Complement depletion with cobra venom factor alleviates acute hepatic injury induced by ischemia­reperfusion.

Increasing evidence has demonstrated that complement activation is required for ischemia­reperfusion injury (IRI)­induced hepatic damage, and cobra venom factor (CVF) can deplete the complement components. The aim of the current study was to investigate the effect and intrinsic mechanism of CVF pretreatment on IRI­induced acute hepatic injury in rats. Acute hepatic injury in rats was induced by bone fracture to simulate trauma, followed by hemorrhage for 90 min, and then the rats were resuscitated for a period of 20 min of reperfusion. The survival times under different CVF treatment doses and schedules for rats with IRI were evaluated. Hepatic tissues and serum samples were analyzed for acute hepatic injury, complement activation, inflammatory mediator release and apoptosis at predetermined times and compared between the IRI group and the CVF pretreatment + IRI groups. Compared to the rats with IRI alone, the survival times were significantly improved among rats with IRI receiving a high­dose or low­dose CVF pretreatment (all P<0.01). Upon histological examination, severe hepatic damage was observed in the rats with IRI, accompanied by liver function deterioration, complement and membrane attack complex activation, inflammatory mediator release and hepatic cell apoptosis. CVF pretreatment significantly attenuated the hepatic injury through depletion of anaphylatoxic C5a and membrane attack complex C5b­9 activation, and subsequent inhibition of inflammatory mediator release and hepatic cell apoptosis (all P<0.05). The results indicated that CVF pretreatment ameliorates IRI­induced acute hepatic injury. However, further studies are required to determine whether this therapy could be a potential agent for the treatment of IRI injuries in clinical settings.

[Effect of WS070117M1 on chronic obstructive pulmonary disease in mice and the underling mechanisms of anti-inflammation].

The aim of this study is to investigate the anti-inflammatory effect of the adenosine derivative N6-(3-hydroxylaniline) adenosine (WS070117M1) on cigarette smoke plus LPS (lipopolysaccharide)-induced chronic obstructive pulmonary disease (COPD) in mice and its mechanism. COPD model was established by exposing male BALB/c mice to cigarette smoke and challenged with LPS inhalation. Supernatants of bronchoalveolar lavage fluid (BALF) were harvested and IL-1ß, IL-6, IL-8 and TGF-ß1 levels were measured by ELISA (enzyme-linked immunesorbent assay). The number of total white blood cells and neutrophils in bronchoalveolar lavage fluid was counted separately. Lung tissue was stained with Mayer 's hematoxylin and eosin for histopathologic examination. pAMPKa protein expression and distribution of lung tissue were analyzed by immunohistochemistry method. In vitro, levels of AMPKα phosphorylation in phorbol-12- myristate-13-acetate (PMA) differentiated THP-1 cells was detected by immunohistochemistry, IL-8 level in supernatants of cigarette smoke condensate stimulating PMA differentiated THP-1 cells was measured by ELISA. The results showed that WS070117M1 treatment significantly activated AMPKa in the lung tissue. It also resulted in down regulation of IL-1ß, IL-6, IL-8 and TGF-ß1 levels in bronchoalveolar lavage fluid and IL-8 level in cigarette smoke condensate stimulating PMA differentiated THP-1 cells. In addition, WS070117M1 could inhibit the recruitment of total white blood cells and neutrophils. These results suggest that WS070117M1 may alleviate the airway inflammation by activating AMPK in the lung tissue.

[Advance in the research on P2X7 and inflammatory respiratory diseases].

P2X7 is the most important subtype of the ATP receptors known so far. Recent investigations showed that the downstream signaling pathway of P2X7 is coupled with several key inflammatory molecules including IL-1beta and IL-18, this suggests P2X7 might have roles in the inflammatory diseases. Moreover, attenuation of P2X7 by selective antagonists in vitro and knockout mice in vivo reducing the inflammatory response indicated that P2X7 is a potential therapeutic target for inflammatory diseases. However, most previous studies on P2X7 were focused on nerve system diseases most, while its effects in inflammatory respiratory diseases, especially in asthma, chronic obstructive pulmonary disease (COPD) and lung cancer have been poorly investigated. In this paper, we reviewed the research progress on the structure, distribution, biological activities of P2X7 and its relationship with inflammatory respiratory diseases including asthma, COPD and lung cancer, along with the development of P2X7 antagonist as therapeutics.

