iNOS Pathway Research Articles

Chicken macrophages infected with Salmonella (S.) Enteritidis or S. Heidelberg produce differential responses in immune and metabolic signaling pathways

Protein kinases act in coordination with phosphatases to control protein phosphorylation and regulate signaling pathways and cellular processes involved in nearly every functions of cell life. Salmonella are known to manipulate the host kinase network to gain entrance and survive inside host cells. The effect of Salmonella infection on the host kinase network has been studied in mammalian cells, but information is largely lacking in chicken immune cells. Our previous study indicated that chicken macrophage cells respond differentially to different Salmonella strains. In order to better understand the interaction between chicken macrophages and Salmonella, we used a peptide array-based kinome analysis to identify cellular process and signaling pathways that may play a critical role in the outcome of Salmonella infection. The kinome assay was performed on chicken HD11 macrophages collected at 1.5, 3, and 7h post-infection (hpi) with either S. Heidelberg or S. Enteritidis. A large number of peptides show significantly changed phosphorylation (p≤0.05) during the infection: 390, 449, and 575 peptides for S. Enteritidis and 185, 470, and 442 for S. Heidelberg at 1.5, 3, and 7 hpi, respectively. Many pathways involved in immunity, signal transduction, cellular process, and metabolism were significantly altered, in some case differentially, during the infection by the two Salmonella strains. Particularly, effects on lysosome process, iNOS, CARD9, NLRP3, and MAPK pathway provide significant insight to the inter play between pathogens and chicken macrophage cells during the infection.

Prenatal stress affects viability, activation, and chemokine signaling in astroglial cultures

CXCL12/SDF-1α and CX3CL1/fractalkine are constitutively expressed in the brain, which indicates their significant functions. Emerging evidence highlights the role of astrocytes and the immune system in the pathophysiology of stress-related disorders. The aim of this study was to assess whether prenatal stress affects chemokine signaling, cell viability/activation, and the iNOS pathway in astroglial cultures. Our results showed that prenatal stress lowered astrocyte viability and simultaneously increased GFAP expression. Furthermore, CX3CL1 production and the CXCL12/CXCR4-7 axis were also altered by prenatal stress. Taken together, malfunctions caused by prenatal stress may adversely influence brain development, leading to long-term effects on adult brain function and behavior.

Indoleamine 2,3-dioxygenase and inducible nitric oxide synthase mediate immune tolerance induced by CTLA4Ig and anti-CD154 hematopoietic stem cell transplantation in a sensitized mouse model

Cytotoxic T-lymphocyte-associated protein 4 immunoglobulin (CTLA4Ig) and anti-cluster of differentiation 154 (anti-CD154) are able to block B7/CD28 and CD40/CD154 co-stimulatory signals in T cells. Additionally, they promote hematopoietic stem cell transplantation (HSCT) in sensitized recipients and are able to induce immune tolerance and complete hematopoietic reconstitution. Indoleamine 2, 3-dioxygenase (IDO) and nitric oxide (NO) have been implicated in T cell immune tolerance. The aim of the present report was to study the in vivo tolerogenic mechanisms by which CTLA4Ig and anti-CD154 induce transplantation survival in mice receiving HSCT. BALB/c mice were sensitized via splenocyte transfusion and pretreated with CTLA4Ig plus anti-CD154 on day-7. IDO and inducible nitric oxide synthase (iNOS) inhibitors were applied on days-7 to 0 and the mice were divided into 4 groups (n=10) and injected with IDO every other day. The mice were sacrificed on day 0, and splenocytes were separated to identify CD11c+ antigen-presenting cells, which were subsequently assessed for IDO expression and activity. The concentration of NO was tested using a nitrate reductase kit. Following the acceptance of allogeneic HSCT, mice were tested for homing and engraftment, as well as survival rate. Application of the IDO inhibitor increased the concentration of NO, whereas a decrease in NO resulted in increased IDO activity. Immune tolerance was abrogated in the presence of both IDO and iNOS inhibitors, whereas this effect was not observed with either compound alone. CTLA4Ig and anti-CD154 may induce immune tolerance by affecting the activity of IDO and iNOS. This tolerance was abrogated in the presence of both IDO and iNOS inhibitors. A cross-regulatory pathway was observed between the IDO and NO pathways, in which the inhibition of IDO stimulated the iNOS pathway and vice versa.

