[Study on anisodamine hydrobromide improves Th17/Treg imbalance in resuscitated pigs].

OBJECTIVE: To investigate whether anisodamine can regulate the ratio of helper T helper cells/regulatory T cells (Th17/Treg) and its protective effect on animals after resuscitation. METHODS: Twenty-four Beijing white minipigs were randomly divided into sham operation group (Sham group), resuscitation and normal saline group (SA group), and resuscitation and anisodamine hydrobromide group (AH group), with 8 pigs in each group. In SA group and AH group, ventricular fibrillation was induced by continuous stimulation with intraventricular electrodes for 8 minutes and then resuscitated to establish ischemia/reperfusion (I/R) model. In SA group, after cardiopulmonary resuscitation (CPR), only normal saline was intravenously infused, while in AH group, normal saline and anisodamine hydrobromide were given intravenously at the same time point. Hemodynamic indexes, arterial blood gas analysis indexes, interleukins (IL-17, IL-10) levels in venous blood and IL-17/IL-10 ratio were recorded at 6 different time points: baseline, immediately after return of spontaneous circulation (ROSC), 1 hour, 2 hours, 4 hours and 6 hours after ROSC. The animals were sacrificed at 6 hours after ROSC, and intestinal lymphatic tissues were taken to observe pathological changes under light microscope. At the same time, the levels of IL-17 and IL-10 in intestinal lymphatic tissue were measured (the ratio of IL-17/IL-10 represents the ratio of Th17/Treg cytokines) to evaluate the immune status of the resuscitated animals. The bacterial translocations of different groups were evaluated by culturing intestinal lymphoid tissue. RESULTS: With the extension of ROSC time, the levels of IL-17 in venous blood and the IL-17/IL-10 ratio in pig blood samples continued to decrease, while the levels of IL-10 continued to increase. From 2 hours after ROSC, the IL-17/IL-10 ratio in AH group was significantly higher than that in SA group continued until at 6 hours after ROSC (0.79±0.05 vs. 0.49±0.08, P < 0.05). Light microscopy showed that the number and size of lymph nodules in the cortex of intestinal lymphatic tissue were less in AH group, compared with SA group. Compared with Sham group, the levels of IL-17 and IL-17/IL-10 ratio also decreased in intestinal lymphatic tissue at 6 hours after ROSC [IL-17 (ng/L): 155.23±0.92, 178.76±7.25 vs. 209.21±19.82, IL-17/IL-10 ratio: 1.43±0.13, 1.92±0.18 vs. 3.30±0.31, all P < 0.05], and IL-10 increased significantly (ng/L: 109.85±11.60, 93.55±81.83 vs. 63.45±0.62, all P < 0.05); IL-17/IL-10 ratio in AH group was significantly higher than that in SA group (1.92±0.18 vs. 1.43±0.13, P < 0.05). Tissue culture indicated the intestinal bacterial translocation after resuscitation, suggesting that the animals had immunosuppression and the increased risk of intestinal secondary infection after resuscitation. Compared with SA group, the risk of bacterial translocation was lower than that in AH group [62.5% (5/8) vs. 87.5% (7/8), P < 0.05]. CONCLUSIONS: Anisodamine plays an immunomodulatory role by affecting the balance of Th17/Treg cytokines in resuscitated animals, so as to reduce the risk of intestinal secondary infection and has an organ protective effect.

Anisodamine hydrobromide ameliorates cardiac damage after resuscitation.