[Effects of modified liangge powder contained serum on LPS stimulated TLR4 expression and release of cytokines in mouse platelets].

OBJECTIVE: To observe the effects of Modified Liangge Powder (MLP) on the expressions of platelet toll like receptor 4 (TLR4) and the release of platelet-derived cytokines interleukin 8 (IL-8), beta platelet globulin (beta-TG), soluble CD40 ligand (sCD40L). METHODS: The modulating effects on the release of cytokines from mice platelets by TLR4 ligand through monoclonal antibody blocking TLR4 on platelet were compared. The stimulated platelet by LPS was incubated with low (0.94 g/mL), medium (1.89 g/mL), and high (2.84 g/mL) dose of MLP contained serum. The changes of the platelet TLR4 expression and platelet-derived cytokines were observed. RESULTS: The positive expression rate of platelet TLR4 obviously decreased (P < 0.01) and the release of sCD40L and beta-TG from platelets significantly increased (P < 0.01) after stimulated by LPS. However, the release of sCD40L and beta-TG from platelets obviously decreased by TLR4 monoclonal antibody (P < 0.05, P < 0.01). There was no statistical difference in IL-8 between before and after LPS stimulation (P > 0.05). Platelet TLR4 positive expression rate was significantly higher after incubated by medium and high doses of MLP contained serum (P < 0.01), and the releasing of sCD40L and beta-TG was lower in the serum contained groups. The inhibitory effects were enhanced in a dose-dependent manner. CONCLUSIONS: LPS induced platelet activation by TLR4 and released sCD40L and beta-TG, while the release of platelet IL-8 was not dependent on platelet TLR4-LPS pathway. MLP could inhibit LPS-stimulated sCD40L and beta-TG, inhibit the binding of platelet TLR4 and LPS in a dose-dependent manner, thus reducing the release of platelet cytokines.

[Platelet parameters and platelet Toll-like receptor 4 (TLR4) expression in patients with sepsis, and the effect of a joint treatment-plan integrating traditional Chinese and western medicine: a clinical study].

OBJECTIVE: To examine the efficacy of a treatment regimen combining modern (Western) medication and a traditional Chinese medicinal recipe Modified Liang-Ge San on sepsis, and its effect on platelet parameters/activation, platelet TLR4 expression and the intensity of inflammatory response in the patients. METHODS: 64 patients with sepsis were randomly assigned to two groups (32 each) to receive Western therapy only (group X) and Western therapy + Modified Liang-Ge San (group L). The values of: platelet count (PLT), mean platelet volume (MPV), platelet distribution width (PDW), the expression of TLR4 and procaspase activating compound-1 (PAC-1) in platelets, plasma concentration of soluble CD40 ligand (sCD40L) and tumor necrosis factor-α (TNF-α) were collected on the day of admission, and 3, 5, 9 days after treatment. Acute physiology and chronic health evaluation II (APACHE II) score, length of stay in intensive care unit (ICU), bleeding events and 28 day mortality in these patients were also analyzed. A group of 15 healthy volunteers (group C) were used as control. RESULTS: Compared to group C, the patients with sepsis have significantly: lower PLT (×10 9/L: 211.37±77.84 vs. 272.33±34.23, P< 0.01 ), increased MPV (fL: 10.24±0.81 vs. 9.64±0.66, P< 0.05) and PDW (fL: 17.79±1.68 vs. 15.61±1.54, P< 0.01), up-regulated platelet TLR4 [(39.93±9.07)% vs. (23.50±4.68)%] and PAC-1 expression [(42.21±8.74)% vs. (21.02±3.49)%], both P < 0.01, and higher level of sCD40L (µg/L: 6.94±1.05 vs. 3.27±0.41)and TNF-α(ng/L: 60.10±9.77 vs. 4.08±3.08), both P< 0.01. Compared to group X, group L had significantly (P< 0.05 or P< 0.01, respectively) lower value in: creatinine (µmol/L: 106.2±34.4 vs. 127.5±43.7); alanine aminotransferase (U/L: 31.7±12.5 vs. 41.9±19.9);aspartate aminotransferase (U/L: 54.1±21.6 vs. 68.5±24.1); TLR4 [(27.14±6.08)% vs. (30.92±5.47)%]; PAC-1 [(27.52±6.51)% vs. (31.24±5.77)%]; sCD40L (3.86±0.69 vs. 4.38±0.73); TNF-α (22.06±7.19 vs. 28.25±8.99), and higher PLT (261.93±55.32 vs. 231.37±63.58, P< 0.05), in the 9 days after treatment. In patients with sepsis, platelet PAC-1 expression correlated significantly to PLT ( r = - 0.409, P< 0.01 ) negatively, and MPV, PDW, platelet TLR4 expression, plasma sCD40L ( r (1) = 0.262, r (2) = 0.318, r (3) = 0.341, r(3) = 0.519, all P< 0.01) positively; sCD40L and TNF-α was positively correlated ( r = 0.542, P < 0.01 ) in these patients. In comparison with group X, the length of stay in ICU (day: 8.06±2.86 vs. 9.31±2.48), the incidence of bleeding (12.5% vs. 21.9%) and APACHE II score (12.75± 4.56 vs. 14.59± 3.97) were significantly lower (all P< 0.05) in group L on the 9 days after treatment. No significant difference was found in 28 day mortality between group L and X (15.63% vs. 18.75%, P> 0.05). CONCLUSION: In patients with sepsis, platelet TLR4 expression is elevated together with platelet activation. The joint application of Western medicine and Modified Liang-Ge San may suppress such up-regulation in TLR4/other inflammatory mediators, and alleviate platelet activation/thrombocytopenia in these patients.

