Copper(II), Nickel(II) and Zinc(II) Complexes of Peptide Fragments of Tau Protein.

Copper(II), nickel(II) and zinc(II) complexes of various peptide fragments of tau protein were studied by potentiometric and spectroscopic techniques. All peptides contained one histidyl residue and represented the sequences of tau(91-97) (Ac-AQPHTEI-NH2), tau(385-390) (Ac-KTDHGA-NH2) and tau(404-409) (Ac-SPRHLS-NH2). Imidazole-N donors of histidine were the primary metal binding sites for all peptides and all metal ions, but in the case of copper(II) and nickel(II), the deprotonated amide groups were also involved in metal binding by increasing pH. The most stable complexes were formed with copper(II) ions, but the presence of prolyl residues resulted in significant changes in the thermodynamic stability and speciation of the systems. It was also demonstrated that nickel(II) and especially zinc(II) complexes have relatively low thermodynamic stability with these peptides. The copper(II)-catalyzed oxidation of the peptides was also studied. In the presence of H2O2, the fragmentation of peptides was detected in all cases. In the simultaneous presence of H2O2 and ascorbic acid, the fragmentation of the peptide is less preferred, and the formation of 2-oxo-histidine also occurs.

Microstrip Antenna Development for Radar Sensor.

Tank level measurement is an important research area in radar technology. Level measurement using radar technology is a safe solution even under extreme process conditions, such as high pressure, temperature, and vapors. Special antennas are required to meet electromagnetic requirements such as high gain, low sidelobe level, and high bandwidth. Another requirement is a small size and good manufacturability of the antenna. One promising solution is the use of microstrip antennas for tank-level measurements. This paper presents a special microstrip antenna, that meets the main requirements in addition to a circular antenna layout, which fills optimally the tank hatch. To do this, we created a special antenna layout with a suitable feeding network and added an optimized ring around the antenna system, which significantly reduces the sidelobe level. The center frequency of the antenna is 25 GHz with a 1 GHz bandwidth.

Deuterium Content of the Organic Compounds in Food Has an Impact on Tumor Growth in Mice.

Research with deuterium-depleted water (DDW) in the last two decades proved that the deuterium/hydrogen ratio has a key role in cell cycle regulation and cellular metabolism. The present study aimed to investigate the possible effect of deuterium-depleted yolk (DDyolk) alone and in combination with DDW on cancer growth in two in vivo mouse models. To produce DDyolk, the drinking water of laying hens was replaced with DDW (25 ppm) for 6 weeks, resulting in a 60 ppm D level in dried egg yolk that was used as a deuterium-depleted food additive. In one model, 4T1, a cell line with a high metastatic capacity to the lung was inoculated in the mice's mammary pad. After three weeks of treatment with DDW and/or DDyolk, the tumor volume in the lungs was smaller in all treated groups vs. controls with natural D levels. Tumor growth and survival in mice transplanted with an MCF-7 breast cancer cell line showed that the anticancer effect of DDW was enhanced by food containing the deuterium-depleted yolk. The study confirmed the importance of the D/H ratio in consumed water and in metabolic water produced by the mitochondria while oxidizing nutrient molecules. This is in line with the concept that the initiation of cell growth requires the cells to generate a higher D/H ratio, but DDW, DDyolk, or the naturally low-D lipids in a ketogenic diet, have a significant effect on tumor growth by preventing the cells from raising the D/H ratio to the threshold.

Blocking the Increase of Intracellular Deuterium Concentration Prevents the Expression of Cancer-Related Genes, Tumor Development, and Tumor Recurrence in Cancer Patients.

The possible role of the naturally occurring deuterium in the regulation of cell division was first described in the 1990s. To investigate the mechanism of influence of deuterium (D) on cell growth, expression of 236 cancer-related and 536 kinase genes were tested in deuterium-depleted (40 and 80 ppm) and deuterium-enriched (300 ppm) media compared to natural D level (150 ppm). Among genes with expression changes exceeding 30% and copy numbers over 30 (124 and 135 genes, respectively) 97.3% of them was upregulated at 300 ppm D-concentration. In mice exposed to chemical carcinogen, one-year survival data showed that deuterium-depleted water (DDW) with 30 ppm D as drinking water prevented tumor development. One quarter of the treated male mice survived 344 days, the females 334 days, while one quarter of the control mice survived only 188 and 156 days, respectively. In our human retrospective study 204 previously treated cancer patients with disease in remission, who consumed DDW, were followed. Cumulative follow-up time was 1024 years, and average follow-up time per patient, 5 years (median: 3.6 years). One hundred and fifty-six patients out of 204 (77.9%) did not relapse during their 803 years cumulative follow-up time. Median survival time (MST) was not calculable due to the extremely low death rate (11 cancer-related deaths, 5.4% of the study population). Importantly, 8 out of 11 deaths occurred several years after stopping DDW consumption, confirming that regular consumption of DDW can prevent recurrence of cancer. These findings point to the likely mechanism in which consumption of DDW keeps D-concentration below natural levels, preventing the D/H ratio from increasing to the threshold required for cell division. This in turn can serve as a key to reduce the relapse rate of cancer patients and/or to reduce cancer incidence in healthy populations.

Full Equilibrium Picture in Aqueous Binary and Ternary Systems Involving Copper(II), 1-Methylimidazole-Containing Hydrazonic Ligands, and the 103-112 Human Prion Protein Fragment.

In this work, we describe two novel 1-methylimidazole N-acylhydyrazonic ligands and their interaction with copper(II) in solution. Binary systems constituted by each of these hydrazones and the metal ion were studied by potentiometric titrations. The magnitude of their affinities for zinc(II) was also determined for the sake of comparison. Additionally, a full evaluation of the copper(II) chelation profile of the new ligands in ternary systems containing a human prion protein fragment was performed. Mixed ligand complexes comprising the HuPrP103-112 fragment, copper(II) ions, and an N-acylhydrazone were characterized by potentiometry, ultraviolet-visible spectroscopy, and circular dichroism. Some of these species were also identified by electrospray ionization mass spectrometry and unequivocally assigned through their isotopic distribution pattern. To the best of our knowledge, this is the first report concerning the stability of ternary complexes involving a hydrazonic metal-protein interaction modulator, copper, and a peptide. The ability of N-acylhydrazones to prevent peptide oxidation was also examined. Both ligands can partially prevent the formation of the doubly oxidized product, a process mediated by copper(II) ions. Oxidative stress is considered an important hallmark of neurodegenerative diseases such as prion-related spongiform encephalopathies. In this context, active intervention with respect to the deleterious copper-catalyzed methionine oxidation could represent an interesting therapeutic approach.

Synthesis of Sucrose-HDI Cooligomers: New Polyols for Novel Polyurethane Networks.

Sucrose-1,6-hexamethylene diisocyanate (HDI) cooligomers were synthesized and used as new polyols for poly(ε-caprolactone) (PCL)-based polyurethanes. The polyaddition reaction of sucrose and HDI was monitored by MALDI-TOF MS. It was found that by selecting appropriate reaction conditions, mostly linear oligomer chains containing 16 sucrose units could be obtained. For the synthesis of polyurethane networks, prepolymers were prepared by the reaction of poly(ε-caprolactone) (PCL, 10 kg/mol) with HDI or 4,4'-methylene diphenyl diisocyanate (MDI) and were reacted with sucrose-HDI cooligomers. The so-obtained sucrose-containing polyurethanes were characterized by means of attenuated total reflectance Fourier-transform infrared spectroscopy (ATR-FT IR), swelling, mechanical (uniaxial tensile tests) and differential scanning calorimetry (DSC).

Isocyanonaphthol Derivatives: Excited-State Proton Transfer and Solvatochromic Properties.

Fluorescent probes that exhibit solvatochromic or excited-state proton-transfer (ESPT) properties are essential tools for the study of complex biological or chemical systems. Herein, the synthesis and characterization of a novel fluorophore that reveals both features, 5-isocyanonaphthalene-1-ol (ICOL), are reported. Various solvatochromic methods, such as Lippert−Mataga and Bilot−Kawski, together with time-dependent density functional theory (TD-DFT) and time-resolved emission spectroscopy (TRES), were applied to gain insights into its excited-state behavior. To make comparisons, the octyloxy derivative of ICOL, 5-isocyano-1-(octyloxy)naphthalene (ICON), was also prepared. We found that internal charge transfer (ICT) takes place between the isocyano and −OH groups of ICOL, and we determined the values of the dipole moments for the ground and excited states of both ICOL and ICON. Furthermore, in the emission spectra of ICOL, a second band at higher wavelengths (green emission) in solvents of higher polarities (dual emission), in addition to the band present at lower wavelengths (blue emission), were observed. The extent of this dual emission increases in the order of 2-propanol < methanol < N,N-dimethylformamide (DMF) < dimethyl sulfoxide (DMSO). The presence of the dual fluorescence of ICOL in these solvents can be ascribed to ESPT. For ICOL, we also determined ground- and excited-state pKa values of 8.4 ± 0.3 and 0.9 ± 0.7, respectively, which indicates a considerable increase in acidity upon excitation. The TRES experiments showed that the excited-state lifetimes of the ICOL and ICON spanned from 10.1 ns to 5.0 ns and from 5.7 ns to 3.8 ns, respectively. In addition, we demonstrated that ICOL can be used as an effective indicator of not only the critical micelle concentration (cmc) of ionic (sodium lauryl sulfate (SLS)) and nonionic surfactants (Tween 80), but also other micellar parameters, such as partition coefficients, as well as to map the microenvironments in the cavities of biomacromolecules (e.g., BSA). It is also pointed out that fluorescence quenching by pyridine can effectively be utilized for the determination of the fractions of ICOL molecules that reside at the water−micelle interface and in the interior spaces of micelles.

Comparison of Homologous and Heterologous Booster SARS-CoV-2 Vaccination in Autoimmune Rheumatic and Musculoskeletal Patients.

Vaccination against SARS-CoV-2 to prevent COVID-19 is highly recommended for immunocompromised patients with autoimmune rheumatic and musculoskeletal diseases (aiRMDs). Little is known about the effect of booster vaccination or infection followed by previously completed two-dose vaccination in aiRMDs. We determined neutralizing anti-SARS-CoV-2 antibody levels and applied flow cytometric immunophenotyping to quantify the SARS-CoV-2 reactive B- and T-cell mediated immunity in aiRMDs receiving homologous or heterologous boosters or acquired infection following vaccination. Patients receiving a heterologous booster had a higher proportion of IgM+ SARS-CoV-2 S+ CD19+CD27+ peripheral memory B-cells in comparison to those who acquired infection. Biologic therapy decreased the number of S+CD19+; S+CD19+CD27+IgG+; and S+CD19+CD27+IgM+ B-cells. The response rate to a booster event in cellular immunity was the highest in the S-, M-, and N-reactive CD4+CD40L+ T-cell subset. Patients with a disease duration of more than 10 years had higher proportions of CD8+TNF-α+ and CD8+IFN-γ+ T-cells in comparison to patients who were diagnosed less than 10 years ago. We detected neutralizing antibodies, S+ reactive peripheral memory B-cells, and five S-, M-, and N-reactive T-cells subsets in our patient cohort showing the importance of booster events. Biologic therapy and <10 years disease duration may confound anti-SARS-CoV-2 specific immunity in aiRMDs.

Investigation of oxidative stress in patients with multifocal motor neuropathy.

Background and purpose: Multifocal motor neuropathy (MMN) is a rare, immune-mediated illness attacking ex-clusively motor nerves. It is known that oxidative stress is present in peripheral neuropathies, but it has not been investigated MMN. Methods: We measured in our prospective study the L-arginine, symmetric and asymmetric dimethylarginine (SDMA, ADMA) serum concentrations of 10 patients and 10 controls before and after intravenous immunoglobulin treatment (IVIG), as markers of the L-arginine/NO pathway involved in chronic inflammation and oxidative stress. The functions of motor nerves were tested in all patients and the serum antiganglioside antibody levels were de-tec-ted, as well. Results: MMN patients showed significantly higher ADMA (p = 0.0048; 0.98 and 0.63, respectively) and SDMA le-vels (p = 0.001; 0.88 and 0.51, respectively) than healthy controls, while L-arginine was not different. Controlling for the covariant age, ADMA (B = -0.474; p = 0.041) or SDMA (B = -0.896; p < 0.0005) serum levels proved to be the significant predictors of the presence of MMN. IVIG therapy decreased significantly ADMA concentrations (p = 0.025; 0.98 and 0.84, respectively) and showed a trend to reduce SDMA levels (p = 0.1; 0.88 and 0.74, respectively). The dimethylamine levels did not correlate with the number of affected nerves, disease duration, or the presence of ganglioside antibodies. The conduction block-related peripheral motor dysfunction improved right after the IVIG treatment. Conclusion: Dimethylamine levels are elevated in the serum and are responsive to IVIG therapy in MMN. These findings support the presence of oxidative stress in MMN.

Deuterium Depletion Inhibits Cell Proliferation, RNA and Nuclear Membrane Turnover to Enhance Survival in Pancreatic Cancer.

The effects of deuterium-depleted water (DDW) containing deuterium (D) at a concentration of 25 parts per million (ppm), 50 ppm, 105 ppm and the control at 150 ppm were monitored in MIA-PaCa-2 pancreatic cancer cells by the real-time cell impedance detection xCELLigence method. The data revealed that lower deuterium concentrations corresponded to lower MiA PaCa-2 growth rate. Nuclear membrane turnover and nucleic acid synthesis rate at different D-concentrations were determined by targeted [1,2-13C2]-D-glucose fate associations. The data showed severely decreased oxidative pentose cycling, RNA ribose 13C labeling from [1,2-13C2]-D-glucose and nuclear membrane lignoceric (C24:0) acid turnover. Here, we treated advanced pancreatic cancer patients with DDW as an extra-mitochondrial deuterium-depleting strategy and evaluated overall patient survival. Eighty-six (36 male and 50 female) pancreatic adenocarcinoma patients were treated with conventional chemotherapy and natural water (control, 30 patients) or 85 ppm DDW (56 patients), which was gradually decreased to preparations with 65 ppm and 45 ppm deuterium content for each 1 to 3 months treatment period. Patient survival curves were calculated by the Kaplan-Meier method and Pearson correlation was taken between medial survival time (MST) and DDW treatment in pancreatic cancer patients. The MST for patients consuming DDW treatment (n = 56) was 19.6 months in comparison with the 6.36 months' MST achieved with chemotherapy alone (n = 30). There was a strong, statistically significant Pearson correlation (r = 0.504, p < 0.001) between survival time and length and frequency of DDW treatment.

Kinetically Equivalent Functionality and Reactivity of Commonly Used Biocompatible Polyurethane Crosslinking Agents.

In this paper, the kinetics of the reaction of phenyl isocyanate with crosslinking agents such as sucrose, sorbitol, and glycerol are reported. Crosslinking agents were used in high molar excess to isocyanate to obtain pseudo-first-order rate dependencies, and the reaction products were separated by high-performance liquid chromatography and detected by UV spectroscopy and mass spectrometry. It was found that the glycerol's primary hydroxyl groups were approximately four times reactive than the secondary ones. However, in the case of sorbitol, the two primary OH groups were found to be the most reactive, and the reactivity of hydroxyl groups decreased in the order of kOH(6)(8.43) > kOH(1)(6.91) > kOH(5)(1.19) > kOH(2)(0.98) > kOH(3)(0.93) > kOH(4)(0.64), where the numbers in the subscript and in the brackets denote the position of OH groups and the pseudo-first-order rate constants, respectively. The Atomic Polar Tenzor (APT) charges of OH groups and dipole moments of monosubstituted sorbitol derivatives calculated by density functional theory (DFT) also confirmed the experimental results. On the other hand, the reactions of phenyl isocyanate with crosslinking agents were also performed using high excess isocyanate in order to determine the number of OH-groups participating effectively in the crosslinking process. However, due to the huge number of derivatives likely formed in these latter reactions, a simplified reaction scheme was introduced to describe the resulting product versus reaction time distributions detected by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS). Based on the results, the kinetically equivalent functionality (fk) of each crosslinking agent was determined and found to be 2.26, 2.6, and 2.96 for glycerol, sorbitol, and sucrose, respectively.

Associations between serum L-arginine and ficolins in the early phase of acute ischemic stroke - A pilot study.

INTRODUCTION: Activation of both the L-arginine and the lectin pathway contributes to the pathophysiology and the outcome of acute ischemic stroke (AIS). However, the interplay between the two systems has not yet been examined. METHODS: A total of 44 patients with AIS were recruited into this study. Serial measurement of serum L-arginine, asymmetric and symmetric dimethylarginine (ADMA, SDMA), and hsCRP, ficolin-2, ficolin-3, MAP-1, MASP-3 and mannose-binding lectin (MBL) were analyzed within 6 h after onset of stroke and 72 h later. Outcomes were assessed as National Institutes of Health Stroke Scale (NIHSS) worsening by 24 h, poststroke infection, and death by 1 month. RESULTS: In the hyperacute stage of AIS, ficolin-3, MAP-1 and MBL were positively correlated with L-arginine within 6 h after onset of symptoms (p<0.05 respectively). Significantly lower ficolin-3 and MASP-3 levels were found at 72 h in patients, who developed post-stroke infection after day 4, when compared to patients without post-stroke infections (p=0.03 and p=0.009). At 72 hours, ficolin-3 levels negatively correlated with S100B (p=0.01). Ficolin-3 at 72 post-stroke hours remained an independent predictor of post-stroke infection, while only hsCRP was an independent predictor of 30-day mortality. CONCLUSION: Early consumption of ficolin-3 is associated with complications such as post-stroke infections. In the hyperacute phase of AIS, the positive correlation between ficolins and the NO donor L-arginine may reflect the protective role of L-arginine presumably by improving the cerebral microcirculation in a prothrombotic environment induced by complement activation.

Conversion of Isocyanide to Amine in The Presence of Water and Hg(II) Ions: Kinetics and Mechanism as Detected by Fluorescence Spectroscopy and Mass Spectrometry.

Aromatic isocyanides including isocyanonaphthalene derivatives have been proven to be very effective fluorescent sensors for the quantification of Hg(II) ions in water. Thus, the reaction of 1,5-isocyanoaminonaphthalene (1,5-ICAN), which is one of the most important members of this family, with water and HgCl2 as the oxidation agents, was studied by fluorescence spectroscopy and mass spectrometry in order to get deeper insight into the kinetics and mechanistic details of this complex reaction. The reactions of 1,5-ICAN with water and HgCl2 were performed in various water/co-solvent mixtures of different compositions. The co-solvents used in this study were both aprotic solvents including tetrahydrofuran, acetonitrile and N,N-dimethylformamide and protic solvents, such as ethanol and 2-propanol. It was found that in aprotic solvents the conversion of the isocyano group to amino moiety takes place, while in protic solvents the corresponding carbamate (urethane) group is formed in addition to the amino moiety. The variation of the resulting fluorescence intensities versus time curves were described using an irreversible, consecutive reaction model, in which the formation of isocyanate and carbamic acid intermediates, as well as diamino and carbamate (in the case of protic solvents) products were assumed. The formation of these intermediates and products was unambiguously confirmed by mass spectrometric measurements. Furthermore, by fitting the model to the experimental fluorescence versus time curves, the corresponding rate coefficients were determined. It was observed that the overall rate of transformation of the isocyano group to amino moiety increased with the water concentration and the polarity of the co-solvent. It was also supported that formation of diamino and carbamate derivatives in protic solvents takes place simultaneously and that the ratio of the amino to the carbamate function increased with the increasing water concentration. In addition, with an extension, the model presented herein proved to be capable of describing the kinetics of the transformation of 1,5-diisocyanonaphthalene (1,5-DIN) into 1,5-diaminonaphthalene (1,5-DAN) in the mixtures of water/aprotic solvents.

Encoding Information into Polyethylene Glycol Using an Alcohol-Isocyanate "Click" Reaction.

In this article, the capability of encoding information using a homologous series of monodisperse monomethoxypolyethylene glycols (mPEG), with a number of ethylene oxide units ranging from nEO = 5 to 8, and monodisperse linear aliphatic isocyanates containing a number of CH2 units from 3 to 7, is demonstrated. The "click" reaction of the two corresponding homologous series yielded 20 different isocyanate end-capped polyethylene glycol derivatives (mPEG-OCONHR) whose sodiated adduct ion's nominal m/z values spanned from 360 to 548, providing an average ca. 8 m/z unit for the storage of one-bit information. These mPEG-OCONHR oligomers were then used to encode information in binary sequences using a 384-well MALDI sample plate and employing the common dried-droplet sample preparation method capable of encoding 20 bit, i.e., 2.5 byte information in one spot, was employed. The information stored in the spots was read by MALDI-TOF MS using the m/z value of the corresponding mPEG-OCONHR oligomers. The capability of the method to store data was demonstrated by writing and reading a text file, visualizing a small picture and capturing a short audio file written in Musical Instrument Digital Interface (MIDI) sequence. Due to the very large similarities in the chemical structures of the encoding oligomers and their "easy to be ionized" property, as well as their very similar ionization efficiencies, the MALDI-TOF MS signal intensities from each compound was so strong and unambiguous that complete decoding could be performed in each case. In addition, the set of the proposed encoding oligomers can be further extended to attain higher bit "densities".

Imidazo[1,2-b]pyrazole-7-Carboxamide Derivative Induces Differentiation-Coupled Apoptosis of Immature Myeloid Cells Such as Acute Myeloid Leukemia and Myeloid-Derived Suppressor Cells.

Chemotherapy-induced differentiation of immature myeloid progenitors, such as acute myeloid leukemia (AML) cells or myeloid-derived suppressor cells (MDSCs), has remained a challenge for the clinicians. Testing our imidazo[1,2-b]pyrazole-7-carboxamide derivative on HL-60 cells, we obtained ERK phosphorylation as an early survival response to treatment followed by the increase of the percentage of the Bcl-xlbright and pAktbright cells. Following the induction of Vav1 and the AP-1 complex, a driver of cellular differentiation, FOS, JUN, JUNB, and JUND were elevated on a concentration and time-dependent manner. As a proof of granulocytic differentiation, the cells remained non-adherent, the expression of CD33 decreased; the granularity, CD11b expression, and MPO activity of HL-60 cells increased upon treatment. Finally, viability of HL-60 cells was hampered shown by the depolarization of mitochondria, activation of caspase-3, cleavage of Z-DEVD-aLUC, appearance of the sub-G1 population, and the leakage of the lactate-dehydrogenase into the supernatant. We confirmed the differentiating effect of our drug candidate on human patient-derived AML cells shown by the increase of CD11b and decrease of CD33+, CD7+, CD206+, and CD38bright cells followed apoptosis (IC50: 80 nM) after treatment ex vivo. Our compound reduced both CD11b+/Ly6C+ and CD11b+/Ly6G+ splenic MDSCs from the murine 4T1 breast cancer model ex vivo.

Impact of pyridine-2-carboxaldehyde-derived aroylhydrazones on the copper-catalyzed oxidation of the M112A PrP103-112 mutant fragment.

Misfolded prion protein (PrPSc) is known for its role in fatal neurodegenerative conditions, such as Creutzfeldt-Jakob disease. PrP fragments and their mutants represent important tools in the investigation of the neurotoxic mechanisms and in the evaluation of new compounds that can interfere with the processes involved in neuronal death. Metal-catalyzed oxidation of PrP has been implicated as a trigger for the conformational changes in protein structure, which, in turn, lead to misfolding. Targeting redox-active biometals copper and iron is relevant in the context of protection against the oxidation of biomolecules and the generation of oxidative stress, observed in several conditions and considered an event that might promote sporadic prion diseases as well as other neurodegenerative disorders. In this context, ortho-pyridine aroylhydrazones are of interest, as they can act as moderate tridentate ligands towards divalent metal ions such as copper(II). In the present work, we explore the potentiality of this chemical class as peptide protecting agents against the deleterious metal-catalyzed oxidation in the M112A mutant fragment of human PrP, which mimics relevant structural features that may play an important role in the neurotoxicity observed in prion pathologies. The compounds inhere studied, especially HPCFur, showed an improved stability in aqueous solution compared to our patented lead hydrazone INHHQ, displaying a very interesting protective effect toward the oxidation of methionine and histidine, processes that are related to both physiological and pathological aging.

Dysregulation of the endothelial nitric oxide pathway is associated with airway inflammation in COPD.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is related to endothelial dysfunction and the impaired generation of nitric oxide (NO) from L-arginine by the endothelial NO synthase (eNOS). The relationship between eNOS dysfunctionality and airway inflammation is unknown. We assessed serum asymmetric and symmetric dimethylarginine (ADMA and SDMA) and nitrite/nitrate concentrations, indicators of eNOS function, in patients with COPD and correlated them with markers of inflammation. METHODS: We recruited 15 control smokers, 29 patients with stable and 32 patients with exacerbated COPD requiring hospitalization (20 of them were measured both at admission and discharge). Serum L-arginine, ADMA, SDMA, nitrite and nitrate were measured and correlated with airway inflammatory markers (fractional exhaled nitric oxide concentration - FENO, sputum nitrite and nitrate, sputum cellularity), serum C-reactive protein - CRP, white blood cell count, lung function and blood gases. ANOVA, t-tests and Pearson correlation were used (mean ± SD or geometric mean ± geometric SD for nitrite/nitrate). RESULTS: Serum L-arginine/ADMA, a marker of substrate availability for eNOS, was lower in stable (214 ± 58, p < 0.01) and exacerbated COPD (231 ± 68, p < 0.05) than in controls (287 ± 64). The serum concentration of SDMA, a competitor of L-arginine transport, was elevated during an exacerbation (0.78 ± 0.39 µM) compared to stable patients (0.53 ± 0.14 µM, p < 0.01) and controls (0.45 ± 0.14 µM, p < 0.001). ADMA correlated with blood neutrophil percentage (r = 0.36, p < 0.01), FENO (r = 0.42, p < 0.01) and a tendency for positive association was observed to sputum neutrophil count (r = 0.33, p = 0.07). SDMA correlated with total sputum inflammatory cell count (r = 0.61, p < 0.01) and sputum neutrophil count (r = 0.62, p < 0.01). Markers were not related to lung function, blood gases or CRP. L-arginine/ADMA was unchanged, but serum SDMA level decreased (0.57 ± 0.42 µM, p < 0.05) after systemic steroid treatment of the exacerbation. Serum nitrite level increased in stable and exacerbated disease (4.11 ± 2.12 and 4.03 ± 1.77 vs. control: 1.61 ± 1.84 µM, both p < 0.001). CONCLUSIONS: Our data suggest impaired eNOS function in stable COPD, which is transiently aggravated during an exacerbation and partly reversed by systemic steroid treatment. Serum ADMA and SDMA correlate with airway inflammatory markers implying a possible effect of anti-inflammatory therapy on endothelial dysfunction. Serum nitrite can serve as a compensatory pool for impaired endothelial NO generation.

Single Cell Mass Cytometry Revealed the Immunomodulatory Effect of Cisplatin Via Downregulation of Splenic CD44+, IL-17A+ MDSCs and Promotion of Circulating IFN-γ+ Myeloid Cells in the 4T1 Metastatic Breast Cancer Model.

The treatment of metastatic breast cancer remained a challenge despite the recent breakthrough in the immunotherapy regimens. Here, we addressed the multidimensional immunophenotyping of 4T1 metastatic breast cancer by the state-of-the-art single cell mass cytometry (CyTOF). We determined the dose and time dependent cytotoxicity of cisplatin on 4T1 cells by the xCelligence real-time electronic sensing assay. Cisplatin treatment reduced tumor growth, number of lung metastasis, and the splenomegaly of 4T1 tumor bearing mice. We showed that cisplatin inhibited the tumor stroma formation, the polarization of carcinoma-associated fibroblasts by the diminished proteolytic activity of fibroblast activating protein. The CyTOF analysis revealed the emergence of CD11b+/Gr-1+/CD44+ or CD11b+/Gr-1+/IL-17A+ myeloid-derived suppressor cells (MDSCs) and the absence of B220+ or CD62L+ B-cells, the CD62L+/CD4+ and CD62L+/CD8+ T-cells in the spleen of advanced cancer. We could show the immunomodulatory effect of cisplatin via the suppression of splenic MDSCs and via the promotion of peripheral IFN-γ+ myeloid cells. Our data could support the use of low dose chemotherapy with cisplatin as an immunomodulatory agent for metastatic triple negative breast cancer.

The Anti-Inflammatory Role of Mannich Curcuminoids; Special Focus on Colitis.

The incidence of inflammatory bowel disease (IBD) increases gradually in Western countries with high need for novel therapeutic interventions. Mannich curcuminoids, C142 or C150 synthetized in our laboratory, have been tested for anti-inflammatory activity in a rat model of TNBS (2,4,6-trinitrobenzenesulphonic acid) induced colitis. Treatment with C142 or C150 reduced leukocyte infiltration to the submucosa and muscular propria of the inflamed gut. C142 or C150 rescued the loss of body weight and C150 decreased the weight of standard colon preparations proportional with 20% less tissue oedema. Both C142 and C150 curcumin analogues caused 25% decrease in the severity of colonic inflammation and haemorrhagic lesion size. Colonic MPO (myeloperoxidase) enzyme activity as an indicator of intense neutrophil infiltration was 50% decreased either by C142 or C150 Mannich curcuminoids. Lipopolysaccharide (LPS) co-treatment with Mannich curcuminoids inhibited NF-κB (nuclear factor kappa B) activity on a concentration-dependent manner in an NF-κB-driven luciferase expressing reporter cell line. Co-treatment with LPS and curcuminoids, C142 or C150, resulted in NF-κB inhibition with 3.57 µM or 1.6 µM half maximal effective concentration (EC50) values, respectively. C150 exerted a profound inhibition of the expression of inflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-4 (IL-4) in human PBMCs (peripheral blood mononuclear cells) upon LPS stimulus. Mannich curcuminoids reported herein possess a powerful anti-inflammatory activity.

L-arginine pathway metabolites can discriminate paroxysmal from permanent atrial fibrillation in acute ischemic stroke.

BACKGROUND AND PURPOSE: Atrial fibrillation (AF) is the most common arrhythmia diagnosed in clinical practice. We aimed to measure the L-arginine pathway metabolites as well as their ratios in patients with different types of AF or sinus rhythm and to explore the relationship among the markers and clinical variables in the subacute phase of acute ischemic stroke (AIS). METHODS: A total of 46 patients with AIS were prospectively enrolled. The patients were divided into three groups based on diagnosis of either sinus rhythm, paroxysmal or permanent AF. Plasma concentration of the L-arginine pathway metabolites were analyzed at post-stroke 24 hours in the three rhythm groups. Besides, clinical variables and laboratory data were recorded. RESULTS: Asymmetric dimetylarginine (ADMA) was significantly higher in patients with permanent AF compared to sinus rhythm (p<0.001). Both ADMA (p<0.001) and symmetric dimethylarginine (SDMA) (p<0.002) at 24 hours were significantly higher among patients with permanent AF compared to those with paroxysmal AF. The L-arginine/SDMA (p<0.031) ratios at 24 hours were significantly higher among patients with sinus rhythm compared to those with permanent AF. ROC analysis also revealed that plasma SDMA cut-off level over 0.639 µmol/L discriminated permanent AF from paroxysmal AF or sinus rhythm with a 90.9% sensitivity and 77.1% specificity. Neutrophil-lymphocyte ratio also showed significantly higher value in individuals with both paroxysmal and permanent AF (p=0.029). CONCLUSION: Plasma level of SDMA could discriminate permanent from paroxysmal AF in the subacute phase of ischemic stroke. In addition, an increased neutrophil-lymphocyte ratio may suggest inflammatory process in the evolution of atrial fibrillation.