Immunomodulatory Effect Of CCK=8 On Reversing Endotoxic Shock And Preliminary Study Of Its Molecular Mechanisms

Endotoxic shock (ES), resulting from cascade reaction of inflammatory mediators induced by excessive endotoxin, leads to uncontrolled disturbance of organisms, multiple organ failure and lethality. Many kinds of inflammatory mediators including pro-inflammatory cytokines, oxygen-derived free radicals and nitric oxide (NO) play an important role in ES, but the exact pathogenesis remains unclear and the treatment is difficult. Many antibodies against pro-inflammatory cytokines and inhibitors of inducible NO synthase (iNOS) have been produced, but most of them are agents targeting one factor and can't efficiently block the development of systemic inflammation. Different kinds of inflammatory mediators are secreted by immunocytes, which suggests that the essence of inflammation is also a kind of immune response. Alterations in immune function are felt to play a pivotal role in the development of ES. So it has become the focus to search for anti-inflammatory agents that have immunomodulatory effect. Cholecystokinin-octapeptide (CCK-8) is a kind of endogenous brain-gut peptide. Recent studies suggest a regulatory function of CCK-8 in the immune system. Our previous study demonstrated that CCK-8 could protect animals from ES induced by lipopolysaccharide (LPS). However, it is not clear whether this protecting effect of CCK-8 is related to its immunomodulatory properties. Lung and spleen are target organs that are easily damaged during ES. Besides respiratory and metabolic function, lung plays a role in immune system. Spleen, the biggest lymphatic organ in body, has important immune functions. Inflammatory response is first observed in spleen during ES. The mitogen-activated protein kinases (MAPKs) are focal points of discretesignaling cascades for diverse extracellular stimuli, and function to regulate fundamental cellular processes. CCK activates MAPK in pancreatic acinar cells. Cross-talk has been shown recently between Janus tyrosine kinase/signal transducers and activators of transcription (JAK/STAT) and MAPK pathways. STAT proteins are activated by G-protein coupled receptors, while it has not been reported whether CCK receptors, a kind of G-protein coupled receptors, can activate JAK/STAT pathway. The STAT3 mutant mice were highly susceptible to ES with increased serum concentration of inflammatory cytokines such as tumor necrosis factor-a(TNF-a), interleukin-lp (IL-13) and interleukin-6 (IL-6). It has been demonstrated that p38 MAPK is involved in serine phosphorylation and regulation of STAT activation. CCK may play immunomodulatory role in ES by activating STAT through p38 MAPK and regulating gene transcription. At rest, the heterodimeric Rel/NF- K B complex is located in the cytoplasm bound to an inhibitory factor, I K B. Upon stimulation, I K B is phosphorylated and degraded, free NF- K B then translocates into the nucleus where it binds to K B site to regulate transcription. NF- K. B can be activated by LPS and regulate transcription of many pro-inflammatory cytokines. Cong et al proved that CCK-8 inhibited LPS-induced NF- K B activity in lung of rats. The degradation of I K B and dissociation from NF-K B complex are essential process for activation of NF- K B. There is no report about whether CCK-8 inhibits NF- K B activity through inhibiting LPS-induced degradation of I K B. The present study was designed to observe the regulatory effect of CCK-8 on changes of immune functions induced by LPS, the receptor mechanism and signal transduction mechanism involving MAPK, STAT3 and I K B at different levels including in vivo and in vitro, in order to study its immunomodulatory effect on reversing ES and its molecular mechanisms. 1 Effect of CCK-8 on changes in lung and spleen of ES rats 1.1 CCK-8 attenuates morphologic and functional changes in lung and spleen of ES ratsTo observe the effect of CCK-8 on decrease of mean arterial pressure (MAP) induced by LPS and ultrastructure changes in lung and spleen of ES rats, animals were randomly assigned to four groups injected different agents via tail...