Metallacage-based enhanced PDT strategy for bacterial elimination via inhibiting endogenous NO production.

Antibiotics are among the most used weapons in fighting microbial infections and have greatly improved the quality of human life. However, bacteria can eventually evolve to exhibit antibiotic resistance to almost all prescribed antibiotic drugs. Photodynamic therapy (PDT) develops little antibiotic resistance and has become a promising strategy in fighting bacterial infection. To augment the killing effect of PDT, the conventional strategy is introducing excess ROS in various ways, such as applying high light doses, high photosensitizer concentrations, and exogenous oxygen. In this study, we report a metallacage-based PDT strategy that minimizes the use of ROS by jointly using gallium-metal organic framework rods to inhibit the production of bacterial endogenous NO, amplify ROS stress, and enhance the killing effect. The augmented bactericidal effect was demonstrated both in vitro and in vivo. This proposed enhanced PDT strategy will provide a new option for bacterial ablation.

Hepialiamides A-C: Aminated Fusaric Acid Derivatives and Related Metabolites with Anti-Inflammatory Activity from the Deep-Sea-Derived Fungus Samsoniella hepiali W7.

A systematic investigation combined with a Global Natural Products Social (GNPS) molecular networking approach, was conducted on the metabolites of the deep-sea-derived fungus Samsoniella hepiali W7, leading to the isolation of three new fusaric acid derivatives, hepialiamides A-C (1-3) and one novel hybrid polyketide hepialide (4), together with 18 known miscellaneous compounds (5-22). The structures of the new compounds were elucidated through detailed spectroscopic analysis. as well as TD-DFT-based ECD calculation. All isolates were tested for anti-inflammatory activity in vitro. Under a concentration of 1 µM, compounds 8, 11, 13, 21, and 22 showed potent inhibitory activity against nitric oxide production in lipopolysaccharide (LPS)-activated BV-2 microglia cells, with inhibition rates of 34.2%, 30.7%, 32.9%, 38.6%, and 58.2%, respectively. Of particularly note is compound 22, which exhibited the most remarkable inhibitory activity, with an IC50 value of 426.2 nM.

Endothelial Cell Behavior and Nitric Oxide Production on a-C:H:SiOx-Coated Ti-6Al-4V Substrate.

This paper focuses on the surface modification of the Ti-6Al-4V alloy substrate via a-C:H:SiOx coating deposition. Research results concern the a-C:H:SiOx coating structure, investigated using transmission electron microscopy and in vitro endothelization to study the coating. Based on the analysis of the atomic radial distribution function, a model is proposed for the atomic short-range order structure of the a-C:H:SiOx coating, and chemical bonds (C-O, C-C, Si-C, Si-O, and Si-Si) are identified. It is shown that the a-C:H:SiOx coating does not possess prolonged cytotoxicity in relation to EA.hy926 endothelial cells. In vitro investigations showed that the adhesion, cell number, and nitric oxide production by EA.hy926 endothelial cells on the a-C:H:SiOx-coated Ti-6Al-4V substrate are significantly lower than those on the uncoated surface. The findings suggest that the a-C:H:SiOx coating can reduce the risk of endothelial cell hyperproliferation on implants and medical devices, including mechanical prosthetic heart valves, endovascular stents, and mechanical circulatory support devices.

Nine New Pregnane Glycosides from the Cultivated Medicinal Plant Marsdenia tenacissima.

The ethnobotanical plant Marsdenia tenacissima has been used for hundreds of years for Dai people in Yunnan Province, China. Previously, chemical investigations on this plant have revealed that pregnane glycosides were the main biological constituents. Nine new pregnane glycosides, marsdeosides A-I (1-9), were isolated from cultivated dried stems of the medicinal plant Marsdenia tenacissima in this study. The structures were analyzed by extensive spectroscopic analysis, including 1D, 2D NMR, HRESIMS, and IR spectroscopic analysis. The absolute configurations of the sugar moieties were identified by comparing the Rf values and specific optical rotations with those of the commercially available standard samples and the data reported in the literature. Marsdeosides A (1) featured an unusual 8,14-seco-pregnane skeleton. Compounds 1, 8, and 9 showed activity against nitric oxide production in lipopolysaccharide-activated macrophage RAW264.7, with IC50 values of 37.5, 38.8, and 42.8 µM (L-NMMA was used as a positive control, IC50 39.3 µM), respectively. This study puts the knowledge of the chemical profile of the botanical plant M. tenacissima one step forward and, thereby, promotes the sustainable utilization of the resources of traditional folk medicinal plants.

