mRNA Expression Levels Of iNOS Research Articles

Artemisia iwayomogi (Dowijigi) inhibits lipopolysaccharide-induced inflammation in RAW264.7 macrophages by suppressing the NF-κB signaling pathway.

Inflammatory diseases are an important health concern and have a growing incidence worldwide. Thus, developing novel and safe drugs to treat these disorders remains an important pursuit. Artemisia iwayomogi, locally known as Dowijigi (DJ), is a perennial herb found primarily in Korea and is used to treat various diseases such as hepatitis, inflammation and immune disorders. In the present study, the anti-inflammatory effects of a polyphenolic extract from the DJ flower (PDJ) in lipopolysaccharide (LPS)-stimulated mouse macrophage RAW264.7 cells were investigated. Cell cytotoxicity was assessed using the MTT assay. The production of nitric oxide (NO) and prostaglandin E2 (PGE2) was measured by Griess and ELISA analysis, respectively. The expression levels of inducible nitric oxide (iNOS) and cyclooxygenase-2 (COX2) were examined by western blot analysis. Reverse transcription-quantitative PCR was performed to detect the mRNA expression levels of pro-inflammatory cytokines, including tumor necrosis factor α (TNFα), interleukin (IL)-6 and IL-1β, as well as COX2 and iNOS. The production of NO and PGE2 was significantly decreased following treatment with PDJ. The mRNA expression levels of TNFα, IL-6, IL-1β, COX2 and iNOS were significantly decreased in LPS-induced PDJ co-treated cells compared with the group treated with LPS alone. Western blot analysis indicated that PDJ downregulated the LPS-induced expression of iNOS and COX2, as well as the expression of NF-κB proteins. In conclusion, the present study demonstrated that PDJ exerted anti-inflammatory effects in LPS-induced macrophage cells by suppressing the NF-κB signaling pathway. Therefore, PDJ may be used as a potential therapeutic agent in inflammation.

Electroacupuncture relieved neuropathic pain by suppressing L-Arg/NO/cGMP pathway in the lumbar spinal cord in rats with spared nerve injury

To observe the effect of electroacupuncture (EA) on changes of expression of L-Arg transporter 2 (CAT-2) mRNA and nitric oxide synthase (iNOS) mRNA and protein and contents of NO and cGMP of L4-L6 segments of spinal cord in rats with spared nerve injury (SNI), so as to reveal its mechanism underlying reducing neuropathic pain. A total of 120 male SD rats were randomly divided into sham operation, model, EA and NOS inhibitor (N omega-Nitro-L-arginine methyl ester hydrochloride, L-NAME) groups, with 30 rats in each group. The neuropathic pain model was established by ligating and cutting the tibial nerve and the common peroneal nerve. EA (2 Hz, 1-3 mA) was applied to "Weizhong" (BL40) and "Huantiao" (GB30)on the damaged hindlimb for 30 min, once daily from day 11 to 17 after SNI. Rats of the L-NAME group received i.p. of L-NAME (60 mg·kg-1·d-1) for 7 consecutive days. The mechanical pain threshold (PT) was determined before and 10 and 16 d after SNI, respectively. The expression le-vels of CAT-2 mRNA and iNOS mRNA, and iNOS protein in the L4-L6 segments of the spinal cord were detected by using reverse transcription - polymerase chain reaction (RT-PCR) and Western blot, respectively, and the contents of NO and cGMP of L4-L6 assayed using nitrate/nitrite reductase method and radioimmunoassay, respectively. After modeling, the PT was significantly decreased on day 10 and 16 after SNI in comparison with the sham operation group and their own baseline data of pre-operation in each group (P<0.01), and remarkably increased in the EA and L-NAME groups relevant to the model group on day 16 (P<0.01, P<0.05). Compared with the sham operation group, the expression levels of CAT-2 mRNA and iNOS mRNA and protein, as well as the contents of NO2－/NO3－and cGMP were signi-ficantly up-regulated in the model group (P<0.05, P<0.01). Following EA intervention, the levels of CAT-2 mRNA and iNOS mRNA and iNOS protein, and NO2－/NO3－and cGMP contents were all reversed in both EA and L-NAME groups (P<0.05, P<0.01). The effect of EA was significantly superior to that of L-NAME in raising the PT on day 16 after SNI (P<0.05), but obviously inferior to that of L-NAME in down-regulating the expression of CAT-2 mRNA and iNOS mRNA and protein (P<0.05). No significant differences were found between the EA and L-NAME groups in down-regulating NO2－/NO3－ andcGMP contents (P>0.05). EA intervention can effectively relieve neuropathic pain in SNI rats, which may be closely related to its function in suppressing L-Arg/NO/cGMP pathway in the lumbar spinal cord.

