Sepsis Control Group Research Articles

Early administration of milrinone ameliorates lung and kidney injury during sepsis in juvenile rats.

A sepsis model was created, induced by cecal ligation and puncture (CLP), in juvenile rat groups. Milrinone (MIL), which is known to have a modulatory effect on pro-inflammatory cytokines, was administered to the designated rat groups in the early period before severe sepsis developed. The study was aimed at investigating the possible protective effects of milrinone on the lung and kidney tissues of rats in the late phase of sepsis. The rat pups were divided into seven groups with six animals in each group: (1) healthy rats who received no drug; (2) CLP-S12 (sacrificed at hour 12); (3) CLP-S24 (sacrificed at hour 24); (4) CLP-MIL1-S12 (administered with 0.5 mg/kg milrinone at hour 1 and sacrificed at hour 12); (5) CLP-MIL1-S24 (administered with 0.5 mg/kg milrinone at hour 1 and sacrificed at hour 24): (6) CLP-MIL12-S24 (administered with 0.5 mg/kg milrinone at hour 12 and sacrificed at hour 24), (7) and CLP-MIL1,12-S24 (administered with 0.5 mg/kg milrinone at hours 1 and 12 and sacrificed at hour 24). Significant differences were found between the early and late administration of milrinone in terms of both molecular and histopathological results. The results showed that the tissues were significantly preserved in the groups in which milrinone had been started in the early period compared to the sepsis control groups and the groups in which milrinone had been started in the late period. In addition to the positive inotropic effects of milrinone, its immunomodulatory properties that result in decreased cytokine storm can be beneficial during early period of sepsis.

The Effects of Chinese Medicine QRD, Antibiotics, and Probiotics on Therapy and Gut Microbiota in Septic Rats.

Sepsis is a common and often treacherous medical emergency with a high mortality and long-term complications in survivors. Though antibiotic therapy can reduce death rate of sepsis significantly, it impairs gut microbiota (GM), which play imperative roles in human health. In this study, we compared the therapeutic effects of antibiotics, probiotics, and Chinese medicine QRD on the survival rates of septic model and observed the GM characteristics of experimental rats via 16S rRNA gene amplicon sequencing. The 72 h survival rates of septic rat demonstrated the significant therapeutic effects in the three groups treated with antibiotics (AT), Chinses medicine QRD (QT), and probiotics (PT), which were elevated from the survival rate of 26.67% for the sepsis control group (ST) to 100.0% for AT, 88.24% for QT, and 58.33% for PT. The original characteristics of GM identified in the sham operation controls (SC) were relatively similar to those in PT and QT; nevertheless, the AT rats were shown dramatically decreased in the GM diversity. In addition, the septic rats in AT were revealed the higher abundances of Escherichia Shigella, Proteus, Morganella, Enterococcus, and Lysinibacillus, but the lower those of Parabacteroides, Alistipes, Desulfovibrio, Bacteroides, Helicobacter, Mucispirillum, Oscillibacter, Lachnospiraceae, and Ruminiclostridium 9, when compared to the PT and QT rats. By contrast, the GM of PT and QT rats shared similar diversity and structure. Our findings indicated that QRD increased the survival rates without impairment of the GM characteristics, which provides novel insights into the role of Chinese medicine in therapy and long-term recovery of sepsis.

Verification of the Effects of Red Light-emitting Diode Therapy on Acute Lung Injury in a Sepsis Model in Rats

The aim of this study was to evaluate the in vivo response of red light-emitting diode (LED) on acute lung injury (ALI) in a sepsis model in rats. Twenty rats were randomly allocated into two experimental groups (n=10): Control Sepsis Group (CS); sepsis and red LED group (SRL). The anterior region of the trachea and ventral regions of the chest (below the ribs), bilaterally were irradiated daily for two consecutive days, starting immediately after the surgery using red (630 nm) LED. The histological results showed that in red LED treated group presented a modulation of the lung inflammatory process, less intense alveolar septum thickening and decrease of the inflammatory cells. Moreover, LED significantly reduced the lung injury score and increased interleukin type 10 (IL-10) protein expression compared SG. These results suggest that LED was efficient in attenuating ALI in a sepsis model in rats by reducing inflammatory cells into lung tissue and enhancing the anti-inflammatory cytokine production.