Sinensiols H-J, three new lignan derivatives from Selaginella sinensis (Desv.) Spring.

One new lignan sinensiol H (1) and two new bisnorlignans, sinensiols I and J (2 and 3), along with three known compounds were isolated from the whole plants of Selaginella sinensis. Their structures were elucidated on the basis of 1D and 2D NMR spectroscopy as well as high-resolution mass spectrometry. The absolute configuration of 1 was established by ECD calculation. Compounds 2 and 3 represent rare examples of naturally occurring 9,9'-bisnorlignans. All the isolated compounds were assayed for their inhibitory effects on LPS-induced nitric oxide production in RAW 264.7 macrophages.

Transforming Growth Factor-induced Protein Promotes NF-κB-mediated Angiogenesis during Postnatal Lung Development.

Pulmonary angiogenesis is a key driver of alveolarization. Our prior studies showed that NF-κB promotes pulmonary angiogenesis during early alveolarization. However, the mechanisms regulating temporal-specific NF-κB activation in the pulmonary vasculature are unknown. To identify mechanisms that activate proangiogenic NF-κB signaling in the developing pulmonary vasculature, proteomic analysis of the lung secretome was performed using two-dimensional difference gel electrophoresis. NF-κB activation and angiogenic function was assessed in primary pulmonary endothelial cells (PECs) and TGFBI (transforming growth factor-ß-induced protein)-regulated genes identified using RNA sequencing. Alveolarization and pulmonary angiogenesis was assessed in wild-type and Tgfbi null mice exposed to normoxia or hyperoxia. Lung TGFBI expression was determined in premature lambs supported by invasive and noninvasive respiratory support. Secreted factors from the early alveolar, but not the late alveolar or adult lung, promoted proliferation and migration in quiescent, adult PECs. Proteomic analysis identified TGFBI as one protein highly expressed by the early alveolar lung that promoted PEC migration by activating NF-κB via αvß3 integrins. RNA sequencing identified Csf3 as a TGFBI-regulated gene that enhances nitric oxide production in PECs. Loss of TGFBI in mice exaggerated the impaired pulmonary angiogenesis induced by chronic hyperoxia, and TGFBI expression was disrupted in premature lambs with impaired alveolarization. Our studies identify TGFBI as a developmentally regulated protein that promotes NF-κB-mediated angiogenesis during early alveolarization by enhancing nitric oxide production. We speculate that dysregulation of TGFBI expression may contribute to diseases marked by impaired alveolar and vascular growth.

Effect of in-vitro heat stress challenge on the function of blood mononuclear cells from dairy cattle ranked as high, average and low immune responders.

BACKGROUND: The warming climate is causing livestock to experience heat stress at an increasing frequency. Holstein cows are particularly susceptible to heat stress because of their high metabolic rate. Heat stress negatively affects immune function, particularly with respect to the cell-mediated immune response, which leads to increased susceptibility to disease. Cattle identified as having enhanced immune response have lower incidence of disease. Therefore, the objective of this study was to evaluate the impact of in vitro heat challenge on blood mononuclear cells from dairy cattle, that had previously been ranked for immune response, in terms of heat shock protein 70 concentration, nitric oxide production, and cell proliferation. RESULTS: Blood mononuclear cells from dairy cattle classified as high immune responders, based on their estimated breeding values for antibody and cell-mediated responses, produced a significantly greater concentration of heat shock protein 70 under most heat stress treatments compared to average and low responders, and greater cell-proliferation across all treatments. Similarly, a trend was observed where high responders displayed greater nitric oxide production compared to average and low responders across heat treatments. CONCLUSION: Overall, these results suggest that blood mononuclear cells from high immune responder dairy cows are more thermotolerant compared to average and low immune responders.