Hepatoprotective potential of 7,8-Dihydroxyflavone against alcohol and high-fat diet induced liver toxicity via attenuation of oxido-nitrosative stress and NF-κB activation.

Fatty liver diseases are the most common and major health concern arises from the modern lifestyle and alcohol (ethanol) abuse. The prevalence of non-alcoholic fatty liver diseases (NAFLD) has been observed prominently in obese and diabetic individuals, while alcoholic liver disease is common in alcoholic persons. Fatty liver disease, such as steatohepatitis, leads to fibrosis, cirrhosis and eventually hepatocellular carcinoma. The present study was designed to investigate the effect of 7,8-Dihydroxyflavone (7,8-DHF) against high-fat diet (HFD) and ethanol (EtOH)-induced hepatotoxicity in rats. Male Wistar rats (150-200 g) were fed HFD (58% calories from fat) and EtOH (3-15% in drinking water) for 12 weeks. 7,8-DHF was administered intraperitoneally at the dose of 5 mg/kg/day for the last four weeks. After 12 weeks, biochemical, ELISA, RT-PCR, and histological studies have been carried out. Biochemical analyses revealed the involvement of oxidative stress and inflammation in the liver of HFD and EtOH-fed rats. 7,8-DHF treatment significantly reduced HFD and EtOH-induced oxidative stress as evidenced by the reduction of lipid peroxidation and augmentation of reduced glutathione level. Moreover, IL-1β level was found significantly reduced in 7,8-DHF treated EtOH, HFD and EtOH+HFD groups. The semi-quantitative RT-PCR results indicated down-regulation of Nrf-2 and HO-1 and up-regulation of NF-κB and iNOS mRNA expression level in the liver of HFD and EtOH-fed rats, which was ameliorated by 7,8-DHF treatment. The present study suggested that 7,8-DHF could be an effective pharmacological intervention in combating HFD and EtOH-induced hepatotoxicity.

Protective Effect of Polygonatum sibiricum Polysaccharides on Apoptosis, Inflammation, and Oxidative Stress in Nucleus Pulposus Cells of Rats with the Degeneration of the Intervertebral Disc

Objective. Polygonatum sibiricum polysaccharide (PSP) has antioxidant activity, immune enhancement, and other biological properties. However, the effect of PSP on intervertebral disc degeneration has not been reported. In this study, we mainly investigated the effect of PSP on the apoptosis, inflammation, and oxidative stress of nucleus pulposus cells (NPCs) during the process of intervertebral disc degeneration. Methods. A rat NPC model induced by H2O2 was constructed. The CCK8 method was used to measure the effects of PSP on the apoptosis of rat NPCs induced by H2O2. The effects on the activity of SOD and content of MDA were also determined. The rat model of intervertebral disc degeneration was treated with PSP for 1 month, and the mRNA expression levels of IL-1β, COX2, iNOS, Col2α1, Col10α1, and MMP3 were measured by qPCR in the tissue of intervertebral disc. NPCs from the degenerated intervertebral discs were separated, and the cell viability was measured by the CCK8 method. The contents of SOD and MDA in NPCs were determined as well. Results. PSP significantly reduced the apoptosis of NPCs induced by H2O2, significantly increased the SOD content, and decreased the content of MDA in H2O2-induced NPCs. The expression level of IL-1β, COX2, and iNOS in the rat model with intervertebral disc degeneration was significantly downregulated after 1 month of PSP treatment. PSP treatment increased the expression of Col2α1 type and significantly decreased the expression of Col10α1 type collagen and MMP3 in rats with disc degeneration. PSP treatment significantly reduced NPC apoptosis and increased its SOD content and reduced MDA content, which is consistent with the results from cell-level experiments. Conclusion. PSP can effectively reduce the apoptosis, inflammation, and oxidative stress of H2O2-induced NPCs in rats with intervertebral disc degeneration and mitigate the progression of intervertebral disc degeneration, which has the potential to be developed as new drugs for the treatment of intervertebral disc degeneration.

