Increased Interleukin-17-Producing γδT Cells in the Brain Exacerbate the Pathogenesis of Sepsis-Associated Encephalopathy and Sepsis-Induced Anxiety in Mice.

Overactivated microglia play a key role in sepsis-associated encephalopathy (SAE), although the involvement of T cells is unclear. γδT cells in the brain and meninges regulate normal fear responses via interleukin (IL)-17 in healthy mice. In our sepsis model, the mice showed exacerbated anxious behavior at 10 days post-induction (dpi). At 8 dpi, IL-17 mRNA was significantly upregulated in the brains of septic mice compared with those of control mice. Simultaneously, the number of γδT cells increased in the brains of septic mice in a severity-dependent manner. Additionally, IL-17-producing γδT cells, expressing both the C-X-C motif receptor (CXCR) 6 and the C-C motif receptor (CCR) 6, increased in mice brains, dependent on the severity of sepsis. The frequency of γδT cells in the meninges fluctuated similarly to that in the brain, peaking at 8 dpi of sepsis. Behavioral tests were performed on septic mice after the continuous administration of anti-γδTCR (α-γδTCR) or anti-IL-17A (α-IL-17A) antibodies to deplete the γδT cells and IL-17A, respectively. Compared with IgG-treated septic mice, α-γδTCR- and α-IL-17A-treated septic mice showed suppressed microglial activation and improvements in anxious behavior. These results suggested that CCR6+CXCR6+ IL-17-producing γδT cells in the brain and meninges promote the exacerbation of SAE and sepsis-induced psychological disorders in mice.

Sepsis causes neutrophil infiltration in muscle leading to muscle atrophy and weakness in mice.

Background: Sepsis-induced muscle atrophy leads to prolonged physical dysfunction. Although the interaction of muscle atrophy and macrophage has been reported in sepsis, the role of neutrophils in muscle atrophy has not been thoroughly investigated. This study sought to investigate the long-term changes in muscle-localized neutrophils after sepsis induction and their possible role in sepsis. Methods: Sepsis was induced in seven-week-old male C57BL/6J mice 8-12 (cecal slurry [CS] model) via intraperitoneal injection of 1 mg/g cecal slurry. The percentage change in body weight and grip strength was evaluated. The tibialis anterior muscles were dissected for microscopic examination of the cross-sectional area of myofibers or Fluorescence-activated cell sorting (FACS) analysis of immune cells. These changes were evaluated in the following conditions: (1) Longitudinal change until day 61, (2) CS concentration-dependent change on day 14 at the low (0.3 mg/g), middle (1.0 mg/g), and high (2.0 mg/g) concentrations, and (3) CS mice on day 14 treated with an anti-Ly6G antibody that depletes neutrophils. Results: Body weight and grip strength were significantly lower in the CS model until day 61 (body weight: 123.1% ± 1.8% vs. 130.3% ± 2.5%, p = 0.04; grip strength: 104.5% ± 3.8% vs. 119.3% ± 5.3%, p = 0.04). Likewise, cross-sectional muscle area gradually decreased until day 61 from the CS induction (895.6 [606.0-1304.9] µm2 vs. 718.8 [536.2-937.0] µm2, p < 0.01). The number of muscle-localized neutrophils increased from 2.3 ± 0.6 cell/mg on day 0 to 22.2 ± 13.0 cell/mg on day 14, and decreased thereafter. In terms of CS concentration-dependent change, cross-sectional area was smaller (484.4 ± 221.2 vs. 825.8 ± 436.2 µm2 [p < 0.001]) and grip strength was lower (71.4% ± 12.8% vs. 116.3% ± 7.4%, p = 0.01) in the CS High group compared with the control, with increased neutrophils (p = 0.03). Ly6G-depleted mice demonstrated significant increase of muscle cross-sectional area and grip strength compared with control mice (p < 0.01). Conclusions: Sepsis causes infiltration of neutrophils in muscles, leading to muscle atrophy and weakness. Depletion of neutrophils in muscle reverses sepsis-induced muscle atrophy and weakness. These results suggest that neutrophils may play a critical role in sepsis-induced muscle atrophy and weakness.

Prevalence and Long-Term Prognosis of Post-Intensive Care Syndrome after Sepsis: A Single-Center Prospective Observational Study.

