Heme oxygenase 1 plays role of neuron-protection by regulating Nrf2-ARE signaling post intracerebral hemorrhage

The NF-E2 related factor 2 (Nrf2) could be activated in intracerebral hemorrhage (ICH), and trigger the expression of ARE regulated heme oxygenase 1 (HO-1) subsequently. This study aims to explore neuroprotection of HO-1 protein in regulating the Nrf2-ARE signaling pathway in ICH. In this study, the femoral artery injection method was used to establish the ICH model. The zinc porphyrin-9 (ZPP-IX) was used to inhibit the HO-1 expression in ICH rats. The ICH rats were randomly divided into 3 groups, ICH group, ZPP-IX (10 mg/kg) + ICH group and DMSO (10 mg/kg) + ICH group. Neurological scores were evaluated for the 3 groups. Double immunofluorescence staining method was employed to observe the co-expression of HO-1, Nrf2, NF-κB and TNF-α and CD11b in glia cells. Western blot and RT-PCR assay were used to detect the total Nrf2, binding Nrf2, HO-1, NF-κB and TNF-α expression. The results indicated that ZPP-IX could aggravate the neurological dyafunstions of ICH rats. The HO-1 level in ZPP-IX group was significantly decreased compared to the ICH group (P < 0.05). The binding-Nrf2 protein was significantly increased in ZPP-IX group compared to ICH group (P < 0.05). The NF-κB and TNF-α level were significantly increased in ZPP-IX group compared to ICH group (P < 0.05). The ZPP-IX significantly inhibited the HO-1 and Nrf2, and enhanced NF-κB and TNF-α co-expressing with the CD11b compared to the ICH group (P < 0.05). In conclusion, HO-1 protein regulates the Nrf2-ARE pathway in ICH model by inhibiting the Nrf2 entering nucleus and activating the NF-κB and TNF-α expression.     

Effects of that ATRA inhibits Nrf2-ARE pathway on glial cells activation after intracerebral hemorrhage

Previous studies indicate that the Nrf2-ARE signaling pathway plays a neruo-protective role in glia cell, however, the mechanism was also elusive. This study aims to explore the inhibitive function of all-trans-retinoic (ATRA) on Nrf2-ARE pathway in intracerebral hemorrhage (ICH), and investigate the mechanism. In this study, the femoral artery injection method was employed to establish ICH model. The model rats were randomly divided into four groups, including Sham group, ICH group, ATRA group and DMSO group. The neurological scores were evaluated for the four groups at different time points. Hematoxylin-Eosin staining was used to stain the CD11b positive glia cells. Double immunofluorescence staining method was utilized to observe the co-expression of HO-1, NF-κB, Nrf2 and TNF-α and CD11b marker in glia cells. Western blot assay was used to detect the Nrf2 protein (total and binding Nrf2), HO-1, NF-κB and TNF-α proteins in every group. The results indicated that neurologiclal scores were significantly decreased in ATRA group compared to ICH gorup (P < 0.05). The glia cells were significantly activated and accumulated in ICH rats. ATRA significantly decreased co-expression of Nrf2, HO-1 and CD11b, and increased co-expression of NF-κB, TNF-α and CD11b of glia cells. ATRA significantly decreased total Nrf2 expression and increased binding Nrf2 expression in ATRA group compared to ICH group (P < 0.05). ATRA decreased anti-oxygen protein Nrf2 and HO-1, and increases inflammatory factors NF-κB and TNF-α. In conclusion, the application of ATRA could inhibit the neuro-protective function effectively by blocking the Nrf2-ARE pathway in glia cells.     