Anti-inflammatory Ellagitannins from Cleidion brevipetiolatum for the Treatment of Rheumatoid Arthritis.

Six new ellagitannins, brevipetins B-G (5 and 7-11), and a new phenolic glucoside, brevipetin A (4), along with six known compounds were isolated from the traditional Chinese medicinal plant Cleidion brevipetiolatum. Their structures and absolute configurations were determined by spectroscopic analyses, chemical methods, and TD-DFT-ECD calculations. Compounds 5-11 exhibited NO inhibitory effects with IC50 values of 1.9-8.2 μM, and 9 showed the most potent inhibitory effect (IC50: 1.9 μM). An in vivo anti-inflammatory assessment of 9 showed that it exerts therapeutic effects in both the carrageenan-induced rat paw edema and collagen-induced arthritis (CIA) models at 50 mg/kg oral administration. The enhanced protein and mRNA expression levels of iNOS (inducible nitric oxide synthase) and COX-2 (cyclooxygenase-2) in LPS-stimulated RAW 264.7 cells were dose-dependently suppressed by 9. An anti-inflammatory mechanistic study revealed that 9 suppressed NF-κB activity by inhibiting IκBα phosphorylation and blocking translocation of p65 from the cytosol to the nucleus. Therefore, 9 might have the potential to be developed as a lead compound for relieving rheumatoid arthritis.

LncRNA SCAL1 inhibits inducible nitric oxide synthase in lung cells under high-glucose conditions.

The lungs are one of the most common target organs of diabetic injury in patients with diabetes. Long non-coding RNA (lncRNA) smoke and cancer-associated lncRNA 1 (SCAL1), also known as lung cancer associated transcript 1 (LUCAT1), is known to have a pivotal role in lung cancer. The aim of the current study was to investigate the potential involvement of SCAL1 in diabetic lung disease. The expression levels of SCAL1 were determined by reverse transcription-quantitative PCR in serum samples from healthy controls (n=40), diabetic patients without lung disease (n=56) and diabetic patients with diabetic lung disease (n=44). Receiver operating characteristic analysis was used to evaluate the diagnostic value of serum SCAL1 in discriminating diabetic patients with diabetic lung disease from diabetic patients without lung disease and healthy controls. Pearson's correlation analysis was performed to examine the correlation between SCAL1 and inducible nitric oxide synthase (iNOS) mRNA expression levels in blood and lung tissue samples. Expression levels of iNOS and NO production following treatment with high (30 mM) glucose were examined by western blot analysis and NO assay, respectively. The expression levels of SCAL1 were significantly downregulated in diabetic patients with diabetic lung disease, and downregulated serum expression levels of SCAL1 effectively distinguished diabetic patients with diabetic lung disease from diabetic patients without lung disease and healthy controls. Treatment with high glucose significantly upregulated SCAL1 expression in normal lung cells. Furthermore, the overexpression of SCAL1 inhibited iNOS protein expression and reduced NO production in cells treated with high glucose. In conclusion, the current study demonstrated that lncRNA SCAL1 inhibits iNOS protein expression in lung cells under high-glucose conditions, which suggests that SCAL1 may have potential in the treatment of patients with diabetic lung disease.

Neonatal Inhibition of Connexin 36 Ameliorates Fetal Brain Injury Induced by Maternal Noninfectious Fever in Mice

Prenatal fever could result in brain function impairments in the offspring. The present study investigated the effect of interleukin-6 (IL-6)-induced maternal fever on the offspring and the involvement of connexin 36 in this process. Pregnant C57BL/6J mice were injected with IL-6 on gestational day 15. The levels of iNOS and COX-2 were measured as an index of neuroinflammation in the brain of newborn pups. Offspring were treated with the connexin 36 (Cx36) inhibitor mefloquine at postnatal day (P)1–P3 or at P40–P42. Rotarod, grip traction, and foot fault tests were carried out to evaluate the motor behavior of adult offspring. Injection of IL-6 led to an elevation of the core temperature in the pregnant dams. Offspring of these dams showed significantly increased COX-2 and iNOS mRNA expression and protein levels in the whole-brain samples and significantly increased Cx36 in the cerebellum. Moreover, offspring of these dams showed motor deficits at an adult age. Neonatal administration of the Cx36 inhibitor mefloquine could prevent these motor deficits. Maternal fever during pregnancy induced by IL-6 injection could lead to neuroinflammation and motor deficits in the offspring. Neonatal inhibition of Cx36 could ameliorate the motor deficits in the offspring, indicating an involvement of Cx36 in the IL-6-induced maternal fever.