β-Patchoulene, isolated from patchouli oil, suppresses inflammatory mediators in LPS-stimulated RAW264.7 macrophages

β-Patchoulene (β-PAE) is a tricyclic sesquiterpene isolated from patchouli oil. According to our previous study, β-PAE has anti-inflammatory activity in vivo; however, its anti-inflammatory response still remains unconfirmed in vitro. Therefore, this study is committed to demonstrate the anti-inflammatory effect of β-PAE on lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. According to our results, pre-treatment with β-PAE significantly decreased the protein and messenger RNA (mRNA) levels of pro-inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-1β while increased the expressions of anti-inflammatory cytokines like IL-10 in a dose-dependent manner. In addition, real-time polymerase chain reaction (PCR) also revealed that β-PAE could interrupt the mRNA expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and thus decreased the levels of nitric oxide (NO) and prostaglandin E2 (PGE2) in LPS-stimulated RAW264.7 macrophages. In conclusion, these results indicated that β-PAE exerted potent anti-inflammatory activity by maintaining the balance between pro- and anti-inflammatory cytokines as well as suppressing iNOS and COX-2 signaling pathways.

Magnesium isoglycyrrhizinate protects against renal‑ischemia‑reperfusion injury in a rat model via anti‑inflammation, anti‑oxidation and anti‑apoptosis.

The present study aimed to investigate whether magnesium isoglycyrrhizinate protects against renal ischemia‑reperfusion injury (RIRI), and to verify the underlying mechanisms. An RIRI rat model was induced by removing the right kidney, and exposing and clamping the left kidney. RIRI model rats were administered 30mg/kg magnesium isoglycyrrhizinate for 3days. Blood urea nitrogen (BUN) and serum creatinine levels in the blood of RIRI model rat were examined, compared with sham‑operated controls. Magnesium isoglycyrrhizinate suppressed the activities of tumor necrosis factor‑α, interleukin (IL)‑1β, IL‑6, superoxide dismutase, glutathione peroxidase, inducible nitric oxide synthase (iNOS) and caspase‑3 in RIRI model rats. Renal iNOS, matrix metalloproteinase (MMP)‑2, phosphorylated‑signal transducers and activators of transcription 3 (STAT3) and intercellular adhesion molecule‑1 (ICAM‑1) protein expression levels were suppressed by magnesium isoglycyrrhizinate treatment in RIRI model rats. These findings suggested that magnesium isoglycyrrhizinate protects RIRI via anti‑inflammatory, ‑oxidative and ‑apoptotic mechanisms in an RIRI rat model. These results implicate magnesium isoglycyrrhizinate pretreatment as a potential approach to protect against RIRI via suppression of the iNOS, ICAM‑1, MMP‑2 and STAT3 signaling pathways.

D-limonene exhibits anti-inflammatory and antioxidant properties in an ulcerative colitis rat model via regulation of iNOS, COX-2, PGE2 and ERK signaling pathways.

D-limonene has been demonstrated to have important immunomodulatory properties, including antitumor effects, and may alleviate asthma and allergies. In the present study, the anti‑inflammatory effects of D‑limonene were investigated in an ulcerative colitis (UC) rat model. Healthy male Sprague‑Dawley rats were randomly divided into control, untreated UC, and treatment with 50or100mg/kg D‑limonene UC groups. In UC rats, disease activity and colonic mucosa damage were significantly reduced by the anti‑inflammatory effects of D‑limonene, via suppression of matrix metalloproteinase (MMP)‑2 and‑9 gene expression. In addition, treatment with D‑limonene significantly increased antioxidant, inducible nitric oxide synthase (iNOS) and cyclooxygenase‑2 (COX‑2) protein expression levels in UC rats. A decrease in prostaglandin E2 (PGE2) production, transforming growth factor‑β (TGF‑β) gene expression and an increase phosphorylated‑extracellular signal regulated kinase (ERK)1/2 expression levelswere observed in UC rats treated with D‑limonene. In conclusion, D‑limonene reduced MMP‑2 and ‑9 mRNA expression levels via regulation of the iNOS, COX‑2, PGE2, TGF‑β and ERK1/2 signaling pathways in a UC rat model, indicating its potential antioxidant and anti‑inflammatory properties.

