Inducible Nitric Oxide Synthase Messenger Research Articles

Efficacy of vibegron, a novel β3-adrenoreceptor agonist, for lower urinary tract dysfunction in mice with spinal cord injury.

To investigate the effect of vibegron, a new clinically approved β3-adrenoceptor agonist in lower urinary tract dysfunction in mice with spinal cord injury. Investigators performed cystometry under awake conditions in 4-week spinal cord injury female mice. Two weeks after spinal cord injury, saline or vibegron (30mg/kg) was orally administered for 2weeks prior to the urodynamic study. Investigators removed L6-S1 dorsal root ganglia from the saline- or vibegron-treated spinal cord injury mice as well as from saline-treated normal (spinal intact) mice to evaluate the levels of transient receptor potential cation channel subfamily V member 1, transient receptor potential cation channel subfamily A member 1, activating transcription factor 3, and inducible nitric oxide synthase transcripts using real-time polymerase chain reaction. In vibegron-treated spinal cord injury mice, nonvoiding contractions during bladder filling, which were increased in spinal cord injury compared to spinal intact mice, were significantly decreased. Micturition pressure or voiding efficiency was not significantly increased in comparison to measurements in saline-treated spinal cord injury mice. The expression of transient receptor potential cation channel subfamily V member 1, transient receptor potential cation channel subfamily A member 1, activating transcription factor 3, and inducible nitric oxide synthase messenger RNA was increased in spinal cord injury mice compared to spinal intact mice, but significantly decreased after vibegron treatment. Vibegron improves spinal cord injury-induced detrusor overactivity in addition to significantly reducing C-fiber afferent receptors such as transient receptor potential cation channel subfamily V member 1, transient receptor potential cation channel subfamily A member 1, and inflammatory cytokines/markers, such as activating transcription factor 3 and inducible nitric oxide synthase, in spinal cord injury mice. Thus, vibegron might be effective in the treatment of storage lower urinary tract dysfunction induced by C-fiber afferent activation after spinal cord injury.

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.

Carvacrol Ameliorates Ligation-Induced Periodontitis in Rats.

This study aims to evaluate the ameliorative effect of carvacrol, an anti-inflammatory monoterpenoid phenol and a major component of Plectranthus amboinicus, on periodontal damage in an experimental rat model of periodontitis. Forty Sprague-Dawley rats were divided into ligation (Lig), non-ligation (n-Lig), and two ligation plus carvacrol (Lig+C) groups. Carvacrol (17.5 or 35.0 mg/kg body weight/day) was administered intragastrically from 1 day before ligation. On day 8, dental alveolar bone loss and gingival inflammation in periodontal specimens were examined by dental radiography, microcomputed tomography, and histology. Expressions of tumor necrosis factor-α, interleukin (IL)-1β, IL-6, and inducible nitric oxide synthase messenger (m)RNAs, and levels of matrix metalloproteinase (MMP)-2 and MMP-9 in gingiva were examined by reverse transcription-polymerase chain reaction and zymography. Dental radiography revealed periodontal bone-supporting ratios in Lig and Lig+C groups were lower than the n-Lig group, with Lig group ratios being lowest. Compared with the n-Lig group, the cemento-enamel junction-bone distance was significantly longer in Lig and Lig+C groups, with Lig+C groups showing shorter distances regardless of radiographic methods used. Histology and histometry showed less inflammatory area and stronger connective tissue attachment in Lig+C groups than in the Lig group. Cytokine/mediator mRNA expression and MMP-9 levels were reduced in the Lig+C groups. Carvacrol reduced tissue damage and bone loss caused by ligation-induced periodontitis. The present results indicate that carvacrol might reduce tissue destruction by downregulating expression of proinflammatory mediators and MMP-9.

