phenanthrenes and Sepsis

Sepsis is a main reason for death in intensive care units, inflammation is closely related to sepsis. Anti-inflammation plays an important role in treating of sepsis. ZT01 is a triptolide derivative with strong anti-inflammatory activity and low toxicity. The purpose of this study is to evaluate the anti-inflammatory activity of ZT01 under the sepsis condition and explore the underlying molecular mechanisms. Two in vivo model of sepsis, caecal ligation and puncture or intraperitoneal injection of LPS in C57BL/6, were used to evaluate the therapeutic effects of ZT01. In vitro, the anti-inflammatory properties of ZT01 were assessed in IFN-γ or LPS-induced macrophages by ELISA, RT-PCR, western blotting and co-immunoprecipitation. Macrophages were used to investigate the polarization phenotype by flow cytometry. The results showed, ZT01 significantly attenuated inflammatory response of sepsis in serum or lung tissue by inhibiting production of pro-inflammatory factors and improved the survival rate of septic mice in vivo. In cultured macrophages, ZT01 not only decreased the levels of TNF-α and IL-6 but also prevented the TKA1-TAB1 complex formation, thereby inhibiting the phosphorylation expression of MKK4 and JNK, which were all stimulated by LPS. Moreover, ZT01 inhibited the LPS-induced polarization of macrophages into pro-inflammatory phenotype. Adoptive transfer ZT01 pretreated bone marrow-derived macrophages obviously reduced the pro-inflammatory factors in mice after LPS challenge. Our findings suggested that ZT01 exhibited anti-inflammation activity via preventing the pro-inflammatory phenotype of macrophages by blocking the formation of the TAK1-TAB1 complex and subsequently phosphorylation of MKK4 and JNK.

Topics: Animals; Anti-Inflammatory Agents; Cells, Cultured; Cytokines; Diterpenes; Epoxy Compounds; Inflammation Mediators; Lipopolysaccharides; Macrophage Activation; Macrophages; Male; MAP Kinase Kinase Kinases; Mice; Models, Biological; Models, Molecular; Phagocytosis; Phenanthrenes; Phosphorylation; Protein Binding; Protein Kinase Inhibitors; RAW 264.7 Cells; Sepsis; Signal Transduction; Structure-Activity Relationship

Dihydrotanshinone (DHT), one of the major ingredients of Salvia miltiorrhiza Bunge (Danshen), displays many bioactivities. However, the activity and underlying mechanism of DHT in anti-inflammation have not yet been elucidated. In this study, we investigated the anti-inflammatory activity and molecular mechanism of action of DHT both in vitro and in vivo. Our data showed that DHT significantly decreased the release of inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-1β in lipopolysaccharide (LPS)-stimulated RAW264.7 cells, THP-1 cells, and bone marrow-derived macrophages (BMDMs), and altered the expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS). In addition, flow cytometry results indicated that DHT reduced the calcium influx, and generation of reactive oxygen species (ROS), and nitric oxide (NO) generation in LPS-stimulated RAW264.7 cells. Moreover, DHT suppressed the transcription of nuclear factor-κB (NF-κB), the expressions of NF-κB proteins, and nuclear translocation of NF-κB/p65, thereby suggesting that the NF-κB pathway played a role in the anti-inflammatory action of DHT. In addition, DHT attenuated LPS-challenged activator protein-1 (AP-1) activity, resulting from interference of the mitogen-activated protein kinase (MAPK) pathway. The molecular docking simulation of DHT to toll-like receptor 4 (TLR4) suggested that DHT binds to the active sites of TLR4 to block TLR4 dimerization, which was further corroborated by cellular thermal shift assay and co-immunoprecipitation (Co-IP) experiments. Furthermore, the recruitment of myeloid differentiation primary response gene 88 (MyD88) and the expression of transforming growth factor-b (TGF-b)-activated kinase 1 (p-TAK1) were disturbed by the inhibition of TLR4 dimerization. Thus, investigating the molecular mechanism of DHT indicated that TLR4-MyD88-NF-κB/MAPK signaling cascades were involved in the anti-inflammatory activity of DHT in vitro. In in vivo mouse models, DHT significantly ameliorated LPS-challenged acute kidney injury, inhibited dimethylbenzene-induced mouse ear oedema, and rescued LPS-induced sepsis in mice. Taken together, our results indicated that DHT exhibited significant anti-inflammatory activity both in vitro and in vivo, suggesting that DHT may be a potential therapeutic agent for inflammatory diseases.