The effects of rotenone-induced toxicity via the NF-κB–iNOS pathway in rat liver

Rotenone has been used as a pesticide for many years, it is an environmental poison reported to cause neurological diseases. However, the effects of rotenone on the rat liver are unclear, as are the mechanisms of toxicity. In the present study, Sprague–Dawley (SD) rats were divided into five groups: control, dimethyl sulfoxide (DMSO), rotenone low-dose (1 mg/kg), rotenone mid-dose (2 mg/kg) and rotenone high-dose (4 mg/kg). The treatments were orally administered daily for 28 days, we assessed health status, mRNA expression levels of inflammatory factors, protein levels, nitric oxide (NO) content and histological changes. The results showed that body weight was significantly decreased in each rotenone group in a dose-dependent manner, compared with the control group. Rotenone significantly increased the mRNA levels of cyclooxygenase-2 (COX-2), nuclear factor kappaB (NF-κB), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS) and tumor necrosis factor (TNF-α) in each rotenone group compared with the control group, except iNOS and TNF-α mRNA expression in the low-dose group. The protein levels of COX-2 were significantly higher in each rotenone group compared with the control group, NF-κB protein expression were significantly higher in the rotenone mid and high-dose groups, but not in the low-dose group, compared with the control group, similar changes were observed in NO content. Additionally, histological analysis revealed that the most severe tissue damage occurred in the high-dose group. These results indicated that rotenone has toxic effect in rat liver relating to inflammatory factors. Our findings provide insight into the mechanisms of rotenone hepatotoxicity.

Aging related erectile dysfunction-potential mechanism to halt or delay its onset.

Erectile dysfunction (ED) will visit every man at some time in his life. The age at when that knock on the door is heard is totally dependent on one’s genetics as well as other extrinsic factors. Unlike guests who come for a visit and then leave, once ED shows up it tends to hang around forever. To add insult to injury, the longer ED hangs around, the worse it will get. It is estimated that by the time a man is in his 40’s, he has about a 40% chance of having some form of ED and this prevalence increases about 10% per decade thereafter. This suggests that the aging related process that leads to ED begins early in life. It turns out that the most common cause of ED, regardless of the patient’s age, is due to a problem with the vascular system of the penis. However, this specific aging related vascular problem is not caused by arterial disease but due to a dysfunction and/or loss of the corporal smooth muscle cells (SMC), the main constituent of the corporal sinusoids. As one gets older, these SMC continue to degrade and disappear. When approximately 15% of these cells have been impacted, it results in an inability of the corporal tissue to retain and/or prevent the blood from “leaking” out of the corporal sinusoids into the systemic veins. However, the corporal SMC themselves begin to combat this aging process by expressing the inducible nitric oxide synthase (iNOS) enzyme to make nitric oxide (NO) in an attempt to quench the high intracellular oxidative stress responsible for the SMC apoptosis. When this iNOS pathway is then pharmacologically upregulated, reversal of these aging related changes in the corpora with correction of the venous leakage is observed. Since we believe that aging related ED is pathologically the same disorder as essential hypertension, the development of a therapeutic regimen that can halt, delay or possibly reverse the cellular processes that lead to aging related ED should also be applicable to those patients diagnosed with essential hypertension.