Effects of prophylactic administration of glutamine on CD4+ T cell polarisation and kidney injury in mice with polymicrobial sepsis.

The present study investigated the effects of glutamine (GLN) pretreatment on CD4+ T cell polarisation and remote kidney injury in mice with gut-derived polymicrobial sepsis. Mice were randomly assigned to three groups: normal control fed with American Institute of Nutrition (AIN)-93G diet and two sepsis groups provided with either AIN-93G-based diet or identical components, except part of casein was replaced by GLN. Mice were given their respective diets for 2 weeks. Then, mice in the sepsis groups were performed with caecal ligation and puncture and were killed 72 h after the surgery. Blood, spleens and kidneys were collected for further examination. The results showed that sepsis resulted in decreased circulating and splenic total Tlymphocyte and CD4+ T cell percentages, whereas IL-4-, and forkhead box p3 (Foxp3)-expressing CD4+ T cells percentages were up-regulated. Compared with the sepsis control group, pretreatment with GLN maintained blood T and CD4+ T cells and reduced percentages of IL-4- and Foxp3-expressing CD4+ T cells. Also, a more pronounced activation and increased anti-apoptotic Bcl-2 gene expression of splenic CD4+ T cells were observed. Concomitant with the decreased plasma IL-6, keratinocyte-derived chemokine (KC) levels, the gene expression of KC, macrophage inflammatory protein-2 and renal injury biomarker kidney injury molecule-1 (Kim-1) were down-regulated when GLN was administered. These findings suggest that antecedent of GLN administration elicit a more balanced blood T helper cell polarisation, sustained T cell populations, prevented splenic CD4+ T cell apoptosis and attenuated kidney injury at late phase of polymicrobial sepsis. GLN may have benefits in subjects at risk of abdominal infection.

Therapeutic effect of phages on extensively drug-resistant Acinetobacter baumannii-induced sepsis in mice

