Bud14 function is crucial for spindle pole body size maintenance.

Background/aim: Spindle pole bodies (SPB), the functional equivalent of centrosomes in yeast, duplicate through generation of a new SPB next to the old one. However, SPBs are dynamic structures that can grow and exchange, and mechanisms that regulate SPB size remain largely unknown. This study aims to elucidate the role of Bud14 in SPB size maintenance in Saccharomyces cerevisiae. Materials and methods: We employed quantitative fluorescence microscopy to assess the relative and absolute amounts of SPB structural proteins at SPBs of wildtype cells and in cells lacking BUD14 (bud14Δ). Quantifications were performed using asynchronous cell cultures, as well as cultures synchronously progressing through the cell cycle and upon different cell cycle arrests. We also utilized mutants that allow the separation of Bud14 functions. Results: Our results indicate that higher levels of SPB inner, outer, and central plaque proteins are present at the SPBs of bud14Δ cells compared to wildtype cells during anaphase, as well as during nocodazole-induced M-phase arrest. However, during α-factor mediated G1 arrest, inner and outer plaque proteins responded differently to the absence of BUD14. A Bud14 mutant that cannot interact with the Protein Phosphatase 1 (Glc7) phenocopied bud14Δ in terms of SPB-bound levels of the inner plaque protein Spc110, whereas disruption of Bud14-Kel1-Kel2 complex did not alter Spc110 levels at SPBs. In cells synchronously released from α-factor arrest, lack of Bud14-Glc7 caused increase of Spc110 at the SPBs at early stages of the cell cycle. Conclusion: We identified Bud14 as a critical protein for SPB size maintenance. The interaction of Bud14 with Glc7, but not with the Kelch proteins, is indispensable for restricting levels of Spc110 incorporated into the SPBs.

Engineering PTS-based glucose metabolism for efficient biosynthesis of bacterial cellulose by Komagataeibacter xylinus.

Bacterial cellulose (BC) is a renewable biomaterial that has attracted significant attention due to its excellent properties and wide applications. Komagataeibacter xylinus CGMCC 2955 is an important BC-producing strain. It primarily produces BC from glucose while simultaneously generating gluconic acid as a by-product, which acidifies the medium and inhibits BC synthesis. To enhance glucose uptake and BC synthesis, we reconstructed the phosphoenolpyruvate-dependent glucose phosphotransferase system (PTSGlc) and strengthened glycolysis by introducing heterologous genes, resulting in a recombinant strain (GX08PTS03; Δgcd::ptsHIcrrE. coli::ptsGE. coli::pfkAE. coli). Strain GX08PTS03 efficiently utilized glucose for BC production without accumulating gluconic acid. Subsequently, the fermentation process was systematically optimized. Under optimal conditions, strain GX08PTS03 produced 7.74 g/L of BC after 6 days of static fermentation, with a BC yield of 0.39 g/g glucose, which were 87.41 % and 77.27 % higher than those of the wild-type strain, respectively. The BC produced by strain GX08PTS03 exhibited a longer fiber diameter along with a lower porosity, significantly higher solid content, crystallinity, tensile strength, and Young's modulus. This study is novel in reporting that the engineered PTSGlc-based glucose metabolism could effectively enhance the production and properties of BC, providing a future outlook for the biopolymer industry.

Isolation and Molecular Characterization of the LTA precursor molecule Glc2-DAG, a potential target for antibiotics.

Bacterial infections present a major global health threat, often displaying resistance to various antibiotics. Lipoteichoic acid (LTA) is a vital component of bacterial cell envelopes of Gram-positive bacteria, crucial for cell integrity, cell division, and host inflammation. Due to its essential role for bacteria, LTA and its biosynthesis are also attractive drug targets, however, there is only scant molecular knowledge on LTA and its precursor molecules in membranes. Here, we report the isolation and molecular characterization of diglucosyldiacylglycerol (Glc2-DAG), the glycolipid precursor molecule that anchors LTA in the bacterial plasma-membrane. Using a tailored growth medium and purification protocols, we isolated 13C-isotope labelled Glc2-DAG from bacteria, which can then be used for high-resolution NMR studies. Using solution-state and solid-state NMR, we show an in-depth molecular characterization of Glc2-DAG, including in native-like membranes. Our approach may help to identify antibiotics that directly target LTA precursor molecules, and it offers a tool for future investigations into the role of Glc2-DAG in bacterial physiology.