The microcirculation is correlated with the prognosis of patients with cardiac arrest and changes after resuscitation. In the present study, the effects of anisodamine hydrobromide (AH) on microcirculation was investigated and its potential mechanisms were explored. A total of 24 pigs were randomly grouped into three groups (n=8): Sham, Saline and AH group. After pigs were anesthetized, intubated and mechanically ventilated, ventricular fibrillation was induced by electrical stimulation. After 8 min, cardiopulmonary resuscitation was given to the restoration of spontaneous circulation (ROSC). Arteriovenous blood was collected at baseline and 0, 1, 2, 4 and 6 h after ROSC to measure blood gas and cytokines. Perfused vessel density (PVD) and microvascular flow index (MFI) were measured to reflect the microcirculation. Continuous cardiac output and global ejection fraction were measured to indicate hemodynamics. Compared with Sham group, PVD and MFI in the intestines and the sublingual regions decreased significantly after resuscitation. The microcirculation recovered faster in the AH group than the SA group. The decrease of intestinal microcirculatory blood flow was closely related to the decrease of sublingual microcirculatory blood flow. The cardiac function was impaired after resuscitation, and a decrease of IFN-γ as well as IL-2 and an increase of IL-4 as well as IL-10 suggested the immune imbalance. The microcirculation changes in sublingual regions were closely related to the changes in intestines. AH could improve the immune imbalance after resuscitation and was beneficial to the recovery of cardiac function.

Protection of intestinal immune barrier against ischemia/reperfusion injury in a swine model using anisodamine hydrobromide

Abstract Intestinal ischemia/reperfusion (I/R) causes barrier impairment and bacterial influx. This study explored the protective effects of anisodamine hydrobromide (AH) on intestinal I/R injury caused by cardiopulmonary resuscitation (CPR) after cardiac arrest (CA). After successful CPR, minipigs were randomly divided into two groups (n = 8): saline and AH (4 mg/kg), and then treated with saline or AH via central venous injection, respectively. The same procedures without ventricular fibrillation initiation were conducted in the Sham group (n = 8). Levels of interferon gamma (IFN-γ) and interleukin 4 (IL-4) were measured at different time points (0, 0.5, 1, 2, 4, and 6 h) in serum and 6 h in gut associated lymphoid tissues (GALTs) after the return of spontaneous circulation (ROSC) to evaluate changes in the proportion of T-helper type 1 (Th1) and T-helper type 2 (Th2). Moreover, the positive culture rates of GALTs were examined to evaluate bacterial translocation. AH treatment markedly alleviated aberrant arterial blood gas and hemodynamics as well as intestinal macroscopic and morphological changes after CPR. Moreover, AH treatment significantly increased IFN-γ and decreased IL-4 in both serum and GALTs. Furthermore, AH treatment dramatically decreased positive bacterial growth in GALTs. AH treatment mitigated immunosuppression caused by intestinal I/R and protected the intestinal immune barrier against bacterial translocation, thereby reducing the risk of secondary intestinal infection

MicroRNA-92a inhibits macrophage antiviral response by targeting retinoic acid inducible gene-I.

MicroRNAs are short, non-coding RNAs that have been shown to regulate a wide range of biological processes, including host antiviral immune responses. In the present study, microRNA-92a (miR-92a) was identified as a negative regulator in macrophage-mediated antiviral responses. Overexpression of miR-92a decreases vesicular stomatitis virus (VSV)-induced production of type-I IFNs and facilitates viral replication in macrophages. The mechanism is that miR-92a directly targets RIG-I and reduces its expression, thereby attenuating VSV-triggered activation of TBK-binding kinase 1 and IRF3, both of which are crucial for initiating transcription of type-I IFN genes. Our results demonstrate for the first time the novel role of miR-92a in suppressing antiviral innate immunity.

[Clinical and laboratory features of macrophage activation syndrome].

OBJECTIVE: To study the clinical and laboratory features of macrophage activation syndrome (MAS) at the early stage of diagnosis, and to explore a method for early identification of MAS. METHODS: A retrospective analysis was performed for the demographic data, clinical and laboratory features, and treatment outcomes of 21 MAS patients. RESULTS: Of the 21 MAS patients, 14 had systemic juvenile idiopathic arthritis, 5 had Kawasaki disease (KD), and 2 had connective tissue disease (CTD) as primary diseases. The median time of MAS onset was 19 days. The KD patients had the shortest time of MAS onset, while the CTD patients had the longest onset time (P=0.009). The top 10 clinical symptoms were fever (95%), rash (86%), lymph node enlargement (67%), hemophagocytic phenomenon in bone marrow (63%), pulmonary disease (62%), serous effusion (62%), hepatomegaly (52%), cerebrospinal fluid abnormalities (50%), central nervous system damage (43%), and splenomegaly (38%). The median of hemoglobin level was lower than the normal value. The medians of C-reactive protein level and erythrocyte sedimentation rate were higher than the normal values. There were significant increases in serum ferritin, glutamic-pyruvic transaminase, aspartate aminotransferase, lactate dehydrogenase, and triglyceride. The median of fibrinogen level was lower than the normal value. There were significant increases in D-dimer, interleukin-6 (IL-6), interleukin-10 (IL-10), and interferon-γ (IFN-γ). Of the 21 patients, 20 were improved and discharged. CONCLUSIONS: If patients with rheumatic disease have persistent fever, hepatic dysfunction, coagulation disorders, multiple organ impairment, significantly increased IL-10 and IFN-γ, and a persistent increase in serum ferritin, the development of MAS should be considered.