Intestinal alkaline phosphatase deficiency leads to dysbiosis and bacterial translocation in the newborn intestine

BackgroundIntestinal alkaline phosphatase (IAP) has been shown to help maintain intestinal homeostasis. Decreased expression of IAP has been linked with pediatric intestinal diseases associated with bacterial overgrowth and subsequent inflammation. We hypothesize that the absence of IAP leads to dysbiosis, with increased inflammation and permeability of the newborn intestine. MethodsSprague–Dawley heterozygote IAP cross-matches were bred. Pups were dam fed ad lib and euthanized at weaning. The microbiotas of terminal ileum (TI) and colon was determined by quantitative real-time polymerase chain reaction (qRT-PCR) of subphylum-specific bacterial 16S ribosomal RNA. RT-PCR was performed on TI for inflammatory cytokines. Intestinal permeability was quantified by fluorescein isothiocyanate–dextran permeability and bacterial translocation by qRT-PCR for bacterial 16S ribosomal RNA in mesenteric lymph nodes. Statistical analysis was done by chi-square analysis. ResultsAll three genotypes had similar concentrations of bacteria in the TI and colon. However, IAP knockout (IAP-KO) had significantly decreased diversity of bacterial species in their colonic stool compared with heterozygous and wild-type (WT). IAP-KO pups had a nonstatistically significant 3.9-fold increased inducible nitric oxide synthase messenger RNA expression compared with WT (IAP-KO, 3.92 ± 1.36; WT, 1.0 ± 0.27; P = 0.03). IAP-KO also had significantly increased bacterial translocation to mesenteric lymph nodes occurred in IAP-KO (IAP-KO, 7625 RFU/g ± 3469; WT, 4957 RFU/g ± 1552; P = 0.04). Furthermore, IAP-KO had increased permeability (IAP-KO, 0.297 mg/mL ± 0.2; WT, 0.189 mg/mL ± 0.15 P = 0.07), but was not statistically significant. ConclusionsDeficiency of IAP in the newborn intestine is associated with dysbiosis and increased inflammation, permeability, and bacterial translocation.

Nitric oxide synthase and changes in oxidative stress levels in embryonic kidney observed in a rabbit model of intrauterine growth restriction

This work aimed to study the effect of uteroplacental circulation restriction on endothelial kidney damage in a fetal rabbit model. New Zealand rabbits were subjected to 40% to 50% of uteroplacental artery ligation at day 25 of pregnancy. After 5 days, surviving fetuses were harvested by cesarean section. The gene and protein expressions of selected enzymes associated with nitric oxide production and oxidative stress were analyzed in fetal kidney homogenates. The placenta weight (6.06 ± 0.27, p < 0.0319) and fetal body (19.90 ± 1.03, p < 0.0001) were significantly reduced in the uteroplacental circulation restriction group. The kidneys from restricted fetuses presented a mild vascular congestion and glomerular capillary congestion, without inflammation or hypertrophy. We found endothelial nitric oxide synthase phosphorylation inhibition (0.23 ± 0.13, p < 0.012) and arginase-2 (0.29 ± 0.14, p < 0.023) protein induction in fetal kidneys of the circulation restriction group. Finally, the kidneys from circulation-restricted fetuses showed increased inducible nitric oxide synthase messenger RNA (mRNA) (2.68 ± 0.24, p < 0.01) and reduced heme oxygenase-1 mRNA (23 ± 1.3, p < 0.003), with increased reactive oxygen species (1.69 ± 0.09, p < 0.001) and nitrotyrosine protein (1.74 ± 0.28, p < 0.003) levels, without changes in Nox mRNA. We describe significant deregulation of vascular activity and oxidative damage in kidneys of fetal rabbits that have been exposed to restriction of the uterine circulation. © 2016 John Wiley & Sons, Ltd.

Adiponectin attenuates liver fibrosis by inducing nitric oxide production of hepatic stellate cells.

