The low solubility of drugs, which poses a serious problem in drug development, can in part be overcome by the use of cyclodextrins (CDs) and their derivatives. Here, the key to solubilisation is ...identified as the formation of inclusion complexes with the drug molecule. If inclusion complexation were the only contribution to drug solubility, it would increase linearly with CD concentration (as per the Higuchi-Connors model); this is because inclusion complexation is a 1 : 1 stoichiometric process. However, solubility curves often deviate from this linearity, whose mechanism is yet to be understood. Here we aim to clarify the origin of such non-linearity, based on the Kirkwood-Buff and the McMillan-Mayer theories of solutions. The rigorous statistical thermodynamic theory shows that non-linearity of solubilisation can be rationalised by two contributions: CD-drug interaction and the drug-induced change of CD-CD interaction.
The low solubility of drugs, which poses a serious problem in drug development, can in part be overcome by the use of cyclodextrins (CDs) and their derivatives.
The solubility of cellulose has been studied as a function of composition in the binary mixture of 1,1,3,3-tetramethylguanidine and propionic acid. In amine-rich compositions, greater quantities of ...cellulose can be dissolved than in the equimolar composition, a.k.a. the protic ionic liquid TMGHOPr. By applying a methodology of a short period of heating followed by cooling, similar concentrations of cellulose can be achieved in a much shorter time period. Finally, regeneration of cellulose from solution can be achieved by altering the acid:amine molar ratio. In comparison to cellulose regenerated from these solutions using water as an antisolvent, cellulose regenerated with propionic acid exhibit a lower crystallinity as inferred from x-ray diffractometry, but a greater average molecular weight as inferred from gel permeation chromatography.
•For IgG detection >14 days after symptoms onset was 100.0 % for all assays.•Specificity for IgG was greater than 98 % for CLIA and LFIA compared to ELISA.•LFIA (NG-Test®) is reliable and accurate to ...diagnose SARS-CoV-2 infection.•Best agreement was observed between CLIA and LFIA assays (97 %; k = 0.936).
The emergence of new SARS-CoV-2 has promoted the development of new serological tests that could be complementary to RT-PCR. Nevertheless, the assessment of clinical performances of available tests is urgently required as their use has just been initiated for diagnose.
The aim of this study was to assess the performance of three immunoassays for the detection of SARS-CoV-2 antibodies.
Two automated immunoassays (Abbott SARS-CoV-2 CLIA IgG and Euroimmun Anti-SARS-CoV-2 ELISA IgG/IgA assays) and one lateral flow immunoassay (LFIA NG-Test® IgG-IgM COVID-19) were tested. 293 specimens were analyzed from patients with a positive RT-PCR response, from patients with symptoms consistent with COVID-19 but exhibiting a negative response to the RT-PCR detection test, and from control group specimens. Days since symptoms onset were collected from clinical information sheet associated with respiratory tract samples.
Overall sensitivity for IgG was equivalent (around 80 %) for CLIA, ELISA and LFIA. Sensitivity for IgG detection, >14 days after onset of symptoms, was 100.0 % for all assays. Overall specificity for IgG was greater for CLIA and LFIA (more than 98 %) compared to ELISA (95.8 %). Specificity was significantly different between IgA ELISA (78.9 %) and IgM LFIA (95.8 %) (p < 0.05). The best agreement was observed between CLIA and LFIA assays (97 %; k = 0.936).
Excellent sensitivity for IgG detection was obtained >14 days after onset of symptoms for all immunoassays. Specificity was also excellent for IgG CLIA and IgG LFIA. Our study shows that NG-Test® is reliable and accurate for routine use in clinical laboratories.
Understanding the gelatinization of starch requires a theoretical framework that can extract microscopic information from the literature's abundant experimental data in a quantitative manner. Recent ...developments in statistical thermodynamics have been adapted here to extract the starch-water and starch-salt affinity changes that occur during gelatinization, in a manner beyond the capabilities of traditional polymer theory. We have clarified the mechanism that leads to the increases of salt binding upon gelatinization at high concentrations. Contrary to previous assumptions, salt-starch affinity has been shown to be predominant in the salt effect on gelatinization over water structure contributions. How the temperature of gelatinization depends on salt concentration can be rationalized by the exclusion of salts at low salt concentrations from the gelatinized state and salt anion accessibility to the inside of the starch granule at increased concentration. Finally, the salt anion accessibility informs whether gelatinization progresses from the hilum or the periphery of the starch granule.
Display omitted
•How salts affect starch gelatinization on a molecular basis is clarified.•Solvation effect beyond the reach of the Flory theory can be clarified directly from calorimetry.•Salt exclusion from starch key at low concentration.•Salts' accessibility to starch and consequent binding crucial at higher salt concentration.•Anions play a major role on salt-starch interaction.
In the study of the cellulose dissolution mechanism opinion is still divided. Here, the solution interaction components of the most prominent hypotheses for the driving force of cellulose dissolution ...were evaluated quantitatively. Combining a rigorous statistical thermodynamic theory and cellobiose solubility data in the presence of chloride salts, whose cations progress in the Hofmeister series (KCl, NaCl, LiCl and ZnCl
), we have shown that cellobiose solubilization is driven by the preferential accumulation of salts around the solutes which is stronger than cellobiose hydration. Yet contrary to the classical chaotropy hypothesis, increasing salt concentration leads to cellobiose dehydration in the presence of the strongest solubilizer ZnCl
. However, thanks to cellobiose dehydration, cellobiose-salt interaction still remains preferential despite weakening salt accumulation. Based on such insights, the previous hypotheses based on hydrophobicity and polymer charging have also been evaluated quantitatively. Thus, our present study successfully paved a way towards identifying the basic driving forces for cellulose solubilization in a quantitative manner for the first time. When combined with unit additivity methods this quantitative information could lead to a full understanding of cellulose solubility.
Classical activation of macrophages (M(LPS+IFNγ)) elicits the expression of inducible nitric oxide synthase (iNOS), generating large amounts of NO and inhibiting mitochondrial respiration. ...Upregulation of glycolysis and a disrupted tricarboxylic acid (TCA) cycle underpin this switch to a pro-inflammatory phenotype. We show that the NOS cofactor tetrahydrobiopterin (BH4) modulates IL-1β production and key aspects of metabolic remodeling in activated murine macrophages via NO production. Using two complementary genetic models, we reveal that NO modulates levels of the essential TCA cycle metabolites citrate and succinate, as well as the inflammatory mediator itaconate. Furthermore, NO regulates macrophage respiratory function via changes in the abundance of critical N-module subunits in Complex I. However, NO-deficient cells can still upregulate glycolysis despite changes in the abundance of glycolytic intermediates and proteins involved in glucose metabolism. Our findings reveal a fundamental role for iNOS-derived NO in regulating metabolic remodeling and cytokine production in the pro-inflammatory macrophage.
Display omitted
•NO orchestrates metabolic remodeling in macrophages responding to LPS+IFNγ•NO regulates itaconate metabolism in two models of infection and inflammation•NO determines Complex I subunit abundance in inflammatory macrophages•Glycolysis is increased in activated NO-deficient cells despite metabolic changes
Metabolic remodeling underpins inflammatory macrophage activation, but the modulatory mechanisms are still being elucidated. Bailey et al. show that NO regulates specific changes in the abundance of TCA cycle metabolites, itaconate, and catalytic subunits of Complex I in the respiratory chain in inflammatory murine macrophages both in vitro and in vivo.
Macrophages are mononuclear phagocytes derived from haematopoietic progenitors that are widely distributed throughout the body. These cells participate in both innate and adaptive immune responses ...and lie central to the processes of inflammation, development, and homeostasis. Macrophage physiology varies depending on the environment in which they reside and they exhibit rapid functional adaption in response to external stimuli. To study macrophages in vitro, cells are typically cultured ex vivo from the peritoneum or alveoli, or differentiated from myeloid bone marrow progenitor cells to form bone marrow-derived macrophages (BMDMs). BMDMs represent an efficient and cost-effective means of studying macrophage biology. However, the inherent sensitivity of macrophages to biochemical stimuli (such as cytokines, metabolic intermediates, and RNS/ROS) makes it imperative to control experimental conditions rigorously. Therefore, the aim of this study was to establish an optimised and standardised method for the isolation and culture of BMDMs. We used classically activated macrophages isolated from WT and nitric oxide (NO)-deficient mice to develop a standardised culture method, whereby the constituents of the culture media are defined. We then methodically compared our standardised protocol to the most commonly used method of BMDM culture to establish an optimal protocol for the study of nitric oxide (NO)-redox biology and immunometabolism in vitro.
•We describe a new and optimised protocol to culture BMDMs.•We used classically activated macrophages isolated from WT and NO-deficient mice.•This study emphasises the importance of measuring culture media components and their effects.•GMCSF priming enhances NO secretion upon inflammatory stimulation.•Protocol standardises conditions of differentiation, culture, and inflammatory stimulation.
Superconducting radio frequency (SRF) cavities with quality factors ~10 10 near 4 K have potential to be cooled using regenerative-cycle cryocoolers. Contrary to using liquid helium, cryogen-free ...operation can be realized by conductively linking a cryocooler to a cavity for extracting the cavity RF dissipation. The cavity-cryocooler thermal link needs careful design as its thermal conductance will control the temperatures of the cavity and the cryocooler. We present a thermal analysis of a conduction-cooled SRF cavity that identifies the link thermal conductance requirement. The analysis uses published or expected RF dissipation characteristics of an Nb 3 Sn coated niobium cavity and measured cooling capacity of a pulse tube cryocooler. We describe the mechanical design of a link that is constructed using commercial high-purity aluminum and facilitates bolted-connection to elliptical SRF cavities. The thermal performance of the link is assessed by finite element simulations, taking into account temperature dependent thermal conductivities and measured thermal contact resistance of aluminum and niobium. The link is shown to support operation at an accelerating gradient of 10 MV/m with the lowest-known `perfect' Nb 3 Sn residual surface resistance (~10 nΩ) and also under non-ideal cases that assume certain static heat leak into the system and non-perfect Nb 3 Sn coatings.
New Findings
What is the central question of this study?
What are the physiological roles of cardiomyocyte‐derived tetrahydrobiopterin (BH4) in cardiac metabolism and stress response?
What is the ...main finding and its importance?
Cardiomyocyte BH4 has a physiological role in cardiac metabolism. There was a shift of substrate preference from fatty acid to glucose in hearts with targeted deletion of BH4 synthesis. The changes in fatty‐acid metabolic profile were associated with a protective effect in response to ischaemia–reperfusion (IR) injury, and reduced infarct size. Manipulating fatty acid metabolism via BH4 availability could play a therapeutic role in limiting IR injury.
Tetrahydrobiopterin (BH4) is an essential cofactor for nitric oxide (NO) synthases in which its production of NO is crucial for cardiac function. However, non‐canonical roles of BH4 have been discovered recently and the cell‐specific role of cardiomyocyte BH4 in cardiac function and metabolism remains to be elucidated. Therefore, we developed a novel mouse model of cardiomyocyte BH4 deficiency, by cardiomyocyte‐specific deletion of Gch1, which encodes guanosine triphosphate cyclohydrolase I, a required enzyme for de novo BH4 synthesis. Cardiomyocyte (cm)Gch1 mRNA expression and BH4 levels from cmGch1 KO mice were significantly reduced compared to Gch1flox/flox (WT) littermates. Transcriptomic analyses and protein assays revealed downregulation of genes involved in fatty acid oxidation in cmGch1 KO hearts compared with WT, accompanied by increased triacylglycerol concentration within the myocardium. Deletion of cardiomyocyte BH4 did not alter basal cardiac function. However, the recovery of left ventricle function was improved in cmGch1 KO hearts when subjected to ex vivo ischaemia–reperfusion (IR) injury, with reduced infarct size compared to WT hearts. Metabolomic analyses of cardiac tissue after IR revealed that long‐chain fatty acids were increased in cmGch1 KO hearts compared to WT, whereas at 5 min reperfusion (post‐35 min ischaemia) fatty acid metabolite levels were higher in WT compared to cmGch1 KO hearts. These results indicate a new role for BH4 in cardiomyocyte fatty acid metabolism, such that reduction of cardiomyocyte BH4 confers a protective effect in response to cardiac IR injury. Manipulating cardiac metabolism via BH4 could play a therapeutic role in limiting IR injury.
Peroxisomal fatty acid α-oxidation is an essential pathway for the degradation of β-carbon methylated fatty acids such as phytanic acid. One enzyme in this pathway is 2-hydroxyacyl CoA lyase (HACL1), ...which is responsible for the cleavage of 2-hydroxyphytanoyl-CoA into pristanal and formyl-CoA. Hacl1 deficient mice do not present with a severe phenotype, unlike mice deficient in other α-oxidation enzymes such as phytanoyl-CoA hydroxylase deficiency (Refsum disease) in which neuropathy and ataxia are present. Tissues from wild-type and
mice fed a high phytol diet were obtained for proteomic and lipidomic analysis. There was no phenotype observed in these mice. Liver, brain, and kidney tissues underwent trypsin digestion for untargeted proteomic liquid chromatography-mass spectrometry analysis, while liver tissues also underwent fatty acid hydrolysis, extraction, and derivatisation for fatty acid gas chromatography-mass spectrometry analysis. The liver fatty acid profile demonstrated an accumulation of phytanic and 2-hydroxyphytanic acid in the
liver and significant decrease in heptadecanoic acid. The liver proteome showed a significant decrease in the abundance of Hacl1 and a significant increase in the abundance of proteins involved in PPAR signalling, peroxisome proliferation, and omega oxidation, particularly Cyp4a10 and Cyp4a14. In addition, the pathway associated with arachidonic acid metabolism was affected; Cyp2c55 was upregulated and Cyp4f14 and Cyp2b9 were downregulated. The kidney proteome revealed fewer significantly upregulated peroxisomal proteins and the brain proteome was not significantly different in
mice. This study demonstrates the powerful insight brought by proteomic and metabolomic profiling of
mice in better understanding disease mechanism in fatty acid α-oxidation disorders.
PDF