Cytokines as the good predictors of refractory Mycoplasma pneumoniae pneumonia in school-aged children.

Excessive immune response against pathogens may play an important role in refracory Mycoplasma pneumoniae pneumonia (RMPP). The aim of this study was to elucidate the associations between cytokines and the prediction of RMPP in school-aged patients. Retrospective analysis was performed on school-aged children with Mycoplasma pneumoniae pneumonia (MPP) hospitalized in our hospital between January 1, 2011 and December 31, 2015. The clinical charcteristics, including the cytokines in serum between the RMPP group and the general Mycoplasma pneumoniae pneumonia (GMPP) group were compared and the predictive values of RMPP were explored. Of total 180 patients, 115 patients were in the GMPP group, 65 were in the RMPP group. We found the levels of cytokines, including nterleukin (IL)-6, IL-10, interferon gamma (IFN-γ) in RMPP group were significantly higher than those in GMPP group (P < 0.01). In ROC curve analysis, IL-10 and IFN-γ were useful for differentiating patients with RMPP from those with GMPP. Logistic regression analysis showed that the IL-10 ≥ 3.65 pg/ml and IFN-γ ≥ 29.05 pg/ml were significant predictors regarding to RMPP. Additionally, a positive correlation between serum IL-10 and IFN-γ concentrations was observed. CONCLUSIONS: IL-10 and IFN-γ could be used as the good predictors of RMPP in school-aged children.

High immunoglobulin E values at admission predict mortality in ED patients with sepsis.

OBJECTIVES: Gamma immunoglobulin E (IgE) is associated with allergic reactions but has not been described as being activated after sepsis. This study aimed at detecting the prognostic value of plasma IgE level in sepsis progression in the emergency department (ED). METHODS: Plasma IgE and related cytokines levels were measured on enrollment, and the Acute Physiology and Chronic Health Evaluation II score, Sequential Organ Failure Assessment score, and Mortality in Emergency Department Sepsis score were calculated on ED admission. A 28-day follow-up was performed for all patients. RESULTS: A total of 480 patients were consecutively enrolled in this study. The results revealed that nonsurvivors were in a more severe critical state, with reflected by higher IgE level and higher scoring systems (P<.001). Multivariate logistic regression analysis showed that IgE level was independent predictor of severe sepsis (odds ratio, 1.034; 95% confidence interval, 1.023-1.044; P<.001) and 28-day mortality (odds ratio, 1.038; 95% confidence interval, 1.027-1.053; P<.001). The areas under the receiver operating characteristic curve (AUC) analysis showed that IgE was a useful parameter in prognosis of severe sepsis (AUC was 0.830; cutoff value was 303.08µg/L) and 28-day mortality (AUC was 0.700; cutoff value was 299.96µg/L), Importantly, the AUC of combination of IgE and Mortality in Emergency Department Sepsis score performed for the most significant prognostic ability than each parameter, respectively, in this cohort (P<.001). CONCLUSIONS: The results of this study indicate that septic patients with higher IgE level present with higher risk of mortality, and a combination of IgE level with scoring systems significantly increased the predictive accuracy for severe sepsis and 28-day mortality.

Diagnostic and prognostic value of neutrophil gelatinase-associated lipocalin, matrix metalloproteinase-9, and tissue inhibitor of matrix metalloproteinases-1 for sepsis in the Emergency Department: an observational study.

