Simultaneous Efficient Photocatalytic Hydrogen Evolution and Degradation of Dye Wastewater without Cocatalysts and Sacrificial Agents Based on g-C3N5 and Hybridized Ni-MOF Derivative-CdS-DETA.

Inspired by energy conversion and waste reuse, hybridized Ni-MOF derivative-CdS-DETA/g-C3N5, a type-II heterojunction photocatalyst, is synthesized by a hydrothermal method for simultaneous and highly efficient photocatalytic degradation and hydrogen evolution in dye wastewater. Without the addition of cocatalysts and sacrificial agents, the optimal MOF-CD(2)/CN5 (i.e. Ni-MOF derivative-CdS-DETA (20 wt.%)/g-C3N5) exhibit good bifunctional catalytic activity, with a H2 evolution rate of 2974.4 µmol g-1 h-1 during the degradation of rhodamine B (RhB), and a removal rate of 99.97% for RhB. In the process of H2-evolution-only, triethanolamine is used as a sacrificial agent, exhibiting a high H2 evolution rate (19663.1 µmol g-1 h-1) in the absence of a cocatalyst, and outperforming most similar related materials (such as MOF/g-C3N5, MOF-CdS, CdS/g-C3N5). With the help of type-II heterojunction, holes are scavenged for the oxidative degradation of RhB, and electrons are used in the decomposition of water for H2 evolution during illumination. This work opens a new path for photocatalysts with dual functions of simultaneous efficient degradation and hydrogen evolution.

Intratracheally Administered Peptide-Modified Lipid Admixture Containing Fasudil and/or DETA NONOate Ameliorates Various Pathologies of Pulmonary Arterial Hypertension.

This study examined the therapeutic potential of a combination therapy using fasudil, a Rho-kinase inhibitor, and DETA NONOate (DN), a nitric oxide donor, delivered as a lipid admixture modified with a cyclic homing peptide known as CAR (CAR-lipid mixture) for the treatment of pulmonary arterial hypertension (PAH). CAR-lipid mixtures were initially prepared via a thin-film hydration method and then combined with fasudil, DN, or a mixture of both. The therapeutic efficacy of this drug-laden lipid mixture was evaluated in a Sugen/Hypoxia (Su/Hx) rat model of PAH by measuring RV systolic pressure (RVSP), mean pulmonary arterial pressure (mPAP), Fulton indices, and assessing right ventricular (RV) functions, as well as evaluating pulmonary vascular morphology. Rats that received no treatment exhibited increases in RVSP, mPAP, Fulton indices, and changes in RV functional parameters. However, the treatment with the CAR-lipid mixture containing either fasudil or DN or a combination of both led to a decline in mPAP, RVSP, and Fulton indices compared to saline-treated rats. Similarly, rats that received these treatments showed concurrent improvement in various echocardiographic parameters such as pulmonary acceleration time (PAT), tricuspid annular plane systolic excursion (TAPSE), and ventricular free wall thickness (RVFWT). A significant decrease in the wall thickness of pulmonary arteries larger than 100 µm was observed with the combination therapy. The findings reveal that fasudil, DN, and their combination in a CAR-modified lipid mixture improved pulmonary hemodynamics, RV functions, and pathological alterations in the pulmonary vasculature. This study underscores the potential of combination therapy and targeted drug delivery in PAH treatment, laying the groundwork for future investigations into the optimization of these treatments, their long-term safety and efficacy, and the underlying mechanism of action of the proposed therapy.

A Comparative Study of the Impact of NO-Related Agents on MK-801- or Scopolamine-Induced Cognitive Impairments in the Morris Water Maze.

Learning and memory deficits accompany numerous brain dysfunctions, including schizophrenia and Alzheimer's disease (AD), and many studies point to the role of nitric oxide (NO) in these processes. The present investigations constitute the follow-up of our previous research, in which we investigated the activity of NO releasers and a selective inhibitor of neuronal NO synthase (nNOS) to prevent short-term memory deficits in novel object recognition and T-maze. Here, the ability of the compounds to prevent the induction of long-term memory deficits by MK-801 or scopolamine administration was investigated. The Morris Water Maze test, a reliable and valid test of spatial learning and memory, was used, in which escape latency in the acquisition phase and nine different parameters in the retention phase were measured. A fast NO releaser (spermine NONOate), a slow NO releaser (DETA NONOate), and a nNOS inhibitor, N(ω)-propyl-L-arginine (NPLA), were used. The compounds were administered i.p. at a dose range of 0.05-0.5 mg/kg. All compounds prevented learning deficits in the acquisition phase and reversed reference memory deficits in the retention phase of the scopolamine-treated mice. Spermine NONOate was the least effective. In contrast, the drugs poorly antagonised MK-801-induced deficits, and only the administration of DETA NONOate induced some improvements in the retention trial.

Engineering Under-Coordinated Active Sites with Tailored Chemical Microenvironments over Mosaic Bismuth Nanosheets for Selective CO2 Electroreduction to Formate.

Selective electrochemical reduction of CO2 into fuels or chemical feedstocks is a promising avenue to achieve carbon-neutral goal, but its development is severely limited by the lack of highly efficient electrocatalysts. Herein, cation-exchange strategy is combined with electrochemical self-reconstruction strategy to successfully develop diethylenetriamine-functionalized mosaic Bi nanosheets (mBi-DETA NSs) for selective electrocatalytic CO2 reduction to formate, delivering a superior formate Faradaic efficiency of 96.87% at a low potential of -0.8 VRHE . Mosaic nanosheet morphology of Bi can sufficiently expose the under-coordinated Bi active sites and promote the activation of CO2 molecules to form the OCHO- * intermediate. Moreover, in situ attenuated total reflectance infrared spectra further corroborate that surface chemical microenvironment modulation of mosaic Bi nanosheets via DETA functionalization can improve CO2 adsorption on the catalyst surface and stabilize the key intermediate (OCHO- *) due to the presence of amine groups, thus facilitate the CO2 -to-HCOO- reaction kinetics and promote formate formation.

Hábitos alimentares das tribos da família Dipsadidae

Feeding habits are important for understanding the behavior of organisms in their respective habitats, and their interactions. Food habit is also an important factor for comparing diet and predation among individuals of the same taxonomic category, such as family, subfamily, tribe, genus, etc.Therefore, the aim of this work is to review and compile data from the literature on the feeding habits of snakes of five Dipsadidae tribes: Dipsadini, Elapomorphini, Philodryadini, Pseudoboini, and Xenodontini. Data on diet, predation, and feeding habits were searched, for understanding the traits of each tribe. The representatives of the tribe Dipsadini for example, have a diet of gastropods, slugs and snails, the tribe Elapomorphini has a specialized diet in ophiophagy, i.e., a diet based on other snakes, while the tribes Philodryadini and Pseudoboini both have a more generalist diet, it can be observed that small lizards and mammals are the most consumed in these two tribes, and the tribe Xenodontini specializes in preying on anurans. These results were obtained based on the food items described in the literature, such as amphibians, birds, gastropods, lizards, mammals, fish, eggs and snakes, so in the present study will also be analyzed, what are the characteristics of specialization and generalization regarding the diets of each tribe.

O hábito alimentar, é importante para compreender comportamentos dos organismos em seus respectivos habitats e as suas interações. O hábito alimentar também é um fator importante para comparar dieta e predação entre indivíduos da mesma categoria taxonômica, como por exemplo, família, subfamília, tribo, gênero e etc. Por isso o intuito deste trabalho foi revisar e compilar dados da literatura em relação a hábitos alimentares de serpentes de cinco tribos da família Dipsadidae, sendo elas Dipsadini, Elapomorphini, Philodryadini, Pseudoboini e Xenodontini. Dados de dieta, predação e hábitos alimentares foram levantados para compreender as tendências e padrões de cada tribo. Os representantes da tribo Dipsadini por exemplo, tem a sua dieta em gastrópodes lesmas e caramujo, na tribo Elapomorphini tem sua dieta especializada em ofiofagia ou seja, dieta a base de outras serpentes, já as tribos Philodryadini e Pseudoboini ambos possuem uma alimentação mais generalista, posso pode se observar que pequenos lagartos e mamíferos são os mais consumidos nestas duas tribos, e a tribo Xenodontini é especialista em predar anuros. Esses resultados foram obtidos com base em itens alimentares descrito na literatura como por exemplo, anfíbios, aves, gastrópodes, lagartos, mamíferos, peixes, ovos e serpentes, portanto no presente estudo será analisado também, quais características de especialização e generalização em relação a dietas de cada tribo.

Prospective, Single-blind, Randomized Controlled Trial to Evaluate the Effectiveness of a Digital Exercise Therapy Application Compared With Conventional Physical Therapy for the Treatment of Nonoperative Knee Conditions.

OBJECTIVE: To evaluate the effectiveness and adherence of a home exercise therapy program using a digital exercise therapy application (DETA) compared with conventional physical therapy (PT). DESIGN: Parallel group, randomized controlled trial. SETTING: Two clinics in a tertiary care academic center. PARTICIPANTS: Participants (N=60) were enrolled within 1 week after a provider visit for knee pain. Inclusion criteria: age 18-75 years, knee pain diagnosis, and clinician-prescribed PT. INTERVENTIONS: Participants were randomized to complete either an 8-week intervention of conventional PT (enrolled n=29; complete n=26) or the DETA (enrolled n=31; completed n=24). MAIN OUTCOME MEASURES: Patient-Reported Outcomes Measurement Information System (PROMIS) Pain Interference (PI) and Physical Function (PF) scores implemented via computer adaptive tests; number of exercise sessions completed per week (adherence). RESULTS: Compared with the PT group, the DETA group reported significant decreases in PROMIS-PI scores (-6.1±6.7 vs -1.5±6.6, P<.05, d=0.78) and increases in PROMIS-PF scores (6.0±6.6 vs -0.8±5.8, P<.01, d=0.89) after 8 weeks. No group differences in adherence were observed (P>.05). CONCLUSIONS: Use of this DETA resulted in greater pain and functional improvements compared with PT, with no differences in adherence. It is possible this application may be a viable alternative to conventional PT in certain cases. A larger sample from various geographic locations is needed to improve generalizability and for subgroup analysis. Further investigation is warranted to determine the factors responsible for the differences observed between the groups.

Research on the Utilization Efficiency of Innovative Resources and Its Change Decomposition of Listed TCM Industrial Enterprises from the Perspective of Total Factors / 中国药房

OBJECTIVE：To prov ide effective improvement paths and policy suggestions for improving the innovation efficiency of TCM industrial enterprises. METHODS ：Based on the input and output data of innovation resources of 38 listed TCM industrial enterprises from 2014 to 2020，an evaluation index system was constructed from the perspective of all factors. First ，the deta envelopment analysis （DEA）model was used to measure the static efficiency of the sample enterprises ，and at the same time ， four patterns of innovative resource utilization were provided ；then the total factor Malmquist index method was used to analyze the decomposition of the changes in utilization efficiency of innovative resources according to different periods and micro-individuals. RESULTS & CONCLUSIONS ：The overall efficiency of innovative resource utilization of listed TCM industrial enterprises was too low，and the average overall efficiency was only 0.293. The input and output of innovation resources of 33 enterprises were unreasonable，and the reasons for the inefficiency are mostly due to insufficient pure technical efficiency. The average value of the all-factor Malmquist index was 0.818 for 38 sample enterprises ，and the overall efficiency was declining. The obstacles to technological progress were a more important reason for the decline in efficiency. It is recommended to improve TCM industrial enterprise’s ability to allocate innovation resources and adjust the proportion of R&D investment in conjunction with the return to scale；clarify the development principles with scientific and technological innovation as the core driving force ，and actively carry out basic research work ；implement the production and research platform of “TCM industrial enterprise-TCM research institution ” to promote the overall technological progress of the industry.

Combining Inulin Multifunctional Polycation and Magnetic Nanoparticles: Redox-Responsive siRNA-Loaded Systems for Magnetofection.

Superparamagnetic Iron Oxide Nanoparticles (SPIONs) are recognized as one of the most promising agents for theranostic applications. Among methods designed for siRNA delivery, magnetofection, that is, nucleic acid cell uptake under the influence of a magnetic field acting on magnetic nucleic acid vectors, is emerging as a unique approach to combining advantages such as strong improvement of the kinetics of the delivery process and the possibility of localizing nucleic acid delivery to an area where the magnetic field is applied. This paper reports on the preparation of siRNA loaded magnetoplexes-named ICD@SS@SPIONs/siRNA-by controlled crosslinking, in the presence of SPIONs, of the polycation INU-C-DETA, synthesized starting from the polysaccharide inulin by grafting diethylenetriamine and cystamine molecules. The obtained ICD@SS@SPIONs/siRNA have suitable chemical-physical characteristics to be employed for iv administration and are also able to release siRNA in a redox-triggered manner thanks to intracellular glutathione (GSH) mediated reduction of disulphide bridges formed during the crosslinking process. Moreover, ICD@SS@SPIONs/siRNA are able to produce magnetic targeting in vitro on breast cancer cells, without appreciable cyto- and hemo-toxic effects, in a wide range of concentrations. Finally, protein binding to nanoparticles revealed that obtained systems are potentially longer circulating and applicable as a smart multifunctional agents for cancer therapy.

Highly Conductive Paper/Textile Electrodes Using Ligand Exchange Reaction-Induced in Situ Metallic Fusion.

Here, we report that metal nanoparticle (NP)-based paper/textile electrodes with bulk metallic conductivity can be prepared via organic linker-modulated ligand exchange reaction and in situ room-temperature metallic fusion without additional chemical or thermal treatments. For this study, amine-functionalized molecule linkers instead of bulky polymer linkers were layer-by-layer (LbL)-assembled with tetraoctylammonium bromide (TOABr)-stabilized Au NPs to form Au NP multilayered films. By conversion of the amine groups of the organic molecule linkers from -NH3+ to the -NH2 groups, as well as by a decrease in the size of the organic linkers, the LbL-assembled Au NPs became highly interconnected and fused during LbL deposition, resulting in Au NP multilayers with adjustable conductivity and transport behavior. These phenomena were also predicted by a density functional theory investigation for the model system. Particularly, LbL-assembled films composed of TOABr-Au NPs and diethylenetriamine ( Mw: ∼104) exhibited a remarkable electrical conductivity of 2.2 × 105 S·cm-1, which was higher than the electrical conductivity of the metal NP-based electrodes as well as the carbon material-based electrodes reported to date. Furthermore, based on our approach, a variety of insulating flexible papers and textiles were successfully converted into real metal-like paper and textile electrodes with high flexibility preserving their highly porous structure. This approach can provide a basis for further improving and controlling the electrical properties of flexible electrodes through the control of organic linkers.

Combination Therapy with DETA/NO and Clopidogrel Inhibits Metastasis in Murine Mammary Gland Cancer Models via Improved Vasoprotection.

Vascular endothelial dysfunction and platelet activation play a key role in tumor metastasis, and therefore, both of these processes are considered important therapeutic targets in cancer. The aim of our studies was to analyze antimetastatic activity of combination therapy using nitric oxide donor DETA/NO and antiplatelet drug clopidogrel. Nitric oxide acts as a vasoprotective mediator, while clopidogrel inhibits ADP-mediated platelet aggregation. 4T1-luc2-tdTomato cell line transplanted intravenously (i.v.) and 4T1 cell line transplanted orthotopically were used as metastatic mammary gland cancer models. Moreover, antiaggregation action of compounds was tested ex vivo on the blood samples taken from breast cancer patients. We have shown that in selected dosage regimes, DETA/NO combined with clopidogrel significantly reduced lung metastatic foci formation in an i.v. model, and such inhibition was transiently observed also in an orthotopic model. The antimetastatic effect was correlated with a significant increase of prostacyclin (PGI2) metabolite and reduction of endothelin-1, sE-selectin, sI-CAM, and TGF-ß plasma levels as well as decreased V-CAM expression on the endothelium. Combination therapy decreased fibrinogen binding to the resting platelets at the early stage of tumor progression (day 14). However, at the later stages (days 21 and 28), the markers of platelet activation were detected (increased JON/A antibody bound, P-selectin level, binding of fibrinogen, and vWf). Decreased aggregation as well as a lower release of TGF-ß were detected in platelets incubated ex vivo with compounds tested from metastatic breast cancer patients. Although combination therapy increases E-cadherin, the increase of N-cadherin and α-SMA in tumor tissue was also observed. The results showed that at the early stages of tumor progression, combined therapy with DETA/NO and clopidogrel improves vasoprotective and antiplatelet activity. However, in advanced tumors, some adverse effects toward platelet activation can be observed.

A novel method to prepare a magnetic carbon-based adsorbent with sugar-containing water as the carbon source and DETA as the modifying reagent.

A novel magnetic heavy metal adsorbent was prepared via diethylenetriamine (DETA) modification on magnetic hydrothermal carbon, with glucose and sugar-containing waste water as the carbon source. The prepared materials were characterized by FT-IR, SEM, TEM, EDXRF, TGA, elemental analysis, XPS, and magnetic moment determination. In this paper, the adsorption mechanism of the modified and unmodified adsorbents was well discussed. Four kinds of waste water (watermelon juice, expired sprite, sugar-pressing waste water, and confectionary waste water) were employed to produce heavy metal ion adsorbents; the chemical properties of hydrothermal carbon derived from the proposed sources were analyzed as well. The maximum uptake capacity for Cu2+, Pb2+, and Cd2+ of the adsorbent produced from glucose was 26.88, 103.09, and 25.38 mg g-1, respectively. After 5 cycles, the adsorption ability was still well preserved. This work represents an efficient new direction for the treatment of heavy metal ions in water and the reuse of sugar-containing waste water. Graphical abstract The schemetic of DETA-modified magnetic carbon preparing from sugar-containing wastewater.

Simultaneous Determination and Investigation of Nine Fungicides in Fruits Using Diethylenetriamine-Functional Magnetic Core-Shell Polymer Modified Graphene Oxide as an Efficient Adsorbent Coupled to UPLC-HRMS.

In this study, diethylenetriamine-functional magnetic core-shell polymer modified graphene oxide (DETA-MPs-GO) was prepared via precipitation polymerization and amidation reaction, and it was characterized by transmission electron microscopy (TEM), Fourier-transformed infrared spectroscopy (FTIR), and X-ray diffractometer (XRD). Subsequently, a magnetic solid-phase extraction (MSPE) procedure was applied to the as-synthesized DETA-MPs-GO for the detection of nine fungicides in fruit samples, prior to ultra-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS). The homogenized fruit samples, spiked with D-labelled internal standards, were firstly extracted by 5 mL of acetonitrile twice and then purified by DETA-MPs-GO adsorbents. The optimization of the adsorption and elution conditions of DETA-MPs-GO toward fungicides was carried out to attain a satisfactory adsorption performance and desorption efficiency. The adsorption mechanism was carefully investigated, and the results revealed that a synergistic adsorption mechanism, including hydrogen bond and a π-π stacking interaction, was confirmed. Moreover, the limits of quantitation (LOQs) of the proposed approach were in the range of 0.01 to 0.30 µg/kg under the optimum conditions. The average recoveries at three spiking levels were 84.9% to 105.2%, with relative standard deviations (RSDs) varying from 0.8% to 8.2% (n = 6). The developed method was successfully utilized for the screening and detection of fungicides in 81 fruit samples purchased from markets. A detailed survey was carried out about the concentration distribution, types of fungicides, and combined use of fungicides in different fruits.

Prolonged exposure of resveratrol induces reactive superoxide species-independent apoptosis in murine prostate cells.

Nitric oxide, a signaling molecule, inhibits mitochondrial respiration by binding with cytochrome c oxidase, resulting in elevated production of reactive superoxide species (reactive oxygen and nitrogen) in the mitochondria and increased susceptibility to cell death. Generation of mitochondrial superoxide species can be suppressed by natural compounds such as resveratrol, a dietary polyphenol found in the skin of red fruits. In various cancer cells, resveratrol shows anti-oxidant and cancer preventive properties. Since, the effect of resveratrol on reactive superoxide species-independent apoptosis in prostate cancer cells is not well illustrated; therefore, we investigated this phenomenon in TRAMP murine prostate cancer cells. To accomplish this, TRAMP cells were incubated with resveratrol, resveratrol + DETA-NONOate, DETA-NONOate (nitric oxide donor), resveratrol + L-NMMA, or L-NMMA (nitric oxide inhibitor) for 48 h, and reactive superoxide species in the mitochondria and culture supernatant were measured. In addition, the mitochondrial membrane potential, cell viability, expression of apoptotic markers (Bax and Bcl2), γ-H2A.x, p53, and caspase-3 was determined. We found that resveratrol suppressed reactive superoxide species such as reactive oxygen species in the mitochondria and nitric oxide in culture supernatant when compared to the DETA-NONOate treatment and disrupted the mitochondrial membrane potential. Resveratrol also reduced cell viability, altered the expression of apoptotic markers (Bax and Bcl2), and increased expression of γ-H2A.x (indicative marker of DNA fragmentation) and p53 (a critical DNA damage response protein). However, there was no appreciable modulation of the caspase-3. Therefore, our data suggest that resveratrol induces superoxide species-independent apoptosis and may act as a therapeutic agent against prostate cancer.

Galactosylated polyaspartamide copolymers for siRNA targeted delivery to hepatocellular carcinoma cells.

The limited efficacy of available treatments for hepatocellular carcinoma (HCC) requires the development of novel therapeutic approaches. We synthesized a novel cationic polymer based on α,ß-poly-(N-2-hydroxyethyl)-d,L-aspartamide (PHEA) for drug delivery to HCC cells. The copolymer was synthesized by subsequent derivatization of PHEA with diethylene triamine (DETA) and with a polyethylene glycol (PEG) derivative bearing galactose (GAL) molecules, obtaining the cationic derivative PHEA-DETA-PEG-GAL. PHEA-DETA-PEG-GAL has suitable chemical-physical characteristics for a potential systemic use and can effectively deliver a siRNA (siE2F1) targeted against the transcription factor E2F1, a gene product involved in HCC. The presence of GAL residues in the polyplexes allows the targeting of HCC cells that express the asialo-glycoprotein receptor (ASGP-R). In these cells, but not in ASGP-R non-expressing cells, PHEA-DETA-PEG-GAL/siE2F1 polyplexes induce the reduction of the mRNA and protein levels of E2F1 and of E2F1-regulated genes, all involved in the promotion of the G1/S phase transition. This results in a decrease of cell proliferation with a G1/G0 phase cells accumulation. Notably, removal of GAL residue almost completely abrogates the targeting capacity of the developed polyplexes. In conclusion, the generated polyplexes demonstrate the potential to effectively contributing to the development of novel anti-HCC therapeutic approaches via a siRNA-targeted delivery.

Arginase-2 is cooperatively up-regulated by nitric oxide and histone deacetylase inhibition in human umbilical artery endothelial cells.

Arginase-2 counteracts endothelial nitric oxide synthase (eNOS) activity in human endothelium, and its expression is negatively controlled by histone deacetylase (HDAC2). Conversely NO inhibits HDAC and previous studies suggest that arginase-2 is up-regulated by NO. We studied whether NO regulates arginase-2 expression in umbilical artery endothelial cells (HUAEC) increasing ARG2 promoter accessibility. HUAEC exposed to NOC-18 (NO donor, 1-100 µM, 0-24 h) showed an increase in arginase-2 but a decrease in eNOS mRNA levels in a time-dependent manner, with a maximal effect at 100 µM (24 h). Conversely NOS inhibition with L-NAME (100 µM) reduced arginase-2 mRNA and protein levels, an effect reverted by co-incubation with NOC-18. Treatment with TSA paralleled the effects of NO on arginase-2 and eNOS at mRNA and protein levels, with maximal effect at 10 µM. Co-incubation of NOC-18 (100 µM) with a sub-maximal concentration of TSA (1 µM) potentiated the increase in arginase-2 mRNA levels, whilst L-NAME prevented TSA-dependent arginase-2 induction. The effects on arginase-2 mRNA were paralleled by changes in chromatin accessibility, as well as increased levels of H3K9 and H4K12 acetylation, at ARG2 proximal (-579 to -367 and -280 to -73 bp from TSS) and core (-121 to +126 bp from TSS) promoter. Finally NO-dependent arginase-2 induction was prevented by pre-incubation for 10 min with the cysteine blocker MMTS (10 mM). These data showed for the first time that NO up-regulates arginase-2 expression in primary cultured human endothelial cells by an epigenetic-mediated mechanism increasing ARG2 promoter accessibility suggesting a negative regulatory loop for eNOS activity.

Development of a simple, biocompatible and cost-effective Inulin-Diethylenetriamine based siRNA delivery system.

Small interfering RNAs (siRNAs) have the potential to be of therapeutic value for many human diseases. So far, however, a serious obstacle to their therapeutic use is represented by the absence of appropriate delivery systems able to protect them from degradation and to allow an efficient cellular uptake. In this work we developed a siRNA delivery system based on inulin (Inu), an abundant and natural polysaccharide. Inu was functionalized via the conjugation with diethylenetriamine (DETA) residues to form the complex Inu-DETA. We studied the size, surface charge and the shape of the Inu-DETA/siRNA complexes; additionally, the cytotoxicity, the silencing efficacy and the cell uptake-mechanisms were studied in the human bronchial epithelial cells (16HBE) and in the hepatocellular carcinoma derived cells (JHH6). The results presented here indicate that Inu-DETA copolymers can effectively bind siRNAs, are highly cytocompatible and, in JHH6, can effectively deliver functional siRNAs. Optimal delivery is observed using a weight ratio Inu-DETA/siRNA of 4 that corresponds to polyplexes with an average size of 600nm and a slightly negative surface charge. Moreover, the uptake and trafficking mechanisms, mainly based on micropinocytosis and clatrin mediated endocytosis, allow the homogeneous diffusion of siRNA within the cytoplasm of JHH6. Notably, in 16 HBE where the trafficking mechanism (caveolae mediated endocytosis) does not allow an even distribution of siRNA within the cell cytoplasm, no significant siRNA activity is observed. In conclusion, we developed a novel inulin-based siRNA delivery system able to efficiently release siRNA in JHH6 with negligible cytotoxicity thus opening the way for further testing in more complex in vivo models.

Peroxiredoxin post-translational modifications by redox messengers.

Peroxiredoxins (Prxs) are a family of thiol peroxidases that participate in hydroperoxide detoxification and regulates H2O2 signaling. In mammals, the four typical 2-Cys Prxs (Prxs 1, 2, 3 and 4) are known to regulate H2O2-mediated intracellular signaling. The 2 catalytic cysteines of 2-Cys Prxs, the so-called peroxidatic and resolving cysteines, are regulatory switches that are prone to react with redox signaling molecules. We investigated the respective modifications induced by H2O2, NO and H2S in the murine macrophage cell line RAW264.7 by mass spectrometry and immunoblotting after separating 2-Cys Prxs by one-dimensional or two-dimensional PAGE. We found that H2S, unlike NO, does not prevent H2O2-mediated sulfinylation of 2-Cys Prxs and that Prx2 is more sensitive to NO-mediated protection against sulfinylation by peroxides. We also observed that cells exposed to exogenous NO, released by Cys-SNO or DETA-NO, or producing NO upon stimulation by IFN-γ and LPS, present an acidic form of Prx1 whose modification is consistent with S-homocysteinylation of its peroxidatic cysteine.

Nitric oxide drives embryonic myogenesis in chicken through the upregulation of myogenic differentiation factors.

The muscle-specific variant of neuronal nitric oxide (NO) synthase (NOS-I), is developmentally regulated in mouse suggesting a role of NO during myogenesis. In chick embryo, a good model of development, we found that the expression of NOS-I is up-regulated, but only in the early phase of development. Through a pharmacological intervention in ovo we found that NO signalling plays a relevant role during embryonic development. The inhibition of NOS-I decreased the growth of embryo, in particular of muscle tissue, while the restoring of physiological NO levels, via administration of a NO donor, reversed this effect. We found a selective action of NO, produced by NOS-I, on regulatory factors involved in myogenic differentiation in the early phase of chick embryo development: inhibition of NO generation leads to a decreased expression of the Myocyte enhancer factor 2a (Mef2a), Mef2c, Myogenin and Myosin, which was reversed by the administration of a NO donor. NO had no effects on Myf5 and MyoD, the myogenic regulatory factors necessary for myogenic determination. The action of NO on the myogenic regulatory factors was mediated via generation of cyclic GMP (cGMP) and activation of the cGMP-dependent protein kinase G (PKG). Finally we found in myoblasts in vitro that the activation of Mef2c was the key event mediating the NO-induced modulation of myogenesis. Our results identify NO produced by NOS-I as a key messenger in the early phase of embryonic development of chicken, acting as a critical determinant of myogenesis through its physiological cGMP/PKG pathway.

Pro-inflammatory cytokines downregulate Hsp27 and cause apoptosis of human retinal capillary endothelial cells.

The formation of acellular capillaries in the retina, a hallmark feature of diabetic retinopathy, is caused by apoptosis of endothelial cells and pericytes. The biochemical mechanism of such apoptosis remains unclear. Small heat shock proteins play an important role in the regulation of apoptosis. In the diabetic retina, pro-inflammatory cytokines are upregulated. In this study, we investigated the effects of pro-inflammatory cytokines on small heat shock protein 27 (Hsp27) in human retinal endothelial cells (HREC). In HREC cultured in the presence of cytokine mixtures (CM), a significant downregulation of Hsp27 at the protein and mRNA level occurred, with no effect on HSF-1, the transcription factor for Hsp27. The presence of high glucose (25mM) amplified the effects of cytokines on Hsp27. CM activated indoleamine 2,3-dioxygenase (IDO) and enhanced the production of kynurenine and ROS. An inhibitor of IDO, 1-methyl tryptophan (MT), inhibited the effects of CM on Hsp27. CM also upregulated NOS2 and, consequently, nitric oxide (NO). A NOS inhibitor, L-NAME, and a ROS scavenger blocked the CM-mediated Hsp27 downregulation. While a NO donor in the culture medium did not decrease the Hsp27 content, a peroxynitrite donor and exogenous peroxynitrite did. The cytokines and high glucose-induced apoptosis of HREC were inhibited by MT and L-NAME. Downregulation of Hsp27 by a siRNA treatment promoted apoptosis in HREC. Together, these data suggest that pro-inflammatory cytokines induce the formation of ROS and NO, which, through the formation of peroxynitrite, reduce the Hsp27 content and bring about apoptosis of retinal capillary endothelial cells.

Effects of DETA/NO on capillary spasm and early brain inj ury after subarachnoid hemorrhage in rats / 西安交通大学学报(医学版)

Objective To explore the effects of diethylenetriamine/nitric oxide (DETA/NO)on capillary spasm and early brain injury (EBI)after subarachnoid hemorrhage (SAH)in rats.Methods Sixty-nine male Sprague-dawley rats were randomized into three groups:sham group,SAH group and DETA/NO group.SAH model was established by wearing out the willis ring with thread and then Garcia neurological score was observed in the general animals.The expressions of alpha smooth muscle actin (αSMA)and PDGFRβwere detected by dual immunofluorescence staining;nitric oxide kit was used for detecting brain tissue NO concentration.Changes in the hemoglobin-stimulated capillaries were observed in rat slices.Results Three days after surgery,neurological deficit score was remarkably improved in DETA/NO group compared with that in SAH group (P<0 .0 5 ). Immunofluorescence results showed that the expressions of peri-capillaryαSMA and PDGFRβwere significantly increased after SAH (P<0.05 ),and that DETA/NO could down-regulate the expressions (P<0.05 ).NO concentration was greatly reduced about 3 hours after SAH and then gradually increased;DETA/NO could maintain the concentration of NO at an early stage (P<0 .0 5 ).The capillary contraction was observed in slices perfused with hemoglobin;DETA/NO could alleviate capillary spasm.Conclusion DETA/NO can alleviate the severity of capillary spasm and EBI after SAH in rats.