Ruthenium complex, TQ‑5, protects against LPS‑induced macrophage inflammation and acute liver injury in mice via downregulating NF‑κB pathways.

A newly synthesized ruthenium metal complex, TQ‑5, exhibited antithrombotic and antiplatelet effects in our previous study. In the present study, the anti‑inflammatory/hepatoprotective effects and mechanisms of action of TQ‑5 were investigated in lipopolysaccharide (LPS)‑induced RAW 264.7 macrophages in vitro and in acute liver injury in mice in vivo. The results demonstrated that TQ‑5 suppressed the LPS‑induced production of nitric oxide, tumor necrosis factor‑α (TNF‑α), interleukin‑1β (IL‑1β) and inducible nitric oxide synthase (iNOS), without inducing cytotoxicity or damaging the morphology of the RAW 264.7 macrophages. In addition, the role of TQ‑5 in mediating mitogen‑activated protein kinases and nuclear factor κB (NF‑κB) pathways involved in the inflammation process of LPS‑stimulated RAW264.7 cells was investigated. Although TQ‑5 did not alter the phosphorylation of extracellular signal‑related kinase, p38 or c‑Jun N‑terminal kinase in LPS‑treated cells, it suppressed the phosphorylation of Akt in a concentration‑dependent manner. TQ‑5 significantly reversed the LPS‑induced degradation of inhibitor of NF‑κBα and phosphorylation of p65. The mRNA expression levels of iNOS, TNF‑α and IL‑1β were also suppressed by TQ‑5. TQ‑5 improved LPS‑induced liver injury in mice by inhibiting the expression of TNF‑α, IL‑1β and iNOS and phosphorylation of NF‑κBp65. These findings suggest that Akt/NF‑κB signaling may be a promising target for TQ‑5 to combat LPS‑induced inflammation. Therefore, TQ‑5 may act as a potential agent for the development of anti‑inflammatory drugs to treat acute liver failure.

Anti-inflammatory activities of Canarium subulatum Guillaumin methanol extract operate by targeting Src and Syk in the NF-κB pathway

Ethnopharmacological relevanceCanarium subulatum Guillaumin is an herbal medicinal plant native to Southeast Asia. Ethnopharmacological evidence suggests that plants of the genus Canarium cure a variety of inflammatory diseases. Aim of the studyThe pharmacological mechanisms of C. subulatum Guillaumin remain poorly understood. In this study, we investigate inflammatory mechanisms and target molecules using C. subulatum Guillaumin methanol extract (Cs-ME) in inflammatory reactions managed by macrophages. Materials and methodsTo identify the anti-inflammatory activities of Cs-ME, lipopolysaccharide (LPS)-stimulated macrophages and a murine HCl/EtOH-induced gastritis model were chosen. The luciferase reporter gene assay, Western blot analysis, overexpression strategy, and the cellular thermal shift assay (CETSA) were employed to investigate the molecular mechanisms and target enzymes of Cs-ME. The active ingredients of this extract were also determined by HPLC. ResultsReleased levels of nitric oxide (NO) and mRNA expression levels of iNOS and IL-6 were downregulated by Cs-ME without exhibiting cytotoxicity. This extract inhibited MyD88-induced promoter activity and the nuclear translocation of nuclear factor (NF)-κB. Moreover, we found that Cs-ME reduced the phosphorylation of NF-κB upstream signaling molecules including IκBα, IKKα/β, Src, and Syk in LPS-stimulated macrophage-like RAW264.7 cells. The results of Western blot and CETSA confirmed that Src and Syk are anti-inflammatory targets of Cs-ME. In addition, orally injected Cs-ME alleviated HCl/EtOH-induced gastric ulcers in mice. HPLC analysis indicated that quercetin, luteolin, and kaempferol are major active components of this extract with anti-inflammatory activity. ConclusionsCs-ME exhibits anti-inflammatory effects in vitro and in vivo by targeting Src and Syk in the NF-κB signaling pathway. Consequently, Cs-ME could be developed as an anti-inflammatory herbal medicine.