Adiponectin protects against liver fibrosis, but the mechanisms have not been fully elucidated. Here, we showed that adiponectin upregulated inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) and protein expression in hepatic non-parenchymal cells, particularly in hepatic stellate cells (HSCs), and increased nitric oxide (NO2-/NO3-) concentration in HSC-conditioned medium. Adiponectin attenuated HSC proliferation and migration but promoted apoptosis in a NO-dependent manner. More advanced liver fibrosis with decreased iNOS/NO levels was observed in adiponectin knockout mice comparing to wide-type mice when administered with CCI4 while NO donor supplementation rescued the phenotype. Further experiments demonstrated that adiponectin-induced iNOS/NO system activation is mediated through adipoR2-AMPK-JNK/Erk1/2-NF-κB signaling. These data suggest that adiponectin inhibits HSC function, further limiting the development of liver fibrosis at least in part through adiponectin-induced NO release. Therefore, adiponectin-mediated NO signaling may be a novel target for the treatment of liver fibrosis. • Adiponectin activates HSC iNOS/NO and SEC eNOS/NO systems. • Adiponectin inhibits HSC proliferation and migration but promotes its apoptosis. • Adiponectin inhibits CCL4-induced liver fibrosis by modulation of liver iNOS/NO. • Adiponectin stimulates HSC iNOS/NO via adipoR2-AMPK-JNK/ErK1/2-NF-κB pathway.

Silica nanoparticles induce oxidative stress, inflammation, and endothelial dysfunction in vitro via activation of the MAPK/Nrf2 pathway and nuclear factor-κB signaling.

Despite the widespread application of silica nanoparticles (SiNPs) in industrial, commercial, and biomedical fields, their response to human cells has not been fully elucidated. Overall, little is known about the toxicological effects of SiNPs on the cardiovascular system. In this study, SiNPs with a 58 nm diameter were used to study their interaction with human umbilical vein endothelial cells (HUVECs). Dose- and time-dependent decrease in cell viability and damage on cell plasma-membrane integrity showed the cytotoxic potential of the SiNPs. SiNPs were found to induce oxidative stress, as evidenced by the significant elevation of reactive oxygen species generation and malondialdehyde production and downregulated activity in glutathione peroxidase. SiNPs also stimulated release of cytoprotective nitric oxide (NO) and upregulated inducible nitric oxide synthase (NOS) messenger ribonucleic acid, while downregulating endothelial NOS and ET-1 messenger ribonucleic acid, suggesting that SiNPs disturbed the NO/NOS system. SiNP-induced oxidative stress and NO/NOS imbalance resulted in endothelial dysfunction. SiNPs induced inflammation characterized by the upregulation of key inflammatory mediators, including IL-1β, IL-6, IL-8, TNFα, ICAM-1, VCAM-1, and MCP-1. In addition, SiNPs triggered the activation of the Nrf2-mediated antioxidant system, as evidenced by the induction of nuclear factor-κB and MAPK pathway activation. Our findings demonstrated that SiNPs could induce oxidative stress, inflammation, and NO/NOS system imbalance, and eventually lead to endothelial dysfunction via activation of the MAPK/Nrf2 pathway and nuclear factor-κB signaling. This study indicated a potential deleterious effect of SiNPs on the vascular endothelium, which warrants more careful assessment of SiNPs before their application.

Inhibition of Lipopolysaccharide (LPS)-induced inflammatory responses by selenium in bovine mammary epithelial cells in primary culture.

Selenium, in the form of selenoproteins, plays a pivotal role in anti-inflammatory processes and antioxidant defense system. The aim of this study was to examine the effects of selenium on lipopolysaccharide (LPS)-induced inflammatory responses in bovine mammary epithelial cells (bMEC) and to investigate the potential mechanism. bMEC viability was measured by MTT assay. TNF-α, IL-1β, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) expressions were evaluated by quantitative real-time polymerase chain reaction (qRT-PCR). The activation of nuclear factor-kappa B (NF-κB) was determined by Western blotting. The results showed that the mRNA expressions of these inflammatory factors were significantly inhibited by selenium in a dose-dependent manner. At protein levels, Western blot analysis demonstrated that selenium dose-dependently decreased NF-κB p65 translocating from the cytoplasm to the nucleus. Taken together, these results suggest that the anti-inflammatory property of selenium in LPS-stimulated primary bMEC may be attributed to the downregulation of NF-κB activation.

Ghrelin attenuates sepsis-associated acute lung injury oxidative stress in rats.