Everolimus is better than rapamycin in attenuating neuroinflammation in kainic acid-induced seizures

BackgroundMicroglia is responsible for neuroinflammation, which may aggravate brain injury in diseases like epilepsy. Mammalian target of rapamycin (mTOR) kinase is related to microglial activation with subsequent neuroinflammation. In the present study, rapamycin and everolimus, both as mTOR inhibitors, were investigated in models of kainic acid (KA)-induced seizure and lipopolysaccharide (LPS)-induced neuroinflammation.MethodsIn vitro, we treated BV2 cells with KA and LPS. In vivo, KA was used to induce seizures on postnatal day 25 in B6.129P-Cx3cr1tm1Litt/J mice. Rapamycin and everolimus were evaluated in their modulation of neuroinflammation detected by real-time PCR, Western blotting, and immunostaining.ResultsEverolimus was significantly more effective than rapamycin in inhibiting iNOS and mTOR signaling pathways in both models of neuroinflammation (LPS) and seizure (KA). Everolimus significantly attenuated the mRNA expression of iNOS by LPS and nitrite production by KA and LPS than that by rapamycin. Only everolimus attenuated the mRNA expression of mTOR by LPS and KA treatment. In the present study, we also found that the modulation of mTOR under LPS and KA treatment was not mediated by Akt pathway but was primarily mediated by ERK phosphorylation, which was more significantly attenuated by everolimus. This inhibition of ERK phosphorylation and microglial activation in the hippocampus by everolimus was also confirmed in KA-treated mice.ConclusionsRapamycin and everolimus can block the activation of inflammation-related molecules and attenuated the microglial activation. Everolimus had better efficacy than rapamycin, possibly mediated by the inhibition of ERK phosphorylation. Taken together, mTOR inhibitor can be a potential pharmacological target of anti-inflammation and seizure treatment.

Patchoulene Epoxide Isolated from Patchouli Oil Suppresses Acute Inflammation through Inhibition of NF-κB and Downregulation of COX-2/iNOS.

According to the GC-MS analysis, compositional variation was observed between samples of patchouli oil, of which an unknown compound identified as patchoulene epoxide (PAO) was found only in the long-stored oil, whose biological activity still remains unknown. Therefore, the present study aimed to evaluate the potential anti-inflammatory activity with three in vivo inflammatory models: xylene-induced ear edema, acetic acid-induced vascular permeability, and carrageenan-induced paw edema. Further investigation into its underlying mechanism on carrageenan-induced paw edema was conducted. Results demonstrated that PAO significantly inhibited the ear edema induced by xylene, lowered vascular permeability induced by acetic acid and decreased the paw edema induced by carrageenan. Moreover, PAO markedly decreased levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), prostaglandin E2 (PGE2), and nitric oxide (NO), but increased levels of interleukin-4 (IL-4) and interleukin-10 (IL-10). PAO was also shown to significantly downregulate the protein and mRNA expressions of cyclooxygenase-2 (COX-2) and inducible nitric-oxide synthase (iNOS). Western blot analysis revealed that PAO remarkably inhibited p50 and p65 translocation from the cytosol to the nucleus by suppressing IKKβ and IκBα phosphorylation. In conclusion, PAO exhibited potent anti-inflammatory activity probably by suppressing the activation of iNOS, COX-2 and NF-κB signaling pathways.

Down-regulation of common NFκB-iNOS pathway by chronic Thalidomide treatment improves Hepatopulmonary Syndrome and Muscle Wasting in rats with Biliary Cirrhosis.

Thalidomide can modulate the TNFα-NFκB and iNOS pathway, which involve in the pathogenesis of hepatopulmonary syndrome (HPS) and muscle wasting in cirrhosis. In bile duct ligated-cirrhotic rats, the increased circulating CD16+ (inflammatory) monocytes and its intracellular TNFα, NFκB, monocyte chemotactic protein (MCP-1) and iNOS levels were associated with increased circulating MCP-1/soluable intercellular cell adehesion molecule-1 (sICAM-1), pulmonary TNFα/NOx, up-regulated M1 polarization, exacerbated angiogenesis and hypoxemia (increased AaPO2) in bronchoalveolar lavage (BAL) fluid and pulmonary homogenates. Meanwhile, a significant correlation was noted between circulating CD16+ monocyte/M1 (%) macrophages in BAL; M1 (%) macrophages in BAL/pulmonary iNOS mRNA expression; pulmonary iNOS mRNA expression/relative pulmonary MVD; pulmonary NOx level/AaPO2; circulating CD16+ monocyte/M1 (%) macrophages in muscle homogenates; 3-nitrotyrosine (representative of peroxynitrite) concentration/M1 (%) macrophages in muscle homogenates. The in vitro data demonstrated an iNOS-dependent inhibition of thalidomide on the TNFα-stimulated angiogenesis and myogenesis in human pulmonary artery endothelial cells (HPAECs) and C2C12 myoblasts. Significantly, the co-culture of CD16+ monocyte from different rats with HPAECs, or co-culture of supernatant of above mixed cultures with HPAECs or C2C12 myoblasts stimulated angiogenesis, migration and myogenesis. Our findings demonstrate that TNFα inhibitor thalidomide markedly diminishes the severity of experimental HPS and muscle wasting by down-regulation of common peripheral and local NFκB-iNOS pathway.