Neuroprotective Effect of 1,4-Naphthoquinones in an In Vitro Model of Paraquat and 6-OHDA-Induced Neurotoxicity.

Targeted screening using the MTT cell viability test with a mini-library of natural and synthetic 1,4-naphthoquinones and their derivatives was performed in order to increase the survival of Neuro-2a neuroblastoma cells in in vitro paraquat and 6-hydroxydopamine models of Parkinson's disease. As a result, 10 compounds were selected that could protect neuronal cells from the cytotoxic effects of both paraquat and 6-hydroxydopamine. The five most active compounds at low concentrations were found to significantly protect the activity of nonspecific esterase from the inhibitory effects of neurotoxins, defend cell biomembranes from lytic destruction in the presence of paraquat and 6-hydroxydopamine, and normalize the cell cycle. The protective effects of these compounds are associated with the suppression of oxidative stress, decreased expression of reactive oxygen species and nitric oxide formation in cells and normalization of mitochondrial function, and restoration of the mitochondrial membrane potential altered by neurotoxins. It was suggested that the neuroprotective activity of the studied 1,4-NQs is attributable to their pronounced antioxidant and free radical scavenging activity and their ability to reduce the amount of reactive oxygen species formed by paraquat and 6-hydroxydopamine action on neuronal cells. The significant correlation between the neuroprotective properties of 1,4-naphthoquinones and Quantitative Structure-Activity Relationship descriptors describing the physicochemical properties of these compounds means that the hydrophobicity, polarity, charge, and shape of the molecules can be of decisive importance in determining the biological activity of studied substances.

Strophasterols E and F: Rearranged ergostane-type sterols from Pleurotus eryngii.

Strophasterols E (1) and F (2) were isolated from the fruiting bodies of Pleurotus eryngii, together with four new ergostane-type sterols (3-6). Single-crystal X-ray diffraction analysis performed on the tris-p-bromobenzoate derivatives of compounds 1 and 2 allowed these two compounds to be identified as the structurally rare (22S,23R)- and (22S,23S)-5α,6α-epoxy-3ß,7ß,23-trihydroxy-15(14 → 22)-abeo-ergost-8-en-14-one, respectively. The inhibitory effects on nitric oxide production of the six new steroids thus isolated from the fruiting bodies of P. eryngii were also evaluated.

EGFR inhibitors from cancer to inflammation: Discovery of 4-fluoro-N-(4-(3-(trifluoromethyl)phenoxy)pyrimidin-5-yl)benzamide as a novel anti-inflammatory EGFR inhibitor.

EGFR inhibitors are well-known as anticancer agents. Quite differently, we report our effort to develop EGFR inhibitors as anti-inflammatory agents. Pyrimidinamide EGFR inhibitors eliciting low micromolar IC50 and the structurally close non-EGFR inhibitor urea analog were synthesized. Comparing their nitric oxide (NO) production inhibitory activity in peritoneal macrophages and RAW 246.7 macrophages indicated that their anti-inflammatory activity in peritoneal macrophages might be a sequence of EGFR inhibition. Further evaluations proved that compound 4d significantly and dose-dependently inhibits LPS-induced iNOS expression and IL-1ß, IL-6, and TNF-α production via NF-κB inactivation in peritoneal macrophages. Compound 4d might serve as a lead compound for development of a novel class of anti-inflammatory EGFR inhibitors.

Bioactive monoterpene indole alkaloids from Nauclea officinalis.

Two new monoterpene indole alkaloids, naucleaoffines A (1) and B (2), together with six known alkaloids (3-8), were isolated from the stems and leaves of Nauclea officinalis. The structures of 1 and 2 were elucidated by extensive spectroscopic methods and the known compounds were identified by comparisons with the data reported in literature. All isolated compounds were evaluated for their anti-inflammatory activities and anti-HIV-1 activities. Compounds 1-8 exhibited significant inhibitory activities on nitric oxide (NO) production induced by lipopolysaccharide in mouse macrophage RAW 264.7 cells in vitro with IC50 values comparable to that of hydrocortisone. In addition, compounds 1-8 showed significant anti-HIV-1 activities with EC50 ranged from 0.06 to 2.08 µM. These findings suggest that the discoveries of these indole alkaloids with significant anti-inflammatory activities and anti-HIV-1 activities isolated from N. officinalis could be of great importance to the development of new anti-inflammatory and anti-HIV agents.