Post-intensive care syndrome (PICS) comprises physical, mental, and cognitive disorders following a severe illness. The impact of PICS on long-term prognosis has not been fully investigated. This study aimed to: (1) clarify the frequency and clinical characteristics of PICS in sepsis patients and (2) explore the relationship between PICS occurrence and 2-year survival. Patients with sepsis admitted to intensive care unit were enrolled. Data on patient background; clinical information since admission; physical, mental, and cognitive impairments at 3-, 6-, and 12-months post-sepsis onset; 2-year survival; and cause of death were obtained from electronic medical records and telephonic interviews with patients and their families. At 3 months, comparisons of variables were undertaken in the PICS group and the non-PICS group. Among the 77 participants, the in-hospital mortality rate was 11% and the 2-year mortality rate was 52%. The frequencies of PICS at 3, 6, and 12 months were 70%, 60%, and 35%, respectively. The 2-year survival was lower in the PICS group than in the non-PICS group (54% vs. 94%, p < 0.01). More than half of the survivors had PICS at 3 and 6 months after sepsis. Among survivors with sepsis, those who developed PICS after 3 months had a lower 2-year survival.

ADAMTS13 activity decreases in the early phase of trauma associated with coagulopathy and systemic inflammation: a prospective observational study.

BACKGROUND: We conducted a prospective observational study for investigating the changes in the 13th member of a disintegrin-like and metalloprotease with thrombospondin type 1 motif (ADAMTS13) and its association with the coagulofibrinolytic response in adult trauma patients. METHODS: In 39 trauma patients hospitalized for longer than 7 days, time-course changes in biomarkers of coagulofibrinolysis and systemic inflammation along with ADAMTS13 activity were examined. The patients were stratified into three groups based on ADAMTS13 activities on admission (day 0): normal group (≥70%), mildly decreased group (≥50 and < 70%) and moderately decreased group (< 50%). RESULTS: Among 39 patients with a median Injury Severity Score (ISS) of 20, 11 patients developed disseminated intravascular coagulation (DIC) and 16 patients required transfusion. Six of 39 patients (15.4%) showed moderate decreased ADAMTS13 activity to < 50%, and 20 patients (51.3%) showed mild drops (≥50 and < 70%). These changes in ADAMTS13 activity on day 0 were significantly correlated with changes in IL-6 and other coagulofibrinolytic markers such as platelet counts, prothrombin time and fibrin/fibrinogen degradation product (FDP). Antithrombin activity (AT) and serum albumin (Alb) level showed significantly positive linear correlations with ADAMTS13 activity (AT: r = 0.513, p < 0.001; Alb: r = 0.647, p < 0.001). Simple logistic regression analyses showed that ADAMTS13 activity, if less than 50%, was significantly correlated with the development of DIC (OR 7.499, 95%CI 1.121-49.242, p = 0.038) and the need for transfusion of fresh frozen plasma (OR 9.000, 95%CI 1.327-61.025, p = 0.028). CONCLUSIONS: ADAMTS13 activity decreased even in the early phase of trauma, which was complicated by coagulopathy and systemic inflammation. Furthermore, the decrease in ADAMTS13 activity was correlated with DIC and plasma transfusion.

A Simple Method for Positioning the Traction Table during Fixation Surgery for a Displaced Femoral Trochanteric Fracture in a Patient Following Ipsilateral Above-the-knee Amputation: A Case Report.

INTRODUCTION: It is difficult to use a traction table during surgery for an ipsilateral displaced femoral trochanteric fracture following above-the-knee amputation. However, there are few reports regarding such cases. We describe the simple method we used for positioning the traction table and reducing fracture site during fixation surgery for a displaced femoral trochanteric fracture in this patient following above-the-knee amputation. CASE REPORT: An 80-year-old man was injured in a head-on collision with an oncoming vehicle. We diagnosed him with traumatic gastric perforation, multiple right lower leg fractures, and right lower leg severe crush wound. We performed right above-the-knee amputation and laparoscopic gastrorrhaphy for lifesaving purposes. Thereafter we performed internal fixation for the right femoral trochanteric fracture on the 5th day after the injury. In that operation, we first inserted a 2.4 mm Kirschner wire under fluoroscopic guidance 3 cm proximal to the femoral cut end and attached a horseshoe and traction rope to it. Then, we tightened the rope to the foot piece of the traction table and secured it. Although shortening of the bone fracture was reduced by traction, rotation control was impossible; therefore, the horseshoe was manually controlled through clean sheets during surgery to reduce rotational displacement. We performed internal fixation surgery using a trochanteric femoral nail in the usual manner. CONCLUSION: In the osteosynthesis surgery of displaced femoral trochanteric fractures following above-the-knee amputation, the method of inserting the Kirschner wire at the distal end of the patient's femur and pulling it through the rope enables surgeons to reduce fracture shortening. Reduction of rotational displacement was possible by controlling the horseshoe by hand. In this way, intramedullary nail fixation could be performed without trouble under fluoroscopic guidance.