Ectopic Expression of C-Type Lectin Mincle Renders Mice Susceptible to Staphylococcal Pneumonia.

Staphylococcus aureus is a prevalent pathogen in pneumonia and harbors glycolipids, which may serve as molecular patterns in Mincle (macrophage-inducible C-type lectin)-dependent pathogen recognition. We examined the role of Mincle in lung defense against S aureus in wild-type (WT), Mincle knockout (KO), and Mincle transgenic (tg) mice. Two glycolipids, glucosyl-diacylglycerol (Glc-DAG) and diglucosyl-diacylglycerol (Glc2-DAG), were purified, of which only Glc-DAG triggered Mincle reporter cell activation and professional phagocyte responses. Proteomic profiling revealed that Glc2-DAG blocked Glc-DAG-induced cytokine responses, thereby acting as inhibitor of Glc-DAG/Mincle signaling. WT mice responded to S aureus with a similar lung pathology as Mincle KO mice, most likely due to Glc2-DAG-dependent inhibition of Glc-DAG/Mincle signaling. In contrast, ectopic Mincle expression caused severe lung pathology in S aureus-infected mice, characterized by bacterial outgrowth and fatal pneumonia. Collectively, Glc2-DAG inhibits Glc-DAG/Mincle-dependent responses in WT mice, whereas sustained Mincle expression overrides Glc2-DAG-mediated inhibitory effects, conferring increased host susceptibility to S aureus.

Potential of ion mobility mass spectrometry in cellulose ether analysis: substitution pattern of hydroxyethyl celluloses.

Ion mobility mass spectrometry (ESI-tims-ToF-MS, syringe pump infusion) has been applied to glucose and oligosaccharide ethers derived from hydroxyethyl-methyl celluloses (HEMC) and hydroxyethyl celluloses (HEC) after permethylation and partial depolymerization: by hydrolysis without or with subsequent reductive amination with m-amino benzoic acid (mABA) or by reductive cleavage. As model compounds without tandem substitution methoxyethylated methylcellulose was used. Regioisomeric glucose ethers were separated according to their ion mobility, and positions of substitution could be assigned. Glucose ethers including isomers with tandem substitution showed additional signals with a smaller collision cross-section (CCS) than core-substituted isomers. Positional isomers of cellobiose ethers were only partly resolved due to too high complexity but showed a characteristic fingerprint that might allow classifying samples. Relative intensities of signals of glucose ether isomers could only be quantified in case of ABA derivatives with its fixed charge, while sodium adducts of methoxyethyl ethers showed an influence of the MeOEt position on ion yield. Results were in very good agreement with reference analysis. [M + Na]+ adducts of α- and ß-anomers of glucose derivatives were separated in IM, complicating position assignment. This could be overcome by reductive cleavage of the permethylated HE(M)C yielding 1,5-anhydroglucitol-terminated oligosaccharides, showing the best resolved fingerprints of the cellobiose ethers of a particular cellulose ether. With this first application of ion mobility MS to the analysis of complex cellulose ethers, the promising potential of this additional separation dimension in mass spectrometry is demonstrated and discussed.

Xiaotan Sanjie recipe, a compound Chinese herbal medicine, inhibits gastric cancer metastasis by regulating GnT-V-mediated E-cadherin glycosylation.