Characterization of Moringa oleifera roots polysaccharide MRP-1 with anti-inflammatory effect.

Moringa oleifera is a mutli-purpose herbal plant which has attained enormous attention as a natural source of nutrients and folk medicine. This work aimed to get a novel polysaccharide, termed MRP-1, which was isolated from Moringa oleifera roots with hot water extraction method followed by ethanol precipitation and purified with DEAE-Sepharose Fast Flow column. Monosaccharide composition analysis based on GC-MS showed that MRP-1 mainly consisted of Rha, Ara, Fru, Xyl, Man and Gal in the molar ratio of 1.5:2.0:3.1:6.0:5.3:1.1. The Roman spectra, FT-IR and NMR analysis showed that the typical features of carbohydrates, such as α-Araf, α-Gly, β-Galp, α-GalpA and β-Gly was contained by MRP-1. The LPS-induced RAW264.7 macrophage cells were used to evaluate the anti-inflammatory activity of MRP-1. The result demonstrated that the increasing of NO and TNF-α production induced by LPS could be prevented by different concentrations of MRP-1 treatment. Moreover, the mRNA expression level of iNOS induced by LPS was decreased significantly (p < 0.05) by MRP-1 treatment while show no obvious effect on the COX-2 mRNA expression. This study may provide new possible application of Moringa oleifera root polysaccharide related to anti-inflammation.

Host–pathogen interaction in Toxoplasma gondii-infected mixed chicken blood cell cultures

Toxoplasma gondii has the ability to infect various nucleated cell types in different hosts. The aim of the present study was to investigate which chicken blood cells were targeted by T. gondii in a mixed blood cell culture similar to in vivo conditions and to evaluate parasite-host cell interactions. The study consisted of two subsequent experiments. In experiment 1, we applied T. gondii tachyzoites (ME49) at a multiplicity of infection of 1tachyzoite per blood cell and examined parasite replication, cytokine, and inducible nitric oxide synthase (iNOS) mRNA expression between 1h and 48h post-infection (p.i.) by quantitative PCR. By using T. gondii RH-GFP tachyzoites expressing the green fluorescent protein (GFP) in experiment 2, we aimed for visualizing infected cells by confocal laser scanning microscopy (CLSM) and flow cytometric analysis at 24h p.i. The parasite replication curve showed a massive decrease of parasite stages until 24h p.i. followed by an approximately plateau phase. We observed mainly significantly increased iNOS mRNA expression levels in T. gondii-infected culture compared to uninfected cells. Flow cytometry and CLSM data confirmed monocytes/macrophages as main target cells for T. gondii. Moreover, different lymphocytes like B cells and cytotoxic T cells seem to be targeted to a low extent. Our findings indicate that monocytes/macrophages play a key role during T. gondii infection in chicken as host cells and triggering of immune response. To the best of our knowledge, this is the first report of a mixed chicken blood cell culture experimentally infected with T. gondii.

Farnesyl thiosalicylic acid prevents iNOS induction triggered by lipopolysaccharide via suppression of iNOS mRNA transcription in murine macrophages

Inducible nitric oxide synthase (iNOS) is a molecule critical for the development of inflammation-associated disorders. Its induction should be tightly controlled in order to maintain cellular homeostasis. Upon lipopolysaccharide (LPS) stimulation, iNOS, in most settings, is induced by the activation of inhibitor of κB-α (IκB-α)-nuclear factor κB (NF-κB) signaling. Farnesyl thiosalicylic acid (FTS), a synthetic small molecule that is considered to detach Ras from the inner cell membrane, has been shown to exhibit numerous anti-inflammatory functions. However, it remains unclear whether and how it affects iNOS induction in macrophages. The present study addressed this issue in cultured macrophages and endotoxemic mice. Results showed that FTS pretreatment significantly prevented LPS-induced increases in iNOS protein and mRNA expression levels in murine cultured macrophages, which were confirmed in organs in vivo from endotoxemic mice, such as the liver and lung. Mechanistic studies revealed that FTS pretreatment did not affect IκB-α degradation and NF-κB activation in LPS-treated macrophages. The nuclear transport of the active NF-κB was also not affected by FTS. But FTS pretreatment reduced the binding of NF-κB to its DNA elements, and reduced NF-κB bindings to iNOS promoter inside LPS-treated macrophages. Finally, our results showed that FST pretreatment increased mouse survival rate compared to LPS alone treatment. Taken together, these results indicate that FTS attenuates iNOS induction in macrophages likely through inhibition of iNOS mRNA transcription, providing further insight into the molecular mechanism of action of FTS in inflammatory disorder therapy.