Peripartum myocardial infarction associated with coronary spasm and acquired protein S deficiency: A case report.

RATIONALE: Coronary angiography (CAG) findings of acute myocardial infarction (AMI) in pregnant women are characterized by a high incidence of normal coronary arteries. This is the first report of AMI with normal coronary arteries during pregnancy, showing coronary spasm and pregnancy-related acquired protein S (PS) deficiency. PATIENT CONCERNS: A 30-year-old Japanese woman was admitted to an emergency department. One hour before admission, she developed sudden onset of precordial discomfort, back pain, and dyspnea. She was a primigravida at 39 weeks' gestation and had no abnormality in the pregnancy thus far. She had no history of heart disease, diabetes, hypertension, dyslipidemia, deep vein thrombosis (DVT), smoking, or oral contraceptive use and no family history of ischemic heart disease, hemostasis disorder, or DVT. She did not take any medication. DIAGNOSIS: Electrocardiography showed ST-segment elevations in leads II, III, aVF, and V2-V6. Heart-type fatty acid-binding protein was positive. Echocardiography showed hypokinesis of the anterior interventricular septum and inferior wall. Continuous intravenous infusion of isosorbide dinitrate was initiated. Coronary computed tomography angiography revealed diffuse narrowing of the apical segment of the left anterior descending coronary artery. Three hours after admission, troponin T became positive, and the following enzymes reached their peak levels: creatine kinase (CK), 1,886 U/L; CK-muscle/brain, 130 U/L. She was diagnosed with transmural AMI due to severe coronary spasm and administered benidipine hydrochloride. Five hours after admission, premature membrane rupture occurred. INTERVENTIONS: Emergency cesarean section was performed. There were no anesthetic or obstetrical complications during the operation. On postpartum day 1, the free PS antigen level was low (29%). On postpartum day 18, she was discharged with no reduction in physical performance. OUTCOMES: Four months after the infarction, CAG showed normal coronary arteries. Acetylcholine provocation test showed diffuse vasospasm in the coronary artery. She was advised that her next pregnancy should be carefully planned. Two years after delivery, free PS antigen level was within normal range, at 86%. She had not experienced recurrence of angina during the 2-year period. Her child was also developing normally. LESSONS: In addition to coronary spasm, pregnancy-related acquired PS deficiency may be involved in AMI etiology.

Anticoagulation Therapy Using rh-Thrombomodulin and/or Antithrombin III Agent is Associated With Reduction in in-Hospital Mortality in Septic Disseminated Intravascular Coagulation: A Nationwide Registry Study.

We analyzed the Nationwide Registry database on sepsis to examine the effects of an anticoagulation therapy, especially with rh-thrombomodulin (rh-TM) and/or antithrombin (AT) III agent, in septic disseminated intravascular coagulation (DIC) patients. In 3,193 patients enrolled after the exclusion, we investigated the association with in-hospital mortality using Cox proportional hazards models. DIC was diagnosed using the Japanese Association of Acute Medicine (JAAM) and the International Society of Thrombosis and Hemostasis (ISTH) criteria. To analyze the separate treatment effects of rh-TM and/or AT III, we excluded the data of 345 patients treated with all available anticoagulation treatments (rh-TM and/or AT III plus "other anticoagulants": protease inhibitors and heparin/heparinoids). The DIC criterion populations were as follows: JAAM DICs, n = 1,891 and ISTH DICs (overt DIC), n = 1,002. Septic DIC patients were divided into 3 groups: Group 1, no anticoagulation therapy for DIC; Group 2, received rh-TM and/or AT III; and Group 3, received only "other anticoagulants." In JAAM DIC patients, Group 2 did not show an independent association with a reduced risk of in-hospital mortality (hazard ratio [HR]: 0.86; 95% confidence interval [CI]: 0.73-1.01]) as compared with Group 1. However, in ISTH DIC patients, Group 2 showed an inverse association with the risk of in-hospital mortality (HR 0.74; 95% CI: 0.60-0.92) as compared with Group 1, but the same was not true for Group 3 (HR 0.73; 95% CI: 0.47-1.14). The present results support previous findings of the beneficial effects of anticoagulation therapies in septic DIC, also expands the importance of using rh-TM and/or AT agent for septic overt DIC.