[The effect of the membrane attack complex C5b-9 on liver cells during traumatic hemorrhagic shock in rat].

OBJECTIVE: To observe whether the membrane attack complex C5b-9 would accumulate in the rats' liver after receiving the assault of traumatic hemorrhagic shock, and whether the membrane attack complex deals an impact on liver apoptosis. METHODS: Fifty male healthy Wistar rats were randomly divided into five groups: normal group, 1, 3, 6, 24 hour model groups. The model of traumatic hemorrhagic shock was reproduced by withdrawal of blood from carotid artery after a bone fracture till the blood pressure lowered to 40 mm Hg (1 mm Hg=0.133 kPa). Plasma membrane attack complex C5b-9 concentration was assayed using enzyme linked immunoadsorbent assay. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in blood was determined by Rate method. Immunohistochemistry was used to detect C5b-9 deposition in the liver. Apoptosis of liver cells was then detected by the terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) assay. The pathological changes in paraffin sections stained with hematoxylin eosin (HE) were observed under light microscope. RESULTS: A small amount of C5b 9 in plasma was found in normal group, and the values (ng/L) of 1, 3, 6 hour models were significantly higher than those of the normal group (272.91 ± 9.56, 192.01 ± 9.04, 156.78 ± 8.37 vs. 25.98 ± 5.87, all <0.05 ). ALT (U/L) in 3 hour model group and AST (U/L) in 1 hour model group were increased significantly (92.90 ± 8.83, 264.83 ± 31.4), peaked at 24 hours (184.30 ± 12.98, 647.36 ± 60.02), and there was significant difference compared with normal group (38.75 ± 5.40, 66.69 ± 19.95, all P <0.05). In the normal group and the 1 hour and 6 hour model groups, no C5b 9 was found in liver, but in the 3 hour model group a large number of liver parenchymal cells in the portal area were found to contain C5b 9 22.60 ± 1.06), however the number decreased significantly in the 24 hour model (2.20 ± 0.60, P<0.05). In normal group there was no apoptotic cell, and in 1, 6, 24 hour model groups there were scattered apoptotic cells (1.20 ± 0.25, 5.60 ± 0.37, 1.60 ± 0.26). In the 3 hour model group apoptosis of hepatic cells around the central vein was increased to the peak (20.60 ± 0.47), and there was significant difference compared with other groups (all P <0.05) . In the model groups the liver cells became edematous, and the integrity of the membrane was lost, and some cells were even lysed.The pathological damage is most serious in 24 hour model group. CONCLUSION: The membrane attack complex C5b-9 insulted the rats' liver after a traumatic hemorrhagic shock, and apoptosis of hepatic cells and the content of C5b-9 peaked in 3 hour model , though they do not occur in the same site. A low level of C5b-9 in blood 3 hours after shock predict a poor prognosis.