This study investigated the effect of ghrelin on oxidative stress in septic rat lung tissue. Male Sprague-Dawley rats were divided into sham-operation, sepsis, and ghrelin groups. Sepsis was induced by cecal ligation and puncture. Ghrelin was administered intraperitoneally at 3 and 15 h post-operation. Bronchoalveolar lavage was performed to collect alveolar macrophages (AMs). Inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) expression in alveolar macrophages and iNOS protein levels were measured by reverse transcription PCR (RT-PCR) and Western blot. Pulmonary pathology was analyzed and nitrotyrosine expression was examined by immunohistochemistry. Plasma superoxide dismutase (SOD) and lung wet/dry weight were measured. In the sepsis group, iNOS mRNA expression in AMs was 1.33 ± 0.05, 1.44 ± 0.08, and 1.57 ± 0.11 at 6, 12, and 20 h post-surgery, respectively, and were higher compared with the sham-operation group (p<0.05). No increase was observed at longer time points. iNOS mRNA expression in the sepsis group was lower compared with the ghrelin group (2.27 ± 0.37) (p<0.05) at 20 h post-surgery. iNOS protein levels in the ghrelin group (0.87 ± 0.03, p<0.05) were lower than in the sepsis group at 20 h. Ghrelin group pathological scores were lower than in the sepsis group (p<0.05). Plasma SOD was slightly non-significantly decreased in the ghrelin group. No difference was observed in lung wet/dry weight ratios between sepsis and ghrelin groups. iNOS mRNA expression in AMs was elevated between 6 and 20 h after cecal ligation and puncture (CLP), but did not progress. Ghrelin attenuated pulmonary iNOS protein expression and tended to increase plasma SOD activity. Ghrelin suppressed pulmonary nitrosative stress in septic rats, but did not improve lung wet/dry weight ratios.

Selective endothelin-A receptor blockade attenuates endotoxin-induced pulmonary hypertension and pulmonary vascular dysfunction.

Endothelin-1 is a potent mediator of sepsis-induced pulmonary hypertension (PH). The pulmonary vascular effects of selective blockade of endothelin receptor subtype A (ETAR) during endotoxemia remain unknown. We hypothesized that selective ETAR antagonism attenuates endotoxin-induced PH and improves pulmonary artery (PA) vasoreactivity. Adult male Sprague-Dawley rats (250-450 g) received lipopolysaccharide (LPS; Salmonella typhimurium; 20 mg/kg intraperitoneally) or vehicle 6 hours before hemodynamic assessment and tissue harvest. The selective ETAR antagonist sitaxsentan (10 or 20 mg/kg) or vehicle was injected intravenously 3 hours after receipt of LPS. Right ventricular systolic pressure, mean arterial pressure (MAP), cardiac output (CO), oxygenation (P/F ratio), and serum bicarbonate were measured. Bronchoalveolar lavage (BAL) cell differential and lung wet-to-dry ratios were obtained. Endothelium-dependent and endothelium-independent vasorelaxations were determined in isolated PA rings. PA interleukin (IL)-1β, IL-6, tumor necrosis factor α (TNF-α), and inducible nitric oxide synthase (iNOS) messenger RNA (mRNA) were measured. LPS caused PH, decreased MAP, CO, and serum bicarbonate, and increased PA IL-1β, IL-6, TNF-α, and iNOS mRNA. Sitaxsentan attenuated sepsis-induced PH and increased MAP. The P/F ratio, CO, serum bicarbonate, and BAL neutrophilia were not affected by sitaxsentan. In isolated PA rings, while not affecting phenylephrine-induced vasocontraction or endothelium-dependent relaxation, sitaxsentan dose-dependently attenuated LPS-induced alterations in endothelium-independent relaxation. PA cytokine mRNA levels were not significantly attenuated by ETAR blockade. We conclude that ETAR blockade attenuates endotoxin-induced alterations in systemic and PA pressures without negatively affecting oxygenation. This protective effect appears to be mediated not by attenuation of sepsis-induced cardiac dysfunction, acidosis, or alveolar inflammation but rather by improved endothelium-independent vasorelaxation.