INTRODUCTION: The aim of this study was to evaluate the early diagnostic, risk stratification and prognostic value of neutrophil gelatinase-associated lipocalin (NGAL), matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), compared with procalcitonin (PCT) and the Mortality in Emergency Department Sepsis (MEDS) score in septic patients in the emergency department (ED). METHODS: In total, 480 consecutive adult patients were enrolled in this study. They fulfilled the systemic inflammatory response syndrome (SIRS) criteria and were admitted to the ED of Beijing Chaoyang Hospital from February 2013 to August 2013. A total of 40 healthy controls comprised the control group. The patients were classified into four groups: SIRS, sepsis, severe sepsis, and septic shock. Serum NGAL, MMP-9, TIMP-1 and PCT were measured, and MEDS score was calculated at enrollment. The prognostic values of NGAL, MMP-9 and TIMP-1 were compared with PCT and MEDS score. A 28-day follow-up was performed for all patients. RESULTS: The median levels of serum NGAL and TIMP-1 increased with sepsis severity. The areas under the receiver operating characteristic (AUC) curves of NGAL or TIMP-1 were greater than those of PCT and MEDS score in diagnosing and predicting 28-day mortality, and the AUC of a combination of NGAL and MEDS score or TIMP-1 and MEDS score was more significant. Serum NGAL, MMP-9 and TIMP-1 levels were significantly higher in non-survivors than survivors at 28 days' follow-up. In addition, the level of NGAL was much higher in septic patients with acute kidney injury (AKI) than those without AKI. NGAL, TIMP-1, MMP-9 and MEDS score were found to be independent predictors of 28-day mortality in septic patients. The levels of serum NGAL and TIMP-1 were positively correlated with PCT and MEDS score in every septic group. CONCLUSIONS: NGAL and TIMP-1 are valuable for the risk stratification, early diagnosis and prognostication of sepsis in the ED. NGAL is also a valuable biomarker for prognosis of septic patients with AKI in the ED.

Prognostic significance of hypothalamic-pituitary-adrenal axis hormones in early sepsis: a study performed in the emergency department.

PURPOSE: The response of the hypothalamic-pituitary-adrenal (HPA) axis to the sustained stress of sepsis has been the focus of study in recent years because the early phase of sepsis is known to be dominated by major alterations in the HPA axis. This prospective observational study aimed at assessing the predictive values of copeptin and HPA hormones in determining sepsis progression and mortality in the emergency department (ED). METHODS: Serum arginine vasopressin (AVP) and copeptin concentrations were measured upon ED admission. Baseline levels of total and free cortisol and adrenocorticotrophic hormone (ACTH) were measured within 24 h of ED admission. Mortality in Emergency Department Sepsis (MEDS) score was calculated at enrollment. RESULTS: Our findings demonstrated that serum copeptin, baseline total cortisol, baseline free cortisol and baseline ACTH concentrations gradually increased, based upon the increasing severity of the disease (p < 0.001). Multivariate logistic regression analysis showed that copeptin and total cortisol baseline concentrations were independent predictors of septic shock (odds ratio = 1.034 and 1.355, respectively) and 28-day mortality (odds ratio = 1.039 and 1.499, respectively). The areas under the receiver operating characteristic curve (AUC) for copeptin level in prediction of septic shock was 0.856 and 28-day mortality was 0.826. Importantly, AUC analysis of the combination of copeptin, total cortisol baseline, MEDS score, and procalcitonin level resulted in a more significant prognostic ability than analysis of each parameter alone (p < 0.001). CONCLUSIONS: Increased copeptin and HPA hormones baseline levels may provide crucial information for risk stratification in a variety of septic states in the ED. Furthermore, measurements of copeptin level and serum baseline cortisol concentration are promising independent prognostic markers for mortality in patients with severe sepsis or septic shock.

Study on the neurotoxic effects of low-level lead exposure in rats.