Macrophages activate iNOS signaling in adventitial fibroblasts and contribute to adventitia fibrosis

A large amount of NO is generated through the inducible nitric oxide synthase (iNOS) pathway from the vascular adventitia in various vascular diseases. However, it is currently not fully understood how the iNOS signaling pathway is activated. In the present study, this question was addressed in the context of adventitial cellular interactions. A rat model of acute hypertension in the contralateral carotid arteries was established through transverse aortic constriction (TAC) surgery. In this model, activated macrophages were found surrounded by a large quantity of iNOS-expressing adventitial fibroblasts (AFs), suggesting a possible causal relationship between macrophages and iNOS activation of the neighboring AFs. In an in vitro model, a macrophage-like cell line RAW 264.7 was first activated by LPS treatment. The supernatant was then harvested and applied to treat primary rat AFs. iNOS in AFs was activated robustly by the supernatant treatment but not by LPS itself. Treating AFs with interleukin-1β (IL-1β) also activated iNOS signaling, suggesting that the IL-1β pathway might be a possible mediator. As a consequence of the iNOS activation, total protein nitration and S-nitrosylation significantly increased in those AFs. Additionally, increased deposition of type I and type III collagens was observed in both in vitro and in vivo models. The collagen deposition was partially restored by an iNOS inhibitor, 1400 W. These findings highlight the importance of iNOS signaling during vascular inflammation, and advance our understanding of its activation through a cellular interaction perspective.

Obesity, Inflammation, and Exercise Training: Relative Contribution of iNOS and eNOS in the Modulation of Vascular Function in the Mouse Aorta.

Background: The understanding of obsesity-related vascular dysfunction remains controversial mainly because of the diseases associated with vascular injury. Exercise training is known to prevent vascular dysfunction. Using an obesity model without comorbidities, we aimed at investigating the underlying mechanism of vascular dysfunction and how exercise interferes with this process.Methods: High-sugar diet was used to induce obesity in mice. Exercise training was performed 5 days/week. Body weight, energy intake, and adipose tissues were assessed; blood metabolic and hormonal parameters were determined; and serum TNFα was measured. Blood pressure and heart rate were assessed by plethysmography. Changes in aortic isometric tension were recorded on myograph. Western blot was used to analyze protein expression. Nitric oxide (NO) was evaluated using fluorescence microscopy. Antisense oligodeoxynucleotides were used for inducible nitric oxide synthase isoform (iNOS) knockdown.Results: Body weight, fat mass, total cholesterol, low-density lipoprotein cholesterol fraction, insulin, and leptin were higher in the sedentary obese group (SD) than in the sedentary control animals (SS). Exercise training prevented these changes. No difference in glucose tolerance, insulin sensitivity, blood pressure, and heart rate was found. Decreased vascular relaxation and reduced endothelial nitric oxide synthase (eNOS) functioning in the SD group were prevented by exercise. Contractile response to phenylephrine was decreased in the aortas of the wild SD mice, compared with that of the SS group; however, no alteration was noted in the SD iNOS−/− animals. The decreased contractility was endothelium-dependent, and was reverted by iNOS inhibition or iNOS silencing. The aortas from the SD group showed increased basal NO production, serum TNFα, TNF receptor-1, and phospho-IκB. Exercise training attenuated iNOS-dependent reduction in contractile response in high-sugar diet–fed animals, decreased iNOS expression, and increased eNOS expression.Conclusion: Obesity caused endothelium dysfunction, TNFα, and iNOS pathway up-regulation, decreasing vascular contractility in the obese animals. Exercise training was an effective therapy to control iNOS-dependent NO production and to preserve endothelial function in obese individuals.