Antitumor effects of dammarane-type saponins from steamed Notoginseng.

Six dammarane-type saponins were extracted from steamed Panax notoginseng. Their chemical structures were identified spectroscopically as ginsenosides Rh1 (1), Rg1 (2), 20 (S)-Rg3 (3), 20 (R)-Rg3 (4), Rb3 (5), and Rb1 (6). Compounds (0.1-10 μM) were tested for inhibition of tumor necrosis factor-α (TNF)-induced nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) luciferase reporter activity using a human kidney 293T cell-based assay. Ginsenoside Rb3 (5) showed the most significant activity with an IC50 of 8.2 μM. This compound also inhibited the induction of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) messenger Ribonucleic acid (mRNA) in a dose-dependent manner after HepG2 cells had been treated with TNF-α (10 ng/mL).

Penehyclidine hydrochloride inhibits the LPS-induced inflammatory response in microglia

BackgroundActivated microglia play an important role in neuroinflammation, which contributes to the neuronal damage found in many neurodegenerative diseases. Penehyclidine hydrochloride (PHC) is an anesthetic used before surgical operations, but also exhibits anti-inflammatory effects on the respiratory and digestive system. In the present study, we investigated whether PHC produces similar anti-inflammatory effects in activated microglia in the central nervous system. Materials and methodsMicroglial cells were incubated with lipopolysaccharide (LPS) in the presence or absence of various concentrations of PHC, SB203580 (p38 mitogen-activated protein kinase [MAPK] inhibitor), and pyrrolidine dithiocarbamate (nuclear factor-kappa B [NF-κB] inhibitor). Markers of inflammation and oxidative stress were measured using enzyme-linked immunosorbent assay and quantitative real-time polymerase chain reaction. The effect of PHC on NF-κB activity was assessed with a NF-κB p50/p65 transcription factor assay kit. The involvement of p38 MAPK phosphorylation in the anti-inflammatory effects of PHC was evaluated with a specific enzyme-linked immunosorbent assay kit for phospho-p38. ResultsPHC significantly inhibited the release of nitric oxide, prostaglandin E2, interleukin 1β, and tumor necrosis factor α while upregulating the expression of inducible nitric oxide synthase messenger RNA in LPS-activated microglia. Moreover, PHC effectively inhibited the translocation of NF-κB from the cytoplasm to the nucleus and the phosphorylation of p38 MAPK. The activities of NF-κB and p38 MAPK in LPS-treated microglia were significantly lowered after pretreatment of PHC. ConclusionsPHC inhibited the LPS-induced release of inflammatory mediators in microglia. These inhibitory effects of PHC may be mediated by blocking p38 MAPK and NF-κB pathways in microglia. These preclinical findings may offer a novel therapeutic option to confine microglial overactivation in neurodegenerative diseases.

Pulmonary Vascular Dysfunction Induced by High Tidal Volume Mechanical Ventilation*

Acute lung injury and acute respiratory distress syndrome are characterized by increased pulmonary artery pressure and ventilation-perfusion mismatch. We analyzed the changes in the pulmonary vascular function in a model of ventilator-induced acute lung injury. Controlled in vivo laboratory study. Animal research laboratory. Anesthetized male Sprague-Dawley rats. Rats were ventilated for 120 minutes using low tidal volume ventilation (control group, tidal volume 9 mL/kg, positive end-expiratory pressure 5 cm H2O, n = 15), high tidal volume ventilation (high tidal volume group, tidal volume 25 mL/kg, zero positive end-expiratory pressure, n = 14), or high tidal volume ventilation plus the poly-(adenosine diphosphate-ribose) polymerase inhibitor 3-aminobenzamide (10 mg/kg IP, high tidal volume group + 3-aminobenzamide group, n = 7). Vascular rings from small pulmonary arteries were mounted in a myograph for isometric tension recording. Lung messenger RNA and protein expression were analyzed by reverse transcriptase-polymerase chain reaction and Western blot, respectively. High tidal volume ventilation impaired phenylephrine- and acetylcholine-induced responses in pulmonary arteries in vitro, which were accompanied by induction of inducible nitric oxide synthase messenger RNA and protein. These effects, as well as hypoxemia and hypotension, were prevented by 3-aminobenzamide. Hypoxic pulmonary vasoconstriction and responses to exogenous sphingomyelinase were increased, whereas the responses to serotonin, Kv current density, and inhibition of Kv currents by hypoxia were unaffected by high tidal volume. High tidal volume ventilation-induced pulmonary vascular dysfunction was characterized by reduced alpha-adrenergic-induced vasoconstriction, reduced endothelium-dependent vasodilatation, and enhanced hypoxic pulmonary vasoconstriction.