Objective To study the therapeutic effect of phages on extensively drug-resistant Acinetobacter baumannii-induced sepsis in mice. Methods (1) Sixty BALB/c mice were divided into blank control group, sepsis control group, antibiotics treatment group, phage treatment group, and phage control group according to the random number table, with 12 mice in each group. Mice in blank control group were intraperitoneally (the same injection position below) injected with 1 mL normal saline. Mice in sepsis control group, antibiotics treatment group, and phage treatment group were injected with 1 mL extensively drug-resistant Acinetobacter baumannii (the strain was isolated from the blood of a severely burned patient hospitalized in our unit) in the concentration of 5×107 colony-forming unit/mL to reproduce sepsis model. Two hours later, mice in sepsis control group, antibiotics treatment group, and phage treatment group were injected with 1 mL saline, 1 mg/mL imipenem/cilastatin, and 1×108 plaque-forming unit (PFU)/mL phages screened based on above-mentioned Acinetobacter baumannii (the same phages below) respectively. Mice in phage control group were injected with 1 mL phages in the titer of 1×108 PFU/mL. The injection was performed continuously for 7 days in each living mouse, and the survival situation of mice was observed each day to calculate the survival ratio in one week. (2) Another 60 BALB/c mice were grouped and treated as in experiment (1), and the injection was performed continuously for 5 days in each living mouse. On experiment day 2, 4, and 6, 3 mice from each group were selected (if the number of survived mouse in any group was less than 3 at sample collecting, all the survived mice were selected), and blood was drawn to determine white blood cell count (WBC, with 3 samples at each time point in each group). On experiment day 2, blood was drawn from the mice that had their blood taken earlier for bacterial culture, and lung, liver, kidney, and spleen tissue was collected from the same mice. The tissue samples were added to the LB solid medium after being homogenized and diluted for bacterial culture. The content of bacteria was calculated after the bacterial colony number was counted. Data were processed Wilcoxon rank sum test, one-way analysis of variance, LSD test and Kruskal-Wallis rank sum test. Results (1) On experiment day 7, there were 12, 8, 10, and 12 mice survived in blank control group, antibiotics treatment group, phage treatment group, and phage control group respectively, while no mouse survived in sepsis control group. Compared with that in sepsis control group, the survival ratio of mice was significantly higher in the other four groups (with Z values from 55.635 to 106.593, P values below 0.05). The survival ratio of mice in phage treatment group was slightly higher than that in antibiotics treatment group, without statistically significant difference (Z=2.797, P>0.05). (2) On experiment day 2, WBC data of mice in blank control group, phage treatment group, and phage control group were close[respectively (5.60±0.94)×109/L, (5.16±0.36)×109/L, and (5.26±1.89)×109/L], all significantly lower than the datum in sepsis control group[(8.64±0.64)×109/L, P 0.05). On experiment day 2, respectively 12, 7, and 2 mice were detected as blood bacterial culture-positive in sepsis control group, antibiotics treatment group, and phage treatment group, while no positive result was detected in the other two groups. Positive ratios of blood bacterial culture of mice in blank control group, phage treatment group, phage control group were significantly lower than the ratio in sepsis control group (with χ2 values from -30.000 to 30.000, P values below 0.01). Positive ratio of blood bacterial culture of mice in antibiotics treatment group was significantly higher than that in blank control group or phage control group (with χ2 values respectively 17.500 and -17.500, P values below 0.05). On experiment day 2, except for the kidney tissue of mice in phage treatment group, the bacteria load in each viscus of mice in blank control group, phage treatment group, and phage control group was significantly lower than that in sepsis control group (with χ2 values from -9.000 to 9.000, P values below 0.01). The bacteria load in kidney of mice in antibiotics treatment group was significantly higher than that in blank control group or phage control group (with χ2 values respectively -7.500 and 7.500, P values below 0.05). Conclusions Phages can significantly improve survival ratio, control inflammation response, and effectively clean bacteria in lung, liver, spleen, and kidney in treating extensively drug-resistant Acinetobacter baumannii-induced sepsis in mice. Key words: Burns; Acinetobacter baumannii; Bacteriophages; Sepsis; Extensively drug-resistant

Effect of siRNA against NF-κB on sepsis‑induced acute lung injury in a mouse model.

The aim of the present study was to explore the protective effect of small interfering RNA (siRNA) against nuclear factor κB (NF-κB) p65 on sepsis-induced acute lung injury (ALI) in mice. In total, 70 male Kunming mice were randomly divided into a healthy control group, a sepsis group, a specific interfering group and a scrambled control group (Sc), and the latter three groups were divided into post-operational 6 and 12 h subgroups, each of which consisted of 10 mice. The mice were administered with NF-κB siRNA, scrambled siRNA and normal saline via tail vein injection. Following 1 h, a mouse model of septic ALI was produced by cecal ligation and puncture (CLP) in the two siRNA groups and the sepsis control group. At 6 and 12 h post-operation, the experimental mice were sacrificed and the lung tissue samples were collected. Histopathological changes, wet/dry ratio of lung weight, NF-κB protein and NF-κB p65 mRNA levels, matrix metalloproteinase-9 (MMP-9) mRNA and protein activity were detected. Compared with the sepsis group and the Sc at the corresponding time, the expression levels of NF-κB p65 mRNA, the lung injury of experimental mice, the wet/dry ratio and the levels of MMP-9 mRNA and protein activity decreased, and significant differences were observed at 6 h post-operation (P<0.05). RNA interference against NF-κB p65 was able to decrease the expression of NF-κB and further inhibit the early phasic excessive inflammatory reaction in sepsis, which may alleviate ALI.