Topics: Acute Kidney Injury; Animals; Anti-Inflammatory Agents; Cytokines; Dimerization; Edema; Furans; HEK293 Cells; Humans; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; NF-kappa B; Phenanthrenes; Quinones; RAW 264.7 Cells; Sepsis; THP-1 Cells; Toll-Like Receptor 4; Xylenes

Dihydrotanshinone, tanshinone I, cryptotanshinone, and tanshinone IIA are major lipid-soluble constituents isolated from Salvia miltiorrhiza Bunge (Danshen). In the present study, a systematic method was developed to simultaneously isolate and purify those compounds using macroporous adsorption resins and semi-preparative HPLC with a dynamic axial compress (DAC) system. The Danshen extract (95% alcohol) was divided into three fractions using different concentrations of alcohol (0%, 45%, and 90%) on D101 column. The content of total tanshinones of 90% alcohol eluent (TTS) was over 97%. Furthermore, the anti-inflammatory effects of those samples were investigated on LPS-stimulated RAW264.7 cells and three animal models. The results showed that the anti-inflammatory effect of TTS in vitro was superior to the one of any other sample including 0% and 45% eluent, and total tanshinones capsules. In addition, TTS exhibited a stronger anti-inflammatory effect than that of dihydrotanshinone, tanshinone IIA, cryptotanshinone, and tanshinone I, respectively. For animal models, TTS could significantly suppress xylene-induced ear oedema and rescue LPS-induced septic death and acute kidney injury in mice. In summary, the separation process developed in the study was high-efficiency, economic, and low-contamination, which was fit to industrial producing. TTS is a potential agent for the treatment of inflammatory diseases.

Topics: Abietanes; Acute Kidney Injury; Animals; Anti-Inflammatory Agents; Chromatography, High Pressure Liquid; Cytokines; Disease Models, Animal; Kidney; Lipopolysaccharides; Macrophages; Mice; Mice, Inbred BALB C; Nitric Oxide; Phenanthrenes; RAW 264.7 Cells; Salvia miltiorrhiza; Sepsis

Cardiac dysfunction, a common consequence of sepsis, is the major contribution to morbidity and mortality in patients. Sodium tanshinone IIA sulfonate (STS) is a water-soluble derivative of Tanshinone IIA (TA), a main active component of Salvia miltiorrhiza Bunge, which has been widely used in China for the treatment of cardiovascular and cerebral system diseases. In the present study, the effect of STS on sepsis-induced cardiac dysfunction was investigated and its effect on survival rate of rats with sepsis was also evaluated. STS treatment could significantly decrease the serum levels of C-reactive protein (CRP), procalcitonin (PCT), cardiac troponin I (cTn-I), cardiac troponin T (cTn-T), and brain natriuretic peptide (BNP) in cecal ligation and puncture (CLP)-induced) septic rats and improve left ventricular function, particularly at 48 and 72 h after CLP. As the pathogenesis of septic myocardial dysfunction is attributable to dysregulated systemic inflammatory responses, several key cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-10 (IL-10) and high mobility group protein B1 (HMGB1), were detected to reveal the possible mechanism of attenuation of septic myocardial dysfunction after being treated by STS. Our study showed that STS, especially at a high dose (15 mg·kg

Topics: Animals; C-Reactive Protein; Cecum; Drugs, Chinese Herbal; Female; Heart; Humans; Interleukin-6; Ligation; Male; Myocardium; Phenanthrenes; Punctures; Rats; Salvia miltiorrhiza; Sepsis; Troponin T; Tumor Necrosis Factor-alpha