Saw Palmetto Extract Enhances Erectile Responses by Inhibition of Phosphodiesterase 5 Activity and Increase in Inducible Nitric Oxide Synthase Messenger Ribonucleic Acid Expression in Rat and Rabbit Corpus Cavernosum

To evaluate whether saw palmetto extract (SPE) relaxes corpus cavernosum and explore the underlying mechanisms. Forty Sprague-Dawley rats and 30 New Zealand rabbits were randomly allocated into 3 SPE-treated groups (low-, middle-, and high-dose) and 1 saline-treated control group. SPE was administered intragastrically for 7 consecutive days. Another 23 rats treated with sildenafil were used to appraise the erectile response to electrical stimulation of nerves in the corpus cavernosum. The erectile functions of rats and rabbits were evaluated 24 hours after the last SPE administration or 15 minutes after intragastric sildenafil. Outcome measures included corpus cavernosum electrical activity recording, phosphodiesterase 5 (PDE5) activity detected by the colorimetric quantitative method, and messenger ribonucleic acid (mRNA) expression level for PDE5 and inducible nitric oxide synthase (iNOS) determined using real-time polymerase chain reaction. In the SPE-treated animals, the relaxant response to electrical stimulation of nerves in the corpus cavernosum, reflected by the amplitude of the electrical activity within the cavernosum, was significantly and dose-dependently augmented. Similar effects were observed in the sildenafil-treated rats. PDE5 activity in rat and rabbit corpus cavernosum tissues was significantly and dose-dependently inhibited in SPE-treated animals, whereas the iNOS mRNA level increased compared with the saline group. PDE5 mRNA, however, was only significantly enhanced in the rats treated with the middle dose of SPE. The results suggest that SPE may have potential application value for the prevention or treatment of erectile dysfunction through an increase in iNOS mRNA expression and inhibition of PDE5 activity in corpus cavernosum smooth muscles.

Enhanced angiogenesis and relaxation of bladder as early response to bladder outlet obstruction

To provide insights into the pathogenesis of bladder insult secondary to bladder outlet obstruction. Six-week-old female Sprague-Dawley rats (n = 80) were divided into eight groups, 10 rats each, according to the duration of bladder outlet obstruction, including 0, 3, 6, 12, 24, 48, 72 h and 1 week. Cystometric parameters were evaluated at 72 h and 1 week after bladder outlet obstruction. Bladder tissues were harvested and Masson's trichrome staining was carried out. Each slide was inspected microscopically and the mean percent collagen area was examined. Changes of collagen deposition and pathological expression of several factors including hypoxia inducible factor-1α, vascular endothelial growth factor, transforming growth factor-β1 and nitric oxide synthase messenger ribonucleic acid of bladders were evaluated. A significant time-dependent increase in the bladder weight after 6 h and the percent of collagen area after 24 h of bladder outlet obstruction were found. Increase in hypoxia inducible factor-1α, transforming growth factor-β1, inducible nitric oxide synthase messenger ribonucleic acid expression, time-dependent increase in vascular endothelial growth factor, neuronal nitric oxide synthase and endothelial nitric oxide synthase messenger ribonucleic acid expression after 6 h of bladder outlet obstruction was found. The intercontraction interval decreased significantly after 72 h of bladder outlet obstruction. Cellular remodeling in the bladder secondary to bladder outlet obstruction starts in the early hours and it includes enhanced angiogenesis and bladder relaxation. Early relief from bladder outlet obstruction is helpful to preserve bladder structure and function.