Xuebijing Protects Rats from Sepsis Challenged withAcinetobacter baumanniiby Promoting Annexin A1 Expression and Inhibiting Proinflammatory Cytokines Secretion

Xuebijing (XBJ) injection is a herbal medicine that has been widely used in the treatment of sepsis in China; however, its role in the development and progression of Acinetobacter baumannii sepsis and the underlying mechanisms remain uninvestigated. In the present study, fifty-four male Wistar rats were randomly assigned to normal-control group, sepsis-control group, and sepsis + XBJ group, each containing three subgroups of different treatment time periods (6, 12, and 24 hrs following injection, resp.). The sepsis model was established by intraperitoneal injection of A. baumannii ATCC 19606. For XBJ treatment, 4 mL/kg XBJ was administrated simultaneously by intravenous injection through caudal vein every 12 hrs. All animals demonstrated ill state, obvious intestinal dysfunction, histopathological lung damages, and overactive inflammatory responses after A. baumannii infection, and these events could be partially reversed by XBJ treatment from the beginning of infection. XBJ induced an increase in the expression of anti-inflammatory mediator annexin A1; however, two proinflammatory cytokines, interleukin-8 (IL-8) and tumor necrosis factor-α (TNF-α), were decreased at the each monitored time point. These findings suggested that XBJ via its cytokine-mediated anti-inflammatory effects might have a potential role in preventing the progression of A. baumannii infection to sepsis by early administration.

Ornithodoros moubata complement inhibitor (OmCI)—A novel and efficient C5 inhibitor in the pig

Complement and CD14/TLR4 are two important upstream components of innate immunity. Inflammation induced by these danger sensors may be excessive and detrimental in conditions such as sepsis. Inhibition of complement or CD14/TLR4 differentially attenuate experimentally induced inflammation. Combined inhibition has a more pronounced effect, almost abolishing Escherichia coli (E. coli)-induced cytokine release, oxidative burst and expression of the cell-surface marker CD11b in vitro in human studies. We recently showed that an anti-CD14 antibody attenuated proinflammatory cytokines, granulocyte activation and hypercoagulation in E. coli-induced sepsis in vivo in pigs. Combination with a complement inhibitor might enhance the antiinflammatory effect. The aim of the present study was to explore the efficacy in pig of the Ornithodoros moubata complement inhibitor (OmCI) that specifically binds C5 and prevents release of C5a and formation of the terminal complement complex (TCC). Porcine serum and whole blood anticoagulated with lepuridin was incubated with increasing doses of OmCI and activated with E. coli. A pilot study was additionally conducted in vivo in pigs, whereby sepsis was induced by E. coli and the effect of OmCI on the inflammatory response was investigated. Inhibition of complement activity, as evaluated by functional assay of the three initial pathways, was complete at a dose of 2.5 μg OmCI/mL in vitro and 1 mg OmCI/kg in vivo. TCC formation induced by E. coli was abolished in vitro at 5 μg OmCI/mL whole blood, while expression of wCD11R3, the pig analog of human CD11b, was reduced by more than 50% at 5 μg OmCI/mL whole blood. In vivo, OmCI attenuated the increase in IL-8 and TNF-α, and partly protected against the fall in leukocytes seen in the sepsis control group. In conclusion, OmCI efficiently inhibited pig complement activation, showed additional anti-inflammatory effects and is a promising candidate inhibitor for further sepsis studies in vivo.

DEPLETION OF NK CELLS IN A MURINE POLYTRAUMA MODEL IS ASSOCIATED WITH IMPROVED OUTCOME AND A MODULATION OF THE INFLAMMATORY RESPONSE

