Oral calcium supplementation versus placebo in mitigating citrate reactions during apheresis: an open-label randomized control trial.

BACKGROUND: Citrate-related hypocalcemia is the most common adverse event linked with peripheral blood progenitor cell apheresis. A previous retrospective study highlighted the prophylactic effectiveness of oral calcium drinks before apheresis, supplemented with intravenous calcium gluconate. Consequently, this study is a randomized controlled trial comparing oral calcium with placebo drinks STUDY DESIGN AND METHODS: Healthy donors were randomized to receive either oral calcium (Cohort A) or placebo (Cohort B) drinks. If symptoms emerged, all donors were given calcium drinks to counteract hypocalcemia. The primary endpoint centered on the incidence of Grade 1 or higher citrate-related symptoms. Analyses were performed using the crude model and doubly robust estimation. RESULTS: Forty-two healthy donors participated from January 2021 to July 2022. Case distribution (Cohort A: Cohort B) stood at 3:7 (Grade 1), 2:2 (Grade 2), and 1:0 (Grade 3); no Grade 4 cases were identified. There was no statistical significance in the incidence of Grade 1 or higher and Grade 3 citrate-related symptoms. DISCUSSION: The cumulative incidence of citrate-related side effects was less pronounced than in the previous research. This could stem from absence of blinding, and the decision to administer calcium drinks to the untreated group upon symptom detection. Although preemptive oral calcium intake before peripheral blood progenitor cell apheresis is not wholly effective, providing calcium-rich beverages to symptomatic donors may stave off symptom intensification.

Protocolized Regional Citrate Anticoagulation during Continuous Renal Replacement Therapy: A Single Center Experience.

Background: Regional citrate anticoagulation (RCA) has emerged as a treatment modality that reduces bleeding risk and filter clotting. With initial experience of using RCA with continuous renal replacement therapy (CRRT), we have formulated a working protocol based on published literature. Objective: The study aimed to evaluate the protocol for routine use of RCA during CRRT requiring anticoagulation and evaluation of filter life. Methodology: It is a single-center, open-label, prospective, non-randomized, non-interventional, single-arm, observational study conducted at a tertiary care hospital between September 2022 and July 2023. All adult patients with acute kidney injury (AKI) or hyperammonemia requiring CRRT and necessitating the use of anticoagulation were enrolled in the study. The study used Prisma Flex M100 AN 69 dialyzer on Prisma Flex (Baxter) CRRT machines during continuous venovenous hemodiafiltration (CVVHDF). The targeted CRRT dose in all the study patients was 25-30 mL/kg/hour. Based on the published literature, we have developed a working protocol (Appendix 1) for managing patients on CRRT using RCA. Results: A total of 159 patients were analyzed for the study. The median [interquartile range (IQR)] filter life using RCA was 30 (12-55) hours. Filter clotting was observed in 33.3% of patients. Citrate accumulation was present in 52.25% of patients, but no CRRT was discontinued as citrate accumulation resolved after following the corrective steps in the protocol. None of the patients had citrate toxicity. Chronic liver disease (CLD) (p ≤ 0.001) and those who were post-living donor liver transplant recipients (p = 0.004) had a statistically significant increase in citrate accumulation. Also, patients who had higher lactate at baseline (6 hours post-CRRT initiation), had a higher chance of citrate accumulation. Conclusion: Our RCA protocol provides a safe approach to regional anticoagulation during CRRT in critically ill patients. How to cite this article: Pachisia AV, Kumar GP, Harne R, Jagadeesh KN, Patel SJ, Pal D, et al. Protocolized Regional Citrate Anticoagulation during Continuous Renal Replacement Therapy: A Single Center Experience. Indian J Crit Care Med 2024;28(9):859-865.

Comparative effectiveness of sildenafil citrate and estradiol valerate as adjuvants during clomiphene citrate-assisted ovarian stimulation cycles in patients with unexplained infertility: a double-blind randomized controlled trial.