Allicin protects traumatic spinal cord injury through regulating the HSP70/Akt/iNOS pathway in mice.

Allicin is a major component of garlic, extracted as an oily liquid. The present study was designed to investigate the beneficial effects of allicin on traumatic spinal cord injury (TSCI) in mice, and whether the effects are mediated via regulation of the heat shock protein 70 (HSP70), v-akt murine thymoma viral oncogene homolog 1 (Akt) and inducible nitric oxide synthase (iNOS) pathways. Adult BALB/c mice (30–40 g) received a laminectomy at the T9 vertebral level as a model of TSCI. In the present study, treatment of the TSCI mice with allicin significantly increased their Basso, Beattie and Bresnahan (BBB) scores (P<0.01) and reduced the spinal cord water content (P<0.01). This protective effect was associated with the inhibition of oxidative stress and inflammatory responses in TSCI mice. Western blot analysis demonstrated that allicin increased the protein levels of HSP70, increased the phosphorylation of Akt and reduced the iNOS protein expression levels in TSCI mice. Additionally, treatment with allicin significantly reduced the levels of ROS and enhanced the NADH levels in TSCI mice. Collectively, these data demonstrate that the effects of allicin on TSCI are mediated via regulation of the HSP70, Akt and iNOS pathways in mice.

Immunohistochemical study of inducible nitric oxide synthase and tumour necrosis factor alpha response in turbot (Scophthalmus maximus) experimentally infected with Aeromonas salmonicida subsp. salmonicida.

Aeromonas salmonicida subsp. salmonicida represents one of the major threats in aquaculture, especially in salmonid fish and turbot farming. In order to fight bacterial infections, fish have an immune system composed by innate and specific cellular and humoral elements analogous to those present in mammals. However, innate immunity plays a primordial role against bacterial infections in teleost fish. Among these non-specific mechanisms, the production of nitric oxide (NO) by inducible nitric oxide synthase (iNOS) pathway and the tumour necrosis factor-alpha (TNFα) produced by mononuclear phagocytes, are two of the main immune effectors to eliminate bacterial pathogens. In this study, the distribution and kinetic of iNOS and TNFα-producing cells of kidney and spleen of turbot experimentally inoculated with A.salmonicida was assessed by immunohistochemistry. In control and challenged fish, individual iNOS(+) and TNFα(+) cells, showing a similar pattern of distribution, were detected. In challenged fish, the number of immunoreactive cells was significantly increased in the evaluated organs, as well as the melanomacrophage centres showed variable positivity for both antigens. These results indicate that A.salmonicida induced an immune response in challenged turbot, which involved the increase of the activity of iNOS and TNFα in the leukocytic population from kidney and spleen.

MODE OF ACTION STUDY OF NSP01-E, A POTENTIAL TREATMENT FOR ALZHEIMER’S DISEASE, IN PROTECTION AGAINST AMYLOID B PEPTIDE (AB) INJURY