Sepsis and associated diseases such as systemic inflammatory response syndrome and multiple organ dysfunction syndrome represent common posttraumatic complications on intensive care units induced by a variety of body defense mechanisms. Natural killer (NK) cells are part of the innate immune system. They are thought to play an important role in the development of such syndromes by interplay with other immune cell types and subsequent activation of the inflammatory cascade. To test this hypothesis, NK cells were depleted by administration of antimouse asialo-GM1 antibody in a murine polytrauma model consisting of femur fracture, hemorrhagic shock, and subsequent sepsis. Mortality and immune parameters such as cytokine expression in lung and liver, lymphocyte phenotyping, lymphocyte apoptosis, and organ pathology were determined 96 h after sepsis induction. Survival values showed 50% in the control sepsis group and 100% after NK cell depletion. Thus, NK cell depletion resulted in 50% mortality reduction. Furthermore, we found reductions in the inflammatory response, represented by IL-6 expression in liver, and a decrease in infiltrating neutrophils in the liver and lung. In addition, lymphocyte apoptosis in spleen was decreased by depletion of NK cells. Taken together, these data demonstrate that NK cells contribute to the pathogenetic pathways in a murine polytrauma model. One main mechanism of action seems to be the induction of systemic inflammatory events. Thus, depletion of NK cells results in attenuated inflammation and an overall improvement in outcome. Therefore, NK cells can be considered as important targets for therapeutic strategies.

N,N′-DIACETYLCHITOBIOSE, AN INHIBITOR OF LYSOZYME, REVERSES MYOCARDIAL DEPRESSION AND LESSENS NOREPINEPHRINE REQUIREMENTS IN ESCHERICHIA COLI SEPSIS IN DOGS

Cardiovascular dysfunction in septic shock (SS) is ascribed to the release of inflammatory mediators. Norepinephrine (NE) is often administered to treat low MAP in SS. We recently found that lysozyme c (Lzm-S) released from leukocytes was a mediator of myocardial depression in an Escherichia coil model of SS in dogs. This effect can be blocked in an in vitro preparation by chitobiose, a competitive inhibitor of Lzm-S. In the present study, we examined whether chitobiose treatment can reverse myocardial depression and obviate NE requirements in two respective canine E. coli preparations. In a 6-h study, we administered chitobiose after 3.5 h of E. coli bacteremia and compared stroke work (SW) and MAP at 6 h with a sepsis control group. In a 12-h study, we determined whether chitobiose treatment can reduce the need for NE requirements during 12 h of bacteremia. In the latter study, either chitobiose or NE was given when MAP decreased approximately 20% from the presepsis value in respective groups. In anesthetized, mechanically ventilated dogs, we monitored hemodynamic parameters during continuous E. coli infusion. In the 6-h study, chitobiose improved SW and MAP at the 6-h period as compared with the nontreated sepsis group. In the 12-h study, SW and MAP increased after chitobiose without the necessity of NE administration. These results suggest that inhibitors of Lzm-S such as chitobiose may improve myocardial depression and reduce the need for NE requirements in SS.

High dose antithrombin III inhibits HMGB1 and improves endotoxin-induced acute lung injury in rats

High mobility group box 1 (HMGB1) is an important factor in the development of sepsis. Previous work suggests that antithrombin III (ATIII) inhibits inflammation, but the mechanism of action is still poorly understood. Prospective controlled animal study in a university laboratory. Rats were randomly divided into a lipopolysaccharide (LPS)-induced sepsis control group and an ATIII-treated experimental group. Animals in the experimental group received a bolus of 250 units/kg of ATIII injected into the tail vein. Animals receiving high-dose ATIII (250 units/kg) had significantly improved lung histopathology and survival compared to the control rats. We measured serum and lung levels of various cytokines and HMGB1 at regular intervals from 0 to 12 h after the induction of sepsis and demonstrated lower HMGB1 levels over time in ATIII-treated animals. In an in vitro experiment, we stimulated the mouse macrophage cell line RAW 264.7 with LPS in the presence or absence of ATIII. Subsequent measurement of HMGB1 concentrations in the supernatant and cell signaling molecules in cell lysates revealed an ATIII dose-dependent decrease in HMGB1 release. Furthermore, inhibition of IkB and p42 phosphorylation was observed with the administration of ATIII, suggestive of downstream signaling pathways. High-dose ATIII decreases lung pathology and reduces mortality in a rat sepsis model. This finding may be mediated by the inhibition of HMGB1.