OBJECTIVE: This study aimed to assess the effect of sildenafil citrate and estradiol valerate as adjuvant therapy during ovarian stimulation cycles with clomiphene citrate in patients with unexplained infertility in Kisangani. METHOD: A double-blind, randomized controlled trial was conducted for two years at two specialized health facilities in Kisangani (University Clinics of Kisangani and "Clinique des Anges Kisangani"). The population included 148 patients, 74 of whom were on clomiphene citrate + sildenafil citrate (CCSC) regimens and 74 of whom were on clomiphene citrate + estradiol valerate (CCEV) regimens for three months. The primary indicator was the conception rate, with secondary outcomes encompassing endometrial thickness, appearance and vascularity, the number of mature follicles and ovulation rate. RESULTS: The two groups were comparable in terms of sociodemographic and clinical characteristics. The mean duration of attempting to conceive was 4.39 years versus 4.36 years (P = 0.839), while the mean AFC was 11.51 versus 11.46 (P = 0.831), in the CCSC group versus CCEV group respectively. Secondary infertility was the most frequent diagnosis in each of the two groups. The biochemical pregnancy rate was comparable between the two groups (P = 0.385), while the clinical pregnancy rate was significantly higher in the CCSC group versus CCEV group (P = 0.04). Both perifollicular flow and the ovulation rate were significantly higher in the CCSC group versus the CCEV group (P = 0.006 and P = 0.002 respectively). However, endometrial vascularity/thickness, and the number of Graafian follicles were not significantly different between the two groups. CONCLUSION: As an adjuvant, sildenafil increases the rate of clinical pregnancy more than does estradiol in patients with unexplained infertility undergoing ovarian stimulation with clomiphene citrate. STUDY REGISTRATION: PACTR 202,310,849,449,401 (Pan African Clinical Trials Registry).

Inhibition of HIV-1 infection with curcumin conjugated PEG-citrate dendrimer; a new nano formulation.

BACKGROUND: Nano-drug delivery systems have become a promising approach to overcoming problems such as low solubility and cellular uptake of drugs. Along with various delivery devices, dendrimers are widely used through their unique features. PEG-citrate dendrimers are biocompatible and nontoxic, with the ability to improve drug solubility. Curcumin, a naturally occurring polyphenol, has multiple beneficial properties, such as antiviral activities. However, its optimum potential has been significantly hampered due to its poor water solubility, which leads to reduced bioavailability. So, the present study attempted to address this issue and investigate its antiviral effects against HIV-1. METHOD: The G2 PEG-citrate dendrimer was synthesized. Then, curcumin was conjugated to it directly. FTIR, HNMR, DLS, and LCMS characterized the structure of products. The conjugate displayed an intense yellow color. In addition, increased aqueous solubility and cell permeability of curcumin were achieved based on flow cytometry results. So, it could be a suitable vehicle for improving the therapeutic applications of curcumin. Moreover, cell toxicity was assessed using XTT method. Ultimately, the SCR HIV system provided an opportunity to evaluate the level of HIV-1 inhibition by the curcumin-dendrimer conjugate using a p24 HIV ELISA kit. RESULTS: The results demonstrated a 50% up to 90% inhibition of HIV proliferation at 12 µm and 60 µm, respectively. Inhibition of HIV-1 at concentrations much lower than CC50 (300 µM) indicates a high potential of curcumin-dendrimer conjugate against this virus. CONCLUSION: Thereby, curcumin-dendrimer conjugate proves to be a promising tool to use in HIV-1 therapy.

Hypokalaemic myopathy associated with iatrogenic hypercorticism in a cat.