Phosphine- and nitrogen-containing Lewis base catalyzed highly regioselective and geometric selective cyclization of isatin derived electron-deficient alkenes with ethyl 2,3-butadienoate.

An interesting phosphine-containing Lewis base catalyzed highly regioselective [3 + 2] cycloaddition and a novel nitrogen-containing Lewis base catalyzed highly geometric selective [4 + 2] cycloaddition of isatin derived α,ß-unsaturated diesters with α-allenic ester have been disclosed to give the corresponding cyclic products in good to excellent yields under mild conditions. A plausible reaction mechanism has also been proposed on the basis of previous literature and our own investigation.

Role of platelet TLR4 expression in pathogensis of septic thrombocytopenia.

BACKGROUND: Infection-induced thrombocytopenia (TCP) is an independent risk factor for death of patients with sepsis, but its mechanism is unknown. This study aimed to explore the underlying mechanism of TCP based on the relationship between TLR4 expression and platelet activation in septic patients. METHODS: A total of 64 patients with sepsis were prospectively studied. Platelet count (PC), mean platelet volume (MPV), platelet distribution width (PDW), platelet TLR4 expression, platelet PAC-1 expression, sCD40L and TNF-α concentrations were compared between the healthy control group (15 volunteers) and sepsis group (64 patients) at admission and on the 3, 5, and 9 days after admission. The changes of MPV and PDW in the TCP and non-TCP subgroups of sepsis before and after treatment were recorded. Prognostic index was analyzed. RESULTS: PC was lower in the sepsis group (P=0.006), and MPV and PDW were higher in the sepsis group than those in the healthy control group (P=0.046, P=0.001). Platelet TLR4 and PAC-1 expressions, and sCD40L and TNF-α levels increased more significantly in the sepsis group (P<0.001). PAC-1 expression and TNF-α level were higher in the TCP group than in the non-TCP group before and after treatment (P=0.023, P=0.011). sCD40L concentration and platelet TLR4 expression were significantly higher in the treated TCP group than in the non-TCP group (P=0.047, P=0.001). Compared to the non-TCP group, the rate of bleeding was higher (P=0.024) and the length of ICU stay was longer (P=0.013). The APACHE II score and the 28-day mortality were higher in the TCP group (P<0.01, P=0.048). CONCLUSIONS: The elevation of platelet TLR4 expression in sepsis along with platelet activation is closely related to the incidence of thrombocytopenia. The occurrence of TCP is a sign of poor prognosis in sepsis patients.

Phosphine-catalyzed highly diastereoselective [3+2] cyclization of isatin derived electron-deficient alkenes with α-allenic esters.

A novel phosphine-catalyzed highly diastereoselective [3+2] cycloaddition of isatin derived α,ß-unsaturated ketones with α-allenic ester has been developed.

[The influences of propofol on corticosteroid and immunity of rats after hemorrhagic shock and resuscitation].

OBJECTIVE: To observe the influence of propofol on corticosteroid, and cytokines in rats after hemorrhagic shock and resuscitation, as well as its protective effects on vital organs. METHODS: Sixty-six male Wistar rats were randomly divided into three groups: control group, hemorrhagic shock and resuscitation group and propofol treatment group. After the model of hemorrhagic shock and resuscitation was reproduced, propofol was infused in propofol treatment group at the speed of 10 mg.kg(-1).h(-1) via femoral vein, while equal amount of normal saline was infused in hemorrhagic shock and resuscitation group. Abdominal aortic blood, lung and small intestinal tissue samples were collected at 30, 60, 120, 180 and 240 minutes, 6 rats at each time point in hemorrhagic shock and resuscitation group and propofol treatment group. Plasma cortisol, interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) concentrations were measured using enzyme linked immunosorbent assay (ELISA). Histopathological changes in lung and small intestine were observed using optical microscope and then histopathological scores were assessed. RESULTS: Comparing with control group, the concentration of cortisol in plasma decreased significantly (all P<0.01), both IFN-gamma and IL-4 increased progressively, and the IFN-gamma/IL-4 ratio decreased gradually with the progress of the course (all P<0.01). Serious injury to lung and small intestinal tissue was observed, and remarkable elevation of histopathological scores was found in hemorrhagic shock and resuscitation group (lung: 3.09+/-0.56 vs. 0.31+/-0.25, small intestine: 7.61+/-1.20 vs. 0.86+/-0.72, both P<0.01). In propofol treatment group, the extent of decrease in corticosteroid level was lessened, the extent of elevation of IFN-gamma and IL-4 was decreased, the downward trend of the IFN-gamma/IL-4 ratio was slowed down (P<0.05 or P<0.01), the extent of injury to lung and small intestinal tissue was alleviated and histopathological scores were reduced remarkably (lung: 1.27+/-0.40, small intestine: 3.69+/-1.28, both P<0.01). CONCLUSION: Propofol can ameliorate the lowering of corticosteroid secretion, moderate inflammatory response, and protect vital organs in rats with hemorrhagic shock.

