase IV, particularly in Gram-negative species 
. The pharmacokinetics 
and pharmacodynamics 
in humans have been well documented. It is well tolerated, distributes rapidly in tissues, and has similar half-lives in mice 
and humans 
, and is readily available in several formulations for oral, intravenous, intraperitoneal, and topical routes. These factors meet the needs for the management of bacterial infection in a mass-casualty scenario. Although it has been used after the anthrax attack in 2001 
, it has never been tested for a treatment of CI.
The wide use of CIP in clinical settings has helped the discovery of its immunomodulatory effect, apart from the antimicrobial activity. Since the stimulation of hematopoiesis highlights immunomodulation by CIP 
, it might combat the CI-induced severe hematologic loss. FQs including CIP were shown ex vivo
to enhance cytokines IL-3 and GM-CSF in mouse splenocytes that gave rise to enhanced hematopoiesis 
, while CIP, together with a mitogen, induced hyper-production of IL-2 in human peripheral lymphocytes 
. Only a subset of FQs produces this induction effect, which appears to be associated with the cyclopropyl moiety present at the N1 position 
. Although the exact mechanism is elusive, it is possible that transcription factors, which are common to increased cytokines, such as activator protein-1 (AP-1), are activated by those FQs 
On the other hand, it has also been reported that FQs including CIP inhibited mammalian topoisomerase II 
. Although this inhibitory effect is about 1,000-fold less than the one directed to bacterial counterparts, it is not negligible when considering sustained accumulation of FQs in the organs 
and intracellularly up to 12-fold greater than extracellular concentrations 
. Inhibition of topoisomerase II resulted in lowered inflammatory cytokines TNF and IL-6 secretion from ex vivo
splenocytes, a majority of which was monocytes, and also lowered MIP-1α in macrophages in vitro
. Therefore, the topoisomerase II inhibitory effect of CIP may be a benefit in treating enhanced inflammation induced by CI.
In the present study, for the first time, we investigated whether CIP mitigated ionizing radiation combined injury (CI) through its effects on survival, cytokine levels, bone marrow cells, and ileum of surviving mice on day 10 after CI. The multiple effects of CIP were observed in several tissues. In the sera, CIP effectively reduced CI-induced pro-inflammatory cytokines IL-6, KC (i.e.
, IL-8 in human), and G-CSF up to 80% and Rantes to basal level (). The enormous induction of IL-6, KC, and G-CSF in serum in CI-mice correlated with the results reported previously from this laboratory on day 7 
. Continuous, high IL-6 expression caused serious secondary damage to the organs rather than its benefits obtained by mild and transient expression 
. It is also well known, for example, that IL-6 is very high in inflammatory bowel disease (IBD) including ulcerative colitis (UC) and Crohn’s disease (CD), and exists in foci specific manner 
, suggesting its major roles in IBD pathology. In fact, IL-6 is considered as a treatment target in IBD and other inflammatory conditions 
. Other major inflammatory cytokines such as KC and Rantes are also known to induce serious tissue damage, if their levels are too high and meet no resolution 
IL-3 is an essential mediator for myeloid lineage development and has been reported to be elevated by CIP treatment 
. In our study, CIP significantly increased IL-3 only in CI-animals (). Further, CIP also accelerated bone-marrow recovery that may be related to increased level in IL-3 (), while the recovery in WBCs and platelets was not yet to be seen (). IL-3 in combination with erythropoietin (EPO) has been reported to increase production of WBC 
. Therefore, the observation of accelerated bone-marrow recovery by CIP after CI warrants future studies on the association of the IL-3, EPO and bone marrow recovery with WBC production.
CIP limited physical damages occurring in the ileum after CI (). Analysis on cell-death type-specific markers revealed that after CI villous epithelial cells underwent apoptosis () while crypt cells displayed elevated formation of autophagosomes (). CIP effectively reduced both types of cell death. A high level of IgA coated the ileal mucosa of CI-mice, particularly in foci exhibiting severe morphological damage in villous tips. This may be a result of massive IL-6 concentration observed in sera, both of which CIP reduced to normal range (). The cell death, which occurred on the villi epithelial cells, was associated with activation of NF-κB in immune effector cells inside villi and a consequence of caspase-3 earlier ( and ). This finding correlates with observations found in human T cells in vitro
and in CD2F1 mice 
. Similarly, the autophagic death, which occurred in crypt cells of CI-mice in our study, was also found in CD2F1 mice 
CIP offers several advantages to be developed further as a drug to treat CI in a mass-casualty scenario: (1) it is included in the Strategic National Stockpile for bacterial infection control; (2) it can be taken orally so that patients can self-administer it; (3) it possesses not only antimicrobial but also favorable immunomodulatory activity; and (4) it is inexpensive.
In summary, CIP significantly increased survival, altered the serum cytokine and chemokine profile, accelerated bone-marrow recovery, inhibited cell death of ileum, and prevented systemic bacterial infection. These effects could be mediated by its capability of inhibiting NF-κB and caspase-3. Therefore, the results suggest that CIP may prove to be beneficial for treating critical sequelae of CI.