OBJECTIVE: Xiaotan Sanjie recipe (XTSJ), a Chinese herbal compound medicine, exerts a significant inhibitory effect on gastric cancer (GC) metastasis. This work investigated the mechanism underlying the XTSJ-mediated inhibition of GC metastasis. METHODS: The effect of XTSJ on GC metastasis and the associated mechanism were investigated in vitro, using GC cell lines, and in vivo, using a GC mouse model, by focusing on the expression of Glc-N-Ac-transferase V (GnT-V; encoded by MGAT5). RESULTS: The migration and invasion ability of GC cells decreased significantly after XTSJ administration, which confirmed the efficacy of XTSJ in treating GC in vitro. XTSJ increased the accumulation of E-cadherin at junctions between GC cells, which was reversed by MGAT5 overexpression. XTSJ administration and MGAT5 knockdown alleviated the structural abnormality of the cell-cell junctions, while MGAT5 overexpression had the opposite effect. MGAT5 knockdown and XTSJ treatment also significantly increased the accumulation of proteins associated with the E-cadherin-mediated adherens junction complex. Furthermore, the expression of MGAT5 was significantly lower in the lungs of BGC-823-MGAT5 + XTSJ mice than in those of BGC-823-MGAT5 + solvent mice, indicating that the ability of gastric tumors to metastasize to the lung was decreased in vivo following XTSJ treatment. CONCLUSION: XTSJ prevented GC metastasis by inhibiting the GnT-V-mediated E-cadherin glycosylation and promoting the E-cadherin accumulation at cell-cell junctions. Please cite this article as: Huang N, He HW, He YY, Gu W, Xu MJ, Liu L. Xiaotan Sanjie recipe, a compound Chinese herbal medicine, inhibits gastric cancer metastasis by regulating GnT-V-mediated E-cadherin glycosylation. J Integr Med. 2023; 21(6): 561-574.

Screening, identification and control of unknown estrogen-like compounds in Maillard reaction products of glucose-arginine/lysine model systems.

It was speculated that estrogen-like compounds may be produced by chemical reactions during food processing, such as Maillard reaction, which would disrupt the endocrine system of organisms. Herein, the Maillard reaction in the process of high temperature for long time was simulated by using model system, and unknown estrogen-like compounds produced during Maillard reaction were screened by colorimetric assay based on dual estrogen receptor (ER)-gold nanoparticles (AuNPs) and enzyme-linked immunosorbent assay (ELISA). Possible structures of estrogen-like compounds were inferred by ultra-performance liquid chromatography-quadrupole time of flight tandem mass spectrometry (UPLC-QTOF/MS) in combination with a mass database, and finally the structure of estrogen-like compound, 2, 4-dihydroxy-1, 4-benzoxazin-3-one-2-o-ß-D-glucopyranoside (DIBOA-glc), was identified by high resolution orbitrap mass spectrometry (Orbitrap HRMS). This is the first study of the screening and identification of unknown estrogen-like compounds produced in Maillard reaction. Additionally, strategy of controlling the formation of DIBOA-glc by adding vitamin B6 in Maillard reaction was proposed, providing effective proposals for the safety control in actual food processing.

Potential Therapeutic Functions of PU-91 and Quercetin in Personalized Cybrids Derived from Patients with Age-Related Macular Degeneration, Keratoconus, and Glaucoma.

The aim of this study is to investigate the therapeutic potential of higher doses of PU-91, quercetin, or in combination on transmitochondrial cybrid cell lines with various mtDNA haplogroups derived from patients with age-related macular degeneration (AMD), glaucoma (Glc), keratoconus (KC), and normal (NL) individuals. Cybrids were treated with PU-91 (P) (200 µM) alone, quercetin (Q) (20 µM) alone, or a combination of PU-91 and quercetin (P+Q) for 48 h. Cellular metabolism and the intracellular levels of reactive oxygen species (ROS) were measured by MTT and H2DCFDA assays, respectively. Quantitative real-time PCR was performed to measure the expression levels of genes associated with mitochondrial biogenesis, antioxidant enzymes, inflammation, apoptosis, and senescence pathways. PU-91(P) (i) improves cellular metabolism in AMD cybrids, (ii) decreases ROS production in AMD cybrids, and (iii) downregulates the expression of LMNB1 in AMD cybrids. Combination treatment of PU-91 plus quercetin (P+Q) (i) improves cellular metabolism in AMD, (ii) induces higher expression levels of TFAM, SOD2, IL6, and BAX in AMD cybrids, and (iii) upregulates CDKN1A genes expression in all disease cybrids. Our study demonstrated that the P+Q combination improves cellular metabolism and mitochondrial biogenesis in AMD cybrids, but senescence is greatly exacerbated in all cybrids regardless of disease type by the P+Q combined treatment.

Development of high sustained anti-drug antibody titers and corresponding clinical decline in a late-onset Pompe disease patient after 11+ years on enzyme replacement therapy.

A late-onset Pompe disease patient developed high sustained antibody titers (HSAT) of ≥51,200 after 11+ years on alglucosidase alfa and previous tolerance. There was a corresponding worsening of motor function and rise in urinary glucose tetrasaccharide (Glc4). Following immunomodulation therapy, HSAT were eliminated with improved clinical outcomes and biomarker trends. This report highlights the importance of continued surveillance of antibody titers and biomarkers, the negative impact of HSAT, and improved outcomes with immunomodulation therapy.

Structural insights into a bacterial ß-glucosidase capable of degrading sesaminol triglucoside to produce sesaminol: toward the understanding of the aglycone recognition mechanism by the C-terminal lid domain.

The sesaminol triglucoside (STG)-hydrolyzing ß-glucosidase from Paenibacillus sp. (PSTG1), which belongs to glycoside hydrolase family 3 (GH3), is a promising catalyst for the industrial production of sesaminol. We determined the X-ray crystal structure of PSTG1 with bound glycerol molecule in the putative active site. PSTG1 monomer contained typical three domains of GH3 with the active site in domain 1 (TIM barrel). In addition, PSTG1 contained an additional domain (domain 4) at the C-terminus that interacts with the active site of the other protomer as a lid in the dimer unit. Interestingly, the interface of domain 4 and the active site forms a hydrophobic cavity probably for recognizing the hydrophobic aglycone moiety of substrate. The short flexible loop region of TIM barrel was found to be approaching the interface of domain 4 and the active site. We found that n-heptyl-ß-D-thioglucopyranoside detergent acts as an inhibitor for PSTG1. Thus, we propose that the recognition of hydrophobic aglycone moiety is important for PSTG1-catalyzed reactions. Domain 4 might be a potential target for elucidating the aglycone recognition mechanism of PSTG1 as well as for engineering PSTG1 to create a further excellent enzyme to degrade STG more efficiently to produce sesaminol.

In vivo and in vitro characterizations of melibiose permease (MelB) conformation-dependent nanobodies reveal sugar-binding mechanisms.

Salmonella enterica serovar Typhimurium melibiose permease (MelBSt) is a prototype of the Na+-coupled major facilitator superfamily transporters, which are important for the cellular uptake of molecules including sugars and small drugs. Although the symport mechanisms have been well-studied, mechanisms of substrate binding and translocation remain enigmatic. We have previously determined the sugar-binding site of outward-facing MelBSt by crystallography. To obtain other key kinetic states, here we raised camelid single-domain nanobodies (Nbs) and carried out a screening against the WT MelBSt under 4 ligand conditions. We applied an in vivo cAMP-dependent two-hybrid assay to detect interactions of Nbs with MelBSt and melibiose transport assays to determine the effects on MelBSt functions. We found that all selected Nbs showed partial to complete inhibitions of MelBSt transport activities, confirming their intracellular interactions. A group of Nbs (714, 725, and 733) was purified, and isothermal titration calorimetry measurements showed that their binding affinities were significantly inhibited by the substrate melibiose. When titrating melibiose to the MelBSt/Nb complexes, Nb also inhibited the sugar-binding. However, the Nb733/MelBSt complex retained binding to the coupling cation Na+ and also to the regulatory enzyme EIIAGlc of the glucose-specific phosphoenolpyruvate/sugar phosphotransferase system. Further, EIIAGlc/MelBSt complex also retained binding to Nb733 and formed a stable supercomplex. All data indicated that MelBSt trapped by Nbs retained its physiological functions and the trapped conformation is similar to that bound by the physiological regulator EIIAGlc. Therefore, these conformational Nbs can be useful tools for further structural, functional, and conformational analyses.

Short-chain fatty acids reprogram metabolic profiles with the induction of reactive oxygen species production in human colorectal adenocarcinoma cells.

Short-chain fatty acids (SCFAs) exhibit anticancer activity in cellular and animal models of colon cancer. Acetate, propionate, and butyrate are the three major SCFAs produced from dietary fiber by gut microbiota fermentation and have beneficial effects on human health. Most previous studies on the antitumor mechanisms of SCFAs have focused on specific metabolites or genes involved in antitumor pathways, such as reactive oxygen species (ROS) biosynthesis. In this study, we performed a systematic and unbiased analysis of the effects of acetate, propionate, and butyrate on ROS levels and metabolic and transcriptomic signatures at physiological concentrations in human colorectal adenocarcinoma cells. We observed significantly elevated levels of ROS in the treated cells. Furthermore, significantly regulated signatures were involved in overlapping pathways at metabolic and transcriptomic levels, including ROS response and metabolism, fatty acid transport and metabolism, glucose response and metabolism, mitochondrial transport and respiratory chain complex, one-carbon metabolism, amino acid transport and metabolism, and glutaminolysis, which are directly or indirectly linked to ROS production. Additionally, metabolic and transcriptomic regulation occurred in a SCFAs types-dependent manner, with an increasing degree from acetate to propionate and then to butyrate. This study provides a comprehensive analysis of how SCFAs induce ROS production and modulate metabolic and transcriptomic levels in colon cancer cells, which is vital for understanding the mechanisms of the effects of SCFAs on antitumor activity in colon cancer.

Mapping of glutamate metabolism using 1H FID-MRSI after oral administration of [1-13C]Glc at 9.4 T.

Glutamate is the major excitatory transmitter in the brain and malfunction of the related metabolism is associated with various neurological diseases and disorders. The observation of labeling changes in the spectra after the administration of a 13C labelled tracer is a common tool to gain better insights into the function of the metabolic system. But so far, only a very few studies presenting the labeling effects in more than two voxels to show the spatial dependence of metabolism. In the present work, the labeling effects were measured in a transversal plane in the human brain using ultra-short TE and TR 1H FID-MRSI. The measurement set-up was most simple: The [1-13C]Glc was administered orally instead of intravenous and the spectra were measured with a pure 1H technique without the need of a 13C channel (as Boumezbeur et al. demonstrated in 2004). Thus, metabolic maps and enrichment curves could be obtained for more metabolites and in more voxels than ever before in human brain. Labeling changes could be observed in [4-13C]glutamate, [3-13C]glutamate+glutamine, [2-13C]glutamate+glutamine, [4-13C]glutamine, and [3-13C]aspartate with a high temporal (3.6 min) and spatial resolution (32 × 32 grid with nominal voxel size of 0.33 µL) in five volunteers.

Ginsenoside 3ß-O-Glc-DM (C3DM) suppressed glioma tumor growth by downregulating the EGFR/PI3K/AKT/mTOR signaling pathway and modulating the tumor microenvironment.

Ginsenosides are the main bioactive constituents of Panax ginseng, which have been broadly studied in cancer treatment. Our previous studies have demonstrated that 3ß-O-Glc-DM (C3DM), a biosynthetic ginsenoside, exhibited antitumor effects in several cancer cell lines with anti-colon cancer activity superior to ginsenoside 20(R)-Rg3 in vivo. However, the efficacy of C3DM on glioma has not been proved yet. In this study, the antitumor activities and underlying mechanisms of C3DM on glioma were investigated in vitro and in vivo. Cell viability, apoptosis, migration, FCM, IHC, RT-qPCR, quantitative proteomics, and western blotting were conducted to evaluate the effect of C3DM on glioma cells. ADP-Glo™ kinase assay was used to validate the interaction between C3DM and EGFR. Co-cultured assays, lactic acid kit, and spatially resolved metabolomics were performed to study the function of C3DM in regulating glioma microenvironment. Both subcutaneously transplanted syngeneic models and orthotopic models of glioma were used to determine the effect of C3DM on tumor growth in vivo. We found that C3DM dose-dependently induced apoptosis, and inhibited the proliferation, migration and angiogenesis of glioma cells. C3DM significantly inhibited tumor growth in both subcutaneous and orthotopic mouse glioma models. Moreover, C3DM attenuated the acidified glioma microenvironment and enhanced T-cell function. Additionally, C3DM inhibited the kinase activity of EGFR and influenced the EGFR/PI3K/AKT/mTOR signaling pathway in glioma. Overall, C3DM might be a promising candidate for glioma prevention and treatment.

Characterization and engineering of the xylose-inducible xylP promoter for use in mold fungal species.

Conditional promoters allowing both induction and silencing of gene expression are indispensable for basic and applied research. The xylP promoter (pxylP) from Penicillium chrysogenum was demonstrated to function in various mold species including Aspergillus fumigatus. pxylP allows high induction by xylan or its degradation product xylose with low basal activity in the absence of an inducer. Here we structurally characterized and engineered pxylP in A. fumigatus to optimize its application. Mutational analysis demonstrated the importance of the putative TATA-box and a pyrimidine-rich region in the core promoter, both copies of a largely duplicated 91-bp sequence (91bpDS), as well as putative binding sites for the transcription factor XlnR and a GATA motif within the 91bpDS. In agreement, pxylP activity was found to depend on XlnR, while glucose repression appeared to be indirect. Truncation of the originally used 1643-bp promoter fragment to 725 bp largely preserved the promoter activity and the regulatory pattern. Integration of a third 91bpDS significantly increased promoter activity particularly under low inducer concentrations. Truncation of pxylP to 199 bp demonstrated that the upstream region including the 91bpDSs mediates not only inducer-dependent activation but also repression in the absence of inducer. Remarkably, the 1579-bp pxylP was found to act bi-bidirectionally with a similar regulatory pattern by driving expression of the upstream-located arabinofuranosidase gene. The latter opens the possibility of dual bidirectional use of pxylP. Comparison with a doxycycline-inducible TetOn system revealed a significantly higher dynamic range of pxylP. Taken together, this study identified functional elements of pxylP and opened new methodological opportunities for its application.

Prospective bacterial and fungal sources of hyaluronic acid: A review.

The unique biological and rheological properties make hyaluronic acid a sought-after material for medicine and cosmetology. Due to very high purity requirements for hyaluronic acid in medical applications, the profitability of streptococcal fermentation is reduced. Production of hyaluronic acid by recombinant systems is considered a promising alternative. Variations in combinations of expressed genes and fermentation conditions alter the yield and molecular weight of produced hyaluronic acid. This review is devoted to the current state of hyaluronic acid production by recombinant bacterial and fungal organisms.

From structure to function - Ligand recognition by myeloid C-type lectin receptors.

The relevance of protein-glycan interactions in immunity has long been underestimated. Yet, the immune system possesses numerous classes of glycan-binding proteins, so-called lectins. Of specific interest is the group of myeloid C-type lectin receptors (CLRs) as they are mainly expressed by myeloid cells and play an important role in the initiation of an immune response. Myeloid CLRs represent a major group amongst pattern recognition receptors (PRRs), placing them at the center of the rapidly growing field of glycoimmunology. CLRs have evolved to encompass a wide range of structures and functions and to recognize a large number of glycans and many other ligands from different classes of biopolymers. This review aims at providing the reader with an overview of myeloid CLRs and selected ligands, while highlighting recent insights into CLR-ligand interactions. Subsequently, methodological approaches in CLR-ligand research will be presented. Finally, this review will discuss how CLR-ligand interactions culminate in immunological functions, how glycan mimicry favors immune escape by pathogens, and in which way immune responses can be affected by CLR-ligand interactions in the long term.

Newborn screening for Pompe disease in Italy: Long-term results and future challenges.

Pompe disease (PD) is a progressive neuromuscular disorder caused by a lysosomal acid α-glucosidase (GAA) deficiency. Enzymatic replacement therapy is available, but early diagnosis by newborn screening (NBS) is essential for early treatment and better outcomes, especially with more severe forms. We present results from 7 years of NBS for PD and the management of infantile-onset (IOPD) and late-onset (LOPD) patients, during which we sought candidate predictive parameters of phenotype severity at baseline and during follow-up. We used a tandem mass spectrometry assay for α-glucosidase activity to screen 206,741 newborns and identified 39 positive neonates (0.019%). Eleven had two pathogenic variants of the GAA gene (3 IOPD, 8 LOPD); six carried variants of uncertain significance (VUS). IOPD patients were treated promptly and had good outcomes. LOPD and infants with VUS were followed; all were asymptomatic at the last visit (mean age 3.4 years, range 0.5-5.5). Urinary glucose tetrasaccharide was a useful and biomarker for rapidly differentiating IOPD from LOPD and monitoring response to therapy during follow-up. Our study, the largest reported to date in Europe, presents data from longstanding NBS for PD, revealing an incidence in North East Italy of 1/18,795 (IOPD 1/68,914; LOPD 1/25,843), and the absence of mortality in IOPD treated from birth. In LOPD, rigorous long-term follow-up is needed to evaluate the best time to start therapy. The high pseudodeficiency frequency, ethical issues with early LOPD diagnosis, and difficulty predicting phenotypes based on biochemical parameters and genotypes, especially in LOPD, need further study.

Abnormal brain oxygen homeostasis in an animal model of liver disease.

Background & Aims: Increased plasma ammonia concentration and consequent disruption of brain energy metabolism could underpin the pathogenesis of hepatic encephalopathy (HE). Brain energy homeostasis relies on effective maintenance of brain oxygenation, and dysregulation impairs neuronal function leading to cognitive impairment. We hypothesised that HE is associated with reduced brain oxygenation and we explored the potential role of ammonia as an underlying pathophysiological factor. Methods: In a rat model of chronic liver disease with minimal HE (mHE; bile duct ligation [BDL]), brain tissue oxygen measurement, and proton magnetic resonance spectroscopy were used to investigate how hyperammonaemia impacts oxygenation and metabolic substrate availability in the central nervous system. Ornithine phenylacetate (OP, OCR-002; Ocera Therapeutics, CA, USA) was used as an experimental treatment to reduce plasma ammonia concentration. Results: In BDL animals, glucose, lactate, and tissue oxygen concentration in the cerebral cortex were significantly lower than those in sham-operated controls. OP treatment corrected the hyperammonaemia and restored brain tissue oxygen. Although BDL animals were hypotensive, cortical tissue oxygen concentration was significantly improved by treatments that increased arterial blood pressure. Cerebrovascular reactivity to exogenously applied CO2 was found to be normal in BDL animals. Conclusions: These data suggest that hyperammonaemia significantly decreases cortical oxygenation, potentially compromising brain energy metabolism. These findings have potential clinical implications for the treatment of patients with mHE. Lay summary: Brain dysfunction is a serious complication of cirrhosis and affects approximately 30% of these patients; however, its treatment continues to be an unmet clinical need. This study shows that oxygen concentration in the brain of an animal model of cirrhosis is markedly reduced. Low arterial blood pressure and increased ammonia (a neurotoxin that accumulates in patients with liver failure) are shown to be the main underlying causes. Experimental correction of these abnormalities restored oxygen concentration in the brain, suggesting potential therapeutic avenues to explore.

Recent developments in Moringa oleifera Lam. polysaccharides: A review of the relationship between extraction methods, structural characteristics and functional activities.

Moringa oleifera Lam. (M. oleifera Lam) is a perennial tropical deciduous tree that belongs to the Moringaceae family. Polysaccharides are one of the major bioactive compounds in M. oleifera Lam and show immunomodulatory, anticancer, antioxidant, intestinal health protection and antidiabetic activities. At present, the structure and functional activities of M. oleifera Lam polysaccharides (MOPs) have been widespread, but the research data are relatively scattered. Moreover, the relationship between the structure and biological activities of MOPs has not been summarized. In this review, the current research on the extraction, purification, structural characteristics and biological activities of polysaccharides from different sources of M. oleifera Lam were summarized, and the structural characteristics of purified polysaccharides were focused on this review. Meanwhile, the biological activities of MOPs were introduced, and some molecular mechanisms were listed. In addition, the relationship between the structure and biological activities of MOPs was discussed. Furthermore, new perspectives and some future research of M. oleifera Lam polysaccharides were proposed in this review.