Anti-Colitic Effects of Ethanol Extract of Persea americana Mill. through Suppression of Pro-Inflammatory Mediators via NF-κB/STAT3 Inactivation in Dextran Sulfate Sodium-Induced Colitis Mice.

Persea americana Mill, cv. Hass, also known as avocado, has been reported to possess hypolipidemic, anti-diabetic, anti-oxidant, cardioprotective, and photoprotective potencies. However, few studies have reported its anti-colitic effects. In this study, we investigated anti-colitic effects of ethanol extract of P. americana (EEP) in dextran sulfate sodium (DSS)-induced colitic mice and the involved molecular mechanisms. EEP effectively improved clinical signs and histological characteristics of DSS-induced colitis mice. In DSS-exposed colonic tissues, EEP reduced expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and pro-inflammatory cytokines such as interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α. Moreover, EEP suppressed DSS-induced activation of nuclear factor-kappa B (NF-κB) and signal transducer and activator of transcription 3 (STAT3). Consistent with in vivo results, EEP also suppressed protein and mRNA expression levels of iNOS, COX-2, and pro-inflammatory cytokines via NF-κB and STAT3 inactivation in LPS-induced RAW 264.7 macrophages. Taken together, our data indicate that ethanol extract of avocado may be used as a promising therapeutic against inflammatory bowel diseases by suppressing the NF-κB and STAT3 signaling pathway.

Immune enhancement activity of a novel polysaccharide produced by Dendrobium officinale endophytic fungus Fusarium solani DO7

A novel polysaccharide DG2 was obtained from the Dendrobium officinale endophytic fungus Fusarium solani DO7. The structural characterization and immunological activity of DG2 were investigated. Results showed that DG2 with the molecular weight of 152.8 kDa was composed of Galp, Manp, Glcp and Araf at a molar ratio of 4.3:1.9:2.8: 1.0. The main linkage types of DG2 were (1 → )-β-D-Glc, (1 → 4)-β-D-Glc, (1 → 3,6)-β-D-Gal and (1 → 6)-β-D-Gal, which demonstrated DG2 was a novel galactoglucan and different from other fungal and botanic polysaccharide. Furthermore, DG2 did not contain deoxynivalenol, and showed no direct toxicity to human embryonic kidney cells and RAW 264.7 cells. Meanwhile, DG2 could increase the TNF-α, IL-6 and iNOs mRNA expression levels of RAW264.7 cells, and further promote the production of TNF-α, IL-6 and NO. These results suggested DG2 could enhance macrophage-mediated immune responses and have the potential to be an attractive functional food supplement candidate for the hypoimmunity therapies.

Glutathione inhibits expression of the proinflammatory biomarker inducible nitric oxide synthase in hepatocytes