Dexmedetomidine attenuates lipopolysaccharide-induced proinflammatory response in primary microglia

BackgroundNeuroinflammation mediated by microglia has been implicated in delirium. Suppression of microglial activation may therefore contribute to alleviate delirium. It has been reported that dexmedetomidine (DEX) has a potent anti-inflammatory property. In the present study, we investigated the effects of DEX on the production of proinflammatory mediators in lipopolysaccharide-stimulated microglia. Materials and methodsThe concentrations of DEX were chosen to correspond to 1, 10, and 100 times of clinically relevant concentration (i.e., 1, 10, and 100ng/mL). The levels of proinflammatory mediators, such as inducible nitric oxide synthase or nitric oxide, prostaglandin E2, interleukin 1β, and tumor necrosis factor α, were measured. ResultsDEX at 1ng/mL did not affect the production of proinflammatory mediators. DEX at 10 and 100ng/mL significantly inhibited the release of nitric oxide, prostaglandin E2, interleukin 1β, and tumor necrosis factor α and the expression of inducible nitric oxide synthase messenger RNA. ConclusionsThese results suggest that DEX is a potent suppressor of lipopolysaccharide-induced inflammation in activated microglia and may be a potential therapeutic agent for the treatment of intensive care unit delirium.

Nitric oxide stress in sporadic inclusion body myositis muscle fibres: inhibition of inducible nitric oxide synthase prevents interleukin-1β-induced accumulation of β-amyloid and cell death

Sporadic inclusion body myositis is a severely disabling myopathy. The design of effective treatment strategies is hampered by insufficient understanding of the complex disease pathology. Particularly, the nature of interrelationships between inflammatory and degenerative pathomechanisms in sporadic inclusion body myositis has remained elusive. In Alzheimer's dementia, accumulation of β-amyloid has been shown to be associated with upregulation of nitric oxide. Using quantitative polymerase chain reaction, an overexpression of inducible nitric oxide synthase was observed in five out of ten patients with sporadic inclusion body myositis, two of eleven with dermatomyositis, three of eight with polymyositis, two of nine with muscular dystrophy and two of ten non-myopathic controls. Immunohistochemistry confirmed protein expression of inducible nitric oxide synthase and demonstrated intracellular nitration of tyrosine, an indicator for intra-fibre production of nitric oxide, in sporadic inclusion body myositis muscle samples, but much less in dermatomyositis or polymyositis, hardly in dystrophic muscle and not in non-myopathic controls. Using fluorescent double-labelling immunohistochemistry, a significant co-localization was observed in sporadic inclusion body myositis muscle between β-amyloid, thioflavine-S and nitrotyrosine. In primary cultures of human myotubes and in myoblasts, exposure to interleukin-1β in combination with interferon-γ induced a robust upregulation of inducible nitric oxide synthase messenger RNA. Using fluorescent detectors of reactive oxygen species and nitric oxide, dichlorofluorescein and diaminofluorescein, respectively, flow cytometry revealed that interleukin-1β combined with interferon-γ induced intracellular production of nitric oxide, which was associated with necrotic cell death in muscle cells. Intracellular nitration of tyrosine was noted, which partly co-localized with amyloid precursor protein, but not with desmin. Pharmacological inhibition of inducible nitric oxide synthase by 1400W reduced intracellular production of nitric oxide and prevented accumulation of β-amyloid, nitration of tyrosine as well as cell death inflicted by interleukin-1β combined with interferon-γ. Collectively, these data suggest that, in skeletal muscle, inducible nitric oxide synthase is a central component of interactions between interleukin-1β and β-amyloid, two of the most relevant molecules in sporadic inclusion body myositis. The data further our understanding of the pathology of sporadic inclusion body myositis and may point to novel treatment strategies.