Successful treatment for disseminated intravascular coagulation (DIC) corresponding to phenotype changes in a heat stroke patient.

BACKGROUND: Heat stroke induces coagulofibrinolytic activation, which leads to life-threatening disseminated intravascular coagulation (DIC). However, treatment strategies for DIC in heat stroke have not yet been established, and also, the time course changes in coagulofibrinolytic markers have not been thoroughly evaluated. We report a severe heat stroke case with DIC who was eventually saved by anti-DIC treatments in accordance with changes in coagulofibrinolytic markers. CASE PRESENTATION: A 45-year-old man was found unconscious outside, and his body temperature was elevated to 41.9 °C. For heat stroke, we performed an immediate tracheal intubation under the general anesthesia along with cooling by iced gastric lavage, cold fluid administration, and an intravascular cooling using Thermogard™. About 4 h after admission, his core temperature fell to 37 °C. We assessed coagulofibrinolytic biomarkers and treated in accordance with changes in these parameters. This case exhibited a biphasic change varying from an enhanced to a suppressed fibrinolytic type of DIC depending on the relative balance between fibrinolytic activation and the level of plasminogen activator inhibitor-1 (PAI-1). In the early phase with consumption coagulopathy and enhanced fibrinolysis, we transfused a large amount of fresh frozen plasma (FFP) and platelets with tranexamic acid, an antifibrinolytic agent, possibly providing relief for the bleeding tendency. Anticoagulant therapy using recombinant human thrombomodulin-α (rh-TM-α) and antithrombin III (ATIII) concentrate was especially effective for DIC with a suppressed fibrinolytic phenotype in the later phase, after which organ failure that included severe hepatic failure was remarkably improved. CONCLUSION: The present case may indicate the clinical significance of monitoring coagulifibrinolytic changes and the potential benefits of anticoagulants for heat stroke-induced DIC.

Decreased antithrombin activity in the early phase of trauma is strongly associated with extravascular leakage, but not with antithrombin consumption: a prospective observational study.

BACKGROUND: We conducted a prospective observational study for investigating coagulofibrinolytic changes and mechanisms of antithrombin (AT) alternations in trauma. METHODS: Trauma patients hospitalized for more than seven days were analyzed for coagulofibrinolytic biomarkers. The patients were stratified into two groups according to AT activity level on admission (day 0), comprising normal AT and low AT patients. RESULTS: Thirty-nine patients (median Injury Severity Score 20) exhibited initial coagulatory activation and triphasic fibrinolytic changes. AT activity did not show a negative linear correlation with levels of thrombin-antithrombin complex (TAT), a marker of coagulation activity and AT consumption, but was strongly correlated with levels of albumin (Alb), an index of vascular permeability, on day 0 (r = 0.702, p <  0.001). Furthermore, Alb was one of the independent predictors for AT on day 0. IL-6 on day 0 and thrombomodulin (TM) levels during the study period, reflecting systemic inflammation and endothelial cell injury, respectively, were significantly higher in the lower AT group (n = 10) than in the normal group (n = 29) (IL-6, p = 0.004; TM, p = 0.017). On days 2 and 4, TAT levels in the lower AT group were significantly higher than in the normal group. CONCLUSIONS: Trauma caused clear triphasic coagulofibrinolytic changes. Decreased AT in the later phase might lead to a prolonged hypercoagulation. AT reduction in the initial phase of trauma is strongly associated with extravascular leakage as suggested by the association of Alb depletion with IL-6 and TM elevation, but not with AT consumption.

Accuracy of screw fixation using the O-arm® and StealthStation® navigation system for unstable pelvic ring fractures.