Case summary: A 12-year-old castrated male domestic shorthair cat was referred for investigation of lethargy, hindlimb weakness with plantigrade stance and ventroflexion of the neck. The cat was fed a balanced diet and had received methylprednisolone acetate at a dose of 20 mg intramuscularly every 6 months for 6 years. On blood work, severe hypokalaemia and marked elevation of muscle enzymes were noticeable. The findings were suggestive of hypokalaemic myopathy. Urine fractional excretion of potassium (FEk) was moderately high (9.04%), and serum aldosterone was below the reference interval. An adrenocorticotropic hormone (ACTH) stimulation test was compatible with adrenal suppression. Upon hospitalisation, the patient was given intravenous (IV) Ringer lactate solution supplemented with potassium chloride and oral potassium citrate. The serum potassium concentration normalised by the fifth day of hospitalisation; therefore, IV potassium supplementation was suspended. The cat was discharged with oral potassium and the dose was gradually reduced over time. After 4 months, the cat was clinically normal; the serum potassium concentration remained within the normal range and the adrenal glands showed some response to ACTH stimulation. Potassium supplementation was therefore discontinued. One month later, the serum potassium concentration was still within normal limits and at the time of writing (7 months after presentation), no clinical signs had reoccurred. Relevance and novel information: This report describes a case of hypokalaemic myopathy associated with iatrogenic hypercorticism in a cat. This condition was successfully treated with supplementation of potassium and a complete clinical remission was achieved within 4 months.

Influence of liquid extender, preservation temperature and time on the sheep sperm quality.

In this study, we evaluated sheep sperm quality after using Tris-citrate-fructose-based extender with and without egg yolk, a Tris-citrate without fructose and with egg yolk and the commercial extender Biladyl®, preserving diluted semen at 15 and 23°C for different times (4, 24, 48 and 72 h). The results showed that the diluents with fructose and egg yolk gave the best results of seminal quality. Moreover, the production of ROS was higher for the temperature of 23°C compared to the temperature of 15°C (control). In addition, VCL and the percentage of spermatozoa with intact acrosome decreased with temperatures of 23°C. Finally, a drastic decrease in sperm quality was observed after 24 hours of preservation for most of the parameters evaluated.

Characterization of Al3+-toxicity responses and molecular mechanisms underlying organic acid efflux in Vigna mungo (L.) Hepper.

Aluminium (Al3+) toxicity in acidic soils poses a significant challenge for crop cultivation and reduces crop productivity. The primary defense mechanism against Al3+ toxicity involves the activation of organic acid secretion. In this study, responses of 9 Vigna mungo cultivars to Al3+ toxicity were investigated, with a particular emphasis on the root system and crucial genes involved in Al3+ tolerance using molecular cloning and expression analysis. Sensitive blackgram-KM2 cultivars exposed to 100-µM Al3+ toxicity for 72 h exhibited a root-growth inhibition of approximately 66.17%. Significant loss of membrane integrity and structural deformative roots were found to be the primary symptoms of Al3+ toxicity in blackgram. MATE (Multidrug and Toxic Compound Extrusion) and ALS3 (Aluminium Sensitive 3) genes were successfully cloned from a sensitive blackgram cv KM2 with phylogenetic analysis revealing their evolutionary relationship to Vigna radiata and Glycine max. The MATE gene is mainly localized in the plasma membrane, and highly expressed under Al3+, thus suggesting its role in transports of citrate-Al3+ complexes, and detoxifying Al3+ within plant cells. In addition, ALS3 was also induced under Al3+ toxicity, which codes the UDP-glucose transporter and is required for the maintenance of ions homeostasis. In summary, this study highlights the understanding of Al3+ toxicity and underlying molecular mechanisms linked to the efflux of organic acid in blackgram, ultimately aiding the framework for the development of strategies to enhance the resilience of blackgram and other pulse crops in Al-rich soils.

Modulation of the Arabidopsis Starch Metabolic Network by the Cytosolic Acetyl-CoA Pathway in the Context of the Diurnal Illumination Cycle.

The starch metabolic network was investigated in relation to other metabolic processes by examining a mutant with altered single-gene expression of ATP citrate lyase (ACL), an enzyme responsible for generating cytosolic acetyl-CoA pool from citrate. Previous research has shown that transgenic antisense plants with reduced ACL activity accumulate abnormally enlarged starch granules. In this study, we explored the underlying molecular mechanisms linking cytosolic acetyl-CoA generation and starch metabolism under short-day photoperiods. We performed transcriptome and quantification of starch accumulation in the leaves of wild-type and antisense seedlings with reduced ACL activity. The antisense-ACLA mutant accumulated more starch than the wild type under short-day conditions. Zymogram analyses were conducted to compare the activities of starch-metabolizing enzymes with transcriptomic changes in the seedling. Differential expression between wild-type and antisense-ACLA plants was detected in genes implicated in starch and acetyl-CoA metabolism, and cell wall metabolism. These analyses revealed a strong correlation between the transcript levels of genes responsible for starch synthesis and degradation, reflecting coordinated regulation at the transcriptomic level. Furthermore, our data provide novel insights into the regulatory links between cytosolic acetyl-CoA metabolism and starch metabolic pathways.