We studied an Asiatic plant traditionally used since decades as tonic and a wellbeing medicine. Thanks to our GREEN extraction platform (Greenform©) we realized and standardized a simplified plant exact (SPE) called NSP01-E well analytically characterized. This standardized specific SPE was tested on primary cell-based pharmacological models of neurodegenerative diseases. In this study, we used an in vitro model of AβO injuries (on primary cortical neurons) and we decided to carefully dissect the mode of action of NSP01-E. Using several antagonists acting on TrK/MEK, Pi3K/AKT, iNOS, PPARγ pathways and apoptosis pathway. For analysing the activation of different pathway, we observed the level of protein with an automatically western blot. We observed a strong neuro-protective and restorative effect against the Aβ induced injuries on cortical primary neurons, a well described Alzheimer’s disease (AD) models. We observed that NSP01-E partly acted though RAS/RAF, and inducible iNOS pathway. Additionally, the PPARγ survival pathway seemed to be activated. But more interestingly, AKT pathway and anti-apoptosis effector BCl2 were essential to its neuroprotective effect. Western blot analyses were also performed and confirmed the activation of these pro-survival pathways (ERK1/2) as well as the reduction of mitochondrial stress effectors (Cytochrome C…). These findings demonstrate that NSP01-E counters Aβ-dependent neuronal cell death and may represent a therapeutic intervention to limit neuronal death in Alzheimer disease.

Trans-Cinnamaldehyde, An Essential Oil in Cinnamon Powder, Ameliorates Cerebral Ischemia-Induced Brain Injury via Inhibition of Neuroinflammation Through Attenuation of iNOS, COX-2 Expression and NFκ-B Signaling Pathway.

Trans-cinnamaldehyde (TCA), an essential oil in cinnamon powder, may have beneficial effects as a treatment for stroke which is the second leading cause of death worldwide. Post-ischemic inflammation induces neuronal cell damage after stroke, and activation of microglia, in particular, has been thought as the main contributor of proinflammatory and neurotoxic factors. The purpose of this study was to investigate the neuroprotective effects of TCA in an animal model of ischemia/reperfusion (I/R)-induced brain injury and the neuroprotective mechanism was verified in LPS-induced inflammation of BV-2 microglial cells. Our results showed that TCA (10-30mg/kg, p.o.) significantly reduced the infarction area, neurological deficit score and decreased iNOS and COX-2 protein expression level in I/R-induced injury brain tissue. It inhibited 0.5µg/ml LPS-induced NO production in BV-2 microglial cells without affecting cell viability, reduced protein expression of iNOS and COX-2, and attenuated inhibition of p53 protein. TCA also suppressed the effects of LPS-induced nuclear translocation of NF-κB p65 and p50 and increased cytosolic IκBα. It also reduced LPS-induced mRNA expression of iNOS, COX-2, and TNFα. We concluded that TCA has a potential neuroprotective effect to against the ischemic stroke, which may be via the inhibition of neuroinflammation through attenuating iNOS, COX-2 expression and NF-κB signaling pathway.

In situ plasma fabrication of ceramic‐like structure on polymeric implant with enhanced surface hardness, cytocompatibility and antibacterial capability

Polymeric materials are commonly found in orthopedic implants due to their unique mechanical properties and biocompatibility but the poor surface hardness and bacterial infection hamper many biomedical applications. In this study, a ceramic-like surface structure doped with silver is produced by successive plasma implantation of silicon (Si) and silver (Ag) into the polyamine 66 (PA66) substrate. Not only the surface hardness and elastic modulus are greatly enhanced due to the partial surface carbonization and the ceramic-like structure produced by the reaction between energetic Si and the carbon chain of PA66, but also the antibacterial activity is improved because of the combined effects rendered by Ag and SiC structure. Furthermore, the modified materials which exhibit good cytocompatibility upregulate bone-related genes and proteins expressions of the contacted bone mesenchymal stem cells (BMSCs). For the first time, it explores out that BMSCs osteogenesis on the antibacterial ceramic-like structure is mediated via the iNOS and nNOS signal pathways. The results reveal that in situ plasma fabrication of an antibacterial ceramic-like structure can endow PA66 with excellent surface hardness, cytocompatibility, as well as antibacterial capability.