PURPOSE: Screw fixation for unstable pelvic ring fractures is generally performed using the C-arm. However, some studies reported erroneous piercing with screws, nerve injuries, and vessel injuries. Recent studies have reported the efficacy of screw fixations using navigation systems. The purpose of this retrospective study was to investigate the accuracy of screw fixation using the O-arm® imaging system and StealthStation® navigation system for unstable pelvic ring fractures. METHODS: The participants were 10 patients with unstable pelvic ring fractures, who underwent screw fixations using the O-arm StealthStation navigation system (nine cases with iliosacral screw and one case with lateral compression screw). We investigated operation duration, bleeding during operation, the presence of complications during operation, and the presence of cortical bone perforation by the screws based on postoperative CT scan images. We also measured the difference in screw tip positions between intraoperative navigation screen shot images and postoperative CT scan images. RESULTS: The average operation duration was 71 min, average bleeding was 12 ml, and there were no nerve or vessel injuries during the operation. There was no cortical bone perforation by the screws. The average difference between intraoperative navigation images and postoperative CT images was 2.5 ± 0.9 mm, for all 18 screws used in this study. CONCLUSION: Our results suggest that the O-arm StealthStation navigation system provides accurate screw fixation for unstable pelvic ring fractures.

Sudden cerebral depression detected by bispectral index monitoring in cryptococcal meningitis with elevated near-fatal cerebrospinal fluid pressure.

Case: An increase in cerebrospinal fluid pressure (CSFP) is usually prominent in cryptococcal meningitis, which has a high mortality rate, so aggressive management to control CSFP is crucial. In this case, a 40-year-old-man survived cryptococcal meningitis treated with continuous spinal drainage under bispectral index (BIS) monitoring. He unexpectedly showed hypertension, went into a coma, and even loss his light reflexes due to CSFP elevation. His BIS values had abruptly dropped before developing these symptoms, but dramatically recovered after lumbar puncture drainage, suggesting that BIS monitoring could reflect cerebral function changes due to CSFP alternations. Outcome: Inducing continuous spinal drainage to control CSFP provided stable control of blood pressure and brain activity, which was continuously monitored by BIS, enabling us to provide prompt treatment. Conclusion: Cerebral depressions due to elevated CSFP may suddenly develop, so continuous spinal drainage is needed for preventing catastrophic events. Bispectral index could be useful for detecting early changes from CSFP elevation in meningitis cases with intracranial hypertension.

Novel Catalysts for Methane Combustion Based on Cobalt-Doped Lanthanum Silicates Having an Apatite-type Structure.

Novel PdO/La10Si6-xCoxO27-δ/γ-Al2O3 catalysts with applications to methane combustion were developed. These materials were based on the use of La10Si6-xCoxO27-δ as a promoter because this compound has an oxide-ion conducting apatite-type structure that allows the smooth migration of active oxygen to the PdO activator. Co3+/2+ ions were also introduced into the original La10Si6O27 lattice to enhance its redox properties. Temperature-programmed reduction measurements revealed that the oxygen supply was facilitated by introducing Co3+/2+, where the reduction was observed at 290 °C for La10Si6-xCoxO27-δ (x = 1.0), whereas no reduction was observed below 460 °C for La10Si6O27. Among the samples synthesized in this work, PdO/La10Si6-xCoxO27-δ/γ-Al2O3 (x = 1.0) exhibited the highest catalytic activity, allowing the complete oxidation of methane at 310 °C, a temperature 80 °C lower than the 390 °C required when employing PdO/La10Si6O27/γ-Al2O3.

Desflurane inhalation before ischemia increases ischemia-reperfusion-induced vascular leakage in isolated rabbit lungs.

BACKGROUND: Isoflurane and sevoflurane protect lungs with ischemia-reperfusion (IR) injury. We examined the influence of desflurane on IR lung injury using isolated rabbit lungs perfused with a physiological salt solution. METHODS: The isolated lungs were divided into three groups: IR, desflurane-treated ischemia-reperfusion (DES-IR), and ventilation/perfusion-continued control (Cont) groups (n = 6 per group). In the DES-IR group, inhalation of desflurane at 1 minimum alveolar concentration (MAC) was conducted in a stable 30-min phase. In the IR and DES-IR groups, ventilation/perfusion was stopped for 75 min after the stable phase. Subsequently, they were resumed. Each lung was placed on a balance, and weighed. Weight changes were measured serially throughout this experiment. The coefficient of filtration (Kfc) was determined immediately before ischemia and 60 min after reperfusion. Furthermore, bronchoalveolar lavage fluid (BALF) was collected from the right bronchus at the completion of the experiment. After the completion of the experiment, the left lung was dried, and the lung wet-to-dry weight ratio (W/D) was calculated. RESULTS: The Kfc values at 60 min after perfusion were 0.40 ± 0.13 ml/min/mmHg/100 g in the DES-IR group, 0.26 ± 0.07 ml/min/mmHg/100 g in the IR group, and 0.22 ± 0.08 (mean ± SD) ml/mmHg/100 g in the Cont group. In the DES-IR group, the Kfc at 60 min after the start of reperfusion was significantly higher than in the other groups. In the DES-IR group, W/D was significantly higher than in the Cont group. In the DES-IR group, the BALF concentrations of nitric oxide metabolites were significantly higher than in the other groups. In the DES-IR group, the total amount of vascular endothelial growth factor in BALF was significantly higher than in the Cont group. CONCLUSIONS: The pre-inhalation of desflurane at 1 MAC exacerbates pulmonary IR injury in isolated/perfused rabbit lungs.