The translation inhibitor pateamine A prevents cachexia-induced muscle wasting in mice

Cachexia, or muscle-wasting syndrome, is one of the major causes of death in patients affected by diseases such as cancer, AIDS and sepsis. However, no effective anti-cachectic treatment is currently available. Here we show that a low dose of pateamine A, an inhibitor of translation initiation, prevents muscle wasting caused by the cytokines interferon γ and tumour necrosis factor α or by C26-adenocarcinoma tumours. Surprisingly, although high doses of pateamine A abrogate general translation, low doses selectively inhibit the expression of pro-cachectic factors such as inducible nitric oxide synthase. This selectivity depends on the 5′UTR of inducible nitric oxide synthase messenger RNA (mRNA) that, unlike the 5′UTR of MyoD mRNA, promotes the recruitment of inducible nitric oxide synthase mRNA to stress granules, where its translation is repressed. Collectively, our data provide a proof of principle that nontoxic doses of compounds such as pateamine A could be used as novel drugs to combat cachexia-induced muscle wasting.

Carbon monoxide induces hypothermia tolerance in Kupffer cells and attenuates liver ischemia/reperfusion injury in rats

Ischemia/reperfusion (I/R) injury in liver grafts, which is initiated by cold preservation and is augmented by reperfusion, is a major problem that complicates graft quality, posttransplant patient care, and outcomes of liver transplantation (LT). Kupffer cells (KCs) play important roles in I/R injury; however, little is known about their changes during cold preservation. We examined whether a pretreatment with carbon monoxide (CO), a cytoprotective product of heme degradation, could influence KC activity during cold storage and protect liver grafts against LT-induced I/R injury. In vitro, primary rat KCs were stimulated for 24 hours under hypothermic conditions (4°C, 20% O(2)), with lipopolysaccharide, or under hypoxic conditions (37°C, 5% O(2)) with or without a CO pretreatment. When rat KCs were exposed to hypothermic conditions, they produced reactive oxygen species (ROS), but they did not produce tumor necrosis factor α (TNF-α) or nitric oxide. The preincubation of KCs with CO up-regulated heat shock protein 70 (HSP70) and inhibited ROS generation. When liver grafts from donor rats exposed to CO (250 ppm) for 24 hours were transplanted after 18 hours of cold preservation in University of Wisconsin solution, HSP70 expression increased in these grafts versus control grafts, and serum aspartate aminotransferase and alanine aminotransferase levels as well as necrotic areas and inflammatory infiltrates were significantly reduced after LT. CO-pretreated liver grafts showed less up-regulation of TNF-α and inducible nitric oxide synthase messenger RNA (mRNA) and reduced expression of proapoptotic B cell lymphoma 2-associated X protein mRNA, cleaved caspase-3, and poly(adenosine diphosphate ribose) polymerase. In conclusion, the pretreatment of donors with CO ameliorates LT-associated I/R injury with increased hepatic HSP70 expression, particularly in the KC population.

Increased inflammation and lethality of Dusp1−/− mice in polymicrobial peritonitis models

The mitogen-activated protein kinase phosphatase Dusp1 (also known as MKP-1) is essential for control of the inflammatory response to systemic challenge with the lipopolysaccharide of Gram-negative bacteria. Here, we have investigated the consequences of Dusp1-deficiency in colon ascendens stent peritonitis (CASP) and caecal ligation and puncture (CLP), two mouse models of septic peritonitis. Following CASP, Dusp1(-/-) mice had increased serum levels of CCL4, interleukin-10 (IL-10) and IL-6, with differences from wild-type mice being dependent on severity of sepsis. These cytokines, along with inducible nitric oxide synthase messenger RNA, were also expressed at higher levels in spleen and liver. Similar over-production of these cytokines was detected in the CLP model, with even larger differences from wild-type mice. Despite the increased inflammatory response, bacterial clearance was impaired in Dusp1(-/-) mice subjected to CASP and CLP. Dusp1(-/-) mice suffered increased lethality in both peritonitis models. Together our data indicate that exaggerated inflammatory responses to gut bacteria introduced into the peritoneum in the absence of Dusp1 do not help to control bacterial replication but are detrimental for the host.