Anti-apoptotic and anti-inflammatory effects of naringin on cisplatin-induced renal injury in the rat

Nephrotoxicity is a common complication of cisplatin chemotherapy and thus limits the use of cisplatin in clinic. Naringin, a natural flavonoid, plays important roles in inflammation and apoptosis in some inflammatory diseases; however, its roles in cisplatin-induced nephrotoxicity remain unclear. In this study, we first assessed the involvement of ROS overproduction and inflammation in cisplatin-induced nephrotoxicity in aged rats, and then we investigated the changes of renal function, histological injury, inflammatory response, and apoptosis in renal tissues after treatment with naringin (20, 50 or 100 mg/kg body weight). Cisplatin resulted in an increase of renal markers, lipid peroxidation, protein and DNA oxidation, and ROS formation. Renal tumor necrosis factor-α (TNF-α) and nitrite levels were also elevated. Expressions of nuclear factor-kappa B (NF-κB), inductible nitric oxide synthase (iNOS), caspase-3 and p53 were up-regulated in renal tissues of Cis-treated rats compared with the normal control group. Histopathological changes were also observed in cisplatin group. Adminstration of naringin at different doses (25, 50 and 100 mg/kg) was able to protect against the deterioration in kidney function, abrogate the decline in antioxidant enzyme activities and suppressed the increase in TBARS, nitrite and TNF-α concentrations. Moreover, naringin inhibited NF-κB and iNOS pathways, caspase-3 and p53 activation and improved the histological changes induced by cisplatin.In conclusion, our studies suggest that oxidative stress and inflammation might play important roles in the development of cisplatin-induced nephrotoxicity and naringin might become an effective therapeutic strategy for this disease.

Abstract 15427: Oxidative Stress From Altered Cardiac β3-adrenergic Receptor-activated Inducible Nitric Oxide Synthase/NO Pathways Promotes Cardiac Aging

Background: There is substantial evidence supporting oxidative stress as an important mechanism to age-related cardiac dysfunction and altered β 3 -adrenergic receptor (AR)-activated nitric oxide synthase (NOS) pathway contributing to this process. However, the NOS isoforms involved are controversial. The mechanism of how β 3 -AR stimulation impacts ROS, SERCA2a, and cardiac function in cardiac aging (CA) is unclear. We tested the hypothesis that oxidative stress from up-regulation of cardiac β 3 -AR-activated iNOS uncoupling promotes CA. Methods: We compared myocyte subtypes of β-ARs, three NOS, peroxynitrite (NT), NADPH, GTPCH and SERCA2a expressions and myocyte contractile and [Ca 2+ ] i transient ([Ca 2+ ] iT ) responses to β-AR stimulation with isoproterenol (ISO, 10 -8 M). LV myocytes were isolated from 2 young (Y) (~4 to 6 m) and 2 aged (A) (~28 to 31 m) groups (5/group) of wild-type (WT) and β 3 -AR knockout (β 3 KO) mice. Results: Compared to YWT, AWT myocytes had significantly increased protein levels of β 3 -AR (0.31 vs 0.16) and iNOS (0.48 vs 0.21) accompanied with increased oxidative stress indicated by significant increases in NT formation (increased 102%) and NADPH (P67 -phox , 35% and p22 -phox , 28%), but decreased GTPCH expression (41%, 0.48 vs 0.82) and activity. AWT myocytes had significantly decreased β 1 -AR protein (0.38 vs 0.59), SERCA2a (0.26 vs 0.72) and the ratios of SERCA2a/PLB. These changes were associated with reduced basal cell contraction (dL/dt max, 76.4 vs 122.8 μm/s), relaxation (dR/dt max , 58.9 vs 94.3 μm/s), and [Ca 2+ ] iT (0.16 vs 0.24) accompanied by diminished β-AR-stimulated positive inotropic response. Importantly, in contrast to age-caused changes in AWT, versus Yβ 3 KO, in Aβ 3 KO myocytes, there were no significant alterations in iNOS (0.19 vs 0.16), NT (increased 9%), β 1 -AR (0.60 vs 0.62), and SERCA2a (1.2 vs 1.4). Aβ 3 KO myocytes had normal cell contraction and relaxation with preserved ISO-stimulated positive inotropic response. Conclusions: Myocardial aging is associated with up-regulation of cardiac β 3 -AR-activated iNOS pathway, which leads to imbalance in NO/superoxide production, favoring subsequent iNOS uncoupling and directly contributes to age-associated deficits in LV myocyte function and [Ca 2+ ] i regulation.