Per a 10 protease activity modulates CD40 expression on dendritic cell surface by nuclear factor-kappaB pathway

Serine protease activity of Per a 10 from Periplaneta americana modulates dendritic cell (DC) functions by a mechanism(s) that remains unclear. In the present study, Per a 10 protease activity on CD40 expression and downstream signalling was evaluated in DCs. Monocyte-derived DCs from cockroach-allergic patients were treated with proteolytically active/heat-inactivated Per a 10. Stimulation with active Per a 10 demonstrated low CD40 expression on DCs surface (P < 0·05), while enhanced soluble CD40 level in the culture supernatant (P < 0·05) compared to the heat-inactivated Per a 10, suggesting cleavage of CD40. Per a 10 activity reduced the interleukin (IL)-12 and interferon (IFN)-γ secretion by DCs (P < 0·05) compared to heat-inactivated Per a 10, indicating that low CD40 expression is associated with low levels of IL-12 secretion. Active Per a 10 stimulation caused low nuclear factor-kappa B (NF-κB) activation in DCs compared to heat-inactivated Per a 10. Inhibition of the NF-κB pathway suppressed the CD40 expression and IL-12 secretion by DCs, further indicating that NF-κB is required for CD40 up-regulation. CD40 expression activated the tumour necrosis factor (TNF) receptor-associated factor 6 (TRAF6), thereby suggesting its involvement in NF-κB activation. Protease activity of Per a 10 induced p38 mitogen-activated protein kinase (MAPK) activation that showed no significant effect on CD40 expression by DCs. However, inhibiting p38 MAPK or NF-κB suppressed the secretion of IL-12, IFN-γ, IL-6 and TNF-α by DCs. Such DCs further reduced the secretion of IL-4, IL-6, IL-12 and TNF-α by CD4⁺ T cells. In conclusion, protease activity of Per a 10 reduces CD40 expression on DCs. CD40 down-regulation leads to low NF-κB levels, thereby modulating DC-mediated immune responses.     

Electroacupuncture modulated the inflammatory reaction in MCAO rats via inhibiting the TLR4/NF-κB signaling pathway in microglia

In this study, we aim to investigate the effects of electroacupuncture on the TLR4/NF-κB signaling pathway in microglia. Male Wistar rat of SPF grade (weighing 200±20 g) were randomly divided into (i): sham control group, which was subjected to sham operation (ii) vehicle group, which underwent the occlusion of middle cerebral artery; (iii-v): acupuncture groups, which were subjected to the occlusion of middle cerebral artery and treated with acupuncture on the Neiguan acupoint (P6), Quchi acupoint (LI11), and Diji acupoint (SP8), respectively. HE staining was performed to detect the necrotic rate of neurons. Mediators of inflammation were measured using ELISA. Immunofluorescence was performed to measure the expression of TLR4, HMGB1, TRAF6, IKKβ and NF-κB p65 in microglia. Severe decrease was noticed in the neurological score, necrotic rates of neuron, expression of IL-1β, IL-6, TLR4, HMGB1, TRAF6, IKKβ and NF-κB p65 in microglia. Compared with the vehicle group, significant decrease was revealed in the neurological score, necrotic rate, IL-1β, TLR4, TRAF6, IKKβ and NF-κB p65 in Neiguan group and Quchi group, respectively. In addition, remarkable decrease was observed in the expression of TNF-α and IL-6 in Quchi group. Compared with the Diji group, the necrotic rate of neurons in hippocampus region was significantly decreased in the Quchi group (P < 0.05). In Neiguan group, the expression of TLR4 and IKKβ was significantly attenuated (P < 0.05). The expression of TRAF6 was remarkably decreased in the Neiguan group and Quchi group, respectively. Electroacupuncture on Neiguan and Quchi could improve the neurological injury, attenuate the inflammation, and inhibit the activity of TLR4/NF-κB signaling pathway in microglia.     

Angiotensin Receptor Blockers and Statins Could Alleviate Atrial Fibrosis via Regulating Platelet-Derived Growth Factor/Rac1 /Nuclear Factor-Kappa B Axis

