Peripheral multiorgan dysfunction induced by sepsis : the role of NFκB pathway and its impact on brain and heart function

Abstract

Sepsis and sepsis-associated multiorgan failure represent a systemic inflammatory state mediated by the innate immune system resulting in an excessive cellular response to severe infection, with high levels of morbidity and mortality. Furthermore, patients who survive sepsis, have long-term cognitive and functional impairment. Animal models are essential to clarify the pathophysiological mechanisms of sepsis. There is mounting evidence that inhibition of nuclear factor NFκB activation can reduce sepsis-associated organ dysfunction and injury. Herein, we deeply characterize an animal model of lipopolysaccharide (LPS)-induced acute inflammation, through an integral functional, behavioural, morphological and molecular evaluation as well as tested different possible therapeutic approaches to determine their efficacy on the modulation of systemic inflammation through the NF-kB pathway. Male and female Wistar rats (12-20 weeks) were injected with LPS (E. coli serotype O127:B8; tail IV) and divided into three groups: LPS 6 (6mg/kg), LPS 12 (12mg/kg) and SHAM (NaCl 0.9%). At 6 h and 24 h after LPS administration, an autonomic evaluation was performed in both conscious animals, with continuous radio-telemetry recording of blood pressure (BP) and heart rate (HR), and anesthetized animals with BP, ECG, HR, tracheal pressure, respiratory frequency (RF) and body temperature continuously monitored. Baro- and chemoreflexes were evaluated with phenylephrine and lobeline, respectively. Behavioural changes were also evaluated through the elevated-plus maze (anxiety), open-field (locomotor/exploratory activity) and Y-maze (short-term spatial working memory) tests. Immunohistochemistry and RT-PCR were executed to determine heart and brain inflammatory state. Serum biomarkers levels for organ disfunction were also measured. For the test of potential therapeutic drugs, animals were exposed with the two LPS doses and treated 15 minutes later with four different drugs: erythropoietin, dexamethasone, IKK 16 and adenosine. The functional, behavioural, and molecular analysis were performed. Overall, the characterization model results showed a rise in BP and HR and elevated RF, indicative of tachycardia and tachypnea. The higher RF was concomitant with a higher chemoreceptor reflex sensitivity. The autonomic data reveals an overexcitation of sympathetic tone with a sympatho-vagal imbalance, concomitant with baroreflex impairment and/or increases in BP, all these were strongly marked in the highest LPS dose. At both time-points, both LPS groups show an anxiety-like behaviour, associated with lower locomotor and exploratory capacity and short-term memory impairment. The molecular studies show reactive astrogliosis and microgliosis, due to inflammatory processes in the hippocampus, as well as, an upregulation of pro-inflammatory factors in the heart and brain. Serum analysis yielded higher levels of biomarkers for renal and liver dysfunction, and pancreatic and neuromuscular injury, mainly in the LPS 12 group. The functional, behavioral, and molecular response between female and male rats was not so different. Both sexes present markedly changes when exposed to higher doses of LPS. Indeed, male animals are more sensitive to endotoxin that females, that can fight inflammation more easily. Of the four therapeutic proposals studied, erythropoietin and dexamethasone were the best to modulate the NF-ΚB pathway in order to reduce the dysfunction of multiple organs as well as attenuate the behavioural changes caused by the exposure to endotoxin. Both IKK 16 and adenosine treatments attenuate the organ dysfunction by reducing the serum renal, hepatic, pancreatic and neuromuscular levels, however these two drugs did not improve the cardiac and behavioral modifications. Further studies are needed in order to better understand the role of these drugs in the early stages of sepsis. Summarizing, higher doses of LPS induces strong alterations in both cardiac and neurological systems, as well as, multiple organ dysfunction, that can be attenuated by the administration of specific NF-κB modulators, such as, erythropoietin and dexamethasone. These findings served to better understand the pathophysiological processes involved in these animal model of acute inflammation as well as showed therapeutic evidence that through NF-κB signalling pathway modulation it is possible to attenuate or even reverse the acute stages before this condition evolves to more aggressive stages, namely septic shock and ultimately dead.