sparstolonin-b and Disease-Models--Animal

It has been previously demonstrated that sparstolonin B (SsnB) inhibits toll‑like receptor (TLR)‑2 and TLR‑4. The present study investigated the effect of SsnB on neuropathic pain (NP). A chronic constriction injury (CCI) model was constructed in rats and the protein expression of TLR‑2 and TLR‑4 was determined by western blot analysis. Rats were divided into the following three groups: Rats with sham surgery (control group); rats with CCI (model group); and rats with CCI and injection of SsnB (SsnB group). The mechanical withdrawal threshold (MWT) was measured by using Von Frey filaments. In addition, the mRNA and protein expression levels of nuclear factor‑κB (NF‑κB) were investigated by reverse transcription‑quantitative polymerase chain reaction and western blot analysis, respectively, and the concentrations of tumor necrosis factor‑α (TNF‑α) and interleukin (IL)‑6 were determined by ELISA. Compared with control rats, the protein expression levels of TLR‑2 and TLR‑4 were increased in model rats (P<0.001). At 7 and 14 days after surgery, the MWTs in the model group were significantly reduced compared with the control group (P<0.001). However, the MWTs in the SsnB group were significantly increased compared with the model group (P<0.001). The results also demonstrated that the mRNA and protein expression levels of NF‑κB, and the protein expression levels of TNF‑α and IL‑6, were increased in model group compared with the control group (P<0.001). Furthermore, these increases in expression were all reduced in the SsnB group compared with the model group. Therefore, the results indicate that SsnB may alleviate NP via suppression of TLR‑2 and TLR‑4, and may be a potential drug for the treatment of NP.

Topics: Animals; Anti-Inflammatory Agents; Cytokines; Disease Models, Animal; Gene Expression; Heterocyclic Compounds, 4 or More Rings; Inflammation Mediators; Male; Neuralgia; NF-kappa B; Rats; RNA, Messenger; Toll-Like Receptor 2; Toll-Like Receptor 4

The present study used a spinal cord injury (SCI) model to evaluate whether sparstolonin B was able to prevent SCI, and to investigate the underlying signaling mechanism. Sparstolonin B attenuated the SCI‑induced Batto, Beattie and Bresnahan score and water content in rats. Sparstolonin B attenuated the mRNA expression of proinflammatory cytokines interleukin (IL)‑18, IL‑6, IL‑1β, and IL‑23, decreased the levels of tumor necrosis factor‑α and interferon‑γ, and decreased caspase‑3 activity and apoptosis regulator Bax protein expression in SCI rats. Similarly, sparstolonin B inhibited monocyte chemoattractant protein‑1 mRNA levels, and Toll‑like receptor (TLR) 4, myeloid differentiation primary response protein MyD88 (MyD88) and nuclear factor (NF)‑κB protein levels in SCI rats. The present results suggested that sparstolonin B may attenuate SCI‑induced inflammation and apoptosis in rats by modulating the TLR4/MyD88/NF‑κB signaling pathway.

Topics: Animals; Caspase 3; Chemokine CCL2; Cytokines; Disease Models, Animal; Enzyme Activation; Heterocyclic Compounds, 4 or More Rings; Inflammation; Inflammation Mediators; Male; Myeloid Differentiation Factor 88; NF-kappa B; Protein Transport; Rats; RNA, Messenger; Spinal Cord Injuries; Toll-Like Receptor 4

Although significant research data exist on the pathophysiology of nonalcoholic steatohepatitis (NASH), finding an efficient treatment regimen for it remains elusive. The present study used sparstolonin B (SsnB), a novel TLR4 antagonist derived from the Chinese herb Sparganium stoloniferum, as a possible drug to mitigate early inflammation in NASH. This study used an early steatohepatitic injury model in high-fat-fed mice with CYP2E1-mediated oxidative stress as a second hit. SsnB was administered for 1 wk along with bromodichloromethane (BDCM), an inducer of CYP2E1-mediated oxidative stress. Results showed that SsnB administration attenuated inflammatory morphology and decreased elevation of the liver enzyme alanine aminotransferase (ALT). Mice administered SsnB also showed decreased mRNA expression of proinflammatory cytokines TNF-α, IFN-γ, IL-1β, and IL-23, while protein levels of both TNF-α and IL-1β were significantly decreased. SsnB significantly decreased Kupffer cell activation as evidenced by reduction in CD68 and monocyte chemoattractant protein-1 (MCP1) mRNA and protein levels with concomitant inhibition of macrophage infiltration in the injured liver. Mechanistically, SsnB decreased TLR4 trafficking to the lipid rafts, a phenomenon described by the colocalization of TLR4 and lipid raft marker flotillin in tissues and immortalized Kupffer cells. Since we have shown previously that NADPH oxidase drives TLR4 trafficking in NASH, we studied the role of SsnB in modulating this pathway. SsnB prevented NADPH oxidase activation in vivo and in vitro as indicated by decreased peroxynitrite formation. In summary, the present study reports a novel use of the TLR4 antagonist SsnB in mitigating inflammation in NASH and in parallel shows a unique molecular mechanism of decreasing nitrative stress.