Short-Term Metformin Therapy in Clomiphene Citrate Resistant PCOS Patients Improves Fertility Outcome by Regulating Follicular Fluid Redox Balance: A Case-Controlled Study.

OBJECTIVES: To determine the effect of short-term metformin administration on follicular fluid (FF) total oxidant status (TOS), total antioxidant status (TAS), oxidative stress index (OSI) and nuclear factor kappa B (NF-kB) in women with clomiphene citrate-resistant polycystic ovary syndrome (PCOS). METHODS: Fifty-eight patients aged 23-34 who were planned to have intracytoplasmic sperm injection due to clomiphene citrate-resistant PCOS were included in the study. Participants were divided into two groups according to whether they used metformin or not. While 30 of 58 PCOS patients were using short-term metformin in combination with controlled ovarian stimulation, 28 PCOS patients were not using metformin. Metformin was started in the mid-luteal period and continued until the day before oocyte retrieval at 850 mg twice daily. To determine FF-NF-kB, TAS, TOS and OSI values, a dominant follicle ≥17-18 mm in diameter was selected for aspiration. RESULTS: The number of mature follicles and fertilization rates of the metformin group were significantly higher than those not taking metformin. FF-TOS and OSI of the metformin group were significantly lower than those of the group not receiving metformin. Patients receiving metformin had higher FF-TAS than the group not receiving metformin. FF-NF-kB levels of the metformin group were significantly lower than the group not receiving metformin. Insulin resistance, FF-NF-kB and FF-TOS were negatively correlated with the number of mature oocytes. FF-TAS was positively correlated with the number of oocytes. CONCLUSIONS: Short-term metformin treatment in clomiphene-resistant PCOS improves the number of mature follicles and fertilization rates by regulating the intra-follicle redox balance.

Ketogenic ß-hydroxybutyrate regulates ß-hydroxybutyrylation of TCA cycle-associated enzymes and attenuates disease-associated pathologies in Alzheimer's mice.

Lysine ß-hydroxybutyrylation (Kbhb) is a post-translational modification that has recently been found to regulate protein functions. However, whether and how protein Kbhb modification participates in Alzheimer's disease (AD) remains unknown. Herein, we carried out 4D label-free ß-hydroxybutylation quantitative proteomics using brain samples of 8-month-old and 2-month-old APP/PS1 AD model mice and wild-type (WT) controls. We identified a series of tricarboxylic acid (TCA) cycle-associated enzymes including citrate synthase (CS) and succinate-CoA ligase subunit alpha (SUCLG1), whose Kbhb modifications were decreased in APP/PS1 mice at pathological stages. Sodium ß-hydroxybutyrate (Na-ß-OHB) treatment markedly increased Kbhb modifications of CS and SUCLG1 and their enzymatic activities, leading to elevated ATP production. We further found that Kbhb modifications at lysine 393 site in CS and at lysine 81 site in SUCLG1 were crucial for their enzymatic activities. Finally, we found that ß-OHB levels were decreased in the brain of APP/PS1 mice at pathological stages. While ketogenic diet not only significantly increased ß-OHB levels, Kbhb modifications and enzymatic activities of CS and SUCLG1, and ATP production, but also dramatically attenuated ß-amyloid plaque pathologies and microgliosis in APP/PS1 mice. Together, our findings indicate the importance of protein Kbhb modification for maintaining normal TCA cycle and ATP production and provide a novel molecular mechanism underlying the beneficial effects of ketogenic diet on energy metabolism and AD intervention.

Comparison and Determination of the Content of Mosapride Citrate by Different qNMR Methods.

As a salt-type compound, mosapride citrate's metabolism and side effects are correlated with its salt-forming ratio. Several techniques were developed in this work to compare various quantitative nuclear magnetic resonance (qNMR) methodologies and to quantitatively examine the content of raw materials. Among the qNMR techniques, methods for 1H NMR and 19F NMR were developed. Appropriate solvents were chosen, and temperature, number of scans, acquisition time, and relaxation delay parameter settings were optimized. Maleic acid was chosen as the internal standard in 1H NMR, and the respective characteristic signals of mosapride and citrate were selected as quantitative peaks. The internal standard in 19F NMR analysis was 4,4'-difluoro diphenylmethanone, and the distinctive signal peak at -116.15 ppm was utilized to quantify mosapride citrate. The precision, repeatability, linearity, stability, accuracy, and robustness of the qNMR methods were all validated according to the ICH guidelines. By contrasting the outcomes with those from high-performance liquid chromatography (HPLC), the accuracy of qNMR was assessed. As a result, we created a quicker and easier qNMR approach to measure the amount of mosapride citrate and evaluated several qNMR techniques to establish a foundation for choosing quantitative peaks for the qNMR method. Concurrently, it is anticipated that various selections of distinct quantitative objects will yield the mosapride citrate salt-forming ratio.

Leaching and vertical distribution of Fe and Zn citrate nanoparticles in Indian red soil.

This research aims to address common constraints in the effective utilization of plant nutrients in soil, such as fixation, mobility, leaching, and reactions with soil colloids. To mitigate these issues, Fe and Zn citrate nanoparticles were synthesized and applied as nanochelators in a reconstructed soil profile column. We evaluated the mobility, release, and leaching behaviors of these nanoparticles. Results revealed that, no leaching of Fe and Zn citrate nanoparticles occurred even after a 90-day incubation. The release profiles exhibited a peak at 60 and 90 days for Fe and Zn respectively. In the mobility studies, Fe and Zn availability was highest in the 0-15 cm soil depth. Fe citrate and Zn citrate nanoparticles demonstrated the highest availability at 264.7 mg/kg and 86.26 mg/kg of soil, respectively as compared with commercial samples. The superior performance of Fe and Zn citrate nanoparticles was observed in terms of reduced leaching and improved accessibility, indicating their potential as efficient and environmentally-friendly plant nutrient sources. The study concludes that Fe and Zn citrate nanoparticles are stable nutrient sources that can enhance plant use efficiency with minimal environmental impact.

O-GlcNAcylation of ATP-citrate lyase couples glucose supply to lipogenesis for rapid tumor cell proliferation.

Elevated lipid synthesis is one of the best-characterized metabolic alterations in cancer and crucial for membrane expansion. As a key rate-limiting enzyme in de novo fatty acid synthesis, ATP-citrate lyase (ACLY) is frequently up-regulated in tumors and regulated by posttranslational modifications (PTMs). Despite emerging evidence showing O-GlcNAcylation on ACLY, its biological function still remains unknown. Here, we observed a significant upregulation of ACLY O-GlcNAcylation in various types of human tumor cells and tissues and identified S979 as a major O-GlcNAcylation site. Importantly, S979 O-GlcNAcylation is required for substrate CoA binding and crucial for ACLY enzymatic activity. Moreover, it is sensitive to glucose fluctuation and decisive for fatty acid synthesis as well as tumor cell proliferation. In response to EGF stimulation, both S979 O-GlcNAcylation and previously characterized S455 phosphorylation played indispensable role in the regulation of ACLY activity and cell proliferation; however, they functioned independently from each other. In vivo, streptozocin treatment- and EGFR overexpression-induced growth of xenograft tumors was mitigated once S979 was mutated. Collectively, this work helps comprehend how cells interrogate the nutrient enrichment for proliferation and suggests that although mammalian cell proliferation is controlled by mitogen signaling, the ancient nutrition-sensing mechanism is conserved and still efficacious in the cells of multicellular organisms.

Ratios of calcium to citrate administration in blood transfusion for traumatic hemorrhage: A retrospective cohort study.

BACKGROUND: Massive transfusion with citrated blood products causes hypocalcemia, which is associated with mortality. Recognition of this problem has led to increased calcium administration; however, the optimal dosing is still unknown. STUDY DESIGN AND METHODS: This retrospective, single-center study included level 1 trauma patients in 2019 and 2020 who underwent an operation within 12 h of arrival and received a transfusion. Preoperative and intraoperative administrations were totaled to calculate the ratio of administered calcium to the number of blood transfusions for each patient. The citrate content of each blood component was estimated to calculate a second ratio, the ratio of administered calcium to administered citrate. Receiver Operating Characteristic (ROC) curves were performed on both ratios to determine the optimal cutoff values for predicting severe hypocalcemia (ionized calcium <0.9 mmol/L) and hypercalcemia (>1.35 mmol/L) at the end of the intraoperative period. RESULTS: A total of 506 trauma activations were included, receiving a mean of 17.4 citrated blood products and 16.3 mmol of calcium (equivalent to 2400 mg of calcium chloride). No ratio was statistically significant in differentiating severely hypocalcemic patients from the rest. A calcium to blood ratio of 0.903 mmol of administered calcium per citrated blood product differentiated hypercalcemic patients from the rest. DISCUSSION: Quantifying received calcium and citrated blood products was insufficient to predict severe hypocalcemia, suggesting other contributions to hypocalcemia. We demonstrated an upper-limit ratio for calcium administration in traumatic hemorrhage; however, further studies are required to determine what calcium dosing regimen results in the best outcomes.

Transcriptomics identify the triggering of citrate export as the key event caused by manganese deficiency in Aspergillus niger.

For over a century, the filamentous Ascomycete fungus Aspergillus niger has played a pivotal role in the industrial production of citric acid. A critical fermentation parameter that sustains high-yield citric acid accumulation is the suboptimal concentration of manganese(II) ions in the culture broth at the early stages of the process. However, the requirement for this deficiency has not been investigated on a functional genomics level. In this study, we compared the transcriptome of the citric acid hyper-producer A. niger NRRL2270 strain grown under citric acid-producing conditions in 6-L scale bioreactors at Mn2+ ion-deficient (5 ppb) and Mn2+ ion-sufficient (100 ppb) conditions at three early time points of cultivation. Of the 11,846 genes in the genome, 963 genes (8.1% of the total) were identified as significantly differentially expressed under these conditions. Disproportionately high number of differentially regulated genes encode predicted extracellular and membrane proteins. The most abundant gene group that was upregulated in Mn2+ ion deficiency condition encodes enzymes acting on polysaccharides. In contrast, six clusters of genes encoding secondary metabolites showed downregulation under manganese deficiency. Mn2+ deficiency also triggers upregulation of the cexA gene, which encodes the citrate exporter. We provide functional evidence that the upregulation of cexA is caused by the intracellular accumulation of citrate or acetyl-CoA and is a major factor in triggering citrate overflow. IMPORTANCE: Citric acid is produced on industrial scale by batch fermentation of the filamentous fungus Aspergillus niger. High-yield citric acid production requires a low (<5 ppb) manganese(II) ion concentration in the culture broth. However, the requirement for this deficiency has not been investigated on a functional genomics level. Here, we compared the transcriptome of a citric acid hyper-producer A. niger strain grown under citric acid-producing conditions in 6-L scale bioreactors at Mn2+ ion-deficient (5 ppb) and Mn2+ ion-sufficient (100 ppb) conditions at three early time points of cultivation. We observed that Mn2+ deficiency triggers an upregulation of the citrate exporter gene cexA and provides functional evidence that this event is responsible for citrate overflow. In addition to the industrial relevance, this is the first study that examined the role of Mn2+ ion deficiency in a heterotrophic eukaryotic cell on a genome-wide scale.

Review on the Polymeric and Chelate Gel Precursor for Li-Ion Battery Cathode Material Synthesis.

The rapid design of advanced materials depends on synthesis parameters and design. A wide range of materials can be synthesized using precursor reactions based on chelated gel and organic polymeric gel pathways. The desire to develop high-performance lithium-ion rechargeable batteries has motivated decades of research on the synthesis of battery active material particles with precise control of composition, phase-purity, and morphology. Among the most common methods reported in the literature to prepare precursors for lithium-ion battery active materials, sol-gel is characterized by simplicity, homogeneous mixing, and tuning of the particle shape. The chelate gel and organic polymeric gel precursor-based sol-gel method is efficient to promote desirable reaction conditions. Both precursor routes are commonly used to synthesize lithium-ion battery cathode active materials from raw materials such as inorganic salts in aqueous solutions or organic solvents. The purpose of this review is to discuss synthesis procedure and summarize the progress that has been made in producing crystalline particles of tunable and complex morphologies by sol-gel synthesis that can be used as active materials for lithium-ion batteries.

Morusin, a novel inhibitor of ACLY, induces mitochondrial apoptosis in hepatocellular carcinoma cells through ROS-mediated mitophagy.

OBJECTIVE: Morusin (Mor), a prenylated flavonoid isolated from the root bark of Morus alba L., exhibits potent anti-tumour effects; however, the molecular target of Mor is still not entirely clear. This study aimed to elucidate the mechanism of Mor against hepatocellular carcinoma (HCC) and identify potential molecular targets. METHODS: Mitochondrial function was assessed by measuring the mitochondrial membrane potential, mitochondrial ultrastructure, oxygen consumption, and ATP levels. Mor-induced mitophagy was confirmed using western blotting, immunofluorescence, and fluorescent probes. Transcriptomics, flow cytometry, western blotting, qRT-PCR and biochemical assays were used to reveal the molecular mechanisms and targets of Mor against HCC. We further validated the interaction between Mor and the target proteins using molecular docking and biolayer interferometry (BLI). The inhibitory effect of Mor in vivo was evaluated using a Hep3B murine xenograft model. RESULTS: Mor significantly reduced the ATP citrate lyase (ACLY) expression and inhibited ACLY activity in HCC cells. BLI analysis demonstrated a direct interaction between Mor and the ACLY active domain. Mor-induced ACLY inhibition led to ROS accumulation in HCC cells, which caused mitochondrial damage, triggered PINK1/Parkin-mediated mitophagy, and ultimately induced mitochondrial apoptosis. We further verified that ROS is crucial in the apoptotic action of Mor through experiments regarding an ROS scavenger. Mor also significantly inhibited tumour xenograft growth in vivo. In addition, analysis of human liver cancer clinical samples revealed elevated ACLY levels positively correlated with histologic grade. CONCLUSION: Collectively, our findings highlight Mor as a potent bioactive inhibitor of ACLY and a promising candidate for HCC therapy.

CaCl2-Citrate Regional Anticoagulation with Continuous Veno-Venous Haemodialysis Leads to Unwanted Chloride Loading Compared to Continuous Veno-Venous Hemofiltration with Systemic Anticoagulation.

INTRODUCTION: Chloride transfers during continuous renal replacement therapy (CRRT) have not been adequately described and may differ based on CRRT technique. We aimed to measure chloride mass transfer (JS,Cl) during CRRT and identify associated determinants. METHODS: We performed a two-centre, prospective, observational study in France and Australia in ICU patients with CRRT initiated for <24 h. Patients received continuous veno-venous hemofiltration (CVVH) or continuous veno-venous haemodialysis (CVVHD, with citrate-CaCl2 regional anticoagulation). Over a 24 h period, plasma and effluent chloride concentrations were measured every 4 h to compute chloride mass transfer (JS,Cl, in mmol.min-1) using a modality-specific model, with negative value indicating chloride transfer towards the patient. Secondary outcomes were the identification of CRRT settings associated with JS,Cl (using multivariate mixed effects regression). Results are presented with median (interquartile range). RESULTS: Between February 2021 and August 2022, we enrolled 37 patients (64 [56-71] years, 67% male), for a total of 20 CVVHD and 20 CVVH sessions. Over 24 h, plasma chloride concentrations were significantly higher, and JS,Cl significantly lower during CVVHD, compared to CVVH (-0.10 [-0.33 to 0.15] vs. 0.01 [-0.10 to 0.13] mmol.min-1, p < 0.05). With both modalities, net ultrafiltration (QUFNET) and plasma chloride concentrations were the principal determinants of JS,Cl, with higher QUFNET being associated with an increase in JS,Cl during CVVHD. Also, CVVHD sessions demonstrated a concentration gradient between the plasma and the effluent chamber of -6 [-9 to -4] mmol.L-1. Finally, CaCl2 reinjection during CVVHD accounted for 35% [32-60%] of total JS,Cl in sessions with a negative JS,Cl. CONCLUSION: Compared to CVVH, CVVHD with regional citrate anticoagulation was associated with greater chloride mass transfer to the patient and higher plasma chloride concentrations. This was due to high dialysate chloride concentrations and CaCl2 reinjection. This effect could only be controlled by high net ultrafiltration flow rates.

A tomato a day keeps the beetle away - the impact of Solanaceae glycoalkaloids on energy management in the mealworm Tenebrio molitor.

Solanine (SOL), chaconine (CHA), and tomatine (TOM) are plant secondary metabolites produced mainly by the species of Solanaceae family, such as tomato Solanum lycopersicum L. These glycoalkaloids (GAs) have a wide range of biological activity, also in insects. However, their mechanisms of action are not precisely understood. The purpose of the study was to investigate how pure GAs and tomato leaf extract (EXT) affect glycolysis, Krebs cycle and ß-oxidation of fatty acid pathways in Tenebrio molitor L. beetle. For this purpose, the larvae were injected with SOL, CHA, TOM, and EXT at two concentrations (10-8 and 10-5 M). For experiments, fat body, gut, and heamolymph samples were collected 2 and 24 h after injection. Then, the changes in the expression level of phosphofructokinase, citrate synthase, and ß-hydroxyacyl-CoA dehydrogenase were measured using the RT-qPCR technique. The catalytic activity of these enzymes and the carbohydrate level in insects after GA treatment were determined by spectrophotometric method. Furthermore, the analysis of the amount of amino acids in tissues was performed with a GC-MS technique. The results obtained show that the GAs changed the activity and expression of the genes encoding key enzymes of crucial metabolic pathways. The effect depends on the type of GA compound, the tissue tested, and the incubation time after treatment. Furthermore, TOM and EXT affected trehalose concentration in the insect hemolymph and led to accumulation of amino acids in the fat body. The observed changes may indicate a protein degradation and/or enhanced catabolism reactions for the production of ATP used in detoxification processes. These results suggest that GAs alter energy metabolism in the mealworm T. molitor. The study contributes to our understanding of the mechanisms of action of secondary metabolites of plants in insects. This knowledge may allow the design of new natural biopesticides against insect pests because proper energy metabolism is necessary for the survival of the organism.

Biochemical and biophysical characterization of Leishmania donovani citrate synthase.

Citrate synthase is a crucial enzyme in the TCA cycle and represents a potential therapeutic target. However, knowledge about this enzyme in Leishmania parasites remains limited. In this study, we have successfully cloned, expressed, and purified citrate synthase from Leishmania donovani (LdCS) using a bacterial system, and characterized it through various biophysical and biochemical methods. Circular dichroism analysis at physiological pH indicates that LdCS is properly folded. Further investigation into its tertiary structure using a quencher reveals that most tryptophan residues are located within the protein's hydrophobic core. Biochemical assays show that the recombinant enzyme is catalytically active, with optimal activity at pH 7.0. Kinetic studies provided parameters such as Km and Vmax. Enzyme inhibition assays revealed that LdCS activity is competitively inhibited by FDA-approved compounds-Abemaciclib, Bazedoxifene, Vorapaxar, and Imatinib-with Ki values ranging from 2 to 3 µM, demonstrating significant binding affinity. This research paves the way for exploring LdCS as a potential drug target for treating leishmaniasis.