To examine whether sodium tanshinone II A sulfonate (STS), the main effective component of Salvia miltiorrhiza is effective in relieving the microcirculatory disturbance of small intestine by suppressing the production of reactive oxygen species (ROS) in rats with sepsis.. A rat model of sepsis was induced by cecal ligation and puncture (CLP). Rats (n =40) were randomly divided into 4 groups: sham-operated group (sham, n =10), sepsis group (CLP, n =10), STS treatment group (STS, n =10) and ROS scavenger dimethylthiourea (DMTU, n =10) group. Animals in the STS group were injected with STS (1 mg/kg) for 10 min through the right external jugular vein after the CLP operation, and animals in the CLP group were given the same volume of normal saline after the CLP operation. Animals in the DMTU group were intraperitoneally injected with 5 mL/kg of 20% DMTU 1 h before CLP. The histopathologic changes in the intestinal tissues and changes of mesenteric microcirculation were observed. The levels of ROS in intestinal tissues from each group were qualitatively evaluated using a fluorescent microscope. The expressions of apoptosis signal-regulating kinase (ASK1), phosphorylated ASK1 (phospho-ASK1), p38 mitogen-activated protein kinases (p38 MAPK), phosphorylated p38 MAPK (phospho-p38 MAPK) and tissue factor (TF) were determined by Western blotting.. It was shown that there were obvious microcirculatory disturbance (P <0.05) and tissue injuries in intestinal tissues after CLP operation. The levels of ROS production, phospho-ASK1, phospho-p38 MAPK and TF were increased. Both STS and DMTU suppressed ROS, phospho-ASK1, phospho-p38 MAPK and TF production, and ameliorated the microcirculatory disturbance and tissues injury (P <0.01).. STS can ameliorate the microcirculatory disturbance of the small intestine by attenuating the production of ROS in rats with sepsis.

Topics: Animals; Intestine, Small; Male; MAP Kinase Kinase Kinase 5; Microcirculation; p38 Mitogen-Activated Protein Kinases; Phenanthrenes; Phosphorylation; Rats, Wistar; Reactive Oxygen Species; Sepsis; Thromboplastin

To explore the protective effect of sodium tanshinone IIA sulfonate (STS) on small: intestine injury in rats with sepsis and its possible mechanism.. According to a random number table, 24 Tats were randomly divided into 3 groups: sham operation group (sham group), sepsis model group (model group) and STS treatment group (STS group), with 8 Tats in each group. A rat model of sepsis was induced by cecal ligation and puncture (CLP) for 5 h. STS (1 mg/kg) was slowly injected through the right external jugular vein after CLP. The histopathologic changes in the intestine tissue were observed under a light microscope, and the intestinal epithelial cell apoptosis was evaluated by terminal deoxynucleoddyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) method. The expressions of Bcl-2, Bax and nuclear factor κB (NF-κB) p65 in the intestinal tissue was determined by Western blot. The levels of tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) in the intestinal tissue were determined using enzyme-linked immuno-sorbent assay (ELISA).. Obvious injuries were observed in the intestinal tissue in the CLP group compared with the sham group. The expression of NF-κB p65 and the levels of TNF-α and IL-6 were up-regulated after CLP, the apoptosis of intestinal epithelial cells was increased after CLP, and the ratio of Bcl-2 to Bax was decreased. STS post-treatment could attenuate the injury on the intestinal tissue induced by CLP, decrease the apoptosis of intestinal treatment epithelial cells and the levels of NF-κB p65, TNF-α and IL-6, and increase the ratio of Bcl-2 to Bax.. STS can protect the small intestine in rats with sepsis, and the mechanism may be associated with the inhibition of intestinal epithelial apoptosis and the reduction of activation of inflammatory cytokines.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Interleukin-6; Intestine, Small; Male; Phenanthrenes; Protective Agents; Rats; Rats, Wistar; Sepsis; Transcription Factor RelA; Tumor Necrosis Factor-alpha

To explore the protective effect of sodium tanshinone IIA sulfonate (STS) on microcirculatory disturbance of small intestine in rats with sepsis, and the possible mechanism, a rat model of sepsis was induced by cecal ligation and puncture (CLP). Rats were randomly divided into 3 groups: sham operated group (S), sepsis group (CLP) and STS treatment group (STS). STS (1 mg/kg) was slowly injected through the right external jugular vein after CLP. The histopathologic changes in the intestinal tissue and changes of mesenteric microcirculation were observed. The levels of tumor necrosis factor-α (TNF-α) in the intestinal tissue were determined by using enzyme-linked immunoabsorbent assay (ELISA). The expression of intercellular adhesion molecule-1 (ICAM-1) in the intestinal tissue was detected by using immunohistochemisty and Western blot, that of nuclear factor κB (NF-κB) and tissue factor (TF) by using Western blot, and the levels of NF-κB mRNA expression by using RT-PCR respectively. The microcirculatory disturbance of the intestine was aggravated after CLP. The injury of the intestinal tissues was obviously aggravated in CLP group as compared with S group. The expression levels of NF-κB p65, ICAM-1, TF and TNF-α were upregulaed after CLP (P<0.01). STS post-treatment could ameliorate the microcirculatory disturbance, attenuate the injury of the intestinal tissues induced by CLP, and decrease the levels of NF-κB, ICAM-1, TF and TNF-α (P<0.01). It is suggested that STS can ameliorate the microcirculatory disturbance of the small intestine in rats with sepsis, and the mechanism may be associated with the inhibition of inflammatory responses and amelioration of coagulation abnormality.