Phosphoenolpyruvate administration protects ischemia-reperfusion injury in isolated rabbit lungs.

Phosphoenolpyruvate (PEP) is an intermediate metabolite of the glycolytic pathway and an in vivo high-energy phosphate compound. We have examined the protective effects of PEP on ischemia-reperfusion lung injury in isolated rabbits lungs perfused with a physiological salt solution. The lungs were divided into three treatment groups: (1) ischemia-reperfusion (IR), (2) ischemia-reperfusion with PEP treatment (PEP-IR), in which 1 mM PEP was pre-administered into the perfusate during the stable period, and (3) ventilation-perfusion continued without interruption (Cont). In the IR and PEP-IR groups, ventilation-perfusion was discontinued for about 60 min after a 30-min stable period and then restarted. The capillary filtration coefficients (K fc) and pyruvate concentration in the perfusate were determined immediately before ischemia and 30 and 60 min after reperfusion. The left lungs were dried at the end of the experiment to calculate the tissue wet-to-dry weight ratio (W/D). The K fc values after reperfusion were significantly higher in the IR group than in the other two groups. Pyruvate concentrations were significantly higher at three time-points in the PEP-IR group than in the other two groups. The W/D was significantly higher in the IR group than in the other two groups. Based on these results, we conclude that the administration of PEP prior to lung ischemia alleviates lung ischemia-reperfusion injury.

[Multiple cerebral infarction by air embolism associated with remarkable low BIS value during lung segmentectomy with video assisted thoracic surgery (VATS) technique: a case report].

A 64-year-old woman (151 cm, 43 kg) with well controlled hypertension was diagnosed as having right lung cancer at S8 segment. She underwent right S8 segmentectomy by video assisted thoracic surgery (VATS) under general anesthesia combined with epidural anesthesia. Her vital signs were stable and BIS value was around 45 before the surgeon injected the air using a syringe with a 22 G needle to confirm the lesion resected. After the injection of air, her systolic blood pressure rapidly increased from 120 to 170 mmHg and the BIS value suddenly decreased to 5. Blood propofol concentration was reduced from 3 microg x ml(-1) to 2 microg x ml(-1) in the target-controlled infusion technique, and thereby the BIS value increased slowly. She did not wake up nor maintain sufficient spontaneous breathing even 2 hours after the discontinuation of opioids, and was transferred to ICU with tracheal intubation. In ICU, she showed clonic convulsions. Urgent CT and MRI confirmed cerebral air embolism. Her vital signs were too unstable to choose hyperbaric oxygen therapy as her first treatment. Her consciousness was recovered and her trachea was extubated on 11th postoperative day. She was discharged with left hemiparalysis from hospital.

Angiotensin type 1 receptor antagonist inhibits lipopolysaccharide-induced stimulation of rat microglial cells by suppressing nuclear factor kappaB and activator protein-1 activation.