Selective Downregulation of Neutrophils by a Phosphatidic Acid Generation Inhibitor in a Porcine Sepsis Model

Effects of lisofylline (1-(5-R-hydroxyhexyl)-3,7-dimethylxanthine), a functional inhibitor of phosphatidic acid (PA) generation derived fromde novosynthesis, on neutrophil function were examined in a porcine sepsis model. Hanford minipigs (18–25 kg) were randomly separated into six groups of six animals each: (1) saline control group; (2) sepsis control group, infused withPseudomonas aeruginosa(1 × 106colony-forming units/kg/min) for 2 h; (3) lisofylline control group, given a 25 mg/kg bolus of lisofylline 30 min prior to time zero, followed by a continuous infusion of 10 mg/kg/h throughout the study; (4) lisofylline pretreatment sepsis group, given lisofylline 30 min prior to sepsis, (5) lisofylline 1-h post-treatment sepsis group, and (6) lisofylline 2-h post-treatment sepsis group. All animals were studied for 6 h. Neutrophils were isolated at −0.5, 2, and 6 h. In the pretreatment and 1-h post-treatment groups, sepsis-induced neutrophil attachment to fibronectin was significantly attenuated. Sepsis-enhanced phagocytic activity was significantly reduced in the lisofylline pretreatment sepsis group, but not in the post-treatment groups. No treatment affected phorbol 12-myristate 13-acetate-induced chemiluminescence and basal filamentous actin content, which increased in sepsis, and cap formation, which declined in sepsis. Sepsis caused neutropenia, pretreatment produced neutrophilia, and 1-h post-treatment caused the neutropenia to recover to control levels. Interestingly, toward the end of the 6-h period, the neutrophil count was higher in the lisofylline control group than in the saline control groups. Thus, the inhibition of PA generation fromde novosynthesis during sepsis not only can selectively downregulate some neutrophil functions but can also reverse neutropenia.

The effects of post-treatment with lisofylline, a phosphatidic acid generation inhibitor, on sepsis-induced acute lung injury in pigs.

The effects of lisofylline [(R)-1-(5-hydroxyhexyl)-3,7-dimethylxanthine] (LSF), an inhibitor of de novo phosphatidic acid (PA) generation, on sepsis-induced acute lung injury was studied using Hanford minipigs weighing 18 to 25 kg. Sepsis was induced by an intravenous infusion of Pseudomonas aeruginosa (1 x 10(6)/colony-forming units/kg/min over 2 h). Saline was used as the control vehicle. Six groups were studied: saline control group (SALINE: n = 5); sepsis control group (SEPSIS: n = 5); LSF control group (LSF: n = 5), which received a 25-mg/kgbolus of LSF 30 min before time zero followed by continuous infusion of 10 mg/kg/h throughout the study; LSF-treated septic groups, which were treated with LSF 30 min prior to sepsis (Pre: n = 5), 1 h postonset (Post-1 h: n = 8) or h postonset (Post-2 h: n = 8) of the bacterial infusion. Hemodynamics PaO2, neutrophil counts, and plasma porcine tumor necrosis factor-alpha concentrations were monitored for 6 h. After the minipigs were killed, lung tissue was sampled to measured wet-to-dry weight ratio (W/D), tissue albumin index (TAI), thiobarbituric acid-reactive material content (TBARM), and myeloperoxidase (MPO) activity. Compared with the SALINE group, the SEPSIS group showed significant systemic hypotension, pulmonary hypertension, arterial hypoxemia, neutropenia, and increase in TNF-alpha, MPO activity, W/D, TBARM, and TAI. LSF treatment attenuated sepsis-induced pulmonary hypertension, neutropenia, and hypoxemia, and increased MPO activity and lung injury measurements in the Pre and Post-1 h groups, but its efficacy was blunted in the Post-2 h group. Plasma TNF-alpha was decreased only in the Pre group. Thus, inhibition of intracellular PA generation through de novo pathways attenuates sepsis-induced acute lung injury.