Topics: Animals; Anti-Inflammatory Agents; Cell Line; Cytochrome P-450 CYP2E1; Cytokines; Disease Models, Animal; Enzyme Activation; Enzyme Induction; Hepatitis; Heterocyclic Compounds, 4 or More Rings; Inflammation Mediators; Kupffer Cells; Liver; Macrophages; Male; Membrane Microdomains; Membrane Proteins; Mice; Mice, Inbred C57BL; MicroRNAs; NADPH Oxidases; Non-alcoholic Fatty Liver Disease; Oxidative Stress; Peroxynitrous Acid; Protein Transport; PTEN Phosphohydrolase; Signal Transduction; Toll-Like Receptor 4

Neuroinflammation plays a key role in intracerebral hemorrhage (ICH)-induced secondary brain injury, but the specific roles of peripheral inflammatory cells such as macrophages and lymphocytes remain unknown. The purpose of this study was to explore the roles of macrophages, T lymphocytes, and the cytokines they secrete as potential targets for treating secondary brain injury after ICH.. Our results showed that peripheral macrophages and T lymphocytes successively infiltrated the brain, with macrophage counts peaking 1 day after ICH and T-lymphocyte counts peaking after 4 days. These peaks in cellular infiltration corresponded to increases in interleukin (IL)-23 and IL-17 expression, respectively. We found that hemoglobin from the hematoma activated IL-23 secretion by infiltrating macrophages by inducing the formation of toll-like receptor (TLR) 2/4 heterodimer. This increased IL-23 expression stimulated γδT-cell production of IL-17, which increased brain edema and neurologic deficits in the model mice as a proinflammatory factor. Finally, we found that sparstolonin B (SsnB) could ameliorate brain edema and neurologic deficits in ICH model mice via inhibition of TLR2/TLR4 heterodimer formation, and notably, SsnB interacted with myeloid differentiation factor 88 Arg196.. Together, our results reveal the importance of the IL-23/IL-17 inflammatory axis in secondary brain injury after ICH and thus provide a new therapeutic target for ICH treatment.

Topics: Animals; Brain Edema; Cerebral Hemorrhage; Disease Models, Animal; Hemoglobins; Heterocyclic Compounds, 4 or More Rings; Interleukin-17; Interleukin-23; Macrophages; Male; Mice; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocytes; Toll-Like Receptor 2; Toll-Like Receptor 4

Sepsis is characterized by an overwhelming systemic inflammation and multiple organ injury. Toll-like receptors (TLRs) 2 and 4 mediate these inflammatory responses. Sparstolonin B (SsnB), isolated from Chinese herb Scirpus yagara, is a new selective TLR2/4 antagonist. Herein, we report that SsnB inhibited the expression of various inflammatory mediators such as tumor necrosis factor (TNF-α), interleukin (IL)-1β, IL-6, and chemokine (C-C motif) ligand 2 (CCL-2) in lipopolysaccharide (LPS)- or Pam3csk4-stimulated macrophages. Moreover, in LPS-stimulated macrophages, the downregulation of peroxisome proliferator-activated receptor γ (PPAR-γ) was reversed by SsnB dose-dependently; and SsnB had synergistic inhibitory effects with rosiglitazone, a PPAR-γ agonist, on TNF-α and IL-6 expression in LPS-stimulated macrophages. The effects of SsnB were further evaluated in a mouse endotoxin shock model. When intraperitoneal injected in mice 2 days before or 1-2h after LPS challenge, SsnB attenuated the body temperature reduction and decreased the mortality. SsnB pre-treatment significantly suppressed LPS-induced increase of TNF-α and IL-6 in serum, lungs and livers, and substantially attenuated lung dysfunction in mice. In vivo toxicity test showed that at doses as high as 500 mg/kg, SsnB did not cause death of mice. These results suggest that SsnB protects mice against endotoxin shock by inhibiting production of multiple cytokines and lung dysfunction. In conclusion, our findings indicate that SsnB may be used in the prevention and treatment of endotoxin shock.

Topics: Animals; Body Temperature; Cell Line; Chemokine CCL2; Disease Models, Animal; Drugs, Chinese Herbal; Female; Heterocyclic Compounds, 4 or More Rings; Inflammation; Interleukin-1beta; Interleukin-6; Lipopeptides; Lipopolysaccharides; Macrophages; Mice; Mice, Inbred BALB C; Mice, Inbred ICR; PPAR gamma; Rosiglitazone; Sepsis; Shock, Septic; Thiazolidinediones; Toll-Like Receptor 2; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha