Sex Differences in Intestinal Carbohydrate Metabolism Promote Food Intake and Sperm Maturation.

Physiology and metabolism are often sexually dimorphic, but the underlying mechanisms remain incompletely understood. Here, we use the intestine of Drosophila melanogaster to investigate how gut-derived signals contribute to sex differences in whole-body physiology. We find that carbohydrate handling is male-biased in a specific portion of the intestine. In contrast to known sexual dimorphisms in invertebrates, the sex differences in intestinal carbohydrate metabolism are extrinsically controlled by the adjacent male gonad, which activates JAK-STAT signaling in enterocytes within this intestinal portion. Sex reversal experiments establish roles for this male-biased intestinal metabolic state in controlling food intake and sperm production through gut-derived citrate. Our work uncovers a male gonad-gut axis coupling diet and sperm production, revealing that metabolic communication across organs is physiologically important. The instructive role of citrate in inter-organ communication might be significant in more biological contexts than previously recognized.

A non-canonical tricarboxylic acid cycle underlies cellular identity.

The tricarboxylic acid (TCA) cycle is a central hub of cellular metabolism, oxidizing nutrients to generate reducing equivalents for energy production and critical metabolites for biosynthetic reactions. Despite the importance of the products of the TCA cycle for cell viability and proliferation, mammalian cells display diversity in TCA-cycle activity1,2. How this diversity is achieved, and whether it is critical for establishing cell fate, remains poorly understood. Here we identify a non-canonical TCA cycle that is required for changes in cell state. Genetic co-essentiality mapping revealed a cluster of genes that is sufficient to compose a biochemical alternative to the canonical TCA cycle, wherein mitochondrially derived citrate exported to the cytoplasm is metabolized by ATP citrate lyase, ultimately regenerating mitochondrial oxaloacetate to complete this non-canonical TCA cycle. Manipulating the expression of ATP citrate lyase or the canonical TCA-cycle enzyme aconitase 2 in mouse myoblasts and embryonic stem cells revealed that changes in the configuration of the TCA cycle accompany cell fate transitions. During exit from pluripotency, embryonic stem cells switch from canonical to non-canonical TCA-cycle metabolism. Accordingly, blocking the non-canonical TCA cycle prevents cells from exiting pluripotency. These results establish a context-dependent alternative to the traditional TCA cycle and reveal that appropriate TCA-cycle engagement is required for changes in cell state.

Structure and inhibition mechanism of the human citrate transporter NaCT.

Citrate is best known as an intermediate in the tricarboxylic acid cycle of the cell. In addition to this essential role in energy metabolism, the tricarboxylate anion also acts as both a precursor and a regulator of fatty acid synthesis1-3. Thus, the rate of fatty acid synthesis correlates directly with the cytosolic concentration of citrate4,5. Liver cells import citrate through the sodium-dependent citrate transporter NaCT (encoded by SLC13A5) and, as a consequence, this protein is a potential target for anti-obesity drugs. Here, to understand the structural basis of its inhibition mechanism, we determined cryo-electron microscopy structures of human NaCT in complexes with citrate or a small-molecule inhibitor. These structures reveal how the inhibitor-which binds to the same site as citrate-arrests the transport cycle of NaCT. The NaCT-inhibitor structure also explains why the compound selectively inhibits NaCT over two homologous human dicarboxylate transporters, and suggests ways to further improve the affinity and selectivity. Finally, the NaCT structures provide a framework for understanding how various mutations abolish the transport activity of NaCT in the brain and thereby cause epilepsy associated with mutations in SLC13A5 in newborns (which is known as SLC13A5-epilepsy)6-8.

Citrate serves as a signal molecule to modulate carbon metabolism and iron homeostasis in Staphylococcus aureus.

Pathogenic bacteria's metabolic adaptation for survival and proliferation within hosts is a crucial aspect of bacterial pathogenesis. Here, we demonstrate that citrate, the first intermediate of the tricarboxylic acid (TCA) cycle, plays a key role as a regulator of gene expression in Staphylococcus aureus. We show that citrate activates the transcriptional regulator CcpE and thus modulates the expression of numerous genes involved in key cellular pathways such as central carbon metabolism, iron uptake and the synthesis and export of virulence factors. Citrate can also suppress the transcriptional regulatory activity of ferric uptake regulator. Moreover, we determined that accumulated intracellular citrate, partly through the activation of CcpE, decreases the pathogenic potential of S. aureus in animal infection models. Therefore, citrate plays a pivotal role in coordinating carbon metabolism, iron homeostasis, and bacterial pathogenicity at the transcriptional level in S. aureus, going beyond its established role as a TCA cycle intermediate.

Dietary fructose feeds hepatic lipogenesis via microbiota-derived acetate.

Consumption of fructose has risen markedly in recent decades owing to the use of sucrose and high-fructose corn syrup in beverages and processed foods1, and this has contributed to increasing rates of obesity and non-alcoholic fatty liver disease2-4. Fructose intake triggers de novo lipogenesis in the liver4-6, in which carbon precursors of acetyl-CoA are converted into fatty acids. The ATP citrate lyase (ACLY) enzyme cleaves cytosolic citrate to generate acetyl-CoA, and is upregulated after consumption of carbohydrates7. Clinical trials are currently pursuing the inhibition of ACLY as a treatment for metabolic diseases8. However, the route from dietary fructose to hepatic acetyl-CoA and lipids remains unknown. Here, using in vivo isotope tracing, we show that liver-specific deletion of Acly in mice is unable to suppress fructose-induced lipogenesis. Dietary fructose is converted to acetate by the gut microbiota9, and this supplies lipogenic acetyl-CoA independently of ACLY10. Depletion of the microbiota or silencing of hepatic ACSS2, which generates acetyl-CoA from acetate, potently suppresses the conversion of bolus fructose into hepatic acetyl-CoA and fatty acids. When fructose is consumed more gradually to facilitate its absorption in the small intestine, both citrate cleavage in hepatocytes and microorganism-derived acetate contribute to lipogenesis. By contrast, the lipogenic transcriptional program is activated in response to fructose in a manner that is independent of acetyl-CoA metabolism. These data reveal a two-pronged mechanism that regulates hepatic lipogenesis, in which fructolysis within hepatocytes provides a signal to promote the expression of lipogenic genes, and the generation of microbial acetate feeds lipogenic pools of acetyl-CoA.

Sfxn5 Regulation of Actin Polymerization for Neutrophil Spreading Depends on a Citrate-Cholesterol-PI(4,5)P2 Pathway.

Cell spreading is an initial and critical step in neutrophil adhesion and migration, leading to neutrophil recruitment to inflammatory tissues. Sideroflexin (Sfxn) family proteins are metabolite transporters located in the mitochondrial membrane. Recombinant SFXN5 protein is a citrate transporter in vitro; however, whether Sfxn5 regulates any cellular behavior or function remains unknown. In this study, we found that small interfering RNA transfection or morpholino injection achieving Sfxn5 deficiency in neutrophils significantly decreased neutrophil recruitment in mice and zebrafish, respectively. Sfxn5 deficiency impaired neutrophil spreading and spreading-associated cellular phenotypes, such as cell adhesion, chemotaxis, and ROS production. Actin polymerization is critical for neutrophil spreading, and we found that actin polymerization in spreading neutrophils was partially inhibited by Sfxn5 deficiency. Mechanistically, we observed that the levels of cytosolic citrate and its downstream metabolic products, acetyl-CoA and cholesterol, were decreased in Sfxn5-deficient neutrophils. The levels of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2), a mediator for the regulation of actin polymerization by cholesterol, were reduced in the plasma membrane of Sfxn5-deficient neutrophils. Exogenous supplementation with citrate or cholesterol partially reversed the reduction in PI(4,5)P2 levels, defective neutrophil actin polymerization, and cell spreading. Altogether, we demonstrated that Sfxn5 maintains cytosolic citrate levels and ensures the synthesis of sufficient cholesterol to promote actin polymerization in a PI(4,5)P2-dependent manner during neutrophil spreading, which is essential for the eventual inflammatory recruitment of neutrophils. Our study revealed the importance of Sfxn5 in neutrophil spreading and migration, thus identifying, to our knowledge, for the first time, the physiological cellular functions of the Sfxn5 gene.

Distinct modes of mitochondrial metabolism uncouple T cell differentiation and function.

Activated CD4 T cells proliferate rapidly and remodel epigenetically before exiting the cell cycle and engaging acquired effector functions. Metabolic reprogramming from the naive state is required throughout these phases of activation1. In CD4 T cells, T-cell-receptor ligation-along with co-stimulatory and cytokine signals-induces a glycolytic anabolic program that is required for biomass generation, rapid proliferation and effector function2. CD4 T cell differentiation (proliferation and epigenetic remodelling) and function are orchestrated coordinately by signal transduction and transcriptional remodelling. However, it remains unclear whether these processes are regulated independently of one another by cellular biochemical composition. Here we demonstrate that distinct modes of mitochondrial metabolism support differentiation and effector functions of mouse T helper 1 (TH1) cells by biochemically uncoupling these two processes. We find that the tricarboxylic acid cycle is required for the terminal effector function of TH1 cells through succinate dehydrogenase (complex II), but that the activity of succinate dehydrogenase suppresses TH1 cell proliferation and histone acetylation. By contrast, we show that complex I of the electron transport chain, the malate-aspartate shuttle and mitochondrial citrate export are required to maintain synthesis of aspartate, which is necessary for the proliferation of T helper cells. Furthermore, we find that mitochondrial citrate export and the malate-aspartate shuttle promote histone acetylation, and specifically regulate the expression of genes involved in T cell activation. Combining genetic, pharmacological and metabolomics approaches, we demonstrate that the differentiation and terminal effector functions of T helper cells are biochemically uncoupled. These findings support a model in which the malate-aspartate shuttle, mitochondrial citrate export and complex I supply the substrates needed for proliferation and epigenetic remodelling early during T cell activation, whereas complex II consumes the substrates of these pathways, which antagonizes differentiation and enforces terminal effector function. Our data suggest that transcriptional programming acts together with a parallel biochemical network to enforce cell state.

An allosteric mechanism for potent inhibition of human ATP-citrate lyase.

ATP-citrate lyase (ACLY) is a central metabolic enzyme and catalyses the ATP-dependent conversion of citrate and coenzyme A (CoA) to oxaloacetate and acetyl-CoA1-5. The acetyl-CoA product is crucial for the metabolism of fatty acids6,7, the biosynthesis of cholesterol8, and the acetylation and prenylation of proteins9,10. There has been considerable interest in ACLY as a target for anti-cancer drugs, because many cancer cells depend on its activity for proliferation2,5,11. ACLY is also a target against dyslipidaemia and hepatic steatosis, with a compound currently in phase 3 clinical trials4,5. Many inhibitors of ACLY have been reported, but most of them have weak activity5. Here we report the development of a series of low nanomolar, small-molecule inhibitors of human ACLY. We have also determined the structure of the full-length human ACLY homo-tetramer in complex with one of these inhibitors (NDI-091143) by cryo-electron microscopy, which reveals an unexpected mechanism of inhibition. The compound is located in an allosteric, mostly hydrophobic cavity next to the citrate-binding site, and requires extensive conformational changes in the enzyme that indirectly disrupt citrate binding. The observed binding mode is supported by and explains the structure-activity relationships of these compounds. This allosteric site greatly enhances the 'druggability' of ACLY and represents an attractive target for the development of new ACLY inhibitors.

A specialized metabolic pathway partitions citrate in hydroxyapatite to impact mineralization of bones and teeth.

Citrate is a critical metabolic substrate and key regulator of energy metabolism in mammalian cells. It has been known for decades that the skeleton contains most (>85%) of the body's citrate, but the question of why and how this metabolite should be partitioned in bone has received singularly little attention. Here, we show that osteoblasts use a specialized metabolic pathway to regulate uptake, endogenous production, and the deposition of citrate into bone. Osteoblasts express high levels of the membranous Na+-dependent citrate transporter solute carrier family 13 member 5 (Slc13a5) gene. Inhibition or genetic disruption of Slc13a5 reduced osteogenic citrate uptake and disrupted mineral nodule formation. Bones from mice lacking Slc13a5 globally, or selectively in osteoblasts, showed equivalent reductions in cortical thickness, with similarly compromised mechanical strength. Surprisingly, citrate content in mineral from Slc13a5-/- osteoblasts was increased fourfold relative to controls, suggesting the engagement of compensatory mechanisms to augment endogenous citrate production. Indeed, through the coordinated functioning of the apical membrane citrate transporter SLC13A5 and a mitochondrial zinc transporter protein (ZIP1; encoded by Slc39a1), a mediator of citrate efflux from the tricarboxylic acid cycle, SLC13A5 mediates citrate entry from blood and its activity exerts homeostatic control of cytoplasmic citrate. Intriguingly, Slc13a5-deficient mice also exhibited defective tooth enamel and dentin formation, a clinical feature, which we show is recapitulated in primary teeth from children with SLC13A5 mutations. Together, our results reveal the components of an osteoblast metabolic pathway, which affects bone strength by regulating citrate deposition into mineral hydroxyapatite.

Carbon sources and pathways for citrate secreted by human prostate cancer cells determined by NMR tracing and metabolic modeling.

Prostate epithelial cells have the unique capacity to secrete large amounts of citrate, but the carbon sources and metabolic pathways that maintain this production are not well known. We mapped potential pathways for citrate carbons in the human prostate cancer metastasis cell lines LNCaP and VCaP, for which we first established that they secrete citrate (For LNCaP 5.6 ± 0.9 nmol/h per 106 cells). Using 13C-labeled substrates, we traced the incorporation of 13C into citrate by NMR of extracellular fluid. Our results provide direct evidence that glucose is a main carbon source for secreted citrate. We also demonstrate that carbons from supplied glutamine flow via oxidative Krebs cycle and reductive carboxylation routes to positions in secreted citrate but likely do not contribute to its net synthesis. The potential anaplerotic carbon sources aspartate and asparagine did not contribute to citrate carbons. We developed a quantitative metabolic model employing the 13C distribution in extracellular citrate after 13C glucose and pyruvate application to assess intracellular pathways of carbons for secreted citrate. From this model, it was estimated that in LNCaP about 21% of pyruvate entering the Krebs cycle is converted via pyruvate carboxylase as an anaplerotic route at a rate more than sufficient to compensate carbon loss of this cycle by citrate secretion. This model provides an estimation of the fraction of molecules, including citrate, leaving the Krebs cycle at every turn. The measured ratios of 13C atoms at different positions in extracellular citrate may serve as biomarkers for (malignant) epithelial cell metabolism.

Alpha-Synuclein Aggregation Mechanism in the Presence of Nanomaterials.

Parkinson's disease (PD) is characterized by the toxic oligomeric and fibrillar phases formed by monomeric alpha-synuclein (α-syn). Certain nanoparticles have been demonstrated to promote protein aggregation, while other nanomaterials have been found to prevent the process. In the current work, we use nuclear magnetic resonance spectroscopy in conjunction with isothermal titration calorimetry to investigate the cause and mechanism of these opposing effects at the amino acid protein level. The interaction of α-syn with two types of nanomaterials was considered: citrate-capped gold nanoparticles (AuNPs) and graphene oxide (GO). In the presence of AuNPs, α-syn aggregation is accelerated, whereas in the presence of GO, aggregation is prevented. The study indicates that GO sequesters the NAC region of α-syn monomers through electrostatic and hydrophobic interactions, leading to a reduced elongation rate, and AuNPs leave the NAC region exposed while binding the N-terminus, leading to higher aggregation. The protein's inclination toward quicker aggregation is explained by the binding of the N-terminus of α-syn with the gold nanoparticles. Conversely, a comparatively stronger interaction with GO causes the nucleation and growth phases to be postponed and inhibits intermolecular interactions. Our finding offers novel experimental insights at the residue level regarding the aggregation of α-syn in the presence of various nanomaterials and creates new opportunities for the development of suitably functionalized nanomaterial-based therapeutic reagents against Parkinson's and other neurodegenerative diseases.

Differential modulation of Target of Rapamycin activity under single and combined iron and sulfur deficiency in tomato plants.

Over the past few decades, a close relationship between sulfur (S) and iron (Fe) in terms of functionality and nutrition was demonstrated in the tomato. However, very little is known about the regulatory mechanisms underlying S/Fe interactions. Recently, the potential role of citrate in plant adaptation to Fe deficiency and combined S and Fe deficiency has been described. It is known that an impaired organic acid metabolism may stimulate a retrograde signal, which has been proven to be linked to the Target of Rapamycin (TOR) signaling in yeast and animal cells. Recent reports provided evidence of TOR involvement in S nutrient sensing in plants. This suggestion prompted us to investigate whether TOR may play a role in the cross-talk of signaling pathway occurring during plant adaptation to combined nutrient deficiency of Fe and S. Our results revealed that Fe deficiency elicited an increase of TOR activity associated with enhanced accumulation of citrate. In contrast, S deficiency resulted in decreased TOR activity and citrate accumulation. Interestingly, citrate accumulated in shoots of plants exposed to combined S/Fe deficiency to values between those found in Fe- and S-deficient plants, again correlated with TOR activity level. Our results suggest that citrate might be involved in establishing a link between plant response to combined S/Fe deficiency and the TOR network.

A citrate efflux transporter important for manganese distribution and phosphorus uptake in rice.

The plant citrate transporters, functional in mineral nutrient uptake and homeostasis, usually belong to the multidrug and toxic compound extrusion transporter family. We identified and functionally characterized a rice (Oryza sativa) citrate transporter, OsCT1, which differs from known plant citrate transporters and is structurally close to rice silicon transporters. Domain analysis depicted that OsCT1 carries a bacterial citrate-metal transporter domain, CitMHS. OsCT1 showed citrate efflux activity when expressed in Xenopus laevis oocytes and is localized to the cell plasma membrane. It is highly expressed in the shoot and reproductive tissues of rice, and its promoter activity was visible in cells surrounding the vasculature. The OsCT1 knockout (KO) lines showed a reduced citrate content in the shoots and the root exudates, whereas overexpression (OE) line showed higher citrate exudation from their roots. Further, the KO and OE lines showed variations in the manganese (Mn) distribution leading to changes in their agronomical traits. Under deficient conditions (Mn-sufficient conditions followed by 8 days of 0 µm MnCl2 · 4H2 O treatment), the supply of manganese towards the newer leaf was found to be obstructed in the KO line. There were no significant differences in phosphorus (P) distribution; however, P uptake was reduced in the KO and increased in OE lines at the vegetative stage. Further, experiments in Xenopus oocytes revealed that OsCT1 could efflux citrate with Mn. In this way, we provide insights into a mechanism of citrate-metal transport in plants and its role in mineral homeostasis, which remains conserved with their bacterial counterparts.

Biocompatible Tough Ionogels with Reversible Supramolecular Adhesion.

Cohesive and interfacial adhesion energies are difficult to balance to obtain reversible adhesives with both high mechanical strength and high adhesion strength, although various methods have been extensively investigated. Here, a biocompatible citric acid/L-(-)-carnitine (CAC)-based ionic liquid was developed as a solvent to prepare tough and high adhesion strength ionogels for reversible engineered and biological adhesives. The prepared ionogels exhibited good mechanical properties, including tensile strength (14.4 MPa), Young's modulus (48.1 MPa), toughness (115.2 MJ m-3), and high adhesion strength on the glass substrate (24.4 MPa). Furthermore, the ionogels can form mechanically matched tough adhesion at the interface of wet biological tissues (interfacial toughness about 191 J m-2) and can be detached by saline solution on demand, thus extending potential applications in various clinical scenarios such as wound adhesion and nondestructive transfer of organs.

A combination of ß-hydroxybutyrate and citrate ameliorates disease progression in a rat model of polycystic kidney disease.

Our research has shown that interventions producing a state of ketosis are highly effective in rat, mouse, and cat models of polycystic kidney disease (PKD), preventing and partially reversing cyst growth and disease progression. The ketone ß-hydroxybutyrate (BHB) appears to underlie this effect. In addition, we have demonstrated that naturally formed microcrystals within kidney tubules trigger a renoprotective response that facilitates tubular obstruction clearance in healthy animals but, alternatively, leads to cyst formation in PKD. The administration of citrate prevents microcrystal formation and slows PKD progression. Juvenile Cy/+ rats, a nonorthologous PKD model, were supplemented from 3 to 8 wk of age with water containing titrated BHB, citrate, or in combination to find minimal effective and optimal dosages, respectively. Adult rats were given a reduced BHB/citrate combination or equimolar control K/NaCl salts from 8 to 12 wk of age. In addition, adult rats were placed in metabolic cages following BHB, citrate, and BHB/citrate administration to determine the impact on mineral, creatinine, and citrate excretion. BHB or citrate alone effectively ameliorates disease progression in juvenile rats, decreasing markers of cystic disease and, in combination, producing a synergistic effect. BHB/citrate leads to partial disease regression in adult rats with established cystic disease, inhibiting cyst formation and kidney injury. BHB/citrate confers benefits via multiple mechanisms, increases creatinine and citrate excretion, and normalizes mineral excretion. BHB and citrate are widely available and generally recognized as safe compounds and, in combination, exhibit high promise for supporting kidney health in polycystic kidney disease.NEW & NOTEWORTHY Combining ß-hydroxybutyrate (BHB) and citrate effectively slows and prevents cyst formation and expansion in young Cy/+ rats using less BHB and citrate than when used alone, demonstrating synergy. In adult rats, the combination causes a partial reversal of existing disease, reducing cyst number and cystic area, preserving glomerular health, and decreasing markers of kidney injury. Our results suggest a safe and feasible strategy for supporting kidney health in polycystic kidney disease (PKD) using a combination of BHB and citrate.

OsCIPK2 mediated rice root microorganisms and metabolites to improve plant nitrogen uptake.

Crop roots are colonized by large numbers of microorganisms, collectively known as the root-microbiome, which modulate plant growth, development and contribute to elemental nutrient uptake. In conditions of nitrogen limitation, the over-expressed Calcineurin B-like interacting protein kinase 2 (OsCIPK2) gene with root-specific promoter (RC) has been shown to enhance growth and nitrogen uptake in rice. Analysis of root-associated bacteria through high-throughput sequencing revealed that OsCIPK2 has a significant impact on the diversity of the root microbial community under low nitrogen stress. The quantification of nifH gene expression demonstrated a significant enhancement in nitrogen-fixing capabilities in the roots of RC transgenetic rice. Synthetic microbial communities (SynCom) consisting of six nitrogen-fixing bacterial strains were observed to be enriched in the roots of RC, leading to a substantial improvement in rice growth and nitrogen uptake in nitrogen-deficient soils. Forty and twenty-three metabolites exhibiting differential abundance were identified in the roots and rhizosphere soils of RC transgenic rice compared to wild-type (WT) rice. These findings suggest that OSCIPK2 plays a role in restructuring the microbial community in the roots through the regulation of metabolite synthesis and secretion. Further experiments involving the exogenous addition of citric acid revealed that an optimal concentration of this compound facilitated the growth of nitrogen-fixing bacteria and substantially augmented their population in the soil, highlighting the importance of citric acid in promoting nitrogen fixation under conditions of low nitrogen availability. These findings suggest that OsCIPK2 plays a role in enhancing nitrogen uptake by rice plants from the soil by influencing the assembly of root microbial communities, thereby offering valuable insights for enhancing nitrogen utilization in rice cultivation.

OsAlR3 regulates aluminum tolerance through promoting the secretion of organic acids and the expression of antioxidant genes in rice.

In acidic soils, aluminum (Al) toxicity inhibits the growth and development of plant roots and affects nutrient and water absorption, leading to reduced yield and quality. Therefore, it is crucial to investigate and identify candidate genes for Al tolerance and elucidate their physiological and molecular mechanisms under Al stress. In this study, we identified a new gene OsAlR3 regulating Al tolerance, and analyzed its mechanism from physiological, transcriptional and metabolic levels. Compared with the WT, malondialdehyde (MDA) and hydrogen peroxide (H2O2) content were significantly increased, superoxide dismutase (SOD) activity and citric acid (CA) content were significantly decreased in the osalr3 mutant lines when exposed to Al stress. Under Al stress, the osalr3 exhibited decreased expression of antioxidant-related genes and lower organic acid content compared with WT. Integrated transcriptome and metabolome analysis showed the phenylpropanoid biosynthetic pathway plays an important role in OsAlR3-mediated Al tolerance. Exogenous CA and oxalic acid (OA) could increase total root length and enhance the antioxidant capacity in the mutant lines under Al stress. Conclusively, we found a new gene OsAlR3 that positively regulates Al tolerance by promoting the chelation of Al ions through the secretion of organic acids, and increasing the expression of antioxidant genes.

Simultaneous Detection of Citric Acid and Oxalic Acid Based on Dual Spectrum and Biomimetic Peroxidase for Urolithiasis Screening with a Fully Automatic Urine Analyzer.

Urolithiasis stands as a prevalent ailment within the urinary system, with hyperoxaluria and hypocitraturia being the most frequent manifestations characterized by excessive oxalic acid (OA) and deficient citric acid (CA) levels in urine. Detecting these compounds in urine quantitatively holds paramount importance for early urolithiasis screening. Existing methodologies fall short in achieving simultaneous and on-site identification of OA and CA, posing challenges for accurate urolithiasis screening. Addressing this concern, the study successfully accomplishes the concurrent identification of OA and CA in urine through a combination of dual-spectral analysis and biomimetic peroxidase utilization. Bovine serum albumin and dithiothreitol-modified copper nanoclusters (BSA-DTT-CuNCs) are employed as biomimetic peroxidases, effectively mitigating interference and enabling the simultaneous determination of OA and CA. The quantification range spans from 0 to 12 mm for OA and 0.5 to 2.5 mm for CA, with detection limits of 0.18 and 0.11 mm, respectively. To facilitate swift and on-location urine analysis, a fully automated urine analyzer (FAUA) is introduced that streamlines the process of biomarker pretreatment and identification within urine samples. Validation with real urine samples from urolithiasis patients demonstrates the method's diagnostic precision, highlighting the dual-spectral technique and analyzer's promising role in urolithiasis screening.

Dissimilar effects of the hydrophilic carbon dots on the amyloid aggregation of two model proteins and the mechanism discussion.

Many proteins could aggregate into amyloid fibrils under certain conditions. However, the aggregation process and morphology of the fibrils may be significantly different because of the distinct protein structure. In this article, the hydrophilic carbon dots (Lys-CA-CDs) were prepared using lysine (Lys) and citric acid (CA) as reactant under the assistance of a microwave. The dissimilar modulation effect of Lys-CA-CDs on the aggregation process of distinct structure protein was further investigated, where bovine serum albumin (BSA) and hen egg white lysozyme (HEWL) were chosen as model proteins. All results showed that Lys-CA-CDs displayed the contrary influence on the aggregation process of BSA and HEWL. Lys-CA-CDs could induce BSA to aggregate into more wormlike fibrils and inhibit the aggregation of HEWL into hair-like fibrils. The influence on the aggregation process of BSA may be assigned to the increased concentration of BSA around the Lys-CA-CDs caused by their interaction. However, inserting of Lys-CA-CDs into the inner structure of HEWL led to the change of protein secondary structure. The change of secondary structure further made it difficult for HEWL to aggregate into fibrils and Lys-CA-CDs showed the inhibition effect on HEWL aggregation.