The effect of host genetics on in vitro performance of bovine monocyte-derived macrophages.

The dynamic interaction between the host and pathogens, along with environmental factors, influences the regulation of mammalian immune responses. Therefore, comprehensive in vivo immune-phenotyping during an active response to a pathogen can be complex and prone to confounding effects. Evaluating critical fundamental aspects of the immune system at a cellular level is an alternative approach to reduce this complexity. Therefore, the objective of the current study was to examine an in vitro model for functional phenotyping of bovine monocyte-derived macrophages (MDM), cells which play a crucial role at all phases of inflammation, as well influence downstream immune responses. As indicators of MDM function, phagocytosis and nitric oxide (NO-) production were tested in MDM of 16 cows in response to 2 common bacterial pathogens of dairy cows, Escherichia coli and Staphylococcus aureus. Notable functional variations were observed among the individuals (coefficient of variation: 33% for phagocytosis and 70% in the production of NO-). The rank correlation analysis revealed a significant, positive, and strong correlation (rho = 0.92) between NO- production in response to E. coli and S. aureus, and a positive but moderate correlation (rho = 0.58) between phagocytosis of E. coli and S. aureus. To gain further insight into this trait, another 58 cows were evaluated solely for NO- response against E. coli. The pedigree of the tested animals was added to the statistical model and the heritability was estimated to be 0.776. Overall, the finding of this study showed a strong effect of host genetics on the in vitro activities of MDM and the possibility of ranking Holstein cows based on the in vitro functional variation of MDM.

Astragaloside IV protects against diabetic nephropathy via activating eNOS in streptozotocin diabetes-induced rats.

BACKGROUND: Astragaloside IV (AS-IV) was reported to play a role in improving diabetic nephropathy (DN), however, the underlying mechanisms still remain unclear. The aim of the present study is to investigate whether AS-IV ameliorates DN via the regulation of endothelial nitric oxide synthase (eNOS). METHODS: DN model was induced in Sprague-Dawley (SD) male rats by intraperitoneal injection of 65 mg/kg streptozotocin (STZ). Rats in the AS-IV treatment group were orally gavaged with 5 mg/kg/day or 10 mg/kg/day AS-IV for eight consecutive weeks. Body weight, blood glucose, blood urea nitrogen (BUN), Serum creatinine (Scr), proteinuria and Glycosylated hemoglobin (HbA1c) levels were measured. Hematoxylin-Eosin (HE) and Periodic Acid-Schiff (PAS) staining were used to detect the renal pathology. The apoptosis status of glomerular cells was measured by TUNEL assay. The phosphorylation and acetylation of eNOS were detected by western blot. The effects of AS-IV on high-glucose (HG)-induced apoptosis and eNOS activity were also investigated in human renal glomerular endothelial cells (HRGECs) in vitro. RESULTS: Treatment with AS-IV apparently reduced DN symptoms in diabetic rats, as evidenced by reduced BUN, Scr, proteinuria, HbA1c levels and expanding mesangial matrix. AS-IV treatment also promoted the synthesis of nitric oxide (NO) in serum and renal tissues and ameliorated the phosphorylation of eNOS at Ser 1177 with decreased eNOS acetylation. Moreover, HG-induced dysfunction of HRGECs including increased cell permeability and apoptosis, impaired eNOS phosphorylation at Ser 1177, and decreased NO production, were all reversed by AS-IV treatment. CONCLUSIONS: These novel findings suggest that AS-IV ameliorates functional abnormalities of DN through inhibiting acetylation of eNOS and activating its phosphorylation at Ser 1177. AS-IV could be served as a potential therapeutic drug for DN.

Marylosides A-G, Norcycloartane Glycosides from Leaves of Cymbidium Great Flower 'Marylaurencin'.

Seven novel norcycloartane glycosides, maryloside A-G (1-7), were isolated from the leaves of Cymbidium Great Flower 'Marylaurencin', along with a known norcycloartane glycoside, cymbidoside (8). These structures were determined on the basis of mainly NMR experiments as well as chemical degradation and X-ray crystallographic analysis. The isolated compounds (1-6 and 8) were evaluated for the inhibitory activity on lipopolysaccharide (LPS) and interferon-γ (IFN-γ)-stimulated nitric oxide (NO) production in RAW 264.7 cells. Consequently, 1 and 3 exhibited moderate activity.

Lignan Glycosides from Urena lobata.

Four new lignan glycosides; urenalignosides A-D (1-4), along with 12 known ones (5-16) were isolated from Urena lobata. Their structures were determined on the basis of extensive spectroscopic and spectrometric data (1D and 2D NMR; IR; CD; and HRESIMS). Compounds 2-4; 6; 7; 10; and 11 showed inhibition of nitric oxide production in lipopolysaccharide-induced RAW 264.7 macrophage cells with IC50 values in the range of 25.5-98.4 µM (positive control; quercetin; IC50 = 7.2 ± 0.2 µM).

Anti-Inflammatory Activity of Some Characteristic Constituents from the Vine Stems of Spatholobus suberectus.

The dried vine stems of Spatholobus suberectus are commonly used in traditional Chinese medicine for treating gynecological and cardiovascular diseases. In this study, five new compounds named spasuberol A (2), homovanillyl-4-oxo-nonanoate (5), spasuberol C (6), spasuberoside A (14), and spasuberoside B (15), together with ten known compounds (1, 3, 4, 7-13), were isolated from the dried vine stems of S. suberectus. Their chemical structures were analyzed using spectroscopic assays. This is the first study interpreting the detailed structural information of 4. The anti-inflammatory activity of these compounds was evaluated by reducing nitric oxide overproduction in RAW264.7 macrophages stimulated by lipopolysaccharide. Compounds 1 and 8-10 showed strong inhibitory activity with half maximal inhibitory concentration (IC50) values of 5.69, 16.34, 16.87, and 6.78 µM, respectively, exhibiting higher activity than the positive drug l-N6-(1-iminoethyl)-lysine (l-NIL) with an IC50 value of 19.08 µM. The IC50 values of inhibitory activity of compounds 2 and 4-6 were 46.26, 40.05, 45.87, and 28.29 µM respectively, which were lower than l-NIL, but better than that of positive drug indomethacin with an IC50 value of 55.44 µM. Quantitative real-time polymerase chain reaction analysis revealed that assayed compounds with good anti-inflammatory activity, such as 1, 6, 9, and 10 at different concentrations, can reduce the messenger RNA (mRNA) expression of some pro-inflammatory cytokines such as tumor necrosis factor α (TNF-α), nitric oxide synthase (iNOS), and cyclooxygenase 2 (COX-2). The anti-inflammatory activity and the possible mechanism of the compounds mentioned in this paper were studied preliminarily.

[Effect of polyethylene terephthalate (Dacron®) on functional potential of human bone-marrow/progenitor stem cells]. / Vliianie poliétilentereftalata (Dakron®) na funktsional'nyi potentsial kostnomozgovykh stvolovykh/progenitornykh kletok cheloveka.

Vascular grafts made of polytetrafluoroethylene and polyethylene terephthalate have widely been used in cardiovascular surgery. The causes of delayed colonization of such grafts by endotheliocytes and mesenchymal stem cells have not been adequately investigated. The authors examined the effect of polyethylene terephthalate on the functional activity of human bone marrow mesenchymal stem cells and endothelial progenitor cells in vitro. Proliferation (MTT assay, real-time cellular impedance), migration (Boyden chamber, real-time cellular impedance), and nitric oxide production (spectrophotometciacally) by progenitor endothelial cells and mesenchymal stem cells were assessed with and without the presence of polyethylene terephthalate. The functional activity of the cells was shown to depend on the presence of polyethylene terephthalate in a well with cells. Thus, polyethylene terephthalate turned out to exhibit a toxic effect on progenitor endothelial and mesenchymal stem cells. Treatment of grafts with gelatine or fibronectin improved colonization of grafts with cells.

DT-13 ameliorates TNF-α-induced nitric oxide production in the endothelium in vivo and in vitro.

The steroidal saponin DT-13 (25(R,S)-ruscogenin-1-O-[ß-d-glucopyranosyl-(1 â†’ 2)][ß-d-xylopyranosyl-(1 â†’ 3)]-ß-d-fucopyranoside), one of the major active compounds of the herb Liriope muscari (Decne.), exhibits significant anti-inflammatory, anti-tumor and cardioprotective effects. This study aimed to explore the protective effect of DT-13 on endothelium through regulating of nitric oxide production induced by Tumor necrosis factor-α (TNF-α). The results demonstrated that DT-13 inhibited inflammatory cell infiltration and thus played a protective effect on endothelial cells in vivo, as shown by hematoxylin-eosin (H&E) staining and immunohistochemical staining. Enzyme-linked immunosorbent assay (ELISA) results demonstrated that DT-13 could suppress the TNF-α-induced upregulation of reactive oxygen species (ROS), tumor necrosis factor receptor (TNFR), interleukin-8 (IL-8), monocyte chemoattractant protein-1 (MCP-1) and nitric oxide in vivo dose-dependently and suppressed production of nitric oxide in vitro as shown by DAF-FMDA. Western blotting results indicated that DT-13 could down-regulate phosphorylation of endothelial nitric oxide synthase (eNOS) significantly in TNF-α-induced human umbilical vein endothelial cells (HUVECs). Taken together, we speculate that DT-13 inhibits endothelium vascular inflammation through regulating nitric oxide production and the expression of ROS, TNFR, IL-8, MCP-1, which are associated with inflammation.

Real time electrochemical investigation of the release, distribution and modulation of nitric oxide in the intestine of individual zebrafish embryos.

Nitric oxide (NO) is an important signaling molecule that has been implicated in a variety of physiological and pathophysiological processes in living organisms. NO plays an important role in embryonic development in vertebrates and has been reported to influence early organ development and plasticity. Quantifying NO in single embryos and their developing organs is challenging because of the small size of the embryos, the low dynamically changing concentration and the short life-time of NO. Here, we measured the distribution of NO in the intestine of live zebrafish (Danio rerio) embryos in physiological conditions and under the influence of therapeutic agents. NO measurements were performed using a miniaturized electrochemical sensor fabricated on a single carbon fiber (CF) which enables quantitative real time in vivo monitoring, and by fluorescence imaging using the 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM-DA) dye. NO production was detected in the middle segment the intestine at a level of 3.78 (±0.64) µM, and at lower levels in the anterior and posterior segments, 1.08 (±0.22) and 1.00 (±0.41) µM respectively. In the presence of resveratrol and rosuvastatin, the intestinal NO concentration decreased by 87% and 84%, demonstrating a downregulating effect. These results indicate the presence of variable micromolar concentrations of NO along the intestine of zebrafish embryos and demonstrate the usefulness of CF microelectrodes to measure quantitatively the NO release at the level of a single organ in individual zebrafish embryos. This work provides a unique approach to study in real time the modulatory role of NO in vivo and contributes to further understanding of the molecular basis of embryonic development for developmental biology and drug screening applications.

Nitric oxide inhibitory constituents from Siegesbeckia pubescens.

Two new sesquiterpenoids (1 and 2) and a new ent-pimarane type diterpenoid (3), together with eighteen known compounds (4-21), were isolated from the whole plants of Siegesbeckia pubescens. The structures of the new compounds were determined on the basis of 1D-, 2D NMR and HRESIMS data. All compounds were evaluated for their inhibitory effects on LPS-induced nitric oxide production in RAW 264.7 macrophages. Of these, highly oxygenated germacrane type sesquiterpenoids (1-2 and 13-14) showed significant inhibitory effects with IC50 values ranging from 3.9 to 16.8 µM.