Background: Intracellular glutathione (GSH) plays an important regulatory role in the host response to liver injury. However, there have been few scientific reports on the anti-inflammatory effects of GSH. In the inflamed liver, proinflammatory cytokines stimulate liver cells, followed by expression of inducible nitric oxide synthase (iNOS). Excessive nitric oxide (NO) levels produced by iNOS are one of the factors involved in liver injury. Therefore, inhibiting iNOS induction is important for preventing liver injury. This study aimed to investigate the protective effects of GSH on the liver by examining interleukin (IL)-1β-stimulated hepatocytes.Methods: Primary cultured rat hepatocytes were treated with IL-1β in the presence or absence of GSH. Induction of iNOS and its signaling pathway were analyzed.Results: Addition of GSH decreased IL-1β-induced iNOS protein and mRNA expression levels, which resulted in inhibition of NO production. GSH also decreased tumor necrosis factor (TNF)-α and IL-6 mRNA expression. GSH blocked “type I IL-1 receptor upregulation”, one of the essential signaling pathways for iNOS induction, through inactivation of an upstream kinase, phosphatidylinositol 3-kinase/Akt. In contrast, GSH had no effects on degradation of IκB and activation of NF-ĸB (nuclear translocation and its DNA binding). Transfection experiments revealed that GSH reduced iNOS mRNA levels at the promoter transactivation and mRNA stabilization steps. Delayed administration of GSH after IL-1β addition also inhibited iNOS induction. Conclusions: Our study suggests that GSH affects induction of inflammatory mediators, including iNOS and TNF-α, indicating its therapeutic potential for organ injuries, especially for the liver.Keywords: glutathione, inducible nitric oxide synthase, liver injury, primary cultured hepatocytes, type I interleukin-1 receptor, tumor necrosis factor-α

The TLR4 Agonist Immunomax Affects the Phenotype of Mouse Lung Macrophages during Respiratory Syncytial Virus Infection

In the study, the effect of the TLR4 agonist Immunomax was investigated in vitro and in vivo. In particular, Immunomax was shown to polarize mouse bone marrow macrophages from the M0 and M2 states into the M1 state (ARG1 and iNOS mRNA expression levels were used to identify the mouse M1 and M2 phenotypes). Next, we investigated the prophylactic antiviral effect of Immunomax in both a model of mouse respiratory syncytial virus (RSV) infection and a model of RSV-induced bronchial asthma (BA) exacerbation. In the experiment with RSV-induced BA exacerbation, Immunomax-treated mice were characterized by a significant decrease of the viral load in lung homogenates, an increased amount of M1 macrophages in the lung, a tendency toward Th2-dependent ovalbumin-specific IgG1 antibodies decrease in blood serum, a significant increase in RSV-activated CD4+ T cells secreting IFNγ (Th1 cells), and a simultaneous significant decrease in the amount of CD4+ cells secreting IL-4 (Th2 cells) in the mouse spleen, which were detected by ELISPOT 1.5 months after experiment. These findings suggest that treatment with the TLR4 agonist Immunomax polarizes the immune response towards antiviral Th1 and may be used for short-term antiviral prophylaxis to prevent acute respiratory viral infections in asthmatics.

Antitumor and anti-inflammatory effects of oligosaccharides from Cistanche deserticola extract on spinal cord injury

In the present study, the pharmacological effects of oligosaccharides from Cistanche deserticola extract on inflammation, oxidative stress, and apoptosis in male albino rats with spinal cord injury were investigated. Lipid peroxidation, GSH, catalase, superoxide dismutase, acetylcholine esterase, GPx, ROS, and nitric acid were significantly altered in the rats with spinal cord injury. The mRNA expression levels of IL-6, TNF-α, cyclooxygenase-2, iNOS, p53, caspase-3, bax, and pro-NGF were reduced by >20% following extract supplementation. Protein expression levels of caspase-3 and pro-NGF were also reduced by >20%. The number of p53 positive cells was 1, 79, 54, 33, and 19 in groups GI–GV, respectively, and the corresponding numbers of caspase-3 positive cells were 2, 87, 51, 23, and 14. Based on the present results, the use of oligosaccharides from Cistanche deserticola extract was effective against inflammation, oxidative stress, and apoptosis in spinal cord injury male albino rats.

Protective effect of avicularin on rheumatoid arthritis and its associated mechanisms.