Topics: Animals; Intestine, Small; Male; Microcirculation; Phenanthrenes; Rats; Rats, Wistar; Sepsis

Pseudomonas aeruginosa is commonly associated with nosocomial pneumonia. Ileal mucosal injury may be induced by severe lung infection. During septic shock, peroxynitrite-mediated DNA strand-breaks activate the enzyme poly-(ADP)-ribose polymerase (PARP) resulting in cellular energetic suppression and cell dysfunction. The aim of this study was to determine whether gut injury could be demonstrated in sepsis induced by P. aeruginosa and the effects of a PARP inhibitor (PJ34) on the associated gut injury.. After baseline measurements, 20 rabbits were randomized into three groups: Sham (n = 5): transtracheally inoculated (TI) with 2 ml of phosphate buffer solution (PBS); P. aeruginosa + saline (n = 8), TI with 4 x 10(12) CFU/ml of P. aeruginosa in 2 ml/kg of PBS + i.v. saline; and P. aeruginosa + PJ34 (n = 7), TI with 4 x 10(12) CFU/ml of P. aeruginosa and i.v. treatment with PJ34.. P. aeruginosa caused a hyperdynamic response with increased blood flow also in the superior mesenteric artery. No significant differences were found in luminal gut lactate concentrations or PCO(2)-gap between groups. Histological specimens showed moderate or diffuse alveolar infiltrate in the P. aeruginosa + saline group (6/8) and in the P. aeruginosa + PJ34 group (6/7). Gut wet-to-dry weight ratio was significantly higher in the P. aeruginosa + saline group than in Shams (7.5 +/- 0.8 versus 6.4 +/- 0.7, P < 0.05) and significantly lower in the P. aeruginosa + PJ34 group (6.1 + 0.5, P < 0.05 versus the other groups). Blood cultures were positive in 1/5 (Sham), 8/8 (P. aeruginosa + saline group) and 4/7 (P. aeruginosa + PJ34 group) (RR 0.57 CI 95% 0.30-1.08).. Pharmacological inhibition of PARP reduces gut inflammation and may limit bacterial translocation.

Topics: Animals; Aorta; Blood Pressure; Enteritis; Enzyme Inhibitors; Intestines; Mesenteric Artery, Superior; Phenanthrenes; Pneumonia, Bacterial; Poly(ADP-ribose) Polymerase Inhibitors; Pseudomonas aeruginosa; Pseudomonas Infections; Rabbits; Regional Blood Flow; Sepsis; Specific Pathogen-Free Organisms; Trachea

The lack of efficacy of anti-inflammatory drugs, anti-coagulants, anti-oxidants, etc. in critically ill patients has shifted interest towards developing alternative treatments. Since inhibitors of the nuclear enzyme poly-(ADP-ribose) polymerase (PARP) were found to be beneficial in many pathophysiological conditions associated with oxidative stress and PARP-1 knock-out mice proved to be resistant to bacterial lipopolysaccharide (LPS)-induced septic shock, PARP inhibitors are candidates for such a role. In this study, the mechanism of the protective effect of a potent PARP-1 inhibitor, PJ34 was studied in LPS-induced (20mg/kg, i.p.) septic shock in mice. We demonstrated a significant inflammatory response by magnetic resonance imaging in the dorsal subcutaneous region, in the abdominal regions around the kidneys and in the inter-intestinal cavities. We have found necrotic and apoptotic histological changes as well as obstructed blood vessels in the liver and small intestine. Additionally, we have detected elevated tumor necrosis factor-alpha levels in the serum and nuclear factor kappa B activation in liver of LPS-treated mice. Pre-treating the animals with PJ34 (10mg/kg, i.p.), before the LPS challenge, besides rescuing the animals from LPS-induced death, attenuated all these changes presumably by activating the phosphatidylinositol 3-kinase-Akt/protein kinase B cytoprotective pathway.

Topics: Animals; Base Sequence; Binding Sites; Consensus Sequence; Endotoxins; Enzyme Inhibitors; Escherichia coli; Inflammation; Lipopolysaccharides; Mice; Mice, Inbred BALB C; NF-kappa B; Phenanthrenes; Poly(ADP-ribose) Polymerase Inhibitors; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Sepsis; Shock, Septic