OBJECTIVE: To investigate effects of developmental lead exposure on nitric oxide synthase (NOS) activity in different brain regions and on N-methyl-D-aspartate (NMDA) receptor mRNA expression in the hippocampus of rats. On the basis of these observations, we explored possible mechanisms by which lead exposure leads to impaired learning and memorizing abilities in children. METHODS: A series of rat animal models exposed to low levels of lead during the developing period was established (drinking water containing 0.025%, 0.05% and 0.075% lead acetate). NOS activities in the hippocampus, the cerebral cortex, the cerebellum and the brain stem were determined with fluorescence measurement and levels of mRNA expression of the NMDA receptor 2A (NR2A) subunit and NMDA receptor 2B (NR2B) subunit in the rat hippocampus were measured with Retro-translation (RT-PCR). RESULTS: There were no differences in the body weight of rat pups between any of the groups at any given time (P>0.05). The blood lead level of Pb-exposed rat pups showed a systematic pattern of change: at 14 d of age, it was lower than that at 7 d of age, then rising to the peak level at 21 d and finally falling to lower levels at 28 d. The hippocampal NOS activities of lead-exposed groups were all lower than that of the control group on the 21st and 28th day (P<0.01). NOS activities in the cerebellum of lead-exposed groups were all lower than that of the control group on the 21st and 28th day (P<0.001) and the NOS activity of the 0.025% group was significantly lower than that of the 0.05% and 0.075% groups on the 28th day (P<0.05). NOS activity in the cerebral cortex of the 0.075% group was significantly lower than that of the control, 0.025% and 0.05% groups on the four day spans (P<0.001). There was no significant difference of NOS activity in the brain stem between any lead-exposed group and the control group on the four day spans. In the 0.05% and the 0.075% groups, the level of NR2A mRNA expression was higher than that in the control group at 7 d and 14 d of age (P<0.05). In the 0.025% group, the level of NR2A was found to be higher than that in the control group at 7 d of age only (P<0.05). No significant differences were found for the levels of NR2B mRNA expression between any of the groups at any given time. CONCLUSIONS: NOS activity in the hippocampus, the cerebral cortex and the cerebellum are inhibited by lead exposure. The degree of the inhibitory effect depends on the time span of exposure and the lead concentration. Developmental low-level lead exposure was found to raise the level of NR2A mRNA expression in the hippocampus of rats. Developmental low-level lead exposure does not affect the level of NR2B mRNA expression in the hippocampus.

[Effects of lead exposure on nitric oxide synthase activity in different brain regions of developmental rat].

OBJECTIVE: To observe the influence of lead exposure on the activity of nitric oxide synthase (NOS) in different brain regions of rat. METHODS: By establishing a series of rat models exposed to different low levels of lead (drinking water containing 0.025%, 0.050%, 0.075% of lead acetate) during developing period, NOS activities in hippocampus, cerebellum, cerebral cortex and brain stem were studied. RESULTS: On the 21st day after birth, NOS activities in hippocampus of three levels of lead exposed groups [(1.53 +/- 0.20), (1.66 +/- 0.23), (1.88 +/- 0.32) U/mg pro respectively], and in cerebellum [(0.87 +/- 0.24), (0.85 +/- 0.09), (0.91 +/- 0.18) U/mg pro respectively] were significantly lower than those of control group [(2.36 +/- 0.18), (1.41 +/- 0.18) U/mg pro, respectively, P < 0.01]. NOS activities in cerebral cortex of 0.075% group [at 7, 14, 21 d of age [(1.29 +/- 0.14), (1.03 +/- 0.15), (0.69 +/- 0.10) U/mg pro] were significantly lower than those in control group [(2.54 +/- 0.31), (1.64 +/- 0.22), (1.24 +/- 0.14) U/mg pro respectively], and 0.025% group [(2.42 +/- 0.19), (1.59 +/- 0.17), (1.27 +/- 0.12) U/mg pro respectively], and 0.050% group [(2.56 +/- 0.53), (1.77 +/- 0.19), (1.24 +/- 0.10) U/mg pro respectively, P < 0.05]. There were no significant differences among control, 0.025%, and 0.050% groups (P > 0.05). Lead exposure had no influence on NOS activity in brain stem at the same age (P > 0.05). CONCLUSION: NOS activities in hippocampus, cerebellum and cerebral cortex were inhibited by low level lead exposure and the degree of the effect was related to Pb exposure time and/or level of Pb exposed.