The present study aimed to investigate the effect of avicularin on rheumatoid arthritis (RA) in vitro, and additionally explore the molecular mechanism. To perform this investigation, an in vitro model of RA was established by treatment of the human RA synovial MH7A cell line with tumor necrosis factor-α (TNF-α). MH7A cells were then treated with various concentrations (10, 30, 100 and 300 µM) of avicularin. Then, the levels of inflammatory factors [interleukin (IL)-1β, IL-6, IL-8, matrix metalloproteinase (MMP)-1 and MMP-13] were measured by ELISA. Cell viability and apoptosis were detected using an MTT assay and flow cytometry, respectively. In addition, the expression levels of genes and proteins were determined reverse transcription quantitative polymerase chain reaction and western blot analysis. The results of the present study indicated that avicularin significantly decreased the levels of inflammatory factors (IL-1β, IL-6, IL-8, MMP-1 and MMP-13), previously increased by TNF-α, in a dose-dependent manner. Concurrently, avicularin inhibited the mRNA and protein expression levels of iNOS and COX-2 increased by TNF-α. It was also identified that TNF-α administration significantly promoted MH7A cell viability and inhibited cell apoptosis, and avicularin treatment dose-dependently inhibited MH7A cell viability and induced cell apoptosis. In addition, these data suggested that avicularin prevented the activation of the mitogen-activated protein kinase kinase (MEK)/nuclear factor kappa light-chain-enhancer of activated B-cells (NF-κB) pathway activated by TNF-α. Taken together, these results demonstrated that avicularin may inhibit the inflammatory response, prevent cell viability and induce apoptosis in human RA synovial cells through preventing the activation of the MEK/NF-κB pathway.

Ameliorative effect of hesperidin on ligation-induced periodontitis in rats.

This study evaluated the ameliorative effect of hesperidin (HES), an anti-inflammatory flavanone, in rats with ligation (Lig)-induced periodontitis. A total of 48 rats were randomly divided into non-ligation group (NL), Lig group, and two ligation-plus-HES groups (L+H). HES was administered immediately after ligature placement at a dose of 75 or 150mg/kg by intragastric feeding. Destruction of the ligated maxillary second and mandibular first molars were evaluated by dental radiography, microcomputed tomography (micro-CT), and histometry performed after sacrificing the rats on the seventh day. The expression levels of interleukin (IL)-1β, IL-6, and inducible nitric oxide synthase (iNOS) messenger (m)RNAs in the gingiva were determined by reverse-transcription polymerase chain reaction. The expression of iNOS was examined by immunohistochemistry. The dental radiography and micro-CT findings revealed significantly increased alveolar bone loss in the Lig group, which was significantly prevented by HES. The histometry results revealed less gingival inflammation and connective tissue loss in the L+H groups compared with that in the Lig group. The mRNA expression levels of IL-6, IL-1 β, and iNOS were significantly increased in the Lig group but were reduced in the L+H groups. The immunostaining results showed that the ligation-induced iNOS expression was also decreased by HES. Oral administration of HES promotes an ameliorative effect against the ligation-induced alveolar bone loss and effectively inhibits the production of proinflammatory mediators in rats with experimentally induced periodontitis. Therefore, HES may be a good candidate for modulating oral inflammatory diseases.

Curcumin ameliorates atherosclerosis in apolipoprotein E deficient asthmatic mice by regulating the balance of Th2/Treg cells

BackgroundAllergic asthma and atherosclerosis represent different directions of inflammatory responses of CD4+ T cells, and allergic asthma accelerates atherosclerosis formation. Curcumin could ameliorate the progression of both atherosclerosis and allergic asthma. PurposeWe aimed to investigate the roles of curcumin in asthma-accelerated atherosclerosis plaque formation, and the change of CD4+ T-cell subsets in this process. MethodsSix to eight-week-old apolipoprotein E−/− (apoE−/−) mice were sensitized and challenged by ovalbumin (OVA) to establish an allergic asthma model, and then received curcumin or vehicle treatment for 8 weeks. ResultsThe accelerated atherosclerosis was induced by allergic asthma accompanied by increased T helper cell (Th)2 and Th17 cells and decreased regulatory T cells (Tregs) in the spleen. After the 8-week treatment with curcumin, the lesion areas in the aortic root in asthmatic mice significantly improved, and the elevated Th2 and Th17 cells significantly decreased, but Tregs markedly increased. Although curcumin treatment markedly reduced the interleukin (IL)-4 and IL-13 in serum and spleen, the elevated IL-17A did not decrease. Moreover, Th1 cells showed no significant change between different groups. The mRNA expression levels of M1 macrophage-related inflammatory factors IL-6, iNOS and IL-1β were markedly elevated in the spleens of asthmatic mice, but significantly decreased after the 8-week treatment with curcumin. ConclusionCurcumin ameliorated the aggravation of atherosclerotic lesions and stabilised plaque by modulating the balance of Th2/Tregs in asthmatic apoE−/− mice.