We investigated whether angiotensin (ANG) II and its receptors contribute to lipopolysaccharide (LPS)-induced microglial activation through activation of the proinflammatory transcription factors nuclear factor kappaB (NF-kappaB) and activator protein-1 (AP-1). Using primary microglial cell cultures, we examined whether losartan [ANG type 1 receptor (AT(1)) antagonist] alters the effects of LPS on: the production of interleukin-1 (IL-1) and nitric oxide, cell morphology, and NF-kappaB and AP-1 activities. Reverse transcription-polymerase chain reaction revealed that LPS-stimulated microglial cells exhibited marked mRNA expression for AT(1), ANG type 2 receptor (AT(2)) and the ANG II precursor angiotensinogen, whereas non-stimulated microglial cells expressed only those for AT(2) and angiotensinogen. We further demonstrated marked peptide/protein expression for AT(1) and ANG II in LPS-activated microglial cells. LPS (100 ng/mL)-stimulated microglial cells showed increased concentrations of IL-1 and nitrite (a relatively stable metabolite of nitric oxide), and increased expression of IL-1 mRNA as well as a morphological change from an amoeboid shape to a multipolar (mostly bipolar but sometimes tripolar) rod shape. These effects were all significantly inhibited by losartan treatment (10(-5) M or less). NF-kappaB and AP-1 activities were enhanced in LPS-stimulated microglial cells, effects that were significantly suppressed by losartan (10(-5) M). ANG II application enhanced the LPS-induced increases in IL-1 and nitrite concentrations, as well as the LPS-induced morphological changes and AP-1 activation, and these enhancements were inhibited by losartan (10(-5) M). These results suggest that endogenous ANG II enhances LPS-induced microglial activities through stimulation of the microglial AT(1), which itself evokes activation of the transcription factors NF-kappaB and AP-1.

ANP inhibits LPS-induced stimulation of rat microglial cells by suppressing NF-kappaB and AP-1 activations.

Atrial natriuretic peptide (ANP) contributes to the inhibition of such causes of inflammation as the lipopolysaccharide (LPS)-induced productions of nitric oxide (NO) and proinflammatory cytokines [including interleukin-1 (IL-1)] in macrophages. In the present study we used primary cultures of rat brain macrophage-like cells (i.e., microglial cells) to investigate whether ANP binding to its receptors inhibits LPS-induced microglial activation via effects on the activation of the proinflammatory transcription factors NF-kappaB and AP-1. The productions of NO and IL-1, as well as morphological changes, were examined to assess LPS-induced activation of microglial cells. Our RT-PCR study revealed that rat microglial cells express the mRNAs for ANP receptors (types A, B, and C) and that for the ANP molecule. LPS (100 ng/ml)-stimulated microglial cells showed increases in nitrite (a relatively stable metabolite of NO) and IL-1 concentrations, and in the expression of IL-1 mRNA, as well as a morphological change from an amoeboid shape to a multipolar (mostly bipolar, but sometimes tripolar) rod shape. These effects were all significantly inhibited by treatment with ANP (at 10(-6)M or less). The inhibition by ANP of the LPS-induced nitrite response was abrogated by a NP-receptor antagonist, HS-142-1 (100 ng/ml). NF-kappaB and AP-1 activities were enhanced in LPS-stimulated microglial cells, and these enhancements were significantly suppressed by ANP (10(-6)M). These results suggest that ANP inhibits LPS-stimulated activities in microglial cells through activation of microglial ANP receptors, leading to inhibitions of NF-kappaB and AP-1.

Systemic administration of polymyxin B induces hypothermia in rats via an inhibitory effect on metabolic rate.

Polymyxin B, a cyclic cationic polypeptide antibiotic, binds to the lipid A of bacterial endotoxin (lipopolysaccharide; LPS) to inhibit LPS-induced fever. On the basis of a casual observation, we hypothesised that in rats (unlike in rabbits and goats), intravenous (i.v.) polymyxin B would decrease resting body temperature. A single i.v. injection of polymyxin B (10, 100 or 1000 microg/kg) induced a rapid, marked drop in body temperature in a dose-related manner, with no change in physical activity. However, the highest dose (1000 microg/kg) seemed to impair heat-loss mechanisms and/or functions controlling the animal's day-night cycle [because the day-time body temperature remained elevated for two days after the injection (versus the pre-injection level)]. By contrast, rats given 100 or 10 microg/kg of the drug showed a normal day-night cycle after recovery from the initial hypothermic effect of the drug. Therefore, we used the middle dose of polymyxin B (100 microg/kg) in the subsequent experiments. In these experiments, significant decreases in metabolic rate and heat-loss responses were observed immediately after an i.v. injection of polymyxin B (100 microg/kg). By contrast, intracerebroventricular injection of polymyxin B (3 microg) had no effect on resting body temperature. These results suggest that the observed decrease in metabolic rate is responsible for the polymyxin-B-induced hypothermia. Further, rats may react with a reduction in heat-loss responses so as to prevent the body temperature decreasing too far in response to polymyxin B. Thus, polymyxin B modulates or interferes with the peripheral mechanisms underlying body temperature regulation in rats.