[Evaluation of nuclear factor-KappaB and I Kappa B mRNA expression in determining the prognosis of multiple organ dysfunction syndrome].

OBJECTIVE: To observe the value of nuclear factor-KappaB(NF-KappaB) and I Kappa B mRNA expression estimation on determining the prognosis of patients with multiple organ dysfunction syndrome (MODS). METHODS: Forty-three MODS patients were divided into two groups based on their prognosis: the survivor group (n =28) and the non-survivor group (n=15). Another group of 10 healthy persons served as normal control group. The expression of NF-Kappa B and I Kappa B-alpha mRNA levels in monocytes/neutrophils of patients and controls were detected by reverse transcription-polymerase reaction (RT-PCR), and the results were compared among groups. RESULTS: The expression of NF-KappaB mRNA levels of MODS patients was higher than that of normal control group(1. 35+/-0.53 vs. 0.74+/-0.25, P<0.01),and the expression of I Kappa B-alpha mRNA levels were lower than those of the control group (1. 24+/-0.60 vs. 1. 97+/-0.71,P<0.01). There was no significant difference in NF-KappaB mRNA levels between the survivor group and the non-survivor group (1. 27+/-0.37 vs. 1.39+/-0.60,P>0.05), but the expression of I Kappa B-alpha mRNA levels in the non-survivor group was significantly lower than that of survivors(0.94+/-0.46 vs. 1. 40+/-0.61, P<0.05). The results suggested that there was a negative correlation between the expression of I Kappa B-alpha mRNA level and acute physiology and chronic health evaluation II (APACHE II)(r=-0.340, P<0.05). Moreover, when the cutoff value of the expression of I Kappa B-alpha mRNA level was 1. 34, the Youden index was 0.51, and it was the highest in all the cutoff values, and the specificity was 90.72%,the sensitivity was 60.75%. CONCLUSION: The expression of I Kappa B-alpha mRNA level can be a useful guide for determining the prognosis of patients with MODS.

[Clinical observation on treatment of multiple organ dysfunction syndrome with auxiliary treatment of Shennong No.33].

OBJECTIVE: To observe the therapeutic effect of Shennong No. 33 (SN33) in treating multiple organ dysfunction syndrome (MODS) by APACHE II and APACHE II scoring. METHODS: One hundred and twenty-nine patients of MODS were randomly divided into the treated group (n = 72) and the control group (n = 57), they were treated with comprehensive therapy and to the treated group, SN33 was given additionally. The changes of APACHE II and APACHE III scores and blood endotoxin level were observed at the time points of within 24 hrs after hospitalization, and the 3rd, 5th and 7th day. RESULTS: In the treated group, 50 patients survived and 22 died, while in the control group, 25 survived and 32 died. The APACHE II and APACHE III scores of the survivors were higher than those of the decedent (P<0.05), which in the treated group was lower than those in the control group (P<0.05). The blood level of endotoxin in the treated group was also lower than that in the control group (P<0.05). CONCLUSION: SN33 in treating MODS could improve patients' condition, lead to the lowering of APACHE scores. APACHE scoring system could be applied as the criteria for evaluating the condition and prognosis of critical patients, and the APACHE III scoring is more accurate.