Aims: To investigate whether the administration of renin-angiotensin system (RAS) inhibitors and statins could alleviate atrial fibrosis via platelet-derived growth factor (PDGF)/Rac1 /nuclear factor-kappa B (NF-κB) axis.Methods and Results: In human left atrium, the degree of atrial fibrosis, as well as the expression levels of PDGF, Rac1 and NF-κB increased 1.5 to 2.9 folds in patients with atrial fibrillation compared to that with sinus rhythm, (P<0.0001). There were strongly positive correlations between angiotensin II (Ang II) or procollagen type III-alpha-1 (COL3A1) with PDGF, Rac1, NF-κB, and among PDGF, Rac1 and NF-κB (all P<0.05). At 3 weeks after the transverse aorta constriction (TAC) operation in rat model and with intervention of irbesartan or/and simvastatin, the collagen volume fraction (CVF) and atrial natriuretic peptide (ANP) values respectively increased 6-folds and 3.5-folds in the TAC group compared to SHAM group (P<0.0001), but these levels decreased by 16% to 63% with following drug intervention (all P<0.0001), the combined treatment was the lowest. Accordingly, the expression levels of PDGF (3-folds), Rac1 (1.6-folds), NF-κB (7-folds) and AngII (12-folds) significantly increased in the TAC group compared to the SHAM group, and these levels were also reduced by 25% to 64% with following drug intervention. The highest reduction could be seen after treatment with irbesartan and simvastatin in combination (all P<0.001).There were strongly positive correlations between AngII or CVF with PDGF, Rac1, NF-κB, and among PDGF, Rac1 and NF-κB (all P<0.05).Conclusions: Irbesartan or/and simvastatin can improve atrial fibrosis by regulating PDGF/Rac1/NF-κB axis.     

Pterostilbene impact on retinal endothelial cells under high glucose environment

Diabetic retinopathy (DR) has complicated pathogenic factors. Studies showed that DR belongs to chronic inflammatory disease, and retinal endothelial cells oxidation by free radicals is one of its mechanisms. Pterostilbene, as the homologous derivative of resveratrol, has obvious antioxidant effect. Its influence on the DR has not been studied. This study intended to investigate the effect and mechanism of pterostilbene on human retinal endothelial cells (hRECs) under high glucose environment to illustrate pterostilbene impact on DR and provide basis for DR clinical treatment. hRECs cultured in high glucose environment were treated by 1.0 mmol/L pterostilbene. MTT assay was applied to test cell proliferation. ELISA was used to detect inflammatory factor TNF-α and IL-1β content. Real time PCR and Western blot were performed to examine NF-κB mRNA and protein expression. ROS and SOD activities were analyzed. Under high glucose environment, hRECs proliferation increased, TNF-α and IL-1β expression elevated, and NF-κB protein level upregulated significantly. On the other side, ROS production increased and SOD activity decreased obviously (P < 0.05). Pterostilbene can suppress hRECs over proliferation, decrease TNF-α and IL-1β, inhibit NF-κB protein expression, reduce ROS production, and increase SOD activity markedly compared with high glucose group (P < 0.05). Pterostilbene may delay DR progress through alleviating inflammation and antioxidation to suppress hRECs over proliferation.     

Performance of the QuantiFERON-Cytomegalovirus (CMV) Assay for Detection and Estimation of the Magnitude and Functionality of the CMV-Specific Gamma Interferon-Producing CD8+ T-Cell Response in Allogeneic Stem Cell Transplant Recipients

The performance of the QuantiFERON-cytomegalovirus (CMV) assay was compared to that of a flow cytometry intracellular cytokine staining (ICS) method for the detection of CMV-specific gamma interferon (IFN-γ)-producing CD8⁺ T-cell responses in allogeneic stem cell transplant (allo-SCT) recipients and for estimations of their magnitude and functionality. A total of 90 whole-blood specimens from 23 allo-SCT recipients was analyzed by both methods. Overall, the percentage of specimens that yielded concordant results by both methods was 68.8% (κ = 0.691; 95% confidence interval [CI], 0.548 to 0.835), and the sensitivity of the QuantiFERON-CMV assay for the detection of positive IFN-γ T-cell responses (>0.2 IU/ml), taking the ICS method as the reference, was 76.3%. The magnitude of IFN-γ-producing CD8⁺ T-cell responses to CMV-specific peptides measured with the QuantiFERON-CMV assay correlated significantly (σ = 0.695; P = <0.001) with that of the total IFN-γ-producing CD8⁺ T cells and dual-functional (IFN-γ/tumor necrosis factor alpha [TNF-α] [σ = 0.652; P = <0.001] and IFN-γ/CD107a [σ = 0.690; P = <0.001]) and trifunctional (IFN-γ/TNF-α/CD107a [σ = 0.679; P = >0.001]) CMV-specific CD8⁺ T-cell responses, as quantitated by ICS. In summary, the data indicated that the QuantiFERON-CMV assay is less sensitive than the ICS method for the detection of CMV-specific IFN-γ-producing CD8⁺ T-cell responses in the allo-SCT setting. Nevertheless, it allowed the estimation of the total and polyfunctional CMV-specific IFN-γ-producing CD8⁺ T-cell responses in specimens that tested positive by both methods.     

P15, MDM2, NF-κB,and Bcl-2 expression in primary bone tumor and correlation with tumor formation and metastasis

Primary bone tumor is one of the most common malignant tumors in skeletal system. It seriously affected bone movement and development with unclear pathogenesis. In this paper, rabbit VX-2 malignant bone tumor model was applied to explore apoptotic genes P15, MDM2, NF-κB and Bcl-2 correlation with primary bone tumor occurrence and metastasis. 0.3 ml rabbit VX-2 tumor cell suspension (1×10⁶/ml) was injected to the marrow cavity of the right tibia condyle to establish the rabbit malignant bone tumor model, while equal amount of the saline was injected to the left tibia as control. Real-time PCR was applied to determine P15, MDM2, NF-κB and Bcl-2 expression level. Immunohistochemistry was performed to detect the abovementioned genes expression in lung, stomach, kidney and bladder. Compared with control, P15 expression level in the inoculation site surrounding tissues decreased obviously following the inoculate time elongation (P<0.05), while Bcl-2, MDM2 and NF-κB expression significantly increased (P<0.05). Bcl-2 showed significant correlation with MDM2 and NF-κB (P<0.05). At the 2, 4, 6 weeks, Bcl-2, MDM2 and NF-κB in lung, Bcl-2 in kidney, and Bcl-2 and MDM2 in bladder positively expressed (P<0.05), whereas P15 gene exhibited no significant positive expression in these tissues (P>0.05). P15, MDM2, NF-κB, and Bcl-2 genes expression levels can effectively reflect malignant bone tumor growth of rabbit tibia. MDM2, NF-κB and Bcl-2 genes involved in primary bone tumors metastasis directly. It has important clinical significance for early diagnosis and treatment of primary bone tumor.     

Protective effects of isorhamnetin on apoptosis and inflammation in TNF-α-induced HUVECs injury

Little is known about the role of isorhamnetin on endothelial cell apoptosis and inflammation when insulted by TNF-α injury. In our study, HUVECs were treated with TNF-α for 6 hours. HUVECs apoptosis were detected using flow cytometry. The expressions of ICAM-1, VCAM-1, E-selectin, NF-κB, AP-1 and eNOS were determined with western blotting or flow cytometry. The results showed TNF-α increased of apoptosis and the expression of ICAM-1, VCAM-1 and E-selectin in HUVECs, accompanied by significant augmentation of NF-κB and AP-1 expression. Pretreatment with isorhamnetin significantly reduced apoptosis in TNF-α-treated HUVECs. Moreover, isorhamnetin significantly attenuated TNF-α-induced upregulation of ICAM-1, VCAM-1, AP-1, E-selectin and NF-κB expression. Meanwhile, isorhamnetin also increased the expression of eNOS. So, isorhamnetin could suppress TNF-α-induced apoptosis and inflammation by blocking NF-κB and AP-1 signaling in HUVECs, which might be one of the underlying mechanisms for treatment of coronary heart disease.     

Hyperbaric oxygen intervention reduces secondary spinal cord injury in rats via regulation of HMGB1/TLR4/NF-κB signaling pathway

Background: To investigate whether hyperbaric oxygen (HBO) intervention affects the expressions of inflammatory cytokines, HMGB1/TLR4/NF-κB, and arrests secondary spinal cord injury (SCI). Methods: One hundred and twenty healthy adult SD rats were randomly divided into four groups: sham, sham + HBO, SCI, and SCI + HBO. Each group was then randomly divided into five subgroups of 6 rats each according to the following time points: 1, 2, 3, 7, and 14 d post injury. Functional recovery of the hindlimb was assessed by Basso, Beattie, and Bresnahan (BBB) scores at different time points after SCI. The expression of HMGB1, TLR4, and NF-κB in the spinal cord tissue was determined by fluorescence quantitative PCR, western blot, immunohistochemistry, and ELISA. Results: The gene expressions of TLR4, HMGB1, and NF-κB (P < 0.01) and the TLR4 protein expression were significantly high after SCI. HBO intervention significantly decreased all the four parameters at 3, 7, and 14 d post injury (P < 0.05). A significant positive correlation (P < 0.01) was observed between the following: HMGB1 mRNA, TLR4 mRNA and TLR4 protein; HMGB1 mRNA and NF-κB mRNA; and TLR4 protein and NF-κB mRNA. BBB score was negatively correlated with HMGB1, TLR4 protein and NF-κB levels. HBO intervention significantly improved the BBB scores at 7 and 14 d post injury (P < 0.05). Conclusions: Hyperbaric oxygen reduced the expressions of HMGB1, TLR4, and NF-κB and reduced secondary SCI as measured using BBB scores.     

Inflammation induced by increased frequency of intermittent hypoxia is attenuated by tempol administration

The levels of serum inflammatory cytokines and the activation of nuclear factor kappa B (NF-κB) and hypoxia inducible factor-1α (HIF-1α) in heart tissues in response to different frequencies of intermittent hypoxia (IH) and the antioxidant tempol were evaluated. Wistar rats (64 males, 200-220 g) were randomly divided into 6 experimental groups and 2 control groups. Four groups were exposed to IH 10, 20, 30, or 40 times/h. The other 2 experimental groups were challenged with IH (30 times/h) plus tempol, either beginning on day 0 (IH30T0) or on day 29 (IH30T29). After 6 weeks of challenge, serum levels of tumor necrosis factor (TNF)-α, intracellular adhesion molecule (ICAM)-1, and interleukin-10 were measured, and western blot analysis was used to detect NF-κB p65 and HIF-1α in myocardial tissues. Serum levels of TNF-α and ICAM-1 and myocardial expression of NF-κB p65 and HIF-1α were all significantly higher in IH rats than in controls (P<0.001). Increased IH frequency resulted in more significant changes. Administration of tempol in IH rats significantly reduced levels of TNF-α, ICAM-1, NF-κB and HIF-1α compared with the non-tempol-treated group (F=16.936, P<0.001). IH induced an inflammatory response in a frequency-dependent manner. Additionally, HIF-1α and NF-κB were increased following IH administration. Importantly, tempol treatment attenuated this effect.     

Raised anti-endothelial cell autoantibodies (AECA), but not anti-neutrophil cytoplasmic autoantibodies (ANCA), in recurrent oral ulceration: modulation of AECA binding by tumour necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ)

Recurrent oral ulceration (ROU) is a common oral mucosal condition of unknown etiology. However, there is evidence to suggest that vasculitis may play a role. Here we investigate the presence in ROU of two autoantibodies associated with vasculitis, AECA and ANCA. AECA target as yet unidentified antigens on the endothelial cell surface and have been identified in patients with vasculitic disorders and inflammatory conditions with a vasculitic component. ANCA target specific neutrophil-associated proteins and are detected in specific vasculitic and chronic inflammatory disorders. AECA and ANCA levels were studied in 20 ROU patients and 20 controls. IgG AECA to the endothelial cell line ECV 304 were detected in 19 ROU patients and four controls. Levels were significantly raised in ROU both to ECV 304 (P < 0.000 05) and to human umbilical vein endothelial cells (HUVEC) (P < 0.005). Although levels were highest during episodes of ulceration, they were also raised between episodes. Stimulation of endothelial cells with TNF-α significantly increased AECA binding of both ROU (P < 0.005) and control samples (P < 0.0001), while IFN-γ decreased binding (ROU P < 0.0001; controls P < 0.05). In contrast, ANCA were detected in only one patient and none of the controls. The presence of raised levels of AECA lends support to the hypothesis that a vasculitic process may underlie ROU. Moreover, these findings suggest that endothelial cell expression of AECA target antigens is increased by TNF-α and decreased by IFN-γ stimulation.     

Enhanced binding of lymphocytes from patients with multiple sclerosis to tumour necrosis factor-alpha (TNF-α)-treated endothelial monolayers: associations with clinical relapse and adhesion molecule expression

This study investigated the adherent properties and adhesion molecule expression of blood mononuclear cells (MNC) from a total of 84 patients with multiple sclerosis (MS). The MNC from MS patients were significantly more adherent than cells from normal healthy subjects to endothelial monolayers pretreated with 0.01 U/ml TNF-α (103% increase; P = 0.002), 0.1 U/ml TNF-α (80% increase; P< 0.01) and 1.0 U/ml TNF-α (41% increase; P< 0.02), and to endothelium pretreated with 10 U/ml IL-1β (44% increase; P< 0.05) and 100 U/ml interferon-gamma (IFN-γ) (100% increase; P< 0.05). This augmented adhesion was a property of the lymphocytes, in particular CD4⁺ cells, and was inversely related to the time of onset of clinical relapse. The percentage of lymphocytes bearing the adhesion molecules CD49d, CD29 and CD62L was increased in MS blood, but the level of CD29 and CD62L expression was reduced. We infer that circulating lymphocytes in MS are predisposed to cross endothelial barriers at sites where inflammation has already commenced.     

NF-kappaB activation during acute Helicobacter pylori infection in mice

Nuclear factor κB (NF-κB) plays a key regulatory role in host cell responses to Helicobacter pylori infection in humans. Although mice are routinely used as a model to study H. pylori pathogenesis, the role of NF-κB in murine cell responses to helicobacters has not been studied in detail. We thus investigated the abilities of different Helicobacter isolates to induce NF-κB-dependent responses in murine gastric epithelial cells (GECs) and in transgenic mice harboring an NF-κB-responsive lacZ reporter gene. H. pylori and Helicobacter felis strains up-regulated the synthesis in mouse GECs of the NF-κB-dependent chemokines KC (CXCL1) and MIP-2 (CXCL2). These responses were cag pathogenicity island (cagPAI) independent and could be abolished by pretreatment with a pharmacological inhibitor of NF-κB. Consistent with the in vitro data, experimental Helicobacter infection of transgenic mice resulted in increased numbers of GECs with nuclear β-galactosidase activity, which is indicative of specific NF-κB activation. The numbers of β-galactosidase-positive cells in mice were significantly increased at day 1 postinoculation with wild-type H. pylori strains harboring or not harboring a functional cagPAI, compared to naive animals (P = 0.007 and P = 0.04, respectively). Strikingly, however, no differences were observed in the levels of gastric NF-κB activation at day 1 postinoculation with H. felis or at day 30 or 135 postinoculation with H. pylori. This work demonstrates for the first time the induction of NF-κB activation within gastric mucosal cells during acute H. pylori infection. Furthermore, the data suggest that helicobacters may be able to regulate NF-κB signaling during chronic infection.     

An assessment of peripheral immunity in patients with sarcoidosis using measurements of serum vitamin D3, cytokines and soluble CD23

The aetiology of the peripheral anergy in sarcoidosis is unclear. To investigate this further we measured the serum levels of several factors important in different aspects of immune regulation to obtain a profile of those factors which promote and inhibit immune activation in sarcoidosis. Thirty-seven patients with sarcoidosis and 20 healthy controls of similar sex and age comprised the study group. Serum IL-10, interferon-gamma (IFN-γ), soluble CD23 (sCD23), IL-8, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-1β and tumour necrosis factor-alpha (TNF-α) were measured using in-house ELISAs. Vitamin D3 was measured using a radioreceptor assay. Serum levels of sCD23 and IL-10 were significantly elevated in patients with sarcoidosis relative to controls (median 13.9 versus 9.5 arbitrary units/ml, P < 0.01 for sCD23, and 9.6 versus 5.0 pg/ml, P < 0.04 for IL-10). Regardless of steroid therapy or disease activity, serum levels of IFN-γ, TNF-α, IL-1β, GM-CSF and IL-8 were no different in patients with sarcoidosis and controls. Vitamin D3 levels were significantly higher in patients with sarcoidosis versus normal controls (medians 78.0 versus 56.0, P < 0.001), active sarcoidosis (n = 20) versus inactive disease (n = 17) (medians 81.5 versus 66.0, P < 0.03) and active sarcoidosis versus controls (medians 81.5 versus 56.0, P < 0.0002). The levels were no different between patients with inactive sarcoidosis and controls. We suggest that IL-10 and vitamin D3 may contribute to the peripheral anergy in sarcoidosis. The elevated serum sCD23 suggests an increase in peripheral humoral immunity. Consistent with a quiescent peripheral immune system, factors capable of monocyte/macrophage activation (TNF-α, IFN-γ, GM-CSF and IL-8) were not elevated in the peripheral circulation.     

Myrtol ameliorates cartilage lesions in an osteoarthritis rat model

Purpose: The aim of this study is to evaluate the effects of myrtol standardized on cartilage lesions in osteoarthritis (OA) rats. Methods: Fifty-six healthy Sprague-Dawley rats were randomly divided into sham group (13 rats) and OA model group (43 rats) with interior meniscus excision. Then serum estradiol (E₂) and glycosaminoglycan (GAG) content in cartilage tissue were measured by radioimmunoassay and toluidine blue staining, respectively. After that, the model rats were randomly divided into low dose myrtol (LDM) group, middle dose myrtol (MDM) group and high dose myrtol (HDM) group (10 rats in each group) with treatment of 450 mg/kg, 300 mg/kg and 150 mg/kg myrtol, respectively. Then, Mankin scores were used to evaluate lesion extent of knee joint cartilage. Expression of tumor necrosis factor α (TNF-α), transforming growth factor β1 (TGF-β1), interleukin (IL)-6, Bax and Bcl-2 were investigated using PCR gel electrophoresis method. Results: Mankin cores were lower in sham group and myrtol group than in model group. There were statistically significant differences (P < 0.01) between sham group and model group in expression of TNF-α, TGF-β1, IL-6, Bax and Bcl-2 in the cartilage tissue. Myrtol significantly reduced the expression of TNF-α, IL-6 and Bax, and increased the expression of TGF-β1 and Bcl-2 in myrtol group, comparing with those in model group (P < 0.01). Conclusions: Myrtol could down-regulate the expression of TNF-α, IL-6 and Bax, and up-regulate the expression of TGF-β1 and Bcl-2. Myrtol standardized is a promising drug to ameliorate knee cartilage lesions in the OA rat model.     

Alterations in Leukocyte Function following Surgical Trauma: Differentiation of Distinct Reaction Types and Association with Tumor Necrosis Factor Gene Polymorphisms

Endotoxin-stimulated blood cytokine responses have been widely used to describe compromised host defense mechanisms after trauma. We investigated whether blood cytokine production after endotoxin stimulation is able to define distinct trauma-induced alteration patterns and whether alteration patterns are associated with tumor necrosis factor (TNF) gene polymorphisms. In 48 patients undergoing joint replacement, the levels of TNF alpha (TNF-α), interleukin 6 (IL-6), and IL-8 production in blood after endotoxin stimulation were measured preoperatively on the day of surgery and 24 h thereafter. Patients were genotyped for the TNF-α position −308 G/A polymorphism and the TNF-β NcoI polymorphism. Postoperative alterations, i.e., increases or decreases of cytokine levels (TNF-α versus IL-6, P = 0.013; TNF-α versus IL-8, P = 0.001; IL-6 versus IL-8, P = 0.007), and relative postoperative changes, i.e., percentages of preoperative cytokine levels (TNF-α versus IL-6, r_s = 0.491, P < 0.001; TNF-α versus IL-8, r_s = 0.591, P < 0.001; IL-6 versus IL-8, r_s = 0.474, P < 0.001 [where r_s is the Spearman rank correlation coefficient]), had significant positive correlations among the cytokines. Overall enhanced postoperative alteration patterns were found in 10 patients, attenuated patterns were found in 18 patients, and mixed patterns were found in 20 patients. Preoperative cytokine production levels differed significantly between these groups (those of the overall enhanced pattern group were less than those of the mixed pattern group, which were less than those of the overall attenuated pattern group). TNF polymorphisms were not associated with overall alteration patterns, but the A*TNFB1 haplotype was associated with a postoperative increase in TNF-α production (P = 0.042). Whole-blood cytokine responses to endotoxin define the following preexisting patterns in leukocyte function: low baseline production and overall enhanced alteration patterns after trauma (type 1), intermediate baseline production and mixed alteration patterns (type 2), and high baseline production and overall attenuated alteration patterns (type 3). TNF gene polymorphisms were associated with changes in TNF-α production but do not explain the overall reaction patterns of cytokine production after trauma. The clinical correlate of these newly defined reaction types remains to be determined.     

Expression of NF-κB and PTEN in primary epithelial ovarian carcinoma and the correlation with chemoresistance

The present study aims to investigate the relationship of NF-κB p65 and PTEN protein with chemotherapy resistance in ovarian cancer by measuring their expression in primary epithelial ovarian cancer, and to explore the correlation of the expression of these two proteins with ovarian carcinoma and their clinical significance. Ovarian cancer patients (n = 161) were divided into two groups: sensitive group (n = 82) and resistant group (n = 79). Expression of NF-κB p65 and PTEN protein in the ovarian cancer tissues was determined using immunohistochemistry to assess the relationship and correlation between the expression levels of these two proteins and chemotherapy resistance of ovarian carcinoma. The Cox model was used to analyze the independent risk factors associated with ovarian cancer prognosis. The expression of NF-κB p65 in the sensitive group (68.29%) was lower than that of the resistant group (94.94%). In contrast, the expression of PTEN protein in the sensitive group (50.00%) was higher than that of the resistant group (17.72%). Expression of NF-κB p65 was negatively correlated with that of PTEN protein in ovarian cancer tissue (rs = -0.246, P = 0.002). Expression of NF-κB p65 or PTEN protein and surgical stage of ovarian cancer were independent risk factors associated with chemoresistance (all P < 0.05). Low expression of PTEN and high expression of NF-κB are significant risk factors for chemotherapy resistance of ovarian cancer patients.     

Bacterial Invasion Is Not Required for Activation of NF-κB in Enterocytes

Pathogenic enteric microorganisms induce the NF-κB-dependent expression of proinflammatory genes in intestinal epithelial cells. The purpose of the present study was to clarify the contribution of microbial invasion to the degradation of the regulatory protein IκBα and the subsequent activation of NF-κB in cultured intestinal epithelial cells. Caco-2BBe cells were incubated with Salmonella dublin, Salmonella typhimurium, or a weakly invasive strain of E. coli. S. dublin and S. typhimurium (10⁷ organisms/ml) induced equivalent concentration-dependent gel mobility shifts of an NF-κB consensus sequence that was preceded by IκBα degradation. E. coli (10⁷ organisms/ml) did not induce IκBα degradation or NF-κB translocation. Pretreatment with cytochalasin D blocked invasion of all three strains but had no effect on IκBα degradation or NF-κB activation. S. dublin and S. typhimurium adhered to Caco-2BBe cells 3- to 10-fold more than E. coli. NF-κB activation was prevented by physical separation of S. dublin from Caco-2BBe cells by a 0.4-μm-pore-size filter. Our results imply that bacterial adhesion, rather than invasion or release of a secreted factor, is sufficient to induce IκBα degradation and NF-κB activation in intestinal epithelial cells. Our data suggest that strategies to reduce enteric inflammation should be directed to the reduction of